ABCMdb

ABCG2 p.Gln141Lys

ClinVar:	c.421C>A , p.Gln141Lys ? , association
Predicted by SNAP2:	A: D (63%), C: D (75%), D: D (80%), E: D (59%), F: D (75%), G: D (80%), H: D (59%), I: D (66%), K: D (80%), L: D (66%), M: D (63%), N: D (66%), P: D (85%), R: D (66%), S: D (63%), T: D (63%), V: D (63%), W: D (71%), Y: D (66%),
Predicted by PROVEAN:	A: D, C: D, D: N, E: N, F: D, G: D, H: N, I: D, K: N, L: D, M: D, N: N, P: D, R: N, S: N, T: D, V: D, W: D, Y: D,

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[hide] C421A polymorphism in the human breast cancer resi... Mol Cancer Ther. 2002 Jun;1(8):611-6. Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, Tsuruo T, Miki Y, Sugimoto Y
C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance.
Mol Cancer Ther. 2002 Jun;1(8):611-6., [PMID:12479221]

Abstract [show]
Breast cancer resistance protein (BCRP) confers multidrug resistance to cancer cells against agents such as SN-38 (an active metabolite of irinotecan), mitoxantrone, and topotecan. Among 59 human tumor cell lines tested, 6 cell lines, A549, NCI-H460, KM-12, HT-29, OVCAR-5, and RPMI8226, showed high BCRP expression. BCRP cDNA was isolated from 11 cancer cell lines and three variant cDNAs [G34A substituting Met for Val-12 (V12M), C421A substituting Lys for Gln-141 (Q141K), and 944-949 deletion lacking Ala-315 and Thr-316 (delta315-6)] were identified. G34A and C421A variants were polymorphisms, and 944-949 deletion was a splicing variant. C421A BCRP-transfected PA317 cells showed markedly decreased protein expression and low-level drug resistance compared with wild-type BCRP-transfected cells when transfectants expressed similar levels of BCRP mRNA. G34A or 944-949-deleted BCRP-transfected PA317 cells showed similar or somewhat lower protein expression and drug resistance compared with wild-type BCRP-transfected cells. Of 124 healthy Japanese volunteers, 67 were wild-type, 48 were heterozygous, and 9 were homozygous for the C421A allele. These results suggest that some people possess the C421A polymorphic BCRP gene and express low amounts of Q141K BCRP. In addition to that, C376T polymorphism in exon 4 substituting stop codon for Gln-126 was found in 3 of the 124 general Japanese population. This C376T polymorphism may also have high impact because active BCRP protein will not be expressed from the C376T allele. Therefore, people with C376T and/or C421A polymorphisms may express low amounts of BCRP, and this low BCRP expression might result in hypersensitivity of normal cells to such anticancer drugs as irinotecan and mitoxantrone.

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3 Access the articles at: E-mail alerts related to this article or journal.Sign up to receive free email-alerts Subscriptions Reprints and .pubs@aacr.orgDepartment at To order reprints of this article or to subscribe to the journal, contact the AACR Publications Permissions .permissions@aacr.orgDepartment at To request permission to re-use all or part of this article, contact the AACR Publications C421A Polymorphism in the Human Breast Cancer Resistance Protein Gene Is Associated with Low Expression of Q141K Protein and Low-Level Drug Resistance1 Yasuo Imai, Minoru Nakane, Kumie Kage, Satomi Tsukahara, Etsuko Ishikawa, Takashi Tsuruo, Yoshio Miki, and Yoshikazu Sugimoto2 Division of Molecular Biotherapy [Y. I., M. N., K. K., S. T., E. I., Y. S.] and Division of Experimental Chemotherapy [T. T.], Cancer Chemotherapy Center, and Department of Molecular Diagnosis [Y. M.], Cancer Institute, Japanese Foundation for Cancer Research, Tokyo 170-8455, and Institute of Molecular and Cellular Biosciences, University of Tokyo, Tokyo 113-0032 [T. T.], Japan Abstract Breast cancer resistance protein (BCRP) confers multidrug resistance to cancer cells against agents such as SN-38 (an active metabolite of irinotecan), mitoxantrone, and topotecan. Login to comment
5 BCRP cDNA was isolated from 11 cancer cell lines and three variant cDNAs [G34A substituting Met for Val-12 (V12M), C421A substituting Lys for Gln141 (Q141K), and 944-949 deletion lacking Ala-315 and Thr-316 (⌬315-6)] were identified. Login to comment
10 These results suggest that some people possess the C421A polymorphic BCRP gene and express low amounts of Q141K BCRP. Login to comment
60 PA317 cells transfected with wild-type, G34A, C421A, and 944-949-deleted BCRP cDNAs were designated PA/WT, PA/V12M, PA/Q141K, and PA/⌬315-6 cells, respectively. Login to comment
92 Western blotting of mutant BCRP-transfected PA317 cells demonstrated markedly low expression of Q141K BCRP in PA/Q141K cells compared with other BCRP-transfected cells. Login to comment
96 In contrast, Northern blotting demonstrated similar levels of BCRP mRNA in PA/WT, PA/V12M, PA/ Q141K, and PA/⌬315-6 cells (Fig. 3B). Login to comment
97 These results suggest that Q141K BCRP is unstable when expressed in mammalian cells. Login to comment
104 Table 1 BCRP cDNA variants identified in this study Variant Amino acid change Cell line G34A Val-12 to Met MCF-7a C421A Gln-141 to Lys MDA-MB-231a A549a HCT-116a Deletion of 944-949 Deletion of Ala-315 and Thr-316 MCF-7 A549 HT-29 SK-OV-3 a Heterozygous allele. Login to comment
118 In contrast, PA/Q141K cells showed a 12-fold greater resistance to SN-38 and a 4-fold greater resistance to mitoxantrone (Table 2; Fig. 4A). Login to comment
119 This means that PA/Q141K cells are 2-3 times more sensitive to these drugs compared with PA/WT cells. Login to comment
120 These results support the low expression of BCRP in PA/Q141K cells. Login to comment
126 Increases of mean fluorescence channel number in PA/ WT, PA/V12M, and PA/⌬315-6 cells were 1.5-, 1.6-, and 1.5-fold in the presence of topotecan, respectively. Login to comment
127 There was a stronger peak shift to the right in PA/Q141K cells than PA/WT cells, showing that topotecan uptake in PA/Q141K cells was higher than that in PA/WT cells, and the increase of mean fluorescence channel number in PA/Q141K cells was 2.1-fold in the presence of topotecan (Fig. 5). Login to comment
128 This suggests that the topotecan efflux activity of exogenous BCRP is low in PA/Q141K cells. Login to comment
137 PA317 cells transfected with wild-type, G34A, C421A, and 944-949-deleted BCRP cDNAs were termed PA/WT, PA/V12M, PA/Q141K, and PA/⌬315-6, respectively. Login to comment
145 Table 2 IC50 a (ng/ml) of BCRP-transfected PA317 cells PA317 PA/WT PA/V12M PA/Q141K PA/⌬315-6 SN-38 2.5 98 98 30 55 Mitoxantrone 0.060 0.58 0.63 0.25 0.42 Topotecan 17 Ͼ200 Ͼ200 100 190 a IC50s (drug dose causing 50% inhibition of cell growth) were determined from cell growth curves in each experiment. Login to comment
148 PA317 cells transfected with wild-type, G34A, C421A, and 944-949- deleted BCRP cDNAs were termed PA/WT, PA/V12M, PA/Q141K, and PA/⌬315-6, respectively. Login to comment
149 A, sensitivity to SN-38 (A-1), mitoxantrone (A-2), and topotecan (A-3) of PA317, PA/WT, PA/V12M, and PA/Q141K cells. Login to comment
151 F, PA317; E, PA/WT; ‚, PA/V12M; Ⅺ, PA/Q141K; ᭛, PA/⌬315-6. Login to comment
163 PA/Q141K cells were significantly more sensitive to anticancer agents than the other BCRP transfectants. Login to comment
164 Intracellular topotecan accumulation of PA/Q141K was higher than that in the other BCRP transfectants. Login to comment
165 By Western blotting, BCRP expression in PA/Q141K cells was markedly lower than that in the other BCRP transfectants. Login to comment
166 Another transfection experiment of mutant BCRP cDNAs in KB-3-1 human epidermoid carcinoma cells also revealed markedly lower expression of Q141K BCRP compared with wild-type and V12M BCRP (data not shown). Login to comment
168 In the transfection experiment, the expression of C421A BCRP mRNA was identical to those of mRNAs from wild-type, G34A, and 944-949-deleted BCRP by Northern blotting. Login to comment
169 Therefore, the increased sensitivity was considered to be a result of instability of Q141K BCRP. Login to comment
186 PA317 cells transfected with wild-type, G34A, C421A, and 944-949- deleted BCRP cDNAs were termed PA/WT, PA/V12M, PA/Q141K, and PA/⌬315-6, respectively. Login to comment
188 In parental PA317 cells, a fluorescence peak shift to the right after the incubation with topotecan indicates cellular uptake of topotecan, whereas only marginal shifts occurred in PA/WT, PA/V12M, and PA/⌬315-6 cells. Login to comment
189 There was a stronger fluorescence peak shift to the right in PA/Q141K cells than the other transfectants (arrow). Login to comment
2 Access the articles at: E-mail alerts related to this article or journal.Sign up to receive free email-alerts Subscriptions Reprints and .pubs@aacr.orgDepartment at To order reprints of this article or to subscribe to the journal, contact the AACR Publications Permissions .permissions@aacr.orgDepartment at To request permission to re-use all or part of this article, contact the AACR Publications C421A Polymorphism in the Human Breast Cancer Resistance Protein Gene Is Associated with Low Expression of Q141K Protein and Low-Level Drug Resistance1 Yasuo Imai, Minoru Nakane, Kumie Kage, Satomi Tsukahara, Etsuko Ishikawa, Takashi Tsuruo, Yoshio Miki, and Yoshikazu Sugimoto2 Division of Molecular Biotherapy [Y. I., M. N., K. K., S. T., E. I., Y. S.] and Division of Experimental Chemotherapy [T. T.], Cancer Chemotherapy Center, and Department of Molecular Diagnosis [Y. M.], Cancer Institute, Japanese Foundation for Cancer Research, Tokyo 170-8455, and Institute of Molecular and Cellular Biosciences, University of Tokyo, Tokyo 113-0032 [T. T.], Japan Abstract Breast cancer resistance protein (BCRP) confers multidrug resistance to cancer cells against agents such as SN-38 (an active metabolite of irinotecan), mitoxantrone, and topotecan. Login to comment
4 BCRP cDNA was isolated from 11 cancer cell lines and three variant cDNAs [G34A substituting Met for Val-12 (V12M), C421A substituting Lys for Gln-141 (Q141K), and 944-949 deletion lacking Ala-315 and Thr-316 (⌬315-6)] were identified. Login to comment
9 These results suggest that some people possess the C421A polymorphic BCRP gene and express low amounts of Q141K BCRP. Login to comment
59 PA317 cells transfected with wild-type, G34A, C421A, and 944-949-deleted BCRP cDNAs were designated PA/WT, PA/V12M, PA/Q141K, and PA/⌬315-6 cells, respectively. Login to comment
91 Western blotting of mutant BCRP-transfected PA317 cells demonstrated markedly low expression of Q141K BCRP in PA/Q141K cells compared with other BCRP-transfected cells. Login to comment
95 In contrast, Northern blotting demonstrated similar levels of BCRP mRNA in PA/WT, PA/V12M, PA/ Q141K, and PA/⌬315-6 cells (Fig. 3B). Login to comment
103 Table 1 BCRP cDNA variants identified in this study Variant Amino acid change Cell line G34A Val-12 to Met MCF-7a C421A Gln-141 to Lys MDA-MB-231a A549a HCT-116a Deletion of 944-949 Deletion of Ala-315 and Thr-316 MCF-7 A549 HT-29 SK-OV-3 a Heterozygous allele. Login to comment
117 In contrast, PA/Q141K cells showed a 12-fold greater resistance to SN-38 and a 4-fold greater resistance to mitoxantrone (Table 2; Fig. 4A). Login to comment
136 PA317 cells transfected with wild-type, G34A, C421A, and 944-949-deleted BCRP cDNAs were termed PA/WT, PA/V12M, PA/Q141K, and PA/⌬315-6, respectively. Login to comment
144 Table 2 IC50 a (ng/ml) of BCRP-transfected PA317 cells PA317 PA/WT PA/V12M PA/Q141K PA/⌬315-6 SN-38 2.5 98 98 30 55 Mitoxantrone 0.060 0.58 0.63 0.25 0.42 Topotecan 17 Ͼ200 Ͼ200 100 190 a IC50s (drug dose causing 50% inhibition of cell growth) were determined from cell growth curves in each experiment. Login to comment
147 PA317 cells transfected with wild-type, G34A, C421A, and 944-949- deleted BCRP cDNAs were termed PA/WT, PA/V12M, PA/Q141K, and PA/⌬315-6, respectively. Login to comment
150 F, PA317; E, PA/WT; ‚, PA/V12M; Ⅺ, PA/Q141K; ᭛, PA/⌬315-6. Login to comment
162 PA/Q141K cells were significantly more sensitive to anticancer agents than the other BCRP transfectants. Login to comment
185 PA317 cells transfected with wild-type, G34A, C421A, and 944-949- deleted BCRP cDNAs were termed PA/WT, PA/V12M, PA/Q141K, and PA/⌬315-6, respectively. Login to comment

[hide] Natural allelic variants of breast cancer resistan... Pharmacogenetics. 2003 Jan;13(1):19-28. Zamber CP, Lamba JK, Yasuda K, Farnum J, Thummel K, Schuetz JD, Schuetz EG
Natural allelic variants of breast cancer resistance protein (BCRP) and their relationship to BCRP expression in human intestine.
Pharmacogenetics. 2003 Jan;13(1):19-28., [PMID:12544509]

Abstract [show]
The aim of this study was to identify the extent of genetic variability in breast cancer resistance protein (BCRP) in humans. We first analysed the sequence of BCRP cDNA from human livers and from human intestines phenotyped for expression of intestinal BCRP. We then determined the frequency of all known coding single nucleotide polymorphisms (cSNPs) using DNA from individuals representing 11 different ethnic populations. Nine SNPs including four non-synonymous and three synonymous cSNPs and two intronic SNPs were identified. Of the missense mutations, exon 2 SNP (G34A) resulted in a V12M change; exon 5 SNP (C421A) resulted in a Q141K substitution; exon 6 SNP (A616C) resulted in an I206L amino acid substitution; and exon 15 SNP (A1768T) resulted in a N590Y change in the BCRP protein. The two most frequent polymorphisms identified in the human population studied were the G34A and C421A transitions. There was marked variation in BCRP genotypes and allele frequencies in the different populations. BCRP mRNA was phenotyped in human small bowel intestinal samples by real-time polymerase chain reaction and BCRP protein was analysed on immunoblots of tissue from the same individuals. There was a 78-fold variation in expression of BCRP mRNA and significant variation in BCRP protein expression in human intestine. Expression of intestinal BCRP mRNA and protein was not different between persons expressing the common Gln141 allele compared to the Lys141 allele. Thus, common natural allelic variants of BCRP have been identified, and did not influence interindividual variation in expression of BCRP mRNA in human intestine, but remain to be tested for their effect on BCRP function.

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4 Of the missense mutations, exon 2 SNP (G34A) resulted in a V12M change; exon 5 SNP (C421A) resulted in a Q141K substitution; exon 6 SNP (A616C) resulted in an I206L amino acid substitution; and exon 15 SNP (A1768T) resulted in a N590Y change in the BCRP protein. Login to comment
95 Exon 5 One SNP, a C421A transversion, results in a Gln141 Lys change. Login to comment
119 Table 1 Frequencies of BCRP alleles in different ethnic groups Position in gene AC084732Ã Position in mRNA XM_032424Ã Sequence Region Caucasians African-Americans Japanese (n ¼ 20) Chinese (n ¼ 20) SE Asians (not Chinese or Japanese) (n ¼ 20) Pacific Islanders (n ¼ 14) À18398 À29 gctct(A/G)ttaag Exon 1 0.02 (1.5%)a 0 (0%)e ND ND ND ND 34 34 tccca(G/A)tgtca Exon 2 (V12M) 0.02 (4.7%)b 0.04 (8.3%)f 0.15 (30%) 0.20 (40%) 0.45 (70%) 0.64 (85.7%) 114 114 ttaag(T/C)tttca Exon 2 0.01 (1.2%)b 0 (0%)f 0 (0%) 0 (0%) 0 (0%) 0 (0%) 239 tttta (A/G)tttac Intron 2 0.03 (5.9%)c 0.05 (9.5%)g 0.15 (30%) 0.20 (40%) 0.45 (70%) 0.64 (85.7%) 8184 369 ggtta(C/T)gtggt Exon 4 0 (0%)a 0.07 (13.3%)e ND ND ND ND 8825 421 actta(C/A)agttc Exon 5 (Q141K) 0.14 (25.9%)d 0 (8%)f 0.35 (50%) 0.35 (60%) 0.15 (20%) 0.14 (28.6%) 18186 attat(A/G)atatt Intron 5 0 (0%)c 0 (8%)g 0 (0%) 0.05 (10%) 0 (0%) 0 (0%) 18286 616 cttcc(A/C)tcttg Exon 6 (I206L) 0 (0%)a 0 (8%)e 0 (0%) 0 (0%) 0 (0%) 0 (0%) 45073 1768 gacaa(A/T)acttc Exon 15 (N590Y) 0.01 (1.5%)a 0 (8%)e ND ND ND ND Position in gene AC084732Ã Position in mRNA XM_032424Ã Sequence Region Mexican-Indians (n ¼ 10) Mexicans (n ¼ 20) Hispanic Livers (n ¼ 10) Middle Eastern (n ¼ 40) Ashkenazi Jewish (n ¼ 20) Africans North of Sahara (n ¼ 14) À18398 À29 gctct(A/G)ttaag Exon 1 ND ND ND ND ND ND 34 34 tccca(G/A)tgtca Exon 2 (V12M) 0.90 (100%) 0.10 (20%) 0.40 (60%) 0.05 (10%) 0.10 (20%) 0.14 (14.3%) 114 114 ttaag(T/C)tttca Exon 2 0 (0%) 0 (0%) 0 (0%) 0 (0%) 0 (0%) 0 (0%) 239 tttta(A/G)tttac Intron 2 0.90 (100%) 0.10 (20%) 0.40 (60%) 0.05 (10%) 0.10 (20%) 0.14 (14.3%) 8184 369 ggtta(C/T)gtggt Exon 4 ND ND ND ND ND ND 8825 421 actta(C/A)agttc Exon 5 (Q141K) 0.10 (20%) 0.05 (10%) 0.10 (20%) 0.13 (25%) 0.05 (10%) 0 (0%) 18186 attat(A/G)atatt Intron 5 0 (0%) 0.10 (20%) 0 (0%) 0 (0%) 0.05 (10%) 0.07 (14.3%) 18286 616 cttcc(A/C)tcttg Exon 6 (I206L) 0 (0%) 0 (0%) 0.10 (20%) 0 (0%) 0 (0%) 0 (0%) 45073 1768 gacaa(A/T)acttc Exon 15 (N590Y) ND ND ND ND ND ND Data reported as: allele frequency (% individuals with at least one variant allele). Login to comment
125 Unauthorized reproduction of this article is prohibited. G34A V12M Exon 2 C71T1 A24V Exon 2 623C1 F208S Exon 6 A616C I206L Exon 6 C496G1 Q166E Exon 5 C421A Q141K Exon 5 A1444G2 R482G Exon 12 G1445C3 R482T Exon 12 A1768T N590Y Exon 15 Walker A motif: amino acids 80-89 Walker B motif: amino acids 206-210 SNPs found in human samples in this study Reported in ABCP1 Drug selected variants, MXR2 and BCRP3 MXR BCRP Fig. 1 BCRP protein topology and the positions of the identified SNPs resulting in missense mutations. Login to comment
127 BCRP mRNA was compared to BCRP SNP C421A; Gln141 Lys. Login to comment
135 However, 16 individuals were heterozygous for the 421C.A transversion, leading to the Gln141 Lys change. Login to comment
146 Unauthorized reproduction of this article is prohibited. 400.0 300.0 200.0 100.0 0.0 BCRPmRNA(%ofHI11as100%) (a) CC CA Q141K 400.0 300.0 200.0 100.0 0.0BCRPmRNA(%ofHI11as100%) (b) Female Male CC - Gln141 CA - Lys141 Fig. 3 Comparison of BCRP genotype and BCRP phenotype in human intestines. Login to comment
147 The BCRP SNP C421A; Gln141 Lys was compared to the (a) amount of BCRP mRNA transcript in all human intestines. Login to comment
173 The Val12 Met and the Gln141 Lys were the most frequent allelic variants. Login to comment
176 The Gln141 Lys was the most prevalent allele in both the Japanese and Chinese populations with an allelic frequency of 35%. Login to comment
189 The most commonly observed SNP detected in these samples was the exon 5 C421A (Gln141 to Lys) with an allelic frequency of 19%. Login to comment

[hide] Single-nucleotide polymorphism (SNP) analysis in t... Cancer Biol Ther. 2002 Nov-Dec;1(6):696-702. Honjo Y, Morisaki K, Huff LM, Robey RW, Hung J, Dean M, Bates SE
Single-nucleotide polymorphism (SNP) analysis in the ABC half-transporter ABCG2 (MXR/BCRP/ABCP1).
Cancer Biol Ther. 2002 Nov-Dec;1(6):696-702., [PMID:12642696]

Abstract [show]
Variations in the amino acid sequence of ABC transporters have been shown to impact substrate specificity. We identified two acquired mutations in ABCG2, the ABC half-transporter overexpressed in mitoxantrone-resistant cell lines. These mutations confer differences in substrate specificity and suggest that naturally occurring variants could also affect substrate specificity. To search for the existence of single nucleotide polymorphisms (SNPs) in ABCG2, we sequenced 90 ethnically diverse DNAs from the Single Nucleotide Polymorphism Discovery Resource representing the spectrum of human genotypes. We identified 3 noncoding SNPs in the untranslated regions, 3 nonsynonymous and 2 synonymous SNPs in the coding region and 7 SNPs in the intron sequences adjacent to the sixteen ABCG2 exons. Nonsynonymous SNPs at nucleotide 238 (V12M; exon 2) and nucleotide 625 (Q141K; exon 5) showed a greater frequency of heterozygosity (22.2% and 10%) than the SNP at 2062 (D620N; exon 16). Heterozygous changes at nucleotide 238 are in linkage disequilibrium with an SNP observed 36 bases downstream from the end of exon 2. No polymorphism at amino acid 482 was identified to correspond to the R to G or R to T mutations previously found in two drug resistant cell lines. Among 23 drug resistant sublines for which sequence at position 482 was determined, no additional mutations were found. Heterozygosity at amino acid 12 allowed us to identify overexpression of a single allele in a subset of drug resistant cell lines, a feature that could be exploited clinically in evaluating the significance of ABCG2 expression in malignancy. We conclude that ABCG2 is well conserved and that described amino acid polymorphisms seem unlikely to alter transporter stability or function.

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6 Nonsynonymous SNPs at nucleotide 238 (V12M; exon 2) and nucleotide 625 (Q141K; exon 5) showed a greater frequency of heterozygosity (22.2% and 10%) than the SNP at 2062 (D620N; exon 16). Login to comment
104 Amplification in the MCF-7 SINGLE-NUCLEOTIDE POLYMORPHISM (SNP) ANALYSIS IN THE ABC HALF-TRANSPORTER ABCG2 (MXR/BCRP/ABCP1) www.landesbioscience.com Cancer Biology & Therapy 699 Table 2 PRIMERS USED IN SEQUENCING 90 DNA SAMPLES Forward Primer (5`-> 3`) Reverse Primer (5`-> 3`) Exon 1 TGCCCACTCAAAAGGTTC CCAACCCACACTTAACACAC Exon 2 TGTCACCTAGTGTTTGCAATC GCCAGTTTCTTGGAAATAGCC Exon 3 AATCCTGCTTTGGTCTCC TCTCCCATTCTTTTTCCTC Exon 4 AGCATGTGTTGGAGGGAAAA ATCAGCCAAAGCACTTACCC Exon 5 GCAGGCTTTGCAGACATCTA TGCTGATCATGATGCTTTCA Exon 6 TCTTACAGGACTGGCACACG CCCCAAGAATATCTGGGACA Exon 7 TCAGGCTGAACTAGAGCAAACA CAAACAGCACTCCTGCAGAC Exon 8 CATGGGAAGAAGAGAGAAAG GTTGACTGGTATCAGAAGAC Exon 9 ACTCCTGACCTCGTAATCC GAAGCAGATGATAACAGAACC Exon 10 TCTAATTGAAACTCTTCCCC AGTTCGAAGCCAGTCTAGC Exon 11 TGAGTTGACTGCGGTGATTT GTAATCCTCCGGATCCCATC Exon 12 GTCTAGCCCTGAGGATGTGG TGCAAAATGGACAGGTGTTT Exon 13 CAGACACAACATTGGAGAC TAAGGGCAAAGAGGAAAG Exon 14 CTGCATGAAATTACTCAAGC CCATCCTCTCATTTACTTCC Exon 15 AAACTGTTTACCTTGCCC GCACCTCACTTCAATCTC Exon 16 GAGTAACATTTGACGGATG CTCTACTCTACCCACAGTTC Table 3 RESULTS OF SNP ANALYSIS OF 16 EXONS ENCODING ABCG2* Wild-type Frequency Frequency Frequency Amino Acid Exon Nucleotide+ allele SNP Wt/Wt Wt/Var Var/Var aa# 1 91 C T 98.9% 1.1% Noncoding 175 A G 97.8% 2.2% Noncoding 2 238 G A 76.7% 22.2% 1.1% 12 Val to Met 5 625 C A 88.9% 10% 1.1% 141 Gln to Lys 9 1302 G A 97.8% 2.2% 366 Glu to Glu 12 1629 A G 98.9% 1.1% 475 Leu to Leu 16 2062 G A 98.9% 1.1% 620 Asp to Asn 2597 C A 98.9% 1.1% Noncoding Intronic Variants 2 +36** A G 76.7% 22.2% 1.1% 6 -16 A G 88.9% 8.9% 2.2% 7 -20 T A 98.9% 1.1% +18 A G 93.3% 5.6% 1.1% 11 +20 A G 63.3% 27.8% 8.9% 12 +49 G T 75.6% 22.2% 2.2% 14 -21 C T 67.8% 28.9% 3.3% *Identified SNPs are recorded in the table with all 16 exons sequenced in 90 DNA samples. Login to comment

[hide] Genetic variation in the ATP-binding cassette tran... Eur J Pharm Sci. 2003 Apr;18(5):359-64. Backstrom G, Taipalensuu J, Melhus H, Brandstrom H, Svensson AC, Artursson P, Kindmark A
Genetic variation in the ATP-binding cassette transporter gene ABCG2 (BCRP) in a Swedish population.
Eur J Pharm Sci. 2003 Apr;18(5):359-64., [PMID:12694888]

Abstract [show]
The ATP-binding cassette transporter ABCG2 (also named breast cancer resistance protein, BCRP) functions as a drug efflux transporter and is expressed at high levels in the human small intestine. The aim of this study was to screen the human ABCG2 gene for genetic variation. The regions of the gene most likely to affect function, namely the coding parts, exon/intron boundaries, 5' untranslated region and 3' untranslated region and the proposed promoter region, were included in the screening. DNA was obtained from 60 Swedish individuals. The screening was performed using a polymerase chain reaction-denaturing high-performance liquid chromatography approach followed by sequence analysis. Eight sites of genetic variation were identified. The sequence variations considered to be most likely to affect transcription level or transport function were a CTCA deletion in the 5' flanking region, a single nucleotide polymorphism (SNP) in a 5' flanking CpG island, two non-synonymous SNPs, changing valine at amino acid position 12 to methionine and glutamine at position 141 to lysine, respectively. Genotyping of these sequence variations revealed linkage between the CTCA deletion and the SNP changing glutamine 141 for lysine. This information forms the basis for future association studies to investigate the genetic basis of differences of drug disposition due to sequence variation in the ABCG2 gene.

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6 The sequence variations considered to be most likely to affect transcription level or transport function were a CTCA deletion in the 59 flanking region, a single nucleotide polymorphism (SNP) in a 59 flanking CpG island, two non-synonymous SNPs, changing valine at amino acid position 12 to methionine and glutamine at position 141 to lysine, respectively. Login to comment
7 Genotyping of these sequence variations revealed linkage between the CTCA deletion and the SNP changing glutamine 141 for lysine. Login to comment
62 Because To determine allele frequencies for the sequence varia- of the highly repetitive sequences surrounding exon 10, Table 2 Sequence variation in the human ABCG2 gene identified in the present study Sequence variant Nucleotide sequence (59 to 39) Affected Effect Identity to a ID DHPLC dbSNP b c Reference sequence Alteration pools g.-19572-19569 actcaCTCAcaaag actca caaag 15 59 Flanking deletion ss 4480605 ]] ]]]]d delCTCA g.-19202G.C gtactGatcag gtactCatcag 5 CpG island SNP rs 2231134 ] ] g.-18845T.C tgagcTcgtcc tgagcCcgtcc 8 59 UTR SNP rs 2231135 ] ] g.-18604delA cggcaAggagg cggca ggagg 1 59 UTR deletion ss 4480606 ] ]d g.34G.A tcccaGtgtca tcccaAtgtca 2 Missense SNP rs 2231137 ] ] Val12Met g.8007G.A ttggaGggaaa ttggaAggaaa 3 Intronic SNP ss 4480607 ] ]d g.8825C.A acttaCagttc acttaAagttc 4 Missense SNP rs 2231142 ] ] Gln141Lys d g.44997G.A ttcttAaaatt ttcttGaaatt 9 Intronic SNP rs 2231164 ] ] a Sequence variant ID in accordance with the nomenclature for sequence variation described at http://www.dmd.nl/mutnomen.html. Login to comment
78 The deletion (g.-19572-19569delCTCA) was identified in the g.8825C.A SNP (Gln141Lys) occurred at a frequency of 59 flanking region. Login to comment
86 A one-base deletion (g.-18604delA) and 1 disequilibrium was observed over the region between SNP (g.-18845T.C) were detected in the untranslated g.-19572-19569delCTCA and g.8825C.A (Gln141Lys) exon 1. Login to comment
115 We also report the allele fre- g.8825C.A and low expression of the Gln141Lys ABCG2 quencies for four identified sequence variations, and the protein (Imai et al., 2002), suggested to be due to the degree of linkage disequilibrium over the region studied. Login to comment
208 Influence of CYP2C9 of Q141K protein and low-level drug resistance. Login to comment

[hide] Multidrug resistance mediated by the breast cancer... Oncogene. 2003 Oct 20;22(47):7340-58. Doyle LA, Ross DD
Multidrug resistance mediated by the breast cancer resistance protein BCRP (ABCG2).
Oncogene. 2003 Oct 20;22(47):7340-58., 2003-10-20 [PMID:14576842]

Abstract [show]
Observations of functional adenosine triphosphate (ATP)-dependent drug efflux in certain multidrug-resistant cancer cell lines without overexpression of P-glycoprotein or multidrug resistance protein (MRP) family members suggested the existence of another ATP-binding cassette (ABC) transporter capable of causing cancer drug resistance. In one such cell line (MCF-7/AdrVp), the overexpression of a novel member of the G subfamily of ABC transporters was found. The new transporter was termed the breast cancer resistance protein (BCRP), because of its identification in MCF-7 human breast carcinoma cells. BCRP is a 655 amino-acid polypeptide, formally designated as ABCG2. Like all members of the ABC G (white) subfamily, BCRP is a half transporter. Transfection and enforced overexpression of BCRP in drug-sensitive MCF-7 or MDA-MB-231 cells recapitulates the drug-resistance phenotype of MCF-7/AdrVp cells, consistent with current evidence suggesting that functional BCRP is a homodimer. BCRP maps to chromosome 4q22, downstream from a TATA-less promoter. The spectrum of anticancer drugs effluxed by BCRP includes mitoxantrone, camptothecin-derived and indolocarbazole topoisomerase I inhibitors, methotrexate, flavopiridol, and quinazoline ErbB1 inhibitors. Transport of anthracyclines is variable and appears to depend on the presence of a BCRP mutation at codon 482. Potent and specific inhibitors of BCRP are now being developed, opening the door to clinical applications of BCRP inhibition. Owing to tissue localization in the placenta, bile canaliculi, colon, small bowel, and brain microvessel endothelium, BCRP may play a role in protecting the organism from potentially harmful xenobiotics. BCRP expression has also been demonstrated in pluripotential "side population" stem cells, responsible for the characteristic ability of these cells to exclude Hoechst 33342 dye, and possibly for the maintenance of the stem cell phenotype. Studies are emerging on the role of BCRP expression in drug resistance in clinical cancers. More prospective studies are needed, preferably combining BCRP protein or mRNA quantification with functional assays, in order to determine the contribution of BCRP to drug resistance in human cancers.

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127 Allelic variation as a result of SNPs results in alterations of the BCRP protein at amino acids 12 (V12M), 141 (Q141K), 206 (I206L), and 590 (N590Y), with the most frequent polymorphisms being the exon 2 SNP (G34A) and the exon 5 SNP (C421A), which produce changes in amino acids 12 and 141 (Honjo et al., 2002; Imai et al., 2002a; Zamber et al., 2003). Login to comment
129 The effect of these polymorphisms on BCRP function remains to be determined, although one study suggested that the C421A/Q141K variant may result in Structure of the Human BCRP (ABCG2) Gene and Promoter (Chromosome 4q22) BCRP Gene Walker A Walker B TM 1,2 TM 3,4 TM 5,6 ABC Signature 1 2 3 4 5 6 7 8 9 10 11121314 15 16 NH2 COOHBCRP Protein -450 -400 -350 -300 -250 -200 -150 - -50 0 50 100 150 BCRP 5` Upstream Region (Promoter) Sp1 Sp1 5` AP1 Predicted Promoter CCAAT box CpG Island >66 kb / 2.4 kb 655 aa 100 (Chromosome 4q22) BCRP Gene Walker A Walker B TM 3,4 TM 5,6 ABC Signature 1 2 3 4 5 6 7 8 9 10 11121314 15 16 COOHBCRP Protein - - - - - BCRP 5` Upstream Region (Promoter) Sp1 Sp1AP1 Predicted Promoter CCAAT box CpG Island >66 kb / 2.4 kb 655 aa Figure 3 Structure of BCRP gene and promoter. Login to comment

[hide] Single nucleotide polymorphisms result in impaired... Int J Cancer. 2004 Mar 20;109(2):238-46. Mizuarai S, Aozasa N, Kotani H
Single nucleotide polymorphisms result in impaired membrane localization and reduced atpase activity in multidrug transporter ABCG2.
Int J Cancer. 2004 Mar 20;109(2):238-46., 2004-03-20 [PMID:14750175]

Abstract [show]
ABCG2/MXR/ABCP1/BCRP is a member of the ATP-binding cassette membrane transporter, which consists of six transmembrane regions and one ATP-binding cassette. The transporter is known to be involved in the efflux of various anticancer compounds such as mitoxantrone, doxorubicin and topoisomerase I inhibitor. In this study, we analyzed the effects of polymorphisms in ABCG2, V12M and Q141K on transporter function. When polarized LLC-PK1 cells were transfected with variant ABCG2, drug-resistance to topoisomerase I inhibitor of cells expressing V12M or Q141K was less than 1/10 that of wild-type ABCG2 transfected cells, and was accompanied by increased drug accumulation and decreased drug efflux in the variant ABCG2-expressing cells. We further elucidated the molecular mechanisms of the transport dysfunction by investigating membrane localization and ATPase activity. Confocal microscopic analysis revealed that apical plasma membrane localization of V12M was disturbed, while the localization of wild-type transporters occurred specifically in the apical plasma membrane of polarized LLC-PK1 cells. Also, ATPase activities measured in the membrane of SF9 cells infected with variant ABCG2 showed that Q141K decreased activity by 1.3 below that of wild-type ABCG2. In addition, kinetic analysis of ATPase activity showed that the K(m) value in Q141K was 1.4-fold higher than that of wild-type ABCG2. These results indicated that naturally occurring SNPs alter transport functions of ABCG2 transporter and analysis of SNPs in ABCG2 may hold great importance in understanding the response/metabolism of chemotherapy compounds that act as substrates for ABCG2.

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2 In this study, we analyzed the effects of polymorphisms in ABCG2, V12M and Q141K on transporter function. Login to comment
3 When polarized LLC-PK1 cells were transfected with variant ABCG2, drug-resistance to topoisomerase I inhibitor of cells expressing V12M or Q141K was less than 1/10 that of wild-type ABCG2 transfected cells, and was accompanied by increased drug accumulation and decreased drug efflux in the variant ABCG2-expressing cells. Login to comment
6 Also, ATPase activities measured in the membrane of SF9 cells infected with variant ABCG2 showed that Q141K decreased activity by 1.3 below that of wild-type ABCG2. Login to comment
7 In addition, kinetic analysis of ATPase activity showed that the Km value in Q141K was 1.4-fold higher than that of wild-type ABCG2. Login to comment
15 Homozygous CC patients expressed lower P-gp in the intestine compared to TT patients, resulting in increased digoxin plasma concentration after orally administered digoxin.21 Another report showed that a naturally occurring mutation of R433S in MRP1 caused increased organic anion transport and decreased doxorubicin resistance.22 Several groups have reported naturally occurring ABCG2 SNPs in various ethnic populations, including Caucasian, Asian and African.23-27 In those reports, polymorphisms at V12M and Q141K occurred at high frequency in most of the ethnic populations. Login to comment
17 For example, Q141K polymorphism has been reported to reduce protein expression level compared to wild-type ABCG2 in vitro;24 however, this observation was not confirmed by in vivo analysis using intestinal samples.25 Mutations at R482 have been well characterized by several groups including our previous studies,17,28-30 demonstrating that they affect drug resistance and are considered mutations acquired during drug selection and not naturally occurring polymorphisms. Login to comment
21 The previously reported polymorphisms,V12M and Q141K, had high frequencies of 10.3% and 9.0%, respectively. Login to comment
22 Drug resistance to the ABCG2 substrate, indolocarbazole topoisomerase I inhibitor, was reduced more than 10-fold in polarized cells that expressed variant ABCG2 with either V12M or Q141K compared Abbreviations: ABC, ATP-binding cassette; ABCP1, placenta-specific ABC transporter; BCRP, breast cancer-resistant protein; GFP, green fluorescent protein; in; HRP, horse radish peroxidase; MDR, multidrug resistance protein; MRP, multidrug resistance-associated prote; MXR, mitoxantrone resistance protein; P-gp, P-glycoprotein; PVDF, polyvinylidene difluoride; Sf9 cells, Spodoptera frugiperda ovarian cells; SNPs, single nucleotide polymorphisms. Login to comment
28 We concluded that the functional impairment of these 2 variants were due to disturbance of apical plasma membrane localization for V12M and reduced ATPase activity for Q141K, indicating ABCG2 gene SNPs may greatly influence resistance to ABCG2 substrate. Login to comment
43 Establishment of LLC-PK1 cells expressing wild-type and mutant ABCG2 Wild-type ABCG2 and mutated ABCG2 genes (G34A for V12M and C421A for Q141K), the point mutations of which were introduced by PCR mutagenesis, were cloned into HindIII and XhoI sites of pcDNA3.1(ϩ) (Invitrogen). Login to comment
61 Drug efflux assay LLC-PK1 cells were incubated with the indicated concentration of indolocarbazole compound for 120 min under energy-depleted TABLE I - SINGLE NUCLEOTIDE POLYMORPHISMS IN ABCG21 SNP Effect Region Domain Frequency in 30 cell lines Frequency in 150 clinical samples Hetero Home Hetero Homo Allele (%) G34A V12M Exon2 N-terminal 5 (16.7%) 0 27 (18.0%) 2 (1.3%) 10.3 Aϩ10G Intron3 ND ND 21 (14.0%) 4 (2.7%) 9.7 C369T Wobble Exon4 ABC 0 0 1 (0.67%) 0 0.3 C376T Q126Term Exon4 ABC 0 1 (3.3%) 0 0 0.0 C421A Q141K Exon5 ABC 5 (16.7%) 1 (3.3%) 22 (15.3%) 2 (1.3%) 9.0 C458T T153M Exon5 ABC 1 (3.3%) 0 0 0 0.0 C474T Wobble Exon5 ABC 0 0 1 (0.67%) 0 0.3 Aϩ20G Intron11 ND ND 34 (22.7%) 10 (6.7%) 18.0 A1444G R482G Exon12 TM3 0 0 0 0 0.0 G1445C R482T Exon12 TM3 0 0 0 0 0.0 C-21T Intron13 ND ND 32 (21.3%) 4 (2.7%) 13.3 A1768T N590Y Exon15 EC3 0 0 1 (0.67%) 0 0.3 G2237T Exon16 3ЈUTR 1 (3.3%) 0 0 0 0.0 G2393T Exon16 3ЈUTR 1 (3.3%) 0 0 0 0.0 The positions of the polymorphisms correspond to that of the ABCG2 cDNA (GenBank accession number AB051855) with the first base of the ATG start codon set to 1. Login to comment
89 Samples of 7 ␮g RNA extracted from HeLa, control C4, WT, V12M and Q141K cells (lanes 1, 2, 3, 4 and 5 respectively) were subjected to Northern hybridization and the blots were probed with a cDNA fragment of ABCG2. Login to comment
92 Vector transformant C4 (a) and stable clones expressing wild-type (b), V12M (c) and Q141K (d) were stained with monoclonal antibody BXP-34. Login to comment
102 Two polymorphisms, V12M and Q141K, had high frequency rates of 10.3% and 9.0%, respectively, in the 150 Caucasian subjects. Login to comment
103 V12M was located at the N-terminal intracellular region and Q141K at the ATP-binding cassette region. Login to comment
108 LLC-PK1 cells expressing WT, V12M and Q141K were incubated with opti-MEM with 50 ␮M (A) or 0.5 ␮M (B) of radiolabeled topoisomerase inhibitor for 180 min. Login to comment
119 TABLE II - RESISTANT PROFILE (IC50) OF ABCG2 TO ANTI-CANCER COMPOUNDS Anti-cancer compound IC50 (␮M)1 Control C4 Wild type V12M Q141K TopoI inhibitor2 0.12 Ͼ50 (420) 0.94 (7.8) 5.9 (49) Mitoxantrone 0.0015 0.029 (19) 0.00093 (0.62) 0.0053 (3.5) Topotecan 0.098 2.1 (22) 0.16 (1.7) 0.48 (4.9) Doxorubicin 0.010 0.039 (3.9) 0.0073 (0.73) 0.014 (1.4) Vincristine 0.0034 0.0053 (1.6) 0.0021 (0.62) 0.0058 (1.7) Camptothecin 0.0087 0.021 (2.4) 0.012 (1.4) 0.027 (3.1) 1 Relative resistances to control cells are described in parentheses.-2 TopoI inhibitor, Indolocarbazole topoisomerase I inhibitor. Login to comment
120 Resistance profile of variant ABCG2 transporters to anticancer compounds Among the polymorphisms detected, V12M and Q141K had a high frequency of amino acid substitutions. Login to comment
123 Northern blotting and immunocytochemical analysis with a monoclonal antibody revealed that approximately equal levels of ABCG2 were expressed in V12M and Q141K clones compared to the expression in the wild-type (WT) clone (Fig. 1). Login to comment
125 Wild-type cells conferred greater than 420-fold higher resistance to an indolocarbazole I topoisomerase inhibitor compared to that of control C4 cells as previously reported.10 In contrast, IC50 values of variant ABCG2 clones, V12M and Q141K, were less than 1/10 that of WT cells. Login to comment
127 The IC50 values of WT cells to mitoxantrone and topotecan increased 19- and 22-fold, respectively, over C4 cells, whereas in the variant V12M and Q141K cells, IC50 values to mitoxantrone and topotecan decreased as observed with the indolocarbazole compound. Login to comment
130 Accumulation and efflux assay of topoisomerase I inhibitor in ABCG2 variant-expressing cells Increased sensitivities of V12M and Q141K cells are thought to arise from changes in the intracellular drug concentration of topoisomerase I inhibitor. Login to comment
134 However, significantly higher indolocarbazole topoisomerase I inhibitor accumulation was observed in both V12M and Q141K cells compared to that in WT cells (2.7and 1.8-fold higher, respectively). Login to comment
135 A drug accumulation assay performed at a low dose (0.5 ␮M) of the compound confirmed that compound accumulations increased in variant cell lines (3.1and 2.8-fold increase for V12M and Q141K, respectively). Login to comment
141 Compared to WT cells, Vmax of V12M and Q141K cells decreased 2.5-and 1.8-fold, respectively, indicating that the increased drug accumulation and consequent reduction in drug resistance were due to the decreased efflux velocity. Login to comment
145 Vmax values of V12M and Q141K decreased 2.2- and 1.7-fold, respectively, which agrees well with the results of drug efflux assays using cell lines. Login to comment
153 In Q141K cells, ABCG2 staining was detected in the apical side, similar to WT cells (Fig. 3d). Login to comment
163 (A) Control C4 cells; (B) and E, WT cells; (C,F), V12M cells; (D,G), Q141K cells. Login to comment
167 In WT and Q141K cells, apical membrane staining of ABCG2 was detected (Fig. 3E,G). Login to comment
170 The expected pattern of apical staining with anti-ABCG2 antibody was observed in wild-type and Q141K variant expressing cell lines, whereas dispersed staining was confirmed in V12M cells (data not shown). Login to comment
172 ATPase activity of variant ABCG2 Since Q141K was mapped in the ATP-binding cassette region, ATPase activity of the variant transporters was measured. Login to comment
177 However, the ATPase activity of Q141K was reduced by about 1.3-fold compared to that of wild-type ABCG2. Login to comment
182 We concluded that Q141K and V12M polymorphisms did not restore drug stimulation, which was observed in R482 mutant. Login to comment
184 In the Q141K membrane, the Vmax value of ATPase activity decreased 1.3-fold in accordance with the values of Figure 4A. Login to comment
185 Unexpectedly, the Km value of Q141K also increased 1.4-fold compared to that of WT (0.64 mM; WT, 0.64 mM; V12M, 0.91 mM; Q141K). Login to comment
188 DISCUSSION In our study, we confirmed the locations and frequencies of SNPs in ABCG2 using 30 cancer cell lines and 150 Caucasian clinical samples, and then characterized the functional effects of the major SNPs, V12M and Q141K (Fig. 5). Login to comment
192 Membrane fractions isolated from Sf9 cells expressing wild-type, V12M and Q141K ABCG2 transporters were subjected to Western blotting with BXP-21 monoclonal antibody. Login to comment
203 the impaired function of ABCG2, membrane localization of transporter for V12M and ATPase activity for Q141K. Login to comment
210 The other variant, Q141K, also reduced drug resistance against previously known ABCG2 substrates when it was expressed in LLC-PK1 cells. Login to comment
212 Several mutagenesis studies on ABC transporters have shown that introduced mutations in the ABC domain completely disrupted ATPase activity.30,38,39 For example, an induced mutation in ABCG2 prevented K86M from transporting ABCG2 substrates due to the loss of ATPase activity.30 When measured in the Sf9 membrane containing ABCG2 transporters, ATPase activity of Q141K was 1.3-fold lower than that of wild-type ABCG2, although the effect on ATPase activity was relatively moderate compared to that of the introduced mutation at K86M. Login to comment
213 While K86M was located at the walker catalytic region in the ABC domain, Q141K was located between the walkerA and walkerB regions. Login to comment
214 In addition to the changes in ATPase activity of Q141K, the Km value of Q141K was different from that of WT by a factor of 1.4. Login to comment
215 The change in Km value infers that the difference in ATPase activity of Q141K and WT might be larger than 1.3-fold at the physiological concentration of ATP. Login to comment
216 Previously, the effects of Q141K on protein and mRNA expression were investigated in an ABCG2-overexpressing murine cell line24 or human clinical intestinal samples.25 The initial report showed that murine PA317 cells expressing Q141K increased intracellular drug accumulation compared to cells expressing WT ABCG2, coupled with reduced protein levels with similar mRNA expression, suggesting that the translation efficiency of ABCG2 was the cause of this phenomenon. Login to comment
217 In contrast, a second study involving 32 human intestinal samples demonstrated no correlation between the amount of ABCG2 protein and Q141K polymorphism. Login to comment
218 Our study, using polarized LLC-PK1, agreed with the latter study in that Q141K did not affect ABCG2 protein expression; instead, it showed impaired Q141K transporting function. Login to comment
219 We also confirmed this observation using Sf9 membranes containing a similar amount of ABCG2 protein, in which WTand Q141K-expressing cells exhibited altered ATPase activity. Login to comment
224 Three identified variants, which affect transporter function, were designated as V12M (1), Q126Term (2) and Q141K (3). Login to comment

[hide] Multidrug resistance in cancer chemotherapy and xe... Curr Med Chem Anticancer Agents. 2004 Jan;4(1):31-42. Han B, Zhang JT
Multidrug resistance in cancer chemotherapy and xenobiotic protection mediated by the half ATP-binding cassette transporter ABCG2.
Curr Med Chem Anticancer Agents. 2004 Jan;4(1):31-42., [PMID:14754410]

Abstract [show]
ABCG2, also termed BCRP/MXR/ABCP, is a half ATP-binding cassette (ABC) transporter expressed on plasma membranes. ABCG2 was independently cloned from placenta as well as cell lines selected for resistance to mitoxantrone or anthracyclines. ABCG2 consists of a nucleotide-binding domain (NBD) at the amino terminus and a transmembrane domain (TMD) at the carboxyl terminus and it is postulated to form a homodimer to perform its biological functions. Over-expression of ABCG2 in cell lines confers resistance on a wide variety of anticancer drugs including mitoxantrone, daunorubicin, doxorubicin, topotecan and epirubicin. The expression of ABCG2 has been implicated in multidrug resistance (MDR) of acute myeloid leukemia and some solid tumors. In addition, ABCG2 can transport several fluorescent dyes or toxins. ABCG2 is found to be expressed in epithelial cells of intestine and colon, liver canaliculi, and renal tubules, where it serves to eliminate the plasma level of orally administered anticancer drugs as well as ingested toxins. ABCG2 is found to be highly expressed in placenta and the luminal surface of microvessel endothelium blood-brain barrier where it may play a role in limiting the penetration of drugs, such as topotecan from the maternal plasma into the fetus and from blood to brain. A variety of inhibitors for ABCG2 including GF120918 may prove useful for sensitizing cancer cells to chemotherapy or altering the distribution of orally administered drug substrates of ABCG2. Interestingly, ABCG2 is also expressed highly in hematopoietic stem cells. However, the function of ABCG2 in stem cells is currently unknown, although it may provide protection to stem cells from a variety of xenobiotics.

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108 Nonsynonymous SNPs at nucleotide 238 (Val12 Met; exon 2) and nucleotide 625 (Gln141 Lys; exon 5) showed a greater frequency of heterozygosity (22.2% and 10%, respectively) than the SNP at 2062 (Asp620 Asn; exon 16). Login to comment
113 Two non-synonymous SNPs, Val12 Met and Gln141 Lys, were also identified, which are consistent with the study by Honjo et al. [25]. Login to comment
115 Genotyping of these sequence variations revealed linkage between the CTCA deletion and the SNP of Gln141 Lys. Login to comment

[hide] ABC transporters and the blood-brain barrier. Curr Pharm Des. 2004;10(12):1295-312. Begley DJ
ABC transporters and the blood-brain barrier.
Curr Pharm Des. 2004;10(12):1295-312., [PMID:15134482]

Abstract [show]
The blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier (BCSFB) form a very effective barrier to the free diffusion of many polar solutes into the brain. Many metabolites that are polar have their brain entry facilitated by specific inwardly-directed transport mechanisms. In general the more lipid soluble a molecule or drug is, the more readily it will tend to partition into brain tissue. However, a very significant number of lipid soluble molecules, among them many useful therapeutic drugs have lower brain permeability than would be predicted from a determination of their lipid solubility. These molecules are substrates for the ABC efflux transporters which are present in the BBB and BCSB and the activity of these transporters very efficiently removes the drug from the CNS, thus limiting brain uptake. P-glycoprotein (Pgp) was the first of these ABC transporters to be described, followed by the multidrug resistance-associated proteins (MRP) and more recently breast cancer resistance protein (BCRP). All are expressed in the BBB and BCSFB and combine to reduce the brain penetration of many drugs. This phenomenon of "multidrug resistance" is a major hurdle when it comes to the delivery of therapeutics to the brain, not to mention the problem of cancer chemotherapy in general. Therefore, the development of strategies for bypassing the influence of these ABC transporters and for the design of effective drugs that are not substrates and the development of inhibitors for the ABC transporters becomes a high imperative for the pharmaceutical industry.

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294 Also a C421A polymorphism has been reported which substitutes lysine for glutamine at position 141 of the transporter. Login to comment

[hide] ABCG2 -- a transporter for all seasons. FEBS Lett. 2004 Jun 1;567(1):116-20. Sarkadi B, Ozvegy-Laczka C, Nemet K, Varadi A
ABCG2 -- a transporter for all seasons.
FEBS Lett. 2004 Jun 1;567(1):116-20., 2004-06-01 [PMID:15165903]

Abstract [show]
The human ABCG2 (ABCP/MXR/BCRP) protein is a recently recognized ABC half-transporter, which forms homodimers in the plasma membrane and actively extrudes a wide variety of chemically unrelated compounds from the cells. This protein protects our cells and tissues against various xenobiotics, with a crucial role in the intestine, liver, placenta, and the blood-brain barrier. Moreover, ABCG2 seems to have a key function in stem cell protection/regulation, and also in hypoxic defense mechanisms. Widely occurring single nucleotide polymorphisms in ABCG2 may affect absorption and distribution, altering the effectiveness and toxicity of drugs in large populations. At the clinics, overexpression of ABCG2 in tumor cells confers cancer multidrug resistance to a variety of newly developed anticancer agents. On the other hand, specific substrate mutants of ABCG2 are advocated for use as selectable markers in stem-cell based gene therapy.

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127 These SNPs yield ABCG2 variants containing V12M and Q141K. Login to comment

[hide] Novel camptothecin analogues that circumvent ABCG2... Int J Cancer. 2004 Jul 20;110(6):921-7. Yoshikawa M, Ikegami Y, Hayasaka S, Ishii K, Ito A, Sano K, Suzuki T, Togawa T, Yoshida H, Soda H, Oka M, Kohno S, Sawada S, Ishikawa T, Tanabe S
Novel camptothecin analogues that circumvent ABCG2-associated drug resistance in human tumor cells.
Int J Cancer. 2004 Jul 20;110(6):921-7., 2004-07-20 [PMID:15170677]

Abstract [show]
Irinotecan (7-ethyl-10-[4-(1-piperidino)-1-piperidino]-carbonyloxycamptothecin; CPT-11) is a widely used potent antitumor drug that inhibits mammalian DNA topoisomerase I (Topo I); however, overexpression of ABCG2 (BCRP/MXR/ABCP) can confer cancer cell resistance to SN-38, the active form of CPT-11. We have recently demonstrated that plasma membrane vesicles prepared from ABCG2-overexpressing PC-6/SN2-5H cells transported SN-38 and its glucuronide conjugate in an ATP-dependent manner (Nakatomi et al., Biochem Biophys Res Commun 2001;288:827-32). In the present study, we have characterized a total of 14 new camptothecin (CPT) analogues with respect to both the inhibition of Topo I and the substrate specificity of ABCG2. All of the tested CPT analogues, which have different substitutions at positions 10 and 11, strongly inhibited the Topo I activity in a cell-free system, as did SN-38. Their antitumor activities in the SN-38-resistant PC-6/SN2-5H2 cell line greatly varied, however, being correlated with intracellular accumulation levels. We have examined ATP-dependent transport of those CPT analogues by using plasma membrane vesicles prepared from both PC-6/SN2-5H2 cells and ABCG2-transfected HEK-293 cells. Based on the substrate specificity of ABCG2 thus evaluated, it is strongly suggested that CPT analogues with high polarity are good substrates for ABCG2 and are therefore effectively extruded from cancer cells. In this context, to circumvent ABCG2-associated drug resistance, low-polarity CPT analogues are considered to be potent lead compounds. The present study provides a practical approach to discover new CPT-based drugs for the chemotherapy of drug-resistant human cancer.

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169 This type of variation (Gln141Lys) has hitherto been reported as a naturally occurring SNP of ABCG2. Login to comment
170 The Gln141Lys polymorphism has recently been identified in healthy Japanese volunteers as well.29 We investigated the effects of this SNP on the substrate specificity of ABCG2, using ABCG2-transfected cells with Gln at amino acid 141. Login to comment

[hide] Diflomotecan pharmacokinetics in relation to ABCG2... Clin Pharmacol Ther. 2004 Jul;76(1):38-44. Sparreboom A, Gelderblom H, Marsh S, Ahluwalia R, Obach R, Principe P, Twelves C, Verweij J, McLeod HL
Diflomotecan pharmacokinetics in relation to ABCG2 421C>A genotype.
Clin Pharmacol Ther. 2004 Jul;76(1):38-44., [PMID:15229462]

Abstract [show]
OBJECTIVE: The adenosine triphosphate-binding cassette transporter ABCG2 (breast cancer resistance protein [BCRP]) functions as an efflux transporter for many drugs, including the topoisomerase I inhibitor diflomotecan, and is expressed at high levels in the intestine and liver. We performed an exploratory analysis to evaluate the effects of the natural allelic variant ABCG2 421C>A on the pharmacokinetics of diflomotecan. METHODS: The drug was administered to 22 adult white patients with cancer as a 20-minute infusion (dose, 0.10-0.27 mg), followed 2 weeks later by an oral solution (dose, 0.10-0.35 mg). RESULTS: The ABCG2 421C>A genotype significantly affected the pharmacokinetics of diflomotecan; in 5 patients heterozygous for this allele, plasma levels after intravenous drug administration were 299% (P =.015) of those in 15 patients with wild-type alleles, at mean values of 138 ng x h/mL x mg(-1) (95% confidence interval, 11.3-264 ng x h/mL x mg(-1)) versus 46.1 ng x h/mL x mg(-1) (95% confidence interval, 25.6-66.7 ng x h/mL x mg(-1)), respectively. Diflomotecan levels were not significantly influenced by 11 known variants in the ABCB1, ABCC2, cytochrome P450 (CYP) 3A4, and CYP3A5 genes. CONCLUSION: These findings provide the first evidence linking variant ABCG2 alleles to altered drug exposure and suggest that interindividual variability in substrate drug effects might be influenced, in part, by ABCG2 genotype.

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56 Frequencies for studied variant genes and genotype-phenotype relationships Polymorphism and nomenclature Effect Allele frequencies Ratio (q/p) for intravenous AUC Ratio (q/p) for oral AUC p q Value† P value Value† P value ABCG2 421CϾA Q141K 0.88 0.12 2.98 .015 1.15 .74 ABCC2-24CϾT 5'-UTR 0.89 0.11 0.868 .82 0.890 .78 ABCC2 1249GϾA V417I 0.86 0.14 1.06 .92 2.10 .063 ABCC2 156231AϾG Intron 3 1.00 0.00 NA NA NA NA ABCB1 1236CϾT‡ (ABCB1*8§) G411G 0.40 0.60 0.877 .82 0.585 .31 ABCB1 2677GϾA/T࿣ (ABCB1*7§) A893S/T 0.63 0.34/0.03 0.784 .61 1.27 .55 ABCB1 3435CϾT‡ (ABCB1*6§) I1145I 0.45 0.55 0.978 .97 0.955 .93 CYP3A4 -392AϾG (CYP3A4*1B) Promotor 0.91 0.09 0.731 .63 0.834 .75 CYP3A4 15713TϾC‡ (CYP3A4*2) S222P 1.00 0.00 NA NA NA NA CYP3A4 23172TϾC (CYP3A4*3) M445T 1.00 0.00 NA NA NA NA CYP3A5 22893GϾA‡ (CYP3A5*3) Splice Variant 0.86 0.14 0.808 .77 0.813 .76 CYP3A5 30597GϾA (CYP3A5*6) Splice Variant 1.00 0.00 NA NA NA NA AUC, Area under plasma concentration-time curve normalized to dose (ie, observed AUC/dose in milligrams); NA, not available. Login to comment
78 Plasma concentration-time profiles of diflomotecan as a function of ABCG2 421CϾA (Q141K) genotype (in which open symbols are homozygous wild-type patients [n ϭ 15] and solid symbols are heterozygous variant patients [n ϭ 5] receiving a 20-minute intravenous infusion of diflomotecan at a dose of 0.1-0.27 mg). Login to comment
87 The studied variant in ABCG2 is a single nucleotide polymorphism causing a nonsynonymous change in the protein sequence, in which a 421C to A transition at exon 5 leads to a lysine to glutamine amino acid substitution at codon 141 (Q141K). Login to comment
91 Area under curve (AUC) of diflomotecan as a function of ABCG2 421CϾA (Q141K) genotype. Login to comment

[hide] ABCG2 pharmacogenetics: ethnic differences in alle... Clin Cancer Res. 2004 Sep 1;10(17):5889-94. de Jong FA, Marsh S, Mathijssen RH, King C, Verweij J, Sparreboom A, McLeod HL
ABCG2 pharmacogenetics: ethnic differences in allele frequency and assessment of influence on irinotecan disposition.
Clin Cancer Res. 2004 Sep 1;10(17):5889-94., 2004-09-01 [PMID:15355921]

Abstract [show]
PURPOSE: The ATP-binding cassette transporter ABCG2 (breast cancer resistance protein) is an efflux protein that plays a role in host detoxification of various xenobiotic substrates, including the irinotecan metabolite 7- ethyl-10-hydroxycamptothecin (SN-38). The ABCG2 421C>A polymorphism has been associated with reduced protein expression and altered function in vitro. The aim of this study was to evaluate the ethnic distribution and potential functional consequence of the ABCG2 421C>A genotype in cancer patients treated with irinotecan. EXPERIMENTAL DESIGN: ABCG2 genotyping was performed using Pyrosequencing on DNA from 88 American Caucasians, 94 African Americans, 938 Africans, and 95 Han Chinese, as well as in 84 European Caucasian patients treated with irinotecan undergoing additional blood sampling for pharmacokinetic studies. RESULTS: Significant differences in allele frequencies were observed between the given world populations (P < 0.001), the variant allele being most common in the Han Chinese population with a frequency as high as 34%. The mean area under the curve of irinotecan and SN-38 were 19,851 and 639 ng x hour/mL, respectively. The frequency of the variant allele (10.7%) was in line with results in American Caucasians. No significant changes in irinotecan pharmacokinetics were observed in relation to the ABCG2 421C>A genotype, although one of two homozygous variant allele carriers showed extensive accumulation of SN-38 and SN-38 glucuronide. CONCLUSIONS: The ABCG2 421C>A polymorphism appears to play a limited role in the disposition of irinotecan in European Caucasians. It is likely that the contribution of this genetic variant is obscured by a functional role of other polymorphic proteins.

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23 In particular, a single nucleotide polymorphism in exon 5 of the ABCG2 gene has been described in which a 421CϾA transversion results in an amino acid change of glutamine to lysine at codon 141 (14-16). Login to comment
108 However, the low frequency of ABCG2 421A (Ͻ 1%) in this population is in line with earlier observations in Africans north of the Fig. 1 Boxplot of dose-normalized AUC of irinotecan (A), SN-38 (B), and SN-38G (C) as a function of the ABCG2 421CϾA (Q141K) genotype. Login to comment

[hide] Functional assessment of ABCG2 (BCRP) gene polymor... Drug Metab Dispos. 2005 Jan;33(1):94-101. Epub 2004 Oct 8. Kobayashi D, Ieiri I, Hirota T, Takane H, Maegawa S, Kigawa J, Suzuki H, Nanba E, Oshimura M, Terakawa N, Otsubo K, Mine K, Sugiyama Y
Functional assessment of ABCG2 (BCRP) gene polymorphisms to protein expression in human placenta.
Drug Metab Dispos. 2005 Jan;33(1):94-101. Epub 2004 Oct 8., [PMID:15475413]

Abstract [show]
The aim of the present study was to assess the contribution of polymorphisms in the breast cancer resistance protein/ATP-binding cassette transporter G2 (BCRP/ABCG2) gene to the placental expression from a new perspective, allelic imbalance. Polymorphisms were screened by polymerase chain reaction (PCR)-single-strand conformation polymorphism analysis followed by sequencing with DNA extracted from 100 placentas. To examine whether polymorphisms of the BCRP gene correlate with the placental BCRP expression, we determined mRNA and protein levels by quantitative real-time PCR and Western blotting, respectively. In placentas, G34A (Val(12)Met) and C421A (Gln(141)Lys) were frequently observed (18-36%), but C376T, which creates a stop codon (Gln(126) stop codon), was found with an allelic frequency of 1%. The mean of the BCRP protein level was significantly lower (p < 0.05) in homozygotes for the A421 allele than in those for the C421 allele, and heterozygotes had an intermediate value. To evaluate whether the C421A polymorphism acts as a cis-element in BCRP transcription, allelic imbalance was determined using informative lymphoblasts and 56 samples of placental cDNA. In most of the placental samples we tested, the difference in expression levels between the two alleles was small, and only two samples indicated a monoallelic expression (i.e., preferential expression of one allele). These results suggest that 1) the predominant allelic expression pattern of BCRP in placental samples is biallelic, and 2) the mutation C421A is not a genetic variant acting in cis, but is considered to influence the translation efficiency.

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3 In placentas, G34A (Val12 Met) and C421A (Gln141 Lys) were frequently observed (18-36%), but C376T, which creates a stop codon (Gln126 stop codon), was found with an allelic frequency of 1%. Login to comment
110 Of these, five SNPs resulted in the following amino acid substitutions: G34A (Val12Met), C376T (Gln126stop), C421A (Gln141Lys), G1322A (Ser441Asn), and T1465C (Phe489Leu). Login to comment
112 C376T, which is associated with an amino acid substitution from Gln to a stop codon at codon 126 (Gln126stop), was detected in only two placental samples (1.0%) as TABLE 1 Genetic polymorphism in the BCRP gene in Japanese placentas (n ϭ 100) Location Positiona Reference Alleleb Variant Allele Amino Acid Substitution Genotype Frequency of Variant Allele R/R R/V V/V 5Ј-Flanking region -20445 gtctCctcc gtctTctcc 98 2 0 0.010 -20296 agctAttaa agctGttaa 80 18 2 0.110 -19781 aaaaAttat aaaaGttat 99 1 0 -19572_-19569 ctcaCTCAcaaa ctca--caaa 60 33 7 0.235 Exon 2 34 cccaGtgtc cccaAtgtc Val12Met 70 24 6 0.180 Intron 2 203 ϩ 16 tttaAttta tttaGttta 70 24 6 0.180 Intron 3 263 ϩ 10 tataAgaga tataGgaga 85 14 1 0.080 263 ϩ 72 ttttGtgtg ttttTGtgtg 99 1 0 0.005 Exon 4 376 ggtaCaagt ggtaTaagt Gln126stop 98 2 0 0.010 Exon 5 421 cttaCagtt cttaAagtt Gln141Lys 42 45 13 0.355 Intron 5 532-16 ttatAatat ttatGatat 99 1 0 0.005 Exon 9 1098 aggaGatca aggaAatca Synonymous 98 2 0 0.010 Intron 10 1277 ϩ 95 atagTgtaa atagAgtaa 97 3 0 0.015 Exon 11 1322 agcaGtgtt agcaAtgtt Ser441Asn 99 1 0 0.005 Intron 11 1367 ϩ 20 ttctAggaa ttctGggaa 71 25 4 0.165 Exon 12 1465 tataTttac tataCttac Phe489Leu 99 1 0 0.005 Intron 12 1492 ϩ 49 ctatGggtg ctatCggtg 44 45 11 0.335 Exon 13 1515 atgcCttct atgc-ttct Phe506Ser 99 1 0 0.005 Phe507Leu Val508Leu Met509stop Intron 13 1648-42 tgaaAttac tgaaTttac 99 1 0 0.005 1648-21 gactCttag gactTttag 71 25 4 0.165 Intron 14 1738-46 tcttAaaat tcttGaaat 24 52 24 0.500 3Ј-UTR 2332 cttcAgtct cttcTAgtct 86 14 0 0.070 2364 tgccAttat tgccCttat 99 1 0 0.005 2512 agaaCttac agaaTttac 99 1 0 0.005 R, reference allele; V, variant allele. Login to comment
148 SNP Amino Acid Change Population Genotypes Frequency of Variant Allele R/R R/V V/V G34A Val12Met Japanese (n ϭ 120) 81 37 2 0.17 (0.12-0.22) Caucasian (n ϭ 150) 139 11 0 0.04 (0.02-0.06) African American (n ϭ 150) 132 17 1 0.06 (0.04-0.09) C376T Gln126stop Japanese (n ϭ 120) 118 2 0 0.01 (0.00-0.02) Caucasian (n ϭ 150) 150 0 0 0.00 African American (n ϭ 150) 150 0 0 0.00 C421A Gln141Lys Japanese (n ϭ 120) 61 45 14 0.30 (0.25-0.36) Caucasian (n ϭ 150) 121 25 4 0.11 (0.08-0.15) African American (n ϭ 150) 144 5 1 0.02 (0.01-0.04) R, reference allele; V, variant allele. Login to comment
169 Among the nonsynonymous polymorphisms, G34A (Val12Met) and C421A (Gln141Lys) appeared commonly in Japanese subjects, and allelic frequencies of these polymorphisms were in keeping with those of a previous report (Imai et al., 2002). Login to comment

[hide] Functional analysis of SNPs variants of BCRP/ABCG2... Pharm Res. 2004 Oct;21(10):1895-903. Kondo C, Suzuki H, Itoda M, Ozawa S, Sawada J, Kobayashi D, Ieiri I, Mine K, Ohtsubo K, Sugiyama Y
Functional analysis of SNPs variants of BCRP/ABCG2.
Pharm Res. 2004 Oct;21(10):1895-903., [PMID:15553238]

Abstract [show]
PURPOSE: The aim of the current study was to identify the effect of single nucleotide polymorphisms (SNPs) in breast cancer resistance protein (BCRP/ABCG2) on its localization, expression level, and transport activity. METHODS: The cellular localization was identified using the wild type and seven different SNP variants of BCRP (V12M, Q141K, A149P, R163K, Q166E, P269S, and S441N BCRP) after transfection of their cDNAs in plasmid vector to LLC-PK1 cells. Their expression levels and transport activities were determined using the membrane vesicles from HEK293 cells infected with the recombinant adenoviruses containing these kinds of BCRP cDNAs. RESULTS: Wild type and six different SNP variants of BCRP other than S441N BCRP were expressed on the apical membrane, whereas S441N BCRP showed intracellular localization. The expression levels of Q141K and S441N BCRP proteins were significantly lower compared with the wild type and the other five variants. Furthermore, the transport activity of E1S, DHEAS, MTX, and PAH normalized by the expression level of BCRP protein was almost the same for the wild type, V12M, Q141K, A149P, R163K, Q166E, and P269S BCRP. CONCLUSIONS: These results suggest that Q141K SNPs may associate with a lower expression level, and S441N SNPs may affect both the expression level and cellular localization. It is possible that subjects with these polymorphisms may have lower expression level of BCRP protein and, consequently, a reduced ability to export these substrates.

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3 The cellular localization was identified using the wild type and seven different SNP variants of BCRP (V12M, Q141K, A149P, R163K, Q166E, P269S, and S441N BCRP) after transfection of their cDNAs in plasmid vector to LLC-PK1 cells. Login to comment
7 The expression levels of Q141K and S441N BCRP proteins were significantly lower compared with the wild type and the other five variants. Login to comment
8 Furthermore, the transport activity of E1S, DHEAS, MTX, and PAH normalized by the expression level of BCRP protein was almost the same for the wild type, V12M, Q141K, A149P, R163K, Q166E, and P269S BCRP. Login to comment
10 These results suggest that Q141K SNPs may associate with a lower expression level, and S441N SNPs may affect both the expression level and cellular localization. Login to comment
26 On analyzing the specimens from the 100 Japanese volunteers, 7 kinds of SNPs were identified for the BCRP gene: G34A (V12M), C376T (Q376Stop), C421A (Q141K), G1098A (E366E), G1322A (S441N), T1465C (F489L), and C1515- (AFFVM505-509ASSL Stop). Login to comment
27 The allele frequencies of these SNPs are 18, 1, 36, 1, 0.5, 0.5, and 0.5%, respectively. In the 84 cell lines, 7 kinds of SNPs were identified and their frequency for G34A (V12M), C376T (Q126Stop), C421A (Q141K), G445C (A149P), G488A (R163K), C805T (P269S), and G1098A (E366E) are 22, 3, 29, 1, 0.6, 0.6 and 2%, respectively. Login to comment
29 We constructed expression systems for the wild type and SNPs variants of BCRP (V12M, Q141K, A149P, R163K, Q166E, P269S, S441N BCRP) and examined whether these SNPs variants of BCRP alter its localization, expression level, and transport activity. Login to comment
42 Using site-directed mutagenesis, SNP variants of BCRP (V12M, Q141K, A149P, R163K, Q166E, P269S and S441N BCRP) were constructed on pcDNA3.1 vector (SNPs type BCRP/pcDNA3.1). Login to comment
44 Q141K BCRP was amplified with 5Ј-CGGTGAGAGAAAACT- TAAAGTTCTCAGCAG-3Ј and `-GCTGCTGAGAACTT- TAAGTTTTCTCTCACCG-3Ј. Login to comment
52 For SNPs type BCRPs, viruses were prepared in the same way, resulting in the production of pAd-SNPs BCRP (pAd-V12M, Q141K, A149P, R163K, Q166E, P269S, and S441N BCRP). Login to comment
97 Except for two BCRP variants (Q141K and S441N BCRP), the expression levels of each BCRP SNPs were approximately the same as that of the wild-type BCRP (Fig. 2). Login to comment
98 The expression level of Q141K BCRP was approximately 30-40% of the wild-type BCRP, whereas that of S441N BCRP was much lower, and could not be determined with any accuracy (Fig. 2). Login to comment
110 Except for two SNP variants of BCRP (Q141K and S441N BCRP), the ATP-dependent uptakes per mg membrane protein of SNP variants (V12M, A149P, R163K, Q166E, P269S BCRP) were similar to that of the wild-type BCRP (Fig. 3a). Login to comment
111 The uptake activity of Q141K BCRP per mg membrane protein was approximately 30-40% of the wild-type BCRP, and that of S441N was almost the same as that of the GFP-infected control cells. Login to comment
114 As shown in Fig. 3b, the transport activity of other SNP variants of BCRP (V12M, Q141K, A149P, R163K, Q166E, and P269S BCRP) was almost identical to that of the wild-type BCRP. Login to comment
115 As far as V12M and Q141K BCRP were concerned, these have a high allele frequency in Japanese and we determined the kinetic parameters for the transport of [3 H]E1S. Login to comment
116 As shown in Fig. 4, the ATP-dependent uptake of [3 H]E1S was saturable, and the Km values were 11.6 ± 4.79, 9.07 ± 1.52, and 14.0 ± 7.27 ␮M, and the Vmax values were 13.3 ± 3.3, 13.5 ± 1.29, and 4.57 ± 1.58 nmol min-1 mg-1 protein, for the wild type, V12M, and Q141K BCRP, respectively. In addition to [3 H]E1S, the transport of other BCRP substrates was examined. Login to comment
120 Figure 5a shows the ATP-dependent uptake of DHEAS, PAH, and MTX per mg membrane protein for the wild-type and SNPs BCRP (V12M, Q141K, A149P, R163K, Q166E, P269S, and S441N BCRP). Login to comment
121 Although Q141K BCRP exhibited a lower activity than wild type BCRP, no significant transport was observed for S441N BCRP (Fig. 5a). Login to comment
133 Q141K BCRP showed a lower expression level, which is approximately 30-40% of that of the wild type. Login to comment
134 This result is consistent with the previous report that the transfection of Q141K BCRP cDNA to PA317 cells and KB-3-1 human cells also resulted in lower expression levels (19). Login to comment
137 Although more detailed analysis is required to clarify the mechanism governing the reduced protein expression of Q141K BCRP, immunohistochemical staining revealed that this variant is expressed on the plasma membrane and, therefore, the altered cellular localization may not be related to the reduced protein level. Login to comment
139 In addition, in the human intestine, the expression of BCRP mRNA was similar in subjects with wild type and Q141K BCRP genes, whereas there was no significant correlation between the protein and mRNA levels for BCRP (20). Login to comment
140 It has also been suggested that linkage disequilibrium is present between a CTCA deletion in the 5Ј-flanking region (g-19572-19569) and Q141K in Swedish subjects (21). Login to comment
162 For these compounds, our results indicated that the transport activity per BCRP molecule for 6 kinds of SNP variants (V12M, A149P, R163K, Q166E, P269S, and also Q141K BCRP) is almost the same as that of the wild type BCRP (Figs. Login to comment
173 Saturation of [3 H]E1S transport was determined for the wild-type, V12M, and Q141K BCRP. Login to comment
180 Furthermore, the Km values for E1S were similar between the wild type, V12M and Q141K BCRP (Fig. 4). Login to comment
181 It has been reported that there is a marked ethnic difference in the frequency of Q141K SNPs, and this is the prevalent allele in the Japanese population (19,21). Login to comment
183 In particular, the frequency of the Q141K variant was 29-36% in the Japanese, whereas the frequency in 26 Caucasians subjects was only 8%. Login to comment
188 Since pheophorbide a is also transported by human BCRP (10), it is likely that Q141K and S441N SNPs may be involved in the phototoxicity and protoporphyria induced by the intake of chlorophyll. Login to comment
190 These data suggest that the ability to protect stem cells from some genotoxic xenobiotics might be lower in subjects who have Q141K and S441N SNPs in BCRP gene. Login to comment
197 Since BCRP also transports SN-38 (28), it is possible that subjects who have Q141K or S441N SNPs variants of BCRP are more sensitive to SN-38. Login to comment
198 In conclusion, we have shown that two kinds of SNP variants of BCRP (Q141K and S441N BCRP) are associated with the reduced expression. Login to comment
200 Since the allele frequency of Q141K in Japanese subjects is as high as 29-36%, it is possible that the interindividual variations in in vivo disposition of BCRP substrates may result from the genotype of BCRP. Login to comment

[hide] Linkage disequilibrium and haplotype architecture ... Ann Hum Genet. 2004 Nov;68(Pt 6):563-73. Wang H, Hao B, Zhou K, Chen X, Wu S, Zhou G, Zhu Y, He F
Linkage disequilibrium and haplotype architecture for two ABC transporter genes (ABCC1 and ABCG2) in Chinese population: implications for pharmacogenomic association studies.
Ann Hum Genet. 2004 Nov;68(Pt 6):563-73., [PMID:15598215]

Abstract [show]
Information about linkage disequilibrium (LD) patterns and haplotype structures for candidate genes is instructive for the design and analysis of genetic association studies for complex diseases and drug response. ABCC1 and ABCG2 are genes coding for two multidrug resistance (MDR) associated transporters; they are also related to some pathophysiological traits. To pinpoint the LD profiles of these MDR genes in Chinese, we systemically screened 27 unrelated individuals for single nucleotide polymorphisms (SNPs) in the coding and regulatory regions of these genes, and thereby characterized their haplotype structures. Despite marked variations in haplotype diversity, LD pattern and intragenic recombination intensity between the two genes, both loci could be partitioned into several LD blocks, in which a modest number of haplotypes accounted for a high fraction of the sampled chromosomes. We concluded that each locus has its own genomic LD profile, but that they still share a common segmental LD architecture with low haplotype diversity. Our data will benefit genetic association studies of complex traits and drug response possibly related to these genes.

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57 SNP Nucleotide sequence Minor allele dbSNP ID effect position (major/minor) frequency (%) ABCC1 1 5`FR/-1862 gacccG/Aggcca 44.4 2 5`FR/-1830 atcctA/Gtctac 1.9 3 5`FR/-1680 gaggaG/Aaaaag 1.9 4 5`FR/-471 cggatA/Gctgtc 7.4 5 E2/218 caaaaC/Tcaaaa 3.7 Thr73Ile 6 I2/-26 gttgtG/Aggggg 1.9 rs8187842 7 I3/-66 ctgggT/Cgacaa 37.0 rs4148337 8 I7/+54 ccactC/Actgtg 9.3 rs903880 9 I7/+64 ggcctC/Gaatcc 48.1 rs246232 10 E8/816 cagccG/Agtgaa 1.9 wobble 11 E8/825 aaggtT/Cgtgta 38.9 rs246221 wobble 12 E9/1062 gtgaaT/Cgacac 35.2 rs35587 wobble 13 I9/+8 aggggA/Gcgctg 37.0 rs35588 14 I12/-37 cactcA/Ggggca 20.4 rs35604 15 E13/1684 tggccT/Ctgtgc 20.4 rs35605 wobble 16 I13/+105 ccggtC/Tgggct 20.4 rs35606 17 I14/+105 ccagcC/Tgcttg 1.9 18 I15/+627 gctgtA/Gtttta 25.8 rs35628 19 I15/+669 aatctG/Ttagaa 7.4* rs4148353 20 I15/-967 ctttcT/Ggctgt 37.0 rs152029 21 E16/2007 atcccC/Tgaagg 3.7 rs2301666 wobble 22 E17/2168 tctccG/Aagaaa 5.6 rs4148356 Arg723Gln 23 I18/-30 gcactG/Cacgtg 16.7 rs2074087 24 I22/+62 aattaT/Ctccct 27.8 rs3887893 25 I22/-43 gtcagC/Ttccct 3.7 26 E27/3915 gaggaC/Tctgga 1.9 wobble 27 E28/4002 aagtcG/Atccct 11.1 rs2239330 wobble 28 I28/-35 tcagcA/Gtgaca 27.8 rs212087 29 I30/+30 gcacaG/Atggcc 29.6 rs212088 30 3`UTR/+801 accccC/Gactcc 33.3 rs129081 noncoding 31 3`UTR/+866 tactgT/Atccca 14.8 rs212090 noncoding 32 3`FR/+1513 gttctT/Ctaagg 27.8 ABCG2 1 5`UTR/-407 cgcagC/Tgcctc 1.9 2 5`UTR/-376 ggggaG/Acgctc 1.9 3 E2/34 tcccaG/Atgtca 20.4 rs2231137 Val12Met 4 I2/+36 ttttaA/Gtttac 25.9 rs4148152 5 I3/+10 gtataA/Ggagag 20.4 rs2231138 6 E5/421 acttaC/Agttct 22.2 rs2231142 Gln141Lys 7 E7/805 acgggC/Tctgct 3.7 Pro269Ser 8 I9/-126 agccaT/Gtgagt 7.4 9 I11/+20 gttctA/Gggaac 31.5 rs2231153 10 I12/+49 cctatG/Tggtga 16.7 rs2231156 11 I13/+40 tgtttT/Ctttcc 24.1 rs2231157 12 I13/-21 tgactC/Tttagt 29.6 rs2231162 13 I14/-46 ttcttG/Aaaatt 48.1 rs2725267 SNPs in specific regions, i.e. 5`flanking region (5`FR), 5`untranslated region (5`UTR), intron (I), exon (E), 3`UTR, and 3`FR, are presented as region/+(-): for 5`FR and 5`UTR, n nucleotides upstream (-) from the translation initiation site; for 3`UTR and 3`FR, n nt downstream (+) from the third base of stop codon; for coding regions, n corresponds to positions of their cDNA with the first base of start codon set to 1; and for introns, n nt upstream (-) from 3` site or downstream (+) from 5` site of introns. Login to comment

[hide] Eight novel single nucleotide polymorphisms in ABC... Drug Metab Pharmacokinet. 2003;18(3):212-7. Itoda M, Saito Y, Shirao K, Minami H, Ohtsu A, Yoshida T, Saijo N, Suzuki H, Sugiyama Y, Ozawa S, Sawada J
Eight novel single nucleotide polymorphisms in ABCG2/BCRP in Japanese cancer patients administered irinotacan.
Drug Metab Pharmacokinet. 2003;18(3):212-7., [PMID:15618737]

Abstract [show]
Eight novel single nucleotide polymorphisms (SNPs) were found in the gene encoding the ATP-binding cassette transporter, ABCG2/BCRP, from 60 Japanese individuals administered the anti-cancer drug irinotecan. The detected SNPs were as follows: 1) SNP, MPJ6_AG2005 (IVS2-93T>C); Gene Name, ABCG2; Accession Number, NT_006204; 2) SNP, MPJ6_AG2007 (IVS3+71_72 insT); Gene Name, ABCG2; Accession Number, NT_006204; 3) SNP, MPJ6_AG2012 (IVS6-204C>T); Gene Name, ABCG2; Accession Number, NT_006204; 4) SNP, MPJ6_AG2015 (at nucleotide 1098G>A (exon 9) from the A of the translation initiation codon); Gene Name, ABCG2; Accession Number, NT_006204; 5) SNP, MPJ6_AG2017 (1291T>C (exon 11)); Gene Name, ABCG2; Accession Number, NT_006204; 6) SNP, MPJ6_AG2019 (IVS11-135G>A); Gene Name, ABCG2; Accession Number, NT_006204; 7) SNP, MPJ6_AG2020 (1465T>C (exon 12)); Gene Name, ABCG2; Accession Number, NT_006204; 8) SNP, MPJ6_AG2023 (IVS13+65T>G); Gene Name, ABCG2; Accession Number, NT_006204.MPJ6_AG2015 was a synonymous SNP (E366E). MPJ6_AG2017 and MPJ6_AG2020 resulted in amino acid alterations, F431L and F489L, respectively.

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46 Information on ABCG2WBCRP single nucleotide polymorphisms (SNPs) has been published.8-10) Five naturally occurring nonsynonymous SNPs have been reported in Japanese and Caucasians: V12M, Q126Stop, Q141K, I206L, and N590Y.8-10) SNP Q126Stop was found in 3 out of 124 healthy Japanese subjects.9) Since it may be possible that ABCG2WBCRP polymorphisms are associated with the eŠectiveness and adverse eŠects of irinotecan, ABCG2WBCRP exons and their ‰anking regions were sequenced to identify Japanese speciˆc SNPs. Login to comment
126 Masaya ITODA, et al.SNP18 (216) Novel ABCG2 SNPs SNP19 (217) tion of ABCG2WBCRP haplotypes in conjunction with other frequently found SNPs, including non-synonymous ones (34GÀA (V12M, SNP frequency 19z) and 421CÀA (Q141K, SNP frequency 33z)) in the Japanese population. Login to comment
128 MPJ6äAG2012 was closely associated with the known SNP, IVS1-99GÀA (rs1584481, ssj0001922), but not with the SNPs, 34GÀA (V12M) and 421CÀA (Q141K). Login to comment

[hide] ABCG2-mediated transport of photosensitizers: pote... Cancer Biol Ther. 2005 Feb;4(2):187-94. Epub 2005 Feb 8. Robey RW, Steadman K, Polgar O, Bates SE
ABCG2-mediated transport of photosensitizers: potential impact on photodynamic therapy.
Cancer Biol Ther. 2005 Feb;4(2):187-94. Epub 2005 Feb 8., [PMID:15684613]

Abstract [show]
In photodynamic therapy (PDT), a tumor-selective photosensitizer is administered followed by activation of the photosensitizer by exposure to a light source of a given wavelength. This, in turn, generates reactive oxygen species that induce cellular apoptosis and necrosis in tumor tissue. Based on our earlier finding that the photosensitizer pheophorbide a is an ABCG2 substrate, we explored the ability of ABCG2 to transport photosensitizers with a structure similar to that of pheophorbide a. ABCG2-overexpressing NCI-H1650 MX50 bronchoalveolar carcinoma cells were found to have reduced intracellular accumulation of pyropheophorbide a methyl ester and chlorin e6 compared to parental cells as measured by flow cytometry. The ABCG2 inhibitor fumitremorgin C was found to abrogate ABCG2-mediated transport. Intracellular fluorescence of hematoporphyrin IX, meso-tetra(3-hydroxyphenyl)porphyrin, and meso-tetra(3-hydroxyphenyl)chlorin was not substantially affected by ABCG2. ABCG2-overexpressing cells also displayed decreased intracellular fluorescence of protoporphyrin IX generated by exogenous application of 5-aminolevulinic acid. Mutations at amino acid 482 in the ABCG2 protein known to affect substrate specificity were not found to impact transport of the photosensitizers. In cytotoxicity assays, ABCG2-transfected HEK-293 cells were 11-fold, 30-fold, 4-fold, and >7-fold resistant to PDT with pheophorbide a, pyropheophorbide a methyl ester, chlorin e6, and 5-aminolevulinic acid, respectively. ABCG2-transfected cells were not resistant to PDT with meso-tetra(3-hydroxyphenyl) chlorin. Neither multidrug resistance-associated protein 1 expression nor P-glycoprotein expression appreciably decreased the intracellular fluorescence of any of the photosensitizers examined as determined by flow cytometry. The results presented here implicate ABCG2 as a possible cause for cellular resistance to photodynamic therapy.

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148 We and others described a 421C>A nonsynonymous single nucleotide polymorphism (glutamine to lysine, Q141K, amino acid change) that impairs the transport activity of the protein.40-42 Cells expressing Q141K ABCG2 were shown to be more sensitive to the ABCG2 substrate drugs mitoxantrone, SN-38, topotecan and diflomotecan compared to cells expressing comparable cell surface levels of wild-type protein.42 Expression of the Q141K SNP has also been linked to higher serum levels of diflomotecan in patients receiving the drug orally, suggesting the Q141K polymorphism may affect the pharmacokinetics of ABCG2 substrate drugs.43 In light of the affect of the Q141K polymoprhism on the disposition of ABCG2 substrate drugs, it is tempting to speculate that the Q141K polymorphism may play a role in this observed extended sensitivity due to decreased transporter activity of the resulting ABCG2 protein. Login to comment
154 These results suggest that PDT with photosensitizers that are ABCG2 substrates may be rendered less effective in ABCG2 expressing cancers and that the Q141K polymorphism in ABCG2 may explain increases in the duration of photosensitivity in some patients. Login to comment

[hide] Functional analysis of the human variants of breas... Drug Metab Dispos. 2005 Jun;33(6):697-705. Epub 2005 Mar 2. Vethanayagam RR, Wang H, Gupta A, Zhang Y, Lewis F, Unadkat JD, Mao Q
Functional analysis of the human variants of breast cancer resistance protein: I206L, N590Y, and D620N.
Drug Metab Dispos. 2005 Jun;33(6):697-705. Epub 2005 Mar 2., [PMID:15743976]

Abstract [show]
Previous studies have shown that the V12M and Q141K variants of breast cancer resistance protein (BCRP) can affect expression and function of the transporter. In this study, the effects of the I206L, N590Y, and D620N variants on protein expression, plasma membrane localization, and transport activity of BCRP were investigated. Wild-type BCRP and the three variants were stably expressed in human embryonic kidney (HEK) cells. Confocal microscopy analysis showed that the three variants were predominantly routed to the plasma membrane of HEK cells. The expression level of I206L in the plasma membrane was approximately 45% of that of wild-type protein, whereas the N590Y and D620N levels were increased approximately 3.6-fold and 2.4-fold, respectively, as determined by immunoblotting. All three variants transported mitoxantrone, pheophorbide a, and BODIPY FL-prazosin. After normalization for differences in BCRP expression, I206L, N590Y, and D620N exhibited approximately 2-fold, 0.3-fold, and 0.5-fold wild-type efflux activities, respectively. The variants also conferred resistance to mitoxantrone and topotecan. Mitoxantrone and topotecan resistance by I206L and N590Y was approximately 2-fold and 0.3-fold of the wild-type BCRP resistance levels, respectively. Although D620N conferred a topotecan resistance similar to that of the wild-type protein, its level of mitoxantrone resistance was decreased by 50%. After normalization to BCRP expression levels, ATPase activities of I206L were not significantly different from those of wild-type protein, whereas N590Y and D620N exhibited approximately 30% and 50% of wild-type ATPase activities, respectively. These results suggest that I206L has the lowest protein expression and the highest activity, whereas N590Y and D620N display higher expression and lower activity, relative to wild-type BCRP.

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20 Two variants, V12M and Q141K, occurred at particularly high allele frequencies in Chinese and Japanese populations (30-60%) and at relatively lower allele frequencies in white people and African-Americans (5-26%) (Zamber et al., 2003). Login to comment
28 6 Copyright (c) 2005 by The American Society for Pharmacology and Experimental Therapeutics 3657/3131752 DMD 33:697-705, 2005 Printed in U.S.A. 697 Q141K seems to be associated with reduced protein expression compared with wild-type BCRP in mammalian expression systems (Imai et al., 2002; Kondo et al., 2004). Login to comment
207 Imai et al. (2002) demonstrated that Q141K was expressed at a lower level in transfected murine fibroblast PA317 cells and conferred lower levels of drug resistance compared with wild-type BCRP, whereas V12M exhibited an expression level and drug resistance profiles very similar to those of wild-type protein (Imai et al., 2002). Login to comment
208 Another study (Mizuarai et al., 2004) showed that V12M and Q141K were expressed in the polarized LLC-PK1 porcine kidney cells at levels comparable to that of wild-type protein; however, both V12M and Q141K conferred significantly lower drug resistance than wild-type protein, accompanied with increased drug accumulation and decreased drug efflux. Login to comment
209 Further analysis of the mechanism of transport dysfunction revealed that the apical membrane localization of V12M was disrupted, whereas the expression and apical membrane localization of Q141K were not affected; however, the ATPase activity of Q141K was decreased. Login to comment
210 A recent study illustrated that V12M was expressed in HEK cells at levels similar to that of wild-type BCRP, whereas the Q141K level was significantly decreased compared with wild-type protein; however, the transport activity normalized by the BCRP expression was almost the same for V12M, Q141K and wild-type BCRP (Kondo et al., 2004). Login to comment
212 However, the lower Q141K expression appears to be consistent with in vivo data. Login to comment
213 When 99 Japanese placental samples were analyzed, the subjects who were homozygous for Q141K were found to express significantly lower amounts of BCRP (Kobayashi et al., 2005). Login to comment
214 The lower protein expression and/or transport dysfunction of Q141K could affect drug disposition. Login to comment
215 A recent clinical study has shown that Q141K is associated with significant changes in pharmacokinetics of diflomotecan (Sparreboom et al., 2004). Login to comment
216 In five patients heterozygous for Q141K, the plasma levels of diflomotecan after intravenous administration were 299% of those in 15 patients expressing wild-type BCRP. Login to comment
269 These data suggest that, whereas ATPase activities of BCRP could be affected by mutations in the NBD (e.g., Q141K and I206L), mutations in other regions including the TM segments (e.g., G406L/G410L) and extracellular loops (e.g., N590Y and D620N) can also influence ATP hydrolysis. Login to comment

[hide] Multidrug transporter ABCG2 prevents tumor cell de... Cancer Res. 2005 Mar 1;65(5):1770-7. Elkind NB, Szentpetery Z, Apati A, Ozvegy-Laczka C, Varady G, Ujhelly O, Szabo K, Homolya L, Varadi A, Buday L, Keri G, Nemet K, Sarkadi B
Multidrug transporter ABCG2 prevents tumor cell death induced by the epidermal growth factor receptor inhibitor Iressa (ZD1839, Gefitinib).
Cancer Res. 2005 Mar 1;65(5):1770-7., 2005-03-01 [PMID:15753373]

Abstract [show]
Iressa (ZD1839, Gefitinib), used in clinics to treat non-small cell lung cancer patients, is a tyrosine kinase receptor inhibitor that leads to specific decoupling of epidermal growth factor receptor (EGFR) signaling. Recent data indicate that Iressa is especially effective in tumors with certain EGFR mutations; however, a subset of these tumors does not respond to Iressa. In addition, certain populations have an elevated risk of side effects during Iressa treatment. The human ABCG2 (BCRP/MXR/ABCP) transporter causes cancer drug resistance by actively extruding a variety of cytotoxic drugs, and it functions physiologically to protect our tissues from xenobiotics. Importantly, ABCG2 modifies absorption, distribution, and toxicity of several pharmacologic agents. Previously, we showed that ABCG2 displays a high-affinity interaction with several tyrosine kinase receptor inhibitors, including Iressa. Here, we show that the expression of ABCG2, but not its nonfunctional mutant, protects the EGFR signaling-dependent A431 tumor cells from death on exposure to Iressa. This protection is reversed by the ABCG2-specific inhibitor, Ko143. These data, reinforced with cell biology and biochemical experiments, strongly suggest that ABCG2 can actively pump Iressa. Therefore, variable expression and polymorphisms of ABCG2 may significantly modify the antitumor effect as well as the absorption and tissue distribution of Iressa.

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320 Imai Y, Nakane M, Kage K, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment
317 Imai Y, Nakane M, Kage K, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Pharmacogenetic profiling across the irinotecan pa... Br J Clin Pharmacol. 2005 Apr;59(4):415-24. Zhou Q, Sparreboom A, Tan EH, Cheung YB, Lee A, Poon D, Lee EJ, Chowbay B
Pharmacogenetic profiling across the irinotecan pathway in Asian patients with cancer.
Br J Clin Pharmacol. 2005 Apr;59(4):415-24., [PMID:15801936]

Abstract [show]
AIMS: The aim of this exploratory study was to investigate associations between irinotecan pharmacokinetic parameters and allelic variants in genes encoding for drug transporters and drug metabolizing enzymes that are involved in irinotecan disposition in Asian patients with cancer. METHODS: Irinotecan was administered at 100 mg m(-2) over 90 min on a weekly schedule to 29 nasopharyngeal carcinoma patients and pharmacokinetic analysis was performed during the first cycle. All patients were genotyped for allelic variants in genes encoding drug metabolizing enzymes (CYP3A4, CYP3A5, UGT1A1) and drug transporters (ABCB1, ABCC2 and ABCG2) that are involved in irinotecan disposition. RESULTS: Of the six candidate genes that were analyzed, 11 genetic variants were found. Significant genotypic-phenotypic associations were apparent only for transporter genes. The C(max) of irinotecan was significantly lower in patients carrying the CC genotype at exon 26 of the ABCB1 gene compared with those harbouring at least one variant allele (P = 0.047). Patients harbouring the wild type ABCG2 CTCA genotype were associated with significantly higher values for relative extent of conversion (REC) of irinotecan to SN-38 compared with patients carrying at least one deletion CTCA allele (P = 0.019). CONCLUSIONS: The present exploratory study shows that genetic polymorphisms in drug transporter genes, particularly in ABCB1 and ABCG2 genes, may be important in influencing the pharmacokinetics of irinotecan and its metabolites. The predictive value of the identified allelic variants in the ABCG2 and ABCB1 genes on irinotecan disposition should be further investigated in a larger patient population as well as in other ethnic populations.

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236 29 Imai Y, Nakane M, Kage K, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low level drug resistance. Login to comment

[hide] Single nucleotide polymorphisms modify the transpo... Cancer Chemother Pharmacol. 2005 Aug;56(2):161-72. Epub 2005 Apr 19. Morisaki K, Robey RW, Ozvegy-Laczka C, Honjo Y, Polgar O, Steadman K, Sarkadi B, Bates SE
Single nucleotide polymorphisms modify the transporter activity of ABCG2.
Cancer Chemother Pharmacol. 2005 Aug;56(2):161-72. Epub 2005 Apr 19., [PMID:15838659]

Abstract [show]
Single nucleotide polymorphism (SNP) analyses of the ABCG2 gene have revealed three nonsynonymous SNPs resulting in the amino acid changes at V12M, Q141K and D620N. To determine whether the SNPs have an effect on drug transport, human embryonic kidney cells (HEK-293) were stably transfected with full length ABCG2 coding wild-type or SNP variants of ABCG2. In 4-day cytotoxicity assays with mitoxantrone, topotecan, SN-38 or diflomotecan, cells transfected with wild-type R482 ABCG2 showed IC50 values up to 1.2-fold to 5-fold higher than cells expressing comparable levels of Q141K ABCG2, suggesting that the Q141K SNP affects drug transport. FTC-inhibitable mitoxantrone efflux normalized to ABCG2 surface expression as assayed by the anti-ABCG2 antibody 5D3 was significantly lower in cells transfected with Q141K ABCG2 than in those transfected with wild-type R482 ABCG2 (P = 0.0048). Values for V12M and D620N ABCG2 were comparable to those for wild-type R482 ABCG2. The vanadate-sensitive ATPase activity of ABCG2 was assayed in Sf9 insect cells infected with wild-type or SNP variants of ABCG2. Basal ATPase activity in cells transfected with Q141K ABCG2 was 1.8-fold lower than in cells transfected with wild-type ABCG2, but was comparable among cells expressing wild-type, V12M or D620N ABCG2. Confocal studies of ABCG2 localization revealed higher intracellular staining in the Q141K transfectants than in cells transfected with wild-type or V12M ABCG2. Decreased transport of Hoechst 33342 was observed in Sf9 cells expressing V12M ABCG2; however, this was not true in HEK-293 cells expressing V12M ABCG2. These results suggest that the Q141K SNP affects the transport efficiency of ABCG2 and may result in altered pharmacokinetics or drug-resistance profiles in clinical oncology.

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0 ORIGINAL ARTICLE Kuniaki Morisaki Æ Robert W. Robey Csilla O¨ zvegy-Laczka Æ Yasumasa Honjo Orsolya Polgar Æ Kenneth Steadman Bala´ zs Sarkadi Æ Susan E. Bates Single nucleotide polymorphisms modify the transporter activity of ABCG2 Received: 21 July 2004 / Accepted: 1 October 2004 / Published online: 19 April 2005 Ó Springer-Verlag 2005 Abstract Single nucleotide polymorphism (SNP) analyses of the ABCG2 gene have revealed three nonsynonymous SNPs resulting in the amino acid changes at V12M, Q141K and D620N. Login to comment
2 In 4-day cytotoxicity assays with mitoxantrone, topotecan, SN-38 or diflomotecan, cells transfected with wild-type R482 ABCG2 showed IC50 values up to 1.2-fold to 5-fold higher than cells expressing comparable levels of Q141K ABCG2, suggesting that the Q141K SNP affects drug transport. Login to comment
3 FTC-inhibitable mitoxantrone efflux normalized to ABCG2 surface expression as assayed by the anti-ABCG2 antibody 5D3 was significantly lower in cells transfected with Q141K ABCG2 than in those transfected with wild-type R482 ABCG2 (P=0.0048). Login to comment
6 Basal ATPase activity in cells transfected with Q141K ABCG2 was 1.8-fold lower than in cells transfected with wild-type ABCG2, but was comparable among cells expressing wild-type, V12M or D620N ABCG2. Login to comment
7 Confocal studies of ABCG2 localization revealed higher intracellular staining in the Q141K transfectants than in cells transfected with wild-type or V12M ABCG2. Login to comment
9 These results suggest that the Q141K SNP affects the transport efficiency of ABCG2 and may result in altered pharmacokinetics or drug-resistance profiles in clinical oncology. Login to comment
30 We previously sequenced the ABCG2 gene in 90 genomic DNA samples representing a global genetic diversity and identified three nonsynonymous SNPs -34G fi A, substituting a valine for methionine (V12M); 421C fi A, substituting a glutamine for lysine (Q141K); and 1858C fi A, substituting an aspartic acid for asparagine (D620N)-in the coding region of ABCG2 [18]. Login to comment
33 In a study of SNPs in ABCG2 in the general Japanese population, the V12M and Q141K SNPs were observed at higher allelic frequencies (17.2% and 26.6%, respectively) [19]. Login to comment
34 Similarly, in other studies, the V12M and Q141K SNPs have also been found to be the most frequent polymorphisms in various ethnic and racial groups including Caucasian, Asian, and Swedish populations [3, 48]. Login to comment
37 containing full-length ABCG2 encoding wild-type (R482), mutant (R482T, R482G), or SNP variants (V12M, Q141K, or D620N) of ABCG2. Login to comment
98 Clones were initially screened using the anti-ABCG2 antibody 5D3 and, from the positive clones obtained, 12 clones transfected with V12M, Q141K, D620N, or 1_11delV12M were selected for further study: V12M-12, -13 and -14; Q141K-5, -8, -13 and -16; D620N-2, -3 and -23; and 1_11delV12M-2 and -8. Login to comment
101 Although Northern blot analysis demonstrated higher expression of ABCG2 mRNA in wild-type (482R) ABCG2-transfected clones than in V12M-13 and Q141K-8 clones, immunoblot analysis showed generally comparable (within two- to threefold) ABCG2 protein expression in 482R, V12M-13, and Q141K-8 transfectants. Login to comment
103 Differential resistance among cells transfected with wild-type, V12M and Q141K ABCG2 To determine whether the SNPs affected the transport activity of ABCG2, 4-day cytotoxicity assays were performed with the ABCG2 substrates mitoxantrone, topotecan, SN-38, and diflomotecan (BN80915) on ABCG2-transfected HEK-293 cells expressing comparable amounts of ABCG2. Login to comment
105 Comparable surface expression of ABCG2 in the 482R-2, Q141K-5, Q141K-8 and V12M-13 clones was confirmed using the 5D3 antibody, as shown in Fig. 2a. Login to comment
108 a Northern blot analysis of ABCG2 expression in representative HEK-293 cells transfected with wild-type, V12M, Q141K, or D620N ABCG2. Login to comment
114 Except for the Q141K-5 clone with diflomotecan, P values for this comparison were <0.05. Login to comment
116 The V12M-13 and 482R-2 clones were comparably resistant to mitoxantrone, topotecan, SN-38 and diflomotecan, whereas the Q141K-5 and -8 clones had IC50 values that were 3-fold to 5-fold lower for mitoxantrone and topotecan, 2-fold to 3.4-fold lower for SN-38, and 1.2-fold to 2.3-fold lower for diflomotecan. Login to comment
118 These results suggest that the Q141K SNP has functional consequences in the resulting ABCG2 protein. Login to comment
119 Correlation between FTC-inhibitable mitoxantrone efflux and surface expression in ABCG2 transfectants To confirm the impaired transport of mitoxantrone in cells expressing Q141K ABCG2, we selected two to six Fig. 2 Impact of SNPs on ABCG2-mediated resistance. Login to comment
121 b Cytotoxicity assays were performed with mitoxantrone, topotecan, SN-38, or diflomotecan on HEK-293 cells transfected with empty vector (filled circles), or the 482R-2 (open circles), V12M-13 (filled triangles), Q141K-5 (open squares), and Q141K-8 (hatched squares) clones from a. Login to comment
122 Representative results are shown clones each of ABCG2-transfected cells expressing R482, R482T, R482G, V12M, Q141K or D620N ABCG2. Login to comment
133 Efflux and expression values for cells transfected with V12M and D620N ABCG2 fell close to the line, while values for cells transfected with Q141K ABCG2 fell predominantly below the line. Login to comment
135 Among the transfectants, Q141K variants showed significantly lower values compared to the transfectants with wild-type ABCG2 and the other SNP variants, V12M and D620N (P=0.0048, 0.0005, and 0.0126, respectively), suggesting that Q141K ABCG2 transports mitoxantrone less efficiently than wild-type ABCG2. Login to comment
140 We next examined the ATPase activity of V12M, Q141K, and D620N variants using this system. Login to comment
143 Despite similar levels of ABCG2 expression, the basal ATPase activity was significantly lower in the case of the Q141K variant than in wild-type ABCG2, whereas in the other SNP variants, the ATPase activity was similar to that observed in wild-type ABCG2 (Fig. 5b). Login to comment
146 The lower basal ATPase activity in the Q141K variant persisted in the presence of mitoxantrone (Fig. 5b). Login to comment
148 At 1 lM Clone Mitoxantrone Topotecan SN-38 Diflomotecan IC50 RR IC50 RR IC50 RR IC50 RR pcDNA3-10 0.6±0.3 - 8.7±1.2 - 1.2±0.7 - 0.3±0.2 - 482R-2 34±5.4 57 275±150 32 123±68 103 1.4±0.6 5 V12M-13 44±15 73 250±71 29 100±1 83 1±0.1 3 Q141K-5 7.4±1.8 12 55±7 6 36±11 30 0.6±0.07 2 Q141K-8 10.8±5.3 18 90±14 10 66±11 55 0.9±0.1 3 Table 1 Relative resistance (RR) of ABCG2-transfected cells to ABCG2 substrates. Login to comment
151 In all cases P<0.05 except for the Q141K-5 clone with diflomotecan where P=0.09 (IC50 values are in nanomoles) Ko143, ATPase activity in membrane protein from cells expressing any of the ABCG2 proteins was almost completely abrogated. Login to comment
157 Effect of SNPs on cellular localization of ABCG2 To determine if the SNPs affected membrane localization of ABCG2, we performed immunofluorescence studies on HEK-293 cells stably transfected with wild-type, V12M or Q141K ABCG2. Login to comment
160 In contrast, HEK-293 cells expressing Q141K ABCG2 demonstrated high intracellular staining with the BXP-21 antibody, as well as cell surface staining (Fig. 6c). Login to comment
161 These results, despite the selection of clones expressing comparable levels of cell surface ABCG2, suggest impaired membrane trafficking or incorrect membrane insertion of Q141K ABCG2. Login to comment
163 To examine whether the nonsynonymous SNPs in ABCG2 affect the transport of this compound, Hoechst 33342 dye transport was measured in intact Sf9 cells expressing wild-type, V12M, Q141K, or D620N ABCG2, as well as the nonfunctional mutant, R482G/K86M. Login to comment
164 Immunoblot analysis of protein obtained from the infected cells is shown in Fig. 7a. Hoechst 33342 transport was comparable in cells expressing wild-type, Q141K or D620N ABCG2. Login to comment
172 Representative histograms for 482R-9, 482G-1, V12M-13, D620N-2, Q141K-5, and 1_11delV12M-8 are shown substrate-free medium for 60 min continuing with or without FTC. Login to comment
174 No significant difference in FTC-inhibitable Hoechst efflux was observed between cells expressing wild-type (R482), V12M or Q141K ABCG2 (Fig. 7c, right column). Login to comment
178 Discussion We and others have recently identified several polymorphisms in ABCG2, including three nonsynonymous SNPs resulting in amino acid substitution in the coding region of ABCG2: V12M, Q141K, D620N [4, 19, 20, 50]. Login to comment
180 Our results suggest that the Q141K SNP affects the transport efficiency of ABCG2. Login to comment
189 Values from the experiment in a were obtained for ABCG2-transfected HEK-293 clones expressing varying levels of 482R, R482G, R482T, V12M, Q141K, and D620N ABCG2 and a box plot was generated. Login to comment
199 Imai et al. have previously reported that the Q141K variant is associated with decreased protein expression in transfected cells and therefore results in increased sensitivity to chemotherapeutic agents [20]. Login to comment
200 In contrast, we did not observe decreased expression of ABCG2 in HEK-293 cells transfected with Q141K ABCG2. Login to comment
201 Supporting our findings, Zamber et al. have found no correlation between the Q141K SNP and level of expression of ABCG2 protein [50]. Login to comment
202 The Q141K SNP does not appear to prevent expression of the protein on the cell surface. Login to comment
204 Four-day cytotoxicity assays demonstrated that, among HEK-293 cells transfected with wild-type, V12M, or Q141K ABCG2, those expressing Q141K ABCG2 had IC50 values for mitoxantrone, topotecan, SN-38 and diflomotecan that were as much as fivefold lower than those for cells expressing comparable levels Fig. 5 ATPase activity in Sf9 insect cells infected with ABCG2-bearing baculovirus. Login to comment
205 a Immunoblot detection of human wild-type, D620N, Q141K and V12M ABCG2 expressed in Sf9 insect cells. Login to comment
206 Membranes of Sf9 cells (1.5 lg total protein from V12M/Sf9, Q141K/Sf9 and D620N/Sf9, 1.0 lg from wild-type ABCG2/Sf9, and 1.2 lg from b-galactosidase/Sf9) dissolved in disaggregation buffer were subjected to electrophoresis on 7.5% Laemmli-type gels and blotted onto PVDF membranes, followed by immunodetection with the BXP-21 antibody. b ATPase activity measured in membranes of Sf9 cells expressing the wild-type, V12M, Q141K, and D620N variants of human ABCG2. Login to comment
210 HEK-293 cells transfected with wild-type (482R-2) (a), V12M (V12M-13) (b), or Q141K (Q141K-8) (c) ABCG2 were fixed, permeabilized, and then incubated with the mouse monoclonal anti-ABCG2 antibody, BXP-21. Login to comment
212 When surface ABCG2 expression was used to normalize the FTC-inhibitable mitoxantrone efflux, we found that HEK-293 cells expressing Q141K ABCG2 transported mitoxantrone less efficiently than cells expressing wild-type, V12M or D620N ABCG2. Login to comment
213 These findings are in agreement with those of Mizuarai et al., who found that polarized LLC/ PK1 cells transfected with Q141K ABCG2 are more sensitive to mitoxantrone, topotecan and an indolocarbazole topoisomerase I inhibitor, all of which are known ABCG2 substrates [30]. Login to comment
221 The Q141K SNP has also recently been shown to significantly affect the pharmacokinetics of diflomotecan. Login to comment
223 Despite the relatively poor transport of diflomotecan by ABCG2, Sparreboom et al. have reported that, in patients heterozygous for the Q141K SNP, plasma diflomotecan levels are approximately threefold higher than in patients expressing the wild-type allele [44]. Login to comment
225 These results suggest that the Q141K SNP may alter the pharmacokinetic profile of other ABCG2 substrate drugs. Login to comment
226 Basal ATPase activity was determined to be 1.8-fold lower in membrane protein isolated from Sf9 insect cells infected with recombinant baculoviruses containing full-length Q141K ABCG2 compared to protein from cells expressing wild-type or the other SNP variant ABCG2 forms. Login to comment
227 Our results are in agreement with those of Mizuarai et al. who reported that the ATPase activity of Q141K ABCG2 is 1.3-fold lower than wild-type in the Sf9 system [30]. Login to comment
241 Consistent with other reports, we found that the Q141K polymorphism impaired the activity of the ABCG2 protein. Login to comment
244 Taken together, the results presented here support the hypothesis that the Q141K variant has the potential to alter the pharmacokinetics of ABCG2 substrates. Login to comment
245 Whether the presence of the Q141K polymorphism in clinical tumors could promote chemosensitivity is unknown, but remains a question for further study. Login to comment

[hide] Mechanisms of resistance to anticancer drugs: the ... Pharmacogenomics. 2005 Mar;6(2):115-38. Lepper ER, Nooter K, Verweij J, Acharya MR, Figg WD, Sparreboom A
Mechanisms of resistance to anticancer drugs: the role of the polymorphic ABC transporters ABCB1 and ABCG2.
Pharmacogenomics. 2005 Mar;6(2):115-38., [PMID:15882131]

Abstract [show]
ATP-binding cassette (ABC) genes play a role in the resistance of malignant cells to anticancer agents. The ABC gene products, including ABCB1 (P-glycoprotein) and ABCG2 (breast cancer-resistance protein [BCRP], mitoxantrone-resistance protein [MXR], or ABC transporter in placenta [ABCP]), are also known to influence oral absorption and disposition of a wide variety of drugs. As a result, the expression levels of these proteins in humans have important consequences for an individual's susceptibility to certain drug-induced side effects, interactions, and treatment efficacy. Naturally occurring variants in ABC transporter genes have been identified that might affect the function and expression of the protein. This review focuses on recent advances in the pharmacogenetics of the ABC transporters ABCB1 and ABCG2, and discusses potential implications of genetic variants for the chemotherapeutic treatment of cancer.

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122 The common ABCG2 C421A variant in exon 5, in which the C to A transversion results in an amino acid change of Gln to Lys at codon 141, has been associated with low ABCG2 expression levels [50,85,91-93]. Login to comment
157 Position in gene* Nucleotide‡ Region Wild-type allele Variant allele Amino acid Change -19572 to -19569 5`-Flanking region CTCA - CTCA deletion -19202 5` UTR G C -18845 5` UTR T C -18604 5` UTR A - Deletion -18482 -113 Exon 1 C T Non-coding -18398 -29 Exon 1 A G Non-coding 34 34 Exon 2 G A 12 Val to Met 71 71 Exon 2 C T 24 Ala to Val 114 114 Exon 2 T C 38 Synonymous 239 Intron 2 A G 7268 Intron 2 T C 7420 Intron 3 - T Insertion 8007 Intron 3 G A 8184 369 Exon 4 C T 123 Synonymous 8191 376 Exon 4 C T 126 Gln to Term 8825 421 Exon 5 C A 141 Gln to Lys 8862 458 Exon 5 C T 153 Thr to Met 8878 474 Exon 5 C T 158 Synonymous 8900 496 Exon 5 C G 166 Gln to Glu 18186 Intron 5 A G 18286 616 Exon 6 A C 206 Ile to Leu 18293 623 Exon 6 T C 208 Phe to Ser 21530 Intron 6 C T 21718 Intron 6 A G 21903 Intron 7 A G 24618 Intron 7 T A 26297 1098 Exon 9 G A 366 Synonymous 38389 1291 Exon 11 T C 431 Phe to Leu 38485 Intron 11 A G 40111 Intron 11 G A 40303 1425 Exon 12 A G 475 Synonymous 40322 1444 Exon 12 A G 482 Arg to Gly 40323 1445 Exon 12 G C 482 Arg to Thr 40343 1465 Exon 12 T C 489 Phe to Leu 40419 Intron 12 G T 42314 Intron 13 T G 44997 Intron 14 A G 45022 Intron 14 C T 45073 1768 Exon 15 A T 590 Asn to Tyr 47355 1858 Exon 16 G A 620 Asp to Asn 47734 2237 Exon 16 G T Non-coding 47890 2393 Exon 16 G T Non-coding 47891 2394 Exon 16 C A Non-coding ABC: ATP-binding cassette; UTR: Untranslated region. Login to comment
524 85. Imai Y, Nakane M, Kage K et al.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment
553 Morisaki K, Robey RW, Nadjem T et al.: The Q141K single-nucleotide polymorphism impacts the transporter activity of ABCG2. Login to comment

[hide] Association of the BCRP C421A polymorphism with no... Int J Cancer. 2005 Nov 10;117(3):431-4. Korenaga Y, Naito K, Okayama N, Hirata H, Suehiro Y, Hamanaka Y, Matsuyama H, Hinoda Y
Association of the BCRP C421A polymorphism with nonpapillary renal cell carcinoma.
Int J Cancer. 2005 Nov 10;117(3):431-4., 2005-11-10 [PMID:15906349]

Abstract [show]
Breast cancer resistance protein (BCRP), the second member of the ATP-binding cassette membrane transporter family, has a single nucleotide polymorphism, C421A (resulting in Q141K), that is of functional importance. Our aim was to explore the relationship between this polymorphism of the BCRP gene and the risk of renal cell carcinoma (RCC) development. For a case-control study, DNA samples from 200 nonpapillary RCC patients and 200 healthy control subjects were analyzed using the TaqMan technique. The genotypic frequencies of the BCRP C421A polymorphism were compared between RCC patients and control subjects. The frequency of the C/C genotype was significantly higher in RCC patients than in control subjects (age- and gender-adjusted OR = 1.96, 95% CI 1.32-2.93). No associations were observed between the BCRP C421A polymorphism and clinicopathologic or epidemiologic factors, including age, gender, tumor grade, stage, cigarette smoking, family history of cancer and body mass index. Carriers with the C/C genotype of the BCRP C421A polymorphism are at risk of developing nonpapillary RCC. These data suggest that BCRP is a candidate RCC susceptibility gene.

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0 Association of the BCRP C421A polymorphism with nonpapillary renal cell carcinoma Yoshihito Korenaga1 , Katsusuke Naito1*, Naoko Okayama2 , Hiroshi Hirata1 , Yutaka Suehiro2 , Yuichiro Hamanaka2 , Hideyasu Matsuyama1 and Yuji Hinoda2 1 Department of Urology, Yamaguchi University School of Medicine, Yamaguchi, Japan 2 Department of Clinical Laboratory Science, Yamaguchi University School of Medicine, Yamaguchi, Japan Breast cancer resistance protein (BCRP), the second member of the ATP-binding cassette membrane transporter family, has a single nucleotide polymorphism, C421A (resulting in Q141K), that is of functional importance. Login to comment
33 SNP analysis of the BCRP gene We used the TaqMan technique, which combines DNA amplification and genotype detection in a single assay.17 The primer set used to amplify the exon 5 SNP (C421A), which results in Q141K, was GGCACTGACGGTGAGA (forward) and CATAGTTGTTGCAAGCCGAAGAG (reverse). Login to comment
65 Our data demonstrated that the frequency of the C/C genotype of C421A (resulting in Q141K) was significantly higher in RCC patients than in controls. Login to comment

[hide] Effect of ABCG2 genotype on the oral bioavailabili... Cancer Biol Ther. 2005 Jun;4(6):650-8. Epub 2005 Jun 11. Sparreboom A, Loos WJ, Burger H, Sissung TM, Verweij J, Figg WD, Nooter K, Gelderblom H
Effect of ABCG2 genotype on the oral bioavailability of topotecan.
Cancer Biol Ther. 2005 Jun;4(6):650-8. Epub 2005 Jun 11., [PMID:15908806]

Abstract [show]
ABCG2 (BCRP/MXR/ABCP) functions as an efflux transporter for many agents, including topotecan, and the protein is expressed at high levels in the human intestine. Some individuals possess a nonsynonymous variant in the ABCG2 gene at nucleotide 421, substituting lysine for glutamine on position 141 at exon 5. The present pilot study indicates that this genotype results in a 30% reduced efflux transport of topotecan in vitro compared to the wild-type. In a preliminary fashion, the heterozygous CA allele observed in two patients was associated with a 1.34-fold increased oral bioavailability of topotecan compared to the bioavailability in ten patients with the wild-type allele (42.0% versus 31.4%; p = 0.037). It is suggested that the high frequency of the A allele in certain ethnic groups may have therapeutic implications for individuals treated with topotecan or other ABCG2 substrates.

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13 In recent years, various naturally occurring variants in ABCG2 have been identified that affect the function and/or expression of its encoded protein.4-6 In the present pilot study, we prospectively evaluated the potential functional significance of a common single-nucleotide polymorphism (SNP) in ABCG2 (421C>A; Q141K) in a cohort of cancer patients treated with topotecan. Login to comment
41 Human embryonic kidney cells (HEK293) cells transfected with pcDNA3 (HEK293/Neo, control cells), wild-type ABCG2 (HEK293/R) and an ABCG2 Q141K clone (HEK293/5)10 were provided by Dr. Susan E. Bates (National Cancer Institute, Bethesda, MD). Login to comment
75 The studied variant in ABCG2 is a SNP causing a nonsynonymous change in the protein sequence, in which a 421C to A transition at exon 5 leads to a Glutamine to Lysine amino acid substitution at codon 141 (Q141K). Login to comment
80 In vitro transport studies presented here in HEK293 cells transfected with the Q141K variant showed an increase of 30% in the intracellular accumulation of topotecan relative to wild-type ABCG2. Login to comment
84 (A) Intracellular accumulation of topotecan in human embryonic kidney cells (HEK293) transfected with pcDNA3 (HEK293/Neo, Control), wild-type ABCG2 (HEK293/R) and an ABCG2 Q141K clone (HEK293/5, Q141K-5). Login to comment
86 (B) Western blot data for ABCG2 expression in control, wild-type ABCG2 and the ABCG2 Q141K clone. Login to comment
87 A B www.landesbioscience.com Cancer Biology & Therapy e Topotecan Pharmacogenetics In support of this theory, Mizuarai6 described recently that the ATPase activity of the Q141K variant, which is localized in the ATP-binding cassette region, was reduced approximately 1.3-fold compared to the activity of the wild type ABCG2. Login to comment
89 However, our finding is not entirely conclusive as the sole contributing factor, since the increased accumulation in the Q141K variant compared to the wild type was not statistically significant (P = 0.16). Login to comment
90 Support for additional mechanisms that might be of relevance, including altered protein expression, comes from the presently performed analysis of the duodenal mucosa biopsies; these preliminary data show that the ABCG2 421C>A genotype is possibly related with reduced mRNA expression of ABCG2 and ABCB1 in intestinal enterocytes, although this seems to contradict earlier findings.22 Nonetheless, earlier data obtained in PA317 cells transfected with the Q141K mutant have indicated that intracellular topotecan accumulation was higher than that in cells with wild-type ABCG2, and this was also ascribed in that case to markedly reduced protein expression levels.23 These authors have speculated that substitution of Glutamine for Lysine at amino acid position 141 might alter the tertiary structure of ABCG2, and lead to increased susceptibility to degradation.23 Clearly, further investigation is required to resolve this issue. Login to comment

[hide] Tyrosine kinase inhibitor resistance in cancer: ro... Drug Resist Updat. 2005 Feb-Apr;8(1-2):15-26. Ozvegy-Laczka C, Cserepes J, Elkind NB, Sarkadi B
Tyrosine kinase inhibitor resistance in cancer: role of ABC multidrug transporters.
Drug Resist Updat. 2005 Feb-Apr;8(1-2):15-26., [PMID:15939339]

Abstract [show]
Recent antitumor drug research has seen the development of a large variety of tyrosine kinase inhibitors (TKIs) with increasing specificity and selectivity. These are highly promising agents for specific inhibition of malignant cell growth and metastasis. However, their therapeutic potential also depends on access to their intracellular targets, which may be significantly affected by certain ABC membrane transporters. It has been recently shown that several human multidrug transporter ABC proteins interact with specific TKIs, and the ABCG2 transporter has an especially high affinity for some of these kinase inhibitors. These results indicate that multidrug resistance protein modulation by TKIs may be an important factor in the treatment of cancer patients; moreover, the extrusion of TKIs by multidrug transporters may result in tumor cell TKI resistance. Interaction with multidrug resistance ABC transporters may also significantly modify the pharmacokinetics and toxicity of TKIs in patients.

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215 Of special interest is the ABCG2 variant Q141K (glutamine to lysine replacement). Login to comment

[hide] Involvement of BCRP (ABCG2) in the biliary excreti... Mol Pharmacol. 2005 Sep;68(3):800-7. Epub 2005 Jun 13. Hirano M, Maeda K, Matsushima S, Nozaki Y, Kusuhara H, Sugiyama Y
Involvement of BCRP (ABCG2) in the biliary excretion of pitavastatin.
Mol Pharmacol. 2005 Sep;68(3):800-7. Epub 2005 Jun 13., [PMID:15955871]

Abstract [show]
Pitavastatin, a novel potent 3-hydroxymethylglutaryl coenzyme A reductase inhibitor, is distributed selectively to the liver and excreted into bile in unchanged form in rats. We reported previously that the hepatic uptake is mainly mediated by organic anion transporting polypeptide (OATP) 1B1, whereas the biliary excretion mechanism remains to be clarified. In the present study, we investigated the role of breast cancer resistance protein (BCRP) in the biliary excretion of pitavastatin. The ATP-dependent uptake of pitavastatin by human and mouse BCRP-expressing membrane vesicles was significantly higher compared with that by control vesicles with Km values of 5.73 and 4.77 microM, respectively. The biliary excretion clearance of pitavastatin in Bcrp1-/- mice was decreased to one-tenth of that in control mice. The biliary excretion of pitavastatin was unchanged between control and Eisai hyperbilirubinemic rats, indicating a minor contribution of multidrug resistance-associated protein (Mrp) 2. This observation differs radically from that for a more hydrophilic statin, pravastatin, of which biliary excretion is largely mediated by Mrp2. These data suggest that the biliary clearance of pitavastatin can be largely accounted for by BCRP in mice. In the case of humans, transcellular transport of pitavastatin was determined in the Madin-Darby canine kidney II cells expressing OATP1B1 and human canalicular efflux transporters. A significant basal-to-apical transport of pitavastatin was observed in OATP1B1/MDR1 and OATP1B1/MRP2 double transfectants as well as OATP1B1/BCRP double transfectants, implying the involvement of multiple transporters in the biliary excretion of pitavastatin in humans. This is in contrast to a previous belief that the biliary excretion of statins is mediated mainly by MRP2.

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224 The elimination of diflomotecan from plasma has been found to be delayed in patients with frequently observed SNP in BCRP (C421A/Q141K) (Sparreboom et al., 2004). Login to comment

[hide] Breast cancer resistance protein-mediated efflux o... Cancer Res. 2005 Aug 1;65(15):6640-50. Huss WJ, Gray DR, Greenberg NM, Mohler JL, Smith GJ
Breast cancer resistance protein-mediated efflux of androgen in putative benign and malignant prostate stem cells.
Cancer Res. 2005 Aug 1;65(15):6640-50., 2005-08-01 [PMID:16061644]

Abstract [show]
Malignantly transformed stem cells represent a potential common nidus for the primary cancer and the recurrent cancer that arises after treatment failure. Putative prostate stem cells and prostate tumor stem cells in benign and malignant human prostate tissue, in primary human prostate xenografts, and in the transgenic adenocarcinoma of the mouse prostate (TRAMP) mouse model of prostate cancer, are defined by expression of breast cancer resistance protein (BCRP), a marker of pluripotent hematopoietic, muscle, and neural stem cells, and by an absence of androgen receptor (AR) protein. Inhibition of BCRP-mediated efflux of dihydrotestosterone by novobiocin or fumitremorgin C in a rat prostate progenitor cell line that expresses BCRP and AR mRNAs, but minimal AR protein, results in stabilization and nuclear translocation of AR protein, providing a mechanism for lack of AR protein in BCRP-expressing stem cells. In both benign and malignant human prostate tissue, the rare epithelial cells that express BCRP and lack AR protein are localized in the basal cell compartment, survive androgen deprivation, and maintain proliferative potential in the hypoxic, androgen-deprived prostate. Putative prostate tumor stem cells that express BCRP but not AR protein in TRAMP are the source of a BCRP-negative and AR-negative, Foxa2- and SV40Tag-expressing, transit amplifying compartment that progresses to the poorly differentiated carcinomas that arise rapidly after castration. Therefore, BCRP expression isolates prostate stem/tumor stem cells from the prostate tissue microenvironment through constitutive efflux of androgen, protecting the putative tumor stem cells from androgen deprivation, hypoxia, or adjuvant chemotherapy, and providing the nidus for recurrent prostate cancer.

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371 Clin Cancer Res 2004;10:440-8. 51. Imai Y, Nakane M, Kage K, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment
366 Clin Cancer Res 2004;10:440-8. 51. Imai Y, Nakane M, Kage K, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Breast cancer resistance protein: molecular target... Cancer Sci. 2005 Aug;96(8):457-65. Sugimoto Y, Tsukahara S, Ishikawa E, Mitsuhashi J
Breast cancer resistance protein: molecular target for anticancer drug resistance and pharmacokinetics/pharmacodynamics.
Cancer Sci. 2005 Aug;96(8):457-65., [PMID:16108826]

Abstract [show]
Breast cancer resistance protein (BCRP) is a half-molecule ATP-binding cassette transporter that forms a functional homodimer and pumps out various anticancer agents, such as 7-ethyl-10-hydroxycamptothecin, topotecan, mitoxantrone and flavopiridol, from cells. Estrogens, such as estrone and 17beta-estradiol, have been found to restore drug sensitivity levels in BCRP-transduced cells by increasing the cellular accumulation of such agents. Furthermore, synthetic estrogens, tamoxifen derivatives and phytoestrogens/flavonoids have now been identified that can effectively circumvent BCRP-mediated drug resistance. Transcellular transport experiments have shown that BCRP transports sulfated estrogens and various sulfated steroidal compounds, but not free estrogens. The kinase inhibitor gefitinib inhibited the transporter function of BCRP and reversed BCRP-mediated drug resistance both in vitro and in vivo. BCRP-transduced human epidermoid carcinoma A431 (A431/BCRP) and BCRP-transduced human non-small cell lung cancer PC-9 (PC-9/BCRP) cells showed gefitinib resistance. Physiological concentrations of estrogens (10-100 pM) reduced BCRP protein expression without affecting its mRNA levels. Two functional polymorphisms of the BCRP gene have been identified. The C376T (Q126Stop) polymorphism has a dramatic phenotype as active BCRP protein cannot be expressed from a C376T allele. The C421A (Q141K) polymorphism is also significant as Q141K-BCRP-transfected cells show markedly low protein expression levels and low-level drug resistance. Hence, individuals with C376T or C421A polymorphisms may express low levels of BCRP or none at all, resulting in hypersensitivity of normal cells to BCRP-substrate anticancer agents. In summary, both modulators of BCRP and functional single nucleotide polymorphisms within the BCRP gene affect the transporter function of the protein and thus can modulate drug sensitivity and substrate pharmacokinetics and pharmacodynamics in affected cells and individuals.

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12 The C421A (Q141K) polymorphism is also significant as Q141K-BCRP-transfected cells show markedly low protein expression levels and low-level drug resistance. Login to comment
165 From these analyses, we identified three BCRP coding SNP, G34A (V12M), C376T (Q126Stop) and C421A (Q141K), and a splicing variant, ∆315-6, that lacked nucleotides 944-949 (deletion of A315 and T316) (Fig. Login to comment
174 In addition, Q141K-BCRP-transfected PA317 cells showed markedly lower levels of both BCRP protein expression and drug resistance than wild-type BCRP-transfected cells (Fig. 4a). Login to comment
175 It was noteworthy in this case that Q141K-BCRP-transfectants and wild-type BCRP-transfectants expressed similar levels of BCRP transcripts (Fig. 4a). Login to comment
178 (42) Low expression levels of the Q141K BCRP protein have been confirmed using various experimental systems. Login to comment
179 (43-45) These results suggest that some individuals harbor a C421A polymorphic BCRP gene and express low amounts of Q141K BCRP (Fig. 4b). Login to comment
208 We have identified two important functioning BCRP SNP, C376T (Q126Stop) and C421A (Q141K), that greatly diminish the expression of this protein. Login to comment
210 Furthermore, cells transfected with Q141K-BCRP cDNA express low amounts of BCRP protein and show only low levels of drug resistance. Login to comment
214 Effect of C376T (Q126Stop) and C421A (Q141K) single nucleotide polymorphisms in the breast cancer resistance protein (BCRP) gene on protein expression. Login to comment
216 PA317 cells transfected with wild-type, G34A, C421A and ∆944-949 BCRP cDNAs were termed PA/WT, PA/V12M, PA/Q141K and PA/∆315-6, respectively. Login to comment

[hide] The ATP-binding cassette transporter ABCG2 (BCRP),... J Histochem Cytochem. 2006 Feb;54(2):215-21. Epub 2005 Aug 22. Meissner K, Heydrich B, Jedlitschky G, Meyer Zu Schwabedissen H, Mosyagin I, Dazert P, Eckel L, Vogelgesang S, Warzok RW, Bohm M, Lehmann C, Wendt M, Cascorbi I, Kroemer HK
The ATP-binding cassette transporter ABCG2 (BCRP), a marker for side population stem cells, is expressed in human heart.
J Histochem Cytochem. 2006 Feb;54(2):215-21. Epub 2005 Aug 22., [PMID:16116030]

Abstract [show]
Efforts to improve severely impaired myocardial function include transplantation of autologous hematopoietic side population (SP) stem cells. The transmembrane ABC-type (ATP binding cassette) half-transporter ABCG2 (BCRP) serves as a marker protein for SP cell selection. We have recently shown that other ABC transport proteins such as ABCB1 and ABCC5 are differentially expressed in normal and diseased human heart. Here we investigated localization and individual ABCG2 expression in 15 ventricular (including 10 cardiomyopathic) and 51 auricular heart tissue samples using immunohistochemistry, confocal laser scanning fluorescence microscopy, and real-time RT-PCR. Individual genotypes were assigned using PCR-restriction fragment length polymorphism (RFLP) analysis and subsequently correlated to ABCG2 mRNA levels. ABCG2 was localized in endothelial cells of capillaries and arterioles of all samples. Ventricular samples from cardiomyopathic hearts exhibited significantly increased levels of ABCG2 mRNA (ABCG2/18S rRNA: 1.08 +/- 0.30 x 10(-7); p=0.028 (dilative cardiomyopathy) and 1.16 +/- 0.46 x 10(-7); p=0.009 (ischemic cardiomyopathy) compared with 0.44 +/- 0.26 x 10(-7) in nonfailing hearts). The individual haplotypes were not associated with altered mRNA expression. ABCG2 is variably expressed in endothelial cells of human heart, where it may function as a protective barrier against cardiotoxic drugs such as anthracyclines or mitoxantrone. ABCG2 expression is induced in dilative and ischemic cardiomyopathies.

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126 Among the four identified naturally occurring single nucleotide polymorphisms, 34G.A (V12M) and 421C.A (Q141K) were most common in diverse populations of different ethnic origin in North America (Zamber et al. 2003). Login to comment
127 Among the four identified naturally occurring single nucleotide polymorphisms, 34G.A (V12M) and 421C.A (Q141K) were most common in diverse populations of different ethnic origin in North America (Zamber et al. 2003). Login to comment

[hide] Role of the breast cancer resistance protein (ABCG... AAPS J. 2005 May 11;7(1):E118-33. Mao Q, Unadkat JD
Role of the breast cancer resistance protein (ABCG2) in drug transport.
AAPS J. 2005 May 11;7(1):E118-33., [PMID:16146333]

Abstract [show]
The 72-kDa breast cancer resistance protein (BCRP) is the second member of the subfamily G of the human ATP binding cassette (ABC) transporter superfamily and thus also designated as ABCG2. Unlike P-glycoprotein and MRP1, which are arranged in 2 repeated halves, BCRP is a half-transporter consisting of only 1 nucleotide binding domain followed by 1 membrane-spanning domain. Current experimental evidence suggests that BCRP may function as a homodimer or homotetramer. Overexpression of BCRP is associated with high levels of resistance to a variety of anticancer agents, including anthracyclines, mitoxantrone, and the camptothecins, by enhancing drug efflux. BCRP expression has been detected in a large number of hematological malignancies and solid tumors, indicating that this transporter may play an important role in clinical drug resistance of cancers. In addition to its role to confer resistance against chemotherapeutic agents, BCRP actively transports structurally diverse organic molecules, conjugated or unconjugated, such as estrone-3-sulfate, 17beta-estradiol 17-(beta-D-glucuronide), and methotrexate. BCRP is highly expressed in the placental syncytiotrophoblasts, in the apical membrane of the epithelium in the small intestine, in the liver canalicular membrane, and at the luminal surface of the endothelial cells of human brain microvessels. This strategic and substantial tissue localization indicates that BCRP also plays an important role in absorption, distribution, and elimination of drugs that are BCRP substrates. This review summarizes current knowledge of BCRP and its relevance to multidrug resistance and drug disposition.

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160 A variety of naturally occurring variants of BCRP have been identified in DNA samples of ethnically diverse origins.95-98 Notably, the alterations of BCRP protein at position 12 (V12M) and 141 (Q141K) occur frequently in Asia populations (~30%-60%) and relatively low frequencies in Caucasians and African-Americans (~5%-10%). Login to comment
161 For example, in a Japanese population studied, 39% to 50% are heterozygous and 7% are homozygous for the variant Q141K.95,96 In a Chinese population, 60% are heterozygous for Q141K.95 Several other variants such as I206L, N590Y, and D620N are much less frequent with allele frequencies of ~1%.95,97 For instance, N590Y is present in ~1.5% of Caucasians.95 I206L is found only in Hispanic populations so far.95 D620N is detected in 1.1% of all DNA samples examined with unknown genetic origin.97 In addition, a polymorphism in exon 4 that results in a substitution of stop codon for Gln at position 126 has also been identified.96 Amino acid changes at position 482 that were found in some drug-selected resistant cell lines have so far not been identified in normal populations or in DNA samples from cancer patients.49 In vitro functional characterization of the variants V12M and Q141K produced contradicting results. Login to comment
162 One study reported that Q141K was expressed at lower levels in transfected cells and therefore conferred lower drug resistance compared with the wild-type protein.96 The variant V12M displayed expression levels and drug-resistance properties similar to the wild-type protein.96 Another study reported that V12M and Q141K were expressed at levels comparable to the wild-type protein; however, both V12M and Q141K conferred significantly lower levels of drug resistance relative to the wild-type protein as compared with increased drug accumulation and decreased drug efflux.98 Further analysis of the mechanism of the transport dysfunction revealed that the apical membrane localization of V12M was disrupted and that ATPase activity of Q141K was decreased.98 A recent clinical study by Sparreboom et al73 has shown that the Q141K polymorphism is associated with significant changes of pharmacokinetic properties of diflomotecan, a substrate of BCRP. Login to comment

[hide] The ABC transporter Abcg2/Bcrp: role in hypoxia me... Biometals. 2005 Aug;18(4):349-58. Krishnamurthy P, Schuetz JD
The ABC transporter Abcg2/Bcrp: role in hypoxia mediated survival.
Biometals. 2005 Aug;18(4):349-58., [PMID:16158227]

Abstract [show]
ABC (ATP-binding cassette) transporters have diverse roles in many cellular processes. These diverse roles require the presence of conserved membrane spanning domains and nucleotide binding domains. Bcrp (Abcg2) is a member of the ATP binding cassette family of plasma membrane transporters that was originally discovered for its ability to confer drug resistance in tumor cells. Subsequent studies showed Bcrp expression in normal tissues and high expression in primitive stem cells. Bcrp expression is induced under low oxygen conditions consistent with its high expression in tissues exposed to low oxygen environments. Moreover, Bcrp interacts with heme and other porphyrins. This finding and its regulation by hypoxia suggests it may play a role in protecting cells/tissue from protoporphyrin accumulation under hypoxia. These observations are strengthened by the fact that porphyrins accumulate in tissues of the Bcrp knockout mouse. It is possible that humans with loss of function Bcrp alleles may be more susceptible to porphyrin-induced phototoxicity. We propose that Bcrp plays a role in porphyrin homoeostasis and regulates survival under low oxygen conditions.

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127 The SNPs in Bcrp that produce non-synonymous changes (i.e., amino acid substitutions) are at amino acids 12 (V12M), 141 (Q141K), 206 (I206L), and 590 (N590Y). Login to comment
128 The most frequent polymorphisms being the exon 2 SNP (G34A/ V12M) and the exon 5 SNP (C421A/Q141K), which produce changes in amino acids 12 and 141 (Imai et al. 2002; Mizuarai et al. 2004). Login to comment

[hide] Pharmacogenomics of the human ABC transporter ABCG... Naturwissenschaften. 2005 Oct;92(10):451-63. Ishikawa T, Tamura A, Saito H, Wakabayashi K, Nakagawa H
Pharmacogenomics of the human ABC transporter ABCG2: from functional evaluation to drug molecular design.
Naturwissenschaften. 2005 Oct;92(10):451-63., [PMID:16160819]

Abstract [show]
In the post-genome-sequencing era, emerging genomic technologies are shifting the paradigm for drug discovery and development. Nevertheless, drug discovery and development still remain high-risk and high-stakes ventures with long and costly timelines. Indeed, the attrition of drug candidates in preclinical and development stages is a major problem in drug design. For at least 30% of the candidates, this attrition is due to poor pharmacokinetics and toxicity. Thus, pharmaceutical companies have begun to seriously re-evaluate their current strategies of drug discovery and development. In that light, we propose that a transport mechanism-based design might help to create new, pharmacokinetically advantageous drugs, and as such should be considered an important component of drug design strategy. Performing enzyme- and/or cell-based drug transporter, interaction tests may greatly facilitate drug development and allow the prediction of drug-drug interactions. We recently developed methods for high-speed functional screening and quantitative structure-activity relationship analysis to study the substrate specificity of ABC transporters and to evaluate the effect of genetic polymorphisms on their function. These methods would provide a practical tool to screen synthetic and natural compounds, and these data can be applied to the molecular design of new drugs. In this review article, we present an overview on the genetic polymorphisms of human ABC transporter ABCG2 and new camptothecin analogues that can circumvent AGCG2-associated multidrug resistance of cancer.

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87 Non-synonymous SNPs are located at nucleotides 238 (exon 2) and 625, resulting in amino acid substitutions: V12M and Q141K. Login to comment
94 The Q141K polymorphism located in exon 5 (c.421C>A) leads to the replacement of the negatively charged glutamic acid residue with a positively charged lysine residue. Login to comment
96 The Q141K variant was also detected in all ethnic groups tested: the allele frequency ranged between 0% and 35%, (the Africans in north of Sahara, the Africans sub-Saharan, the African American subjects with low, and the Japanese and Chinese populations with high allele frequencies) (Imai et al. 2002; Zamber et al. 2003; de Jong et al. 2004; Kobayashi et al. 2005). Login to comment
97 Imai et al. (2002) have demonstrated that the ABCG2 Q141K variant-transfected PA317 cells showed a low-level of drug resistance associated with decreased protein expression. Login to comment
99 The SNP (Q141K) was postulated to cause increased sensitivity of normal cells to anticancer agents that are ABCG2 substrates such as topotecan, diflomotecan, and SN-38. Login to comment
100 An investigation of the expression level of ABCG2 in 99 Japanese placenta samples revealed that individuals homozygous for the Q141K variant showed significantly lower expression levels of this transporter protein, while the heterozygous samples displayed an intermediate expression level (Kobayashi et al. 2005). Login to comment
102 Based on these observations, it is assumed that the protein stability of the Q141K variant is significantly reduced without significant changes in its mRNA levels. Login to comment
109 Imai et al. (2002) reported that the Q141K variant of ABCG2, stably expressed in PA317 cells, had a markedly lower expression level than the wild-type ABCG2 or the V12M variant. Login to comment
110 In their study, the Q141K variant-transfected PA317 cells exhibited a low-level drug resistance compared with the wild-type ABCG2-transfected cells. Login to comment
111 On the other hand, Mizuarai et al. (2004) expressed ABCG2 in polarized LLC-PK1 cells, and by using confocal microscopy demonstrated that the wild-type and the Q141K variant of ABCG2 mainly showed apical staining, while the V12M variant showed intracellular localization. Login to comment
112 However, Kondo et al. (2004) demonstrated that both V12M and Q141K variants were localized at the apical membrane in LLC-PK1 cells. Login to comment
113 These contradictory expression and localization data for ABCG2 variants indicate that differences in transfection conditions (transient or stable expression), the copy number of cDNA incorporated in genomic DNA or other cellular determinants may variably Table 2 Frequencies of ABCG2 alleles in different ethnic groups Position Ethnic group Variant allele Allele Reference Amino acid cDNA N Hetero Homo Frequency (%) V12M c.34G>A Japanese 29 9 1 19.0 Imai et al. (2002) Japanese 10 - - 15.0 Zamber et al. (2003) Japanese 220 61 8 17.5 Kobayashi et al. (2005) Chinese 10 - - 20.0 Zamber et al. (2003) Southeast Asians 10 - - 45.0 Zamber et al. (2003) Pacific Islanders 7 - - 64.0 Zamber et al. (2003) Swedish 60 2 0 1.7 B¨ackstr¨om et al. (2003) Dutch 100 11 1 6.5 Bosch et al. (2005) Caucasian 86 - - 2.0 Zamber et al. (2003) Caucasian 150 27 2 10.3 Mizuarai et al. (2004) Caucasian 150 11 0 3.7 Kobayashi et al. (2005) Ashkenazi Jewish 10 - - 10.0 Zamber et al. (2003) Middle Eastern 20 - - 5.0 Zamber et al. (2003) Africans North of Sahara 7 - - 14.0 Zamber et al. (2003) African American 150 17 1 6.3 Kobayashi et al. (2005) Mexicans 10 - - 10.0 Zamber et al. (2003) Hispanic Livers 5 - - 40.0 Zamber et al. (2003) Mexican Indians 5 - - 90.0 Zamber et al. (2003) Q126Stop c.376C>T Japanese 124 3 0 1.2 Imai et al. (2002) Japanese 60 2 0 1.7 Itoda et al. (2003) Japanese 220 4 0 0.9 Kobayashi et al. (2005) Caucasian 150 0 0 0.0 Mizuarai et al. (2004) Caucasian 150 0 0 0.0 Kobayashi et al. (2005) African American 150 0 0 0.0 Kobayashi et al. (2005) Q141K c.421C>A Japanese 124 48 9 26.6 Imai et al. (2002) Japanese 10 - - 35.0 Zamber et al. (2003) Japanese 220 90 27 32.7 Kobayashi et al. (2005) Chinese 95 43 11 34.2 de Jong et al. (2004) Chinese 10 - - 35.0 Zamber et al. (2003) Southeast Asians 10 - - 15.0 Zamber et al. (2003) Pacific Islanders 7 - - 14.0 Zamber et al. (2003) Swedish 60 10 1 10.0 B¨ackstr¨om et al. (2003) Dutch 100 20 2 12.0 Bosch et al. (2005) Caucasian 85 - - 14.0 Zamber et al. (2003) Caucasian 172 33 3 11.3 de Jong et al. (2004) Caucasian 150 22 2 8.7 Mizuarai et al. (2004) Caucasian 150 25 4 11.0 Kobayashi et al. (2005) Ashkenazi Jewish 10 - - 5.0 Zamber et al. (2003) Middle Eastern 20 - - 13.0 Zamber et al. (2003) Africans North of Sahara 7 - - 0.0 Zamber et al. (2003) African, Sub-Saharan 938 14 1 0.9 de Jong et al. (2004) African American 24 - - 0.0 Zamber et al. (2003) African American 150 5 1 2.3 Kobayashi et al. (2005) African American 94 8 1 5.3 de Jong et al. (2004) Mexicans 10 - - 5.0 Zamber et al. (2003) Hispanic Livers 5 - - 10.0 Zamber et al. (2003) Mexican Indians 5 - - 10.0 Zamber et al. (2003) R160Q c.479G>A Dutch 100 1 0 0.5 Bosch et al. (2005) I206L c.616A>C Japanese 10 - - 0.0 Zamber et al. (2003) Chinese 10 - - 0.0 Zamber et al. (2003) Southeast Asians 10 - - 0.0 Zamber et al. (2003) Pacific Islanders 7 - - 0.0 Zamber et al. (2003) Caucasian 65 - - 0.0 Zamber et al. (2003) Table 2 Continued Position Ethnic group Variant allele Allele Reference Amino acid cDNA N Hetero Homo Frequency (%) Ashkenazi Jewish 10 - - 0.0 Zamber et al. (2003) Middle Eastern 20 - - 0.0 Zamber et al. (2003) Africans North of Sahara 7 - - 0.0 Zamber et al. (2003) African American 15 - - 0.0 Zamber et al. (2003) Mexicans 10 - - 0.0 Zamber et al. (2003) Hispanic Livers 5 - - 10.0 Zamber et al. (2003) Mexican Indians 5 - - 0.0 Zamber et al. (2003) F431L c.1291T>C Japanese 60 1 0 0.8 Itoda et al. (2003) S441N c.1322G>A Japanese 100 1 0 0.5 Kobayashi et al. (2005) F489L c.1465T>C Japanese 60 1 0 0.8 Itoda et al. (2003) Japanese 100 1 0 0.5 Kobayashi et al. (2005) R575Stop c.1723C>T Dutch 100 1 0 0.5 Bosch et al. (2005) N590Y c.1768A>T Caucasian 65 - - 1.0 Zamber et al. (2003) Caucasian 150 1 0 0.3 Mizuarai et al. (2004) African Americans 15 - - 0.0 Zamber et al. (2003) D620N c.1858G>A Dutch 100 1 0 0.5 Bosch et al. (2005) affect the cellular processing and sorting of these proteins. Login to comment
114 Detailed studies are needed to clarify the mechanism of a reduced protein expression for Q141K and the altered cellular localization of V12M and Q141K variants. Login to comment
118 For this purpose, we have created variant forms of ABCG2 (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, E334stop, N590Y, D620N, R482G, and R482T) by site-directed mutagenesis. Login to comment
131 Interestingly, the MTX transport activity of the Q141K variant was even higher than the wild type. Login to comment
211 However, one of the two homozygous individuals showed increased accumulation of SN-38 and SN-38 glucuronide, indicating that the Q141K homodimer may have an im- Fig. 5 Molecular structures of newly synthesized CPT analogues and their anticancer activity in ABCG2-transfected HEK293 (HEK293/ABCG2) cells. Login to comment

[hide] Membrane transporters and channels in chemoresista... Cancer Lett. 2006 Aug 8;239(2):168-82. Epub 2005 Oct 5. Huang Y, Sadee W
Membrane transporters and channels in chemoresistance and -sensitivity of tumor cells.
Cancer Lett. 2006 Aug 8;239(2):168-82. Epub 2005 Oct 5., 2006-08-08 [PMID:16169662]

Abstract [show]
Membrane transporters play important roles in mediating chemosensitivity and -resistance of tumor cells. ABC transporters, such as ABCB1/MDR1, ABCC1/MRP1 and ABCG2/BCRP, are frequently associated with decreased cellular accumulation of anticancer drugs and multidrug resistance of tumors. SLC transporters, such as folate, nucleoside, and amino acid transporters, commonly increase chemosensitivity by mediating the cellular uptake of hydrophilic drugs. Ion channels and pumps variably affect sensitivity to anticancer therapy by modulating viability of tumor cells. A pharmacogenomic approach, using correlations between drug potency and transporter gene expression in multiple cancer cell lines, has shown promise for identifying potential drug-transporter relationships and predicting anticancer drug response, in an effort to optimize chemotherapy for individual patients.

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79 Two additional SNPs, i.e. V12M and Q141K, also affect substrate specificity and transport capacity of ABCG2 [26]. Login to comment

[hide] Genetic polymorphisms of ATP-binding cassette tran... Expert Opin Pharmacother. 2005 Nov;6(14):2455-73. Sakurai A, Tamura A, Onishi Y, Ishikawa T
Genetic polymorphisms of ATP-binding cassette transporters ABCB1 and ABCG2: therapeutic implications.
Expert Opin Pharmacother. 2005 Nov;6(14):2455-73., [PMID:16259577]

Abstract [show]
Pharmacogenomics, the study of the influence of genetic factors on drug action, is increasingly important for predicting pharmacokinetics profiles and/or adverse reactions to drugs. Drug transporters, as well as drug metabolism play pivotal roles in determining the pharmacokinetic profiles of drugs and their overall pharmacological effects. There is an increasing number of reports addressing genetic polymorphisms of drug transporters. However, information regarding the functional impact of genetic polymorphisms in drug transporter genes is still limited. Detailed functional analysis in vitro may provide clear insight into the biochemical and therapeutic significance of genetic polymorphisms. This review addresses functional aspects of the genetic polymorphisms of human ATP-binding cassette transporters, ABCB1 and ABCG2, which are critically involved in the pharmacokinetics of drugs.

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210 In different ethnic groups, seven naturally-occurring non-synonymous SNPs have been reported: V12M, Q126Stop, Q141K, I206L, F431L, S441N, F489L, N590Y and D620N. Login to comment
211 Among the above variations, two alterations (c.34G > A [V12M], c.421C > A [Q141K]) affecting the protein structure have been reported to be polymorphic in several populations. Login to comment
215 Non-synonymous SNPs are located at nucleotides 238 (exon 2) and 625 (exon 5), resulting in the amino acid substitutions V12M and Q141K. Login to comment
221 The Q141K polymorphism located in exon 5 (c.421C > A) leads to the replacement of the negatively charged glutamic acid residue with a positively charged lysine residue. Login to comment
223 The Q141K variant was also detected in all ethnic groups tested: the allele frequency was 1 - 35% (the African and African-American subjects had low allele frequencies, whereas those of the Table 4. Login to comment
225 Location Position Allele Amino acid Allele frequency in Caucasian populations Allele frequency in Japanese populatins Allele frequency in African populations n % n % n % Exon 2 34 G A 12 Val 12 Met 546 94.4 5.6 259 82.4 17.6 181 93.7 6.3 Exon 4 376 C T 126 Gln 126 stop 300 100 0 404 98.9 1.1 150 100 0 Exon 5 421 C A 141 Gln 141 Lys 717 89.0 11.0 354 69.4 30.6 1213 98.6 1.4 Exon 5 479 G A 160 Arg 160 Gln 100 99.5 0.5 ND ND ND ND ND ND Exon 11 1291 T C 431 Phe 431 Leu ND ND ND 60 99.2 0.8 ND ND ND Exon 11 1322 G A 441 Ser 441 Asn ND ND ND 100 99.5 0.5 ND ND ND Exon 12 1465 T C 489 Phe 489 Leu ND ND ND 160 99.4 0.6 ND ND ND Exon 14 1723 C T 575 Arg 575 stop 100 99.5 0.5 ND ND ND ND ND ND Exon 15 1768 A T 590 Asn 590 Tyr 215 99.5 0.5 ND ND ND 15 100 0 Exon 16 1858 T A 620 Asp 620 Asp 100 99.5 0.5 ND ND ND ND ND ND Data are from [129-135,137]. Login to comment
229 Imai et al. [129] have demonstrated that the ABCG2 Q141K variant-transfected PA317 cells showed a low-level of drug resistance associated with decreased protein expression. Login to comment
231 The SNP (Q141K) was postulated to cause increased sensitivity of normal cells to anticancer agents that are ABCG2 substrates, such as topotecan, diflomotecan and SN-38. Login to comment
232 An investigation of the expression level of ABCG2 in 99 Japanese placenta samples revealed that individuals homozygous for the Q141K variant showed significantly lower expression levels of this transporter protein, and the heterozygous samples displayed an intermediate expression level [134]. Login to comment
235 It is assumed that the protein stability of the Q141K variant is reduced without significant changes in its mRNA levels. Login to comment
237 It has been postulated that the 376C > T SNP may have higher impact than the 421C > A polymorphism causing Q141K because active ABCG2 protein will not be synthesised from the variant allele. Login to comment
242 Imai et al. [129] reported that the Q141K variant of ABCG2 stably expressed in PA317 cells had a markedly lower expression level than the wild-type ABCG2 or the V12M variant. Login to comment
243 In their study, the Q141K variant-transfected PA317 cells exhibited a low-level drug resistance compared with that of wild-type ABCG2-transfected cells. Login to comment
244 On the other hand, Mizuarai et al. [135] expressed ABCG2 in polarised LLC-PK1 cells, and by using confocal microscopy demonstrated that the wild-type and the Q141K variant of ABCG2 showed mainly apical staining, and the V12M variant showed intracellular localisation. Login to comment
245 However, Kondo et al. [138] demonstrated that both V12M and Q141K variants were localised at the apical membrane in LLC-PK1 cells. Login to comment
250 COOH H2N N590Y V12M G51C Q126stop Q141K T153M Q166E I206L F208S S248P E334stop F431L F489L D620N R482G R482T S441N F571I EXTRACELLULAR INTRACELLULAR R160Q R575stop ATP-binding site (transient or stable expression), the copy number of cDNA incorporated in genomic DNA or other cellular determinants may variably affect the cellular processing and sorting of these proteins. Login to comment
251 Detailed studies are needed to clarify the mechanism of a reduced protein expression for Q141K and the altered cellular localisation of V12M and Q141K variants. Login to comment
255 For this purpose, variant forms of ABCG2 (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, E334stop, N590Y, D620N, R482G and R482T) were created by site-directed mutagenesis (Figure 3). Login to comment
268 Interestingly, the MTX transport activity of the Q141K variant was even higher than the wild type. Login to comment
768 IMAI Y, NAKANE M, KAGE K et al.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Mol. Cancer Ther. (2002) 1(8):611-616. Login to comment
651 IMAI Y, NAKANE M, KAGE K et al.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Mol. Cancer Ther. (2002) 1(8):611-616. 130. Login to comment

[hide] Pharmacogenetics of irinotecan metabolism and tran... Toxicol In Vitro. 2006 Mar;20(2):163-75. Epub 2005 Nov 3. Smith NF, Figg WD, Sparreboom A
Pharmacogenetics of irinotecan metabolism and transport: an update.
Toxicol In Vitro. 2006 Mar;20(2):163-75. Epub 2005 Nov 3., [PMID:16271446]

Abstract [show]
The anticancer agent irinotecan (CPT-11) is converted to SN-38, which is approximately 100 to 1,000-fold more cytotoxic than the parent drug. The pharmacokinetics of irinotecan are extremely complex and have been the subject of intensive investigation in recent years. Irinotecan is subject to extensive metabolism by various polymorphic enzymes, including CES2 to form SN-38, members of the UGT1A subfamily, and CYP3A4 and CYP3A5, which form several pharmacologically inactive oxidation products. Elimination of irinotecan is also dependent on drug-transporting proteins, notably ABCB1 (P-glycoprotein), ABCC2 (cMOAT) and ABCG2 (BCRP), present on the bile canalicular membrane. The various processes mediating drug elimination, either through metabolic breakdown or excretion, likely impact substantially on interindividual variability in drug handling. This report provides an update on current strategies to individualize irinotecan chemotherapy based on each patient's genetic constitution, which may ultimately lead to more selective use of this agent.

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998 Of these, the most extensively studied is the 421C > A transversion, which results in an amino acid change of glutamine to lysine at codon 141 (Q141K). Login to comment

[hide] Functional SNPs of the breast cancer resistance pr... Cancer Lett. 2006 Mar 8;234(1):73-80. Epub 2005 Nov 21. Yanase K, Tsukahara S, Mitsuhashi J, Sugimoto Y
Functional SNPs of the breast cancer resistance protein-therapeutic effects and inhibitor development.
Cancer Lett. 2006 Mar 8;234(1):73-80. Epub 2005 Nov 21., 2006-03-08 [PMID:16303243]

Abstract [show]
Breast cancer resistance protein (BCRP) is a half-molecule ATP-binding cassette transporter that pumps out various anticancer agents such as 7-ethyl-10-hydroxycamptothecin, topotecan and mitoxantrone. We have previously identified three polymorphisms within the BCRP gene, G34A (substituting Met for Val-12), C376T (substituting a stop codon for Gln-126) and C421A (substituting Lys for Gln-141). C421A BCRP-transfected murine fibroblast PA317 cells showed markedly decreased protein expression and low-level drug resistance when compared with wild-type BCRP-transfected cells. In contrast, G34A BCRP-transfected PA317 cells showed a similar protein expression and drug resistance profile to wild-type. The C376T polymorphism would be expected to have a considerable impact as active BCRP protein will not be expressed from a T376 allele. Hence, people with C376T and/or C421A polymorphisms may express low levels of BCRP, resulting in hypersensitivity of normal cells to BCRP-substrate anticancer agents. Estrogens, estrone and 17beta-estradiol, were previously found to restore drug sensitivity levels in BCRP-transduced cells by increasing the cellular accumulation of anticancer agents. BCRP transports sulfated estrogens but not free estrogens and in a series of screening experiments for synthesized and natural estrogenic compounds, several tamoxifen derivatives and phytoestrogens/flavonoids were identified that effectively circumvent BCRP-mediated drug resistance. The kinase inhibitors gefitinib and imatinib mesylate also interact with BCRP. Gefitinib, an inhibitor of epidermal growth factor receptor-tyrosine kinase, inhibits its transporter function and reverses BCRP-mediated drug resistance both in vitro and in vivo. BCRP-transfected human epidermoid carcinoma A431 cells and BCRP-transfected human non-small cell lung cancer PC-9 cells show gefitinib resistance. Imatinib, an inhibitor of BCR-ABL tyrosine kinase, also inhibits BCRP-mediated drug transport. Hence, both functional SNPs and inhibitors of BCRP reduce its transporter function and thus modulate substrate pharmacokinetics and pharmacodynamics.

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1 We have previously identified three polymorphisms within the BCRP gene, G34A (substituting Met for Val-12), C376T (substituting a stop codon for Gln-126) and C421A (substituting Lys for Gln-141). Login to comment
42 C421A (Q141K) BCRP SNP We have previously identified three variant BCRP cDNAs, containing the substitutions G34A (V12M), C421A (Q141K) and a 944-949 deletion lacking Ala-315 and Thr-316 (D315-6) [23]. Login to comment
44 We have subsequently found that C421A BCRP-transfected murine fibroblast PA317 (PA/Q141K) cells show markedly decreased exogenous protein expression and also a low-level of drug resistance when normalized to wild-type BCRP-transfected (PA/WT) cells (Fig. 1 and Table 1). Login to comment
46 We had already shown in our previous study that the intracellular topotecan accumulation in PA/Q141K cells was higher than in other BCRP transfectants [23]. Login to comment
47 Kondo et al. have also reported low Q141K-BCRP protein expression levels using adenovirus-mediated gene transfection [24]. Login to comment
56 This may be associated with a greater susceptibility ofthe resultingBCRP protein (Q141K) to degradation [23]. Login to comment
58 We previously examined the frequency of the C421A SNP in a normal Japanese population and found that 57/124 samples carried the A421 allele and that nine of these were homozygous for this polymorphism [23], indicating that some individuals possess the C421A polymorphic BCRP gene and express low amounts of the Q141K BCRP. Login to comment
74 However, in our Table 1 Drug sensitivities of BCRP-transfected PA317 cells Cells IC50 (ng/ml) SN-38 Topotecan MXR PA317 2.5 0.060 17 PA/WT 98 0.58 O200 PA/V12M 98 0.63 O200 PA/Q141K 30 0.25 100 PA/D315-6 55 0.42 190 Cells were cultured for 5 days in the absence or presence of increasing concentrations of the indicated anticancer agents. Login to comment
92 Therefore, we first Table 3 SNPs within the BCRP gene Variation Region Effect Domain A-1379G 50 -flanking (promoter) - D-654-651 50 -flanking (promoter) - G-286C 50 -flanking (promoter) - T-476C Exon 1 (50 - UTR) - D-235A Exon 1 (50 - UTR) - A-113G Exon 1 (50 - UTR) - A-29G Exon 1 (50 - UTR) - G34A Exon 2 V12M N-terminal T114C Exon 2 No change N-terminal G151T Exon 2 G51C N-terminal C369T Exon 4 No change NBD C376T Exon 4 Q126stop NBD C421A Exon 5 Q141K NBD C458T Exon 5 T153M NBD C474T Exon 5 No change NBD C496G Exon 5 Q166E NBD A564G Exon 6 No change NBD A616C Exon 6 I206L NBD T623C Exon 6 F208S NBD T742C Exon 7 S248P Linker G1000T Exon 9 E334stop Linker G1098A Exon 9 No change Linker T1291C Exon 11 F431L TMD A1425G Exon 12 No change TMD T1465C Exon 12 F489L TMD A1768T Exon 15 N590Y TMD G1858A Exon 16 D620N TMD G2237T Exon 16 (30 - UTR) - G2393T Exon 16 (30 - UTR) - Abbreviations: UTR, untranslated region; NBD, nucleotide-binding domain; TMD, transmembrane domain. Login to comment
187 [23] Y. Imai, M. Nakane, K. Kage, S. Tsukahara, E. Ishikawa, T. Tsuruo, et al., C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance, Mol. Cancer Ther. 1 (2002) 611-616. Login to comment

[hide] Molecular modeling of new camptothecin analogues t... Cancer Lett. 2006 Mar 8;234(1):81-9. Epub 2005 Nov 23. Nakagawa H, Saito H, Ikegami Y, Aida-Hyugaji S, Sawada S, Ishikawa T
Molecular modeling of new camptothecin analogues to circumvent ABCG2-mediated drug resistance in cancer.
Cancer Lett. 2006 Mar 8;234(1):81-9. Epub 2005 Nov 23., 2006-03-08 [PMID:16309825]

Abstract [show]
Irinotecan (CPT-11) is a widely used potent antitumor drug that inhibits mammalian DNA topoisomerase I (Topo I). However, overexpression of ABCG2 (BCRP/MXR/ABCP) reportedly confers cancer cells resistance to SN-38, the active form of CPT-11. To circumvent the ABCG2-associated drug resistance, the structure-activity-relationship (SAR) of 14 new camptothecin (CPT) analogues has been studied with respect to the substrate specificity of ABCG2. While the lactone E ring is a prerequisite for anticancer activity, modifications of the A or B rings do not significantly affect Topo I inhibition. Based on the substrate specificity of ABCG2, it is strongly suggested that CPT analogues with a hydroxyl group at position 10 or 11 of the A ring are recognized by ABCG2 and are thereby effectively extruded from cancer cells. To develop a platform for the molecular modeling to circumvent anticancer drug resistance, we have carried out quantum chemical calculations and neural network SAR analysis. Electrostatic potential iso-surfaces generated by ab initio MO calculations using restricted Hartree-Fock method have revealed that negative potential localized at positions 10 or 11 in the A ring is important for recognition by ABCG2.

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112 However, one of the two homozygous individuals showed increased accumulation of SN-38 and SN-38 glucuronide, indicating that the Q141K homodimer may have an impaired function. Login to comment

[hide] The role of the human ABCG2 multidrug transporter ... Cancer Lett. 2006 Mar 8;234(1):62-72. Epub 2005 Dec 7. Cervenak J, Andrikovics H, Ozvegy-Laczka C, Tordai A, Nemet K, Varadi A, Sarkadi B
The role of the human ABCG2 multidrug transporter and its variants in cancer therapy and toxicology.
Cancer Lett. 2006 Mar 8;234(1):62-72. Epub 2005 Dec 7., 2006-03-08 [PMID:16337740]

Abstract [show]
The human multidrug resistance ABC transporters provide a protective function in our body against a large number of toxic compounds. These proteins, residing in the plasma membrane, perform an active, ATP-dependent extrusion of such xenobiotics. However, the same proteins are also used by the tumor cells to fight various anticancer agents. ABCG2 is an important member of the multidrug resistance proteins, an 'ABC half transporter', which functions as a homodimer in the cell membrane. In this review, we provide a basic overview of ABCG2 function in physiology and drug metabolism, but concentrate on the discussion of mutations and polymorphisms discovered in this protein. Interestingly, a single nucleotide mutation, changing amino acid 482 from arginine to threonine or glycine in ABCG2, results in a major increase in the catalytic activity and a wider drug recognition by this protein. Still, this mutation proved to be an in vitro artifact, produced only in heavily drug-selected cell lines. In contrast, at least two, but possibly more polymorphic variants of ABCG2 were found to be present in large human populations with different ethnic background. However, currently available experimental data regarding the cellular expression, localization and function of these ABCG2 variants are strongly contradictory. Since, the proteins produced by these variant alleles may differently modulate cancer treatment, general drug absorption and toxicity, may represent risk factors in fetal toxicity, or alter the differentiation of stem cells, their exact characterization is a major challenge in this field.

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109 To date, altogether eight non-synonymous (V12M, Q141K, I206L, F431L, S441N, F489L, N590Y, D620N), five synonymous (silent) (c.114TOC, c.369COT, c.474COT, c.1098GOA, c.1425AOG) missense mutations, one nonsense (Q126X), and one frameshift (c.1515delC) mutations were identified in the coding region of ABCG2 in healthy individuals or in patients [43-46,49,63-65]. Login to comment
110 Among the above variations, affecting the protein structure, two alterations [c.34GOA (V12M), c.421CO A (Q141K)] have been reported to be polymorphic in several populations. Login to comment
119 The Q141K polymorphism located in exon 5 (c.421COA) leads to the replacement of the negatively charged glutamic acid residue with a positively charged lysine residue. Login to comment
121 The Q141K variant was also detected in all ethnic groups tested: the allele frequency ranged between 1 and 35%, (the African and African-American subjects with low, while the Japanese and Chinese populations with high allele frequencies) [46,63]. Login to comment
123 On the basis of definitive molecular haplotype analyses (PCR-RFLP) for the three major variants [c.34GOA (V12M), c.376COT (Q126X), c.421COA (Q141K)] in a Japanese population, four haplotypes were identified G-C-C (V-Q-Q), G-C-A (V-Q-K), A-CC (M-Q-Q), and G-T-C (V-X-Q). Login to comment
127 The above results collectively suggest that the V12M, Q126X, and Q141K variants are likely to occur on separate chromosomes. Login to comment
130 Mainly the two major non-synonymous polymorphisms, V12M and Q141K, were investigated. Login to comment
132 Imai et al. [64] and Morisaki et al. [66] in stable mammalian expression systems found that in PA317 or HEK-293 cells the expressed Q141K ABCG2 protein had a lower expression level than the wild-type ABCG2, or the V12M variant. Login to comment
133 Morisaki et al. demonstrated that both the V12M and Q141K ABCG2 could reach the plasma membrane in the HEK-293 cells, while a significant portion of Table 1 Summary of population genetics data on naturally occurring sequence variations, affecting the coding region of the human ABCG2 gene Position in the ABCG2 genea Position in the ABCG2 cDNAb Amino acid substitution Population n C/K C/C AF (%G 95%CI) Ref. Login to comment
134 g.34GOA (exon 2) c.34GOA V12M Caucasian 150 27 2 10.3G3.5 [47] Caucasian 150 11 0 3.7G2.2 [46] Caucasian 86 n.a n.a 2.0Gn.a [49] Swedish 60 2 0 1.7G2.3 [43] Total Caucasian 360 40 2 6.1G1.8 Japanese 220 61 8 17.5G3.6 [46] Japanese 29 9 1 19.0G10.3 [64] Total Japanese 249 70 9 17.7G3.4 African-American 150 17 1 6.3G2.8 [46] g.8191COT (exon 4) c.376COT Q126X Caucasian 150 0 0 0.0 [46] Caucasian 150 0 0 0.0 [47] Total Caucasian 300 0 0 0.0 Japanese 220 4 0 0.9G0.9 [46] Japanese 124 3 0 1.2G1.4 [64] Japanese 60 2 0 1.7G2.3 [45] Total Japanese 404 9 0 1.1G0.7 African-American 150 0 0 0.0 [46] g.8825CO A (exon 5) c.421COA Q141K Caucasian 172 33 3 11.3G3.4 [63] Caucasian 150 25 4 11.0G3.6 [46] Caucasian 150 22 2 8.7G3.2 [47] Caucasian 85 n.a n.a 14.0Gn.a [49] Swedish 60 10 1 10.0G5.5 [43] Total Caucasian 532 90 10 10.3G1.9 Japanese 220 90 27 32.7G4.5 [46] Japanese 124 48 9 26.6G5.6 [64] Chinese 95 43 11 34.2G6.9 [63] Total Asian 439 181 47 31.3G3.1 African, Sub-Saharan 938 14 1 0.9G0.4 [63] African-American 150 5 1 2.3G1.7 [46] African-American 94 8 1 5.3G3.3 [63] Total Africanc 1182 27 3 1.4G0.5 g.40645AO T (exon 12) c.1465TOC F489L Japanese 100 1 0 0.5G1.0 [46] Japanese 60 1 0 0.8G1.7 [45] Total Japanese 160 2 0 0.6G0.9 g.45367AO T (exon 15) c.1768AOT N590Y Caucasian 150 1 0 0.3G0.7 [47] Caucasian 65 1 0 0.8G1.5 [49] Total Caucasian 215 2 0 0.5G0.7 Only those cDNA SNPs were listed that were detected in at least two independent studies. Login to comment
142 J. Cervenak et al. / Cancer Letters 234 (2006) 62-72 67 Q141K (though having a lower expression level), remained intracellular [66]. Login to comment
143 According to other studies, a 30-40% reduction in cell surface expression of the Q141K variant, although having a similar mRNA level than the wild-type ABCG2, was observed [55,64]. Login to comment
144 Recent investigation of the expression level of ABCG2 in 99 Japanese placenta samples revealed that individuals homozygous for the Q141K variant showed significantly lower expression levels of this transporter protein, while the heterozygous samples displayed an intermediate expression level [46]. Login to comment
146 Mizuarai et al. expressed ABCG2 in polarized LLC-PKI cells, and by using confocal microscopy demonstrated that the wtABCG2 and Q141K showed mainly apical staining, while the V12M variant showed intracellular localization [47]. Login to comment
147 In a recent study, similarly LLC-PKI cells where used to express the V12M and Q141K variants and additionally five other polymorphisms (A149P, R163K, Q166E, P269S and S441N [55]). Login to comment
148 Interestingly, they found that all polymorphisms, including V12M and Q141K, had an apical localization, and only the S441N variant showed intracellular staining. Login to comment
151 Clearly, more detailed studies are required to clarify the mechanism of a reduced protein expression for Q141K, and the altered cellular localization found for the V12M and Q141K variants under certain conditions. Login to comment
153 When the functions of the ABCG2 variants were examined in cytotoxicity assays, a 10-fold decrease in drug resistance, as compared to the wild-type ABCG2, was reported by Mizuarai et al., when the V12M or Q141K-transfected LLC-PKI cells were challenged by mitoxantrone, topotecan, or an indolocarbazole topoisomerase I inhibitor [47]. Login to comment
154 In contrast, Morisaki et al. found that only the Q141K variant had a moderately lower level resistance against mitoxantrone, topotecan or SN-38, as compared to the wild-type ABCG2-transfected cells [66]. Login to comment
161 Q141K is mapped in the functionally important ATP-binding cassette region of ABCG2 (Fig. 1) between the Walker A and the signature motifs, therefore, it is possible that the ATPase activity of this variant is altered. Login to comment
162 Two studies compared the vanadate-sensitive ATPase activity of ABCG2 V12M and Q141K variants, using Sf9 (Spodoptera frugiperda) cell membranes [47,66]. Login to comment
163 A reduced, 1.3 and 1.8-fold lower basal ATPase activity was observed by Mizuarai et al. and Morisaki et al., respectively, for the Q141K than for wild-type ABCG2. Login to comment
182 A recent investigation performed an exploratory, retrospective evaluation of the functional consequence of the ABCG2 421COA (Q141K) variant in 20 adult white patients, treated with diflomotecan [72]. Login to comment
185 This observation is in harmony with studies indicating a reduced protein expression and function for the Q141K variant, while seems to contradict the results of de Jong et al. As mentioned above, the allele frequency of ABCG2 421COA (Q141K) varies in diverse populations (Table 1), and in Japan and China this polymorphism appears to be common, with an overall allele frequency of 31.3G3.1%. Login to comment

[hide] Role of pharmacogenetics in irinotecan therapy. Cancer Lett. 2006 Mar 8;234(1):90-106. Epub 2005 Dec 15. de Jong FA, de Jonge MJ, Verweij J, Mathijssen RH
Role of pharmacogenetics in irinotecan therapy.
Cancer Lett. 2006 Mar 8;234(1):90-106. Epub 2005 Dec 15., 2006-03-08 [PMID:16343744]

Abstract [show]
In the treatment of advanced colorectal cancer, irinotecan has become one of the most important drugs, despite its sometimes unpredictable adverse effects. To understand why some patients experience severe adverse effects (diarrhea and neutropenia), while others do not, the metabolic pathways of this drug have to be unraveled in detail. Individual variation in expression of several phase I and phase II metabolizing enzymes and ABC-transporters involved in irinotecan metabolism and excretion, at least partly explains the observed pharmacokinetic interpatient variability. Although the difference in expression-level of these proteins to a certain amount is explained by physiologic and environmental factors, the presence of specific genetic determinants also does influence their expression and function. In this review, the role of genetic polymorphisms in the main enzyme-systems (carboxylesterase, cytochrome P450 3A, and uridine diphosphate-glucuronosyltransferase) and ABC-transporters (ABCB1, ABCC2, and ABCG2) involved in irinotecan metabolism, are discussed. Since at this moment the field of pharmacogenetics and pharmacogenomics is rapidly expanding and simultaneously more rapid and cost-effective screening methods are emerging, a wealth of future data is expected to enrich our knowledge of the genetic basis of irinotecan metabolism. Eventually, this may help to truly individualize the dosing of this (and other) anti-cancer agent(s), using a personal genetic profile of the most relevant enzymes for every patient.

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202 This ABCG2 421COA transversion results in an amino acid change of glutamine to lysine at codon 141 [151-153], and leads to altered substrate specificity and function of the mutant protein [151,154]. Login to comment

[hide] High-speed screening of human ATP-binding cassette... Methods Enzymol. 2005;400:485-510. Ishikawa T, Sakurai A, Kanamori Y, Nagakura M, Hirano H, Takarada Y, Yamada K, Fukushima K, Kitajima M
High-speed screening of human ATP-binding cassette transporter function and genetic polymorphisms: new strategies in pharmacogenomics.
Methods Enzymol. 2005;400:485-510., [PMID:16399366]

Abstract [show]
Drug transporters represent an important mechanism in cellular uptake and efflux of drugs and their metabolites. Hitherto a variety of drug transporter genes have been cloned and classified into either solute carriers or ATP-binding cassette (ABC) transporters. Such drug transporters are expressed in various tissues such as the intestine, brain, liver, kidney, and, importantly, cancer cells, where they play critical roles in the absorption, distribution, and excretion of drugs. We developed high-speed functional screening and quantitative structure-activity relationship analysis methods to study the substrate specificity of ABC transporters and to evaluate the effect of genetic polymorphisms on their function. These methods would provide powerful and practical tools for screening synthetic and natural compounds, and the deduced data can be applied to the molecular design of new drugs. Furthermore, we demonstrate a new "SNP array" method to detect genetic polymorphisms of ABC transporters in human samples.

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115 For this purpose, variant forms (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, E334stop, N590Y, D620N, R482G, and R482T) have been created by site‐ directed mutagenesis with the QuikChange site‐directed mutagensis kit (Stratagene, La Jolla, CA). Login to comment
233 Figure 13 shows the result of SNP array detection where nonsynonymous polymorphisms of ABCG2, that is, Q126stop and Q141K, were analyzed. Login to comment
235 Imai et al. (2002) identified three allelic variants in the ABCG2 gene, of which two were nonsynonymous SNPs (V12M and Q141K) and the third was a splice variant with deletion of nucleotides 944-949 that lacks Ala‐315 and Thr‐316 (Á315‐6). Login to comment
236 Compared with wild‐type transfected cells, ABCG2 Q141K variant‐transfected cells showed a low level of drug resistance that is associated with decreased protein expression. Login to comment
237 The SNP (Q141K) was postulated to cause increased sensitivity of normal cells to anticancer agents that are ABCG2 substrates such as SN‐38. Login to comment
242 It has been postulated that the 376C>T SNP may have a higher impact than the 421C>A polymorphism causing Q141K because active ABCG2 protein will not be synthesized from the variant allele. Login to comment
249 Detection of nonsynonymous polymorphisms (Q126stop and Q141K) of ABCG2 with the SNP array. Login to comment

[hide] Role of ABCG2/BCRP in biology and medicine. Annu Rev Pharmacol Toxicol. 2006;46:381-410. Krishnamurthy P, Schuetz JD
Role of ABCG2/BCRP in biology and medicine.
Annu Rev Pharmacol Toxicol. 2006;46:381-410., [PMID:16402910]

Abstract [show]
The protein variously named ABCG2/BCRP/MXR/ABCP is a recently described ATP-binding cassette (ABC) transporter originally identified by its ability to confer drug resistance that is independent of Mrp1 (multidrug-resistance protein 1) and Pgp (P-glycoprotein). Unlike Mrp1 and Pgp, ABCG2 is a half-transporter that must homodimerize to acquire transport activity. ABCG2 is found in a variety of stem cells and may protect them from exogenous and endogenous toxins. ABCG2 expression is upregulated under low-oxygen conditions, consistent with its high expression in tissues exposed to low-oxygen environments. ABCG2 interacts with heme and other porphyrins and protects cells and/or tissues from protoporphyrin accumulation under hypoxic conditions. Individuals who carry ABCG2 alleles that have impaired function may be more susceptible to porphyrin-induced toxicity. Abcg2 knock-out models have allowed in vivo studies of Abcg2 function in host and cellular defense. In combination with immunohistochemical analyses, these studies have revealed how ABCG2 influences the absorption, distribution, and excretion of drugs and cytotoxins.

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296 The two SNPs most frequently identified were in exon 2 (G34A, resulting in a V12M change) and exon 5 (C421A, resulting in a Q141K substitution). Login to comment

[hide] Topotecan is a substrate for multidrug resistance ... Curr Drug Metab. 2006 Jan;7(1):105-18. Tian Q, Zhang J, Chan SY, Tan TM, Duan W, Huang M, Zhu YZ, Chan E, Yu Q, Nie YQ, Ho PC, Li Q, Ng KY, Yang HY, Wei H, Bian JS, Zhou SF
Topotecan is a substrate for multidrug resistance associated protein 4.
Curr Drug Metab. 2006 Jan;7(1):105-18., [PMID:16454695]

Abstract [show]
Topotecan (TPT) is a semisynthetic water-soluble derivative of camptothecin (CPT) used as second-line therapy in patients with metastatic ovarian carcinoma, small cell lung cancer, and other malignancies. However, both dose-limiting toxicity and tumor resistance hinder the clinical use of TPT. The mechanisms for resistance to TPT are not fully defined, but increased efflux of the drug by multiple drug transporters including P-glycoprotein (PgP), multidrug resistance associated protein 1 (MRP1) and breast cancer resistance protein (BCRP) from tumor cells has been highly implicated. This study aimed to investigate whether overexpression of human MRP4 rendered resistance to TPT by examining the cytotoxicity profiles using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazonium bromide (MTT) assay and cellular accumulation of TPT in HepG2 cells stably overexpressing MRP4. Two kinds of cell lines, HepG2 with insertion of an empty vector plasmid (V/HepG2), HepG2 cells stably expressing MRP4 (MRP4/HepG2), were exposed to TPT for 4 or 48 hr in the absence or presence of various MRP4 inhibitors including DL-buthionine-(S,R)-sulphoximine (BSO), diclofenac, celecoxib, or MK-571. The intracellular accumulation of TPT and paclitaxel (a PgP substrate) by V/HepG2 and MRP4/HepG2 cells was determined by incubation of TPT with the cells and the amounts of the drug in cells were determined by validated HPLC methods. The study demonstrated that MRP4 conferred a 12.03- and 6.86-fold resistance to TPT in the 4- and 48-hr drug-exposure MTT assay, respectively. BSO, MK-571, celecoxib, or diclofenac sensitised MRP4/HepG2 cells to TPT cytotoxicity and partially reversed MRP4-mediated resistance to TPT. In addition, the accumulation of TPT was significantly reduced in MRP4/HepG2 cells compared to V/HepG2 cells, and one-binding site model was found the best fit for the MRP4-mediated efflux of TPT, with an estimated K(m) of 1.66 microM and V(max) of 0.341 ng/min/106 cells. Preincubation of MRP4/HepG2 cells with BSO (200 microM) for 24 hr, celecoxib (50 microM), or MK-571 (100 microM) for 2 hr significantly increased the accumulation of TPT over 10 min in MRP4/HepG2 cells by 28.0%, 37.3% and 32.5% (P < 0.05), respectively. By contrast, there was no significant difference in intracellular accumulation of paclitaxel in V/HepG2 and MRP4/HepG2 cells over 120 min. MRP4 also rendered resistance to adefovir dipivoxil (bis-POM-PMEA) and methotrexate, two reported MRP4 substrates. MRP4 did not exhibit any significant resistance to other model drugs including vinblastine, vincristine, etoposide, carboplatin, cyclosporine and paclitaxel in both long (48 hr) and short (4 hr) drug-exposure MTT assays. These findings indicate that MRP4 confers resistance to TPT and TPT is the substrate for MRP4. Further studies are needed to explore the role of MRP4 in resistance to, toxicity and pharmacokinetics of TPT in cancer patients.

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47 However, resistance levels of TPT are inconsistent in different BCRP overexpressing cell lines [51-59], probably due to the existence of three mutant variants of BCRP resulting in the amino acid changes at V12M, Q141K and D620N [63-67]. Login to comment

[hide] Inhibitors of cancer cell multidrug resistance med... Anticancer Drugs. 2006 Mar;17(3):239-43. Ahmed-Belkacem A, Pozza A, Macalou S, Perez-Victoria JM, Boumendjel A, Di Pietro A
Inhibitors of cancer cell multidrug resistance mediated by breast cancer resistance protein (BCRP/ABCG2).
Anticancer Drugs. 2006 Mar;17(3):239-43., [PMID:16520651]

Abstract [show]
Breast cancer resistance protein (BCRP/ABCG2) belongs to the ATP-binding cassette (ABC) transporter superfamily. It is able to efflux a broad range of anti-cancer drugs through the cellular membrane, thus limiting their anti-proliferative effects. Due to its relatively recent discovery in 1998, and in contrast to the other ABC transporters P-glycoprotein (MDR1/ABCB1) and multidrug resistance-associated protein (MRP1/ABCC1), only a few BCRP inhibitors have been reported. This review summarizes the known classes of inhibitors that are either specific for BCRP or also inhibit the other multidrug resistance ABC transporters. Information is presented on structure-activity relationship aspects and how modulators may interact with BCRP.

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81 Iressa transport/binding appears to be dependent on natural ABCG2 polymorphisms such as Q141K [36]. Login to comment

[hide] Functional validation of the genetic polymorphisms... Mol Pharmacol. 2006 Jul;70(1):287-96. Epub 2006 Apr 11. Tamura A, Watanabe M, Saito H, Nakagawa H, Kamachi T, Okura I, Ishikawa T
Functional validation of the genetic polymorphisms of human ATP-binding cassette (ABC) transporter ABCG2: identification of alleles that are defective in porphyrin transport.
Mol Pharmacol. 2006 Jul;70(1):287-96. Epub 2006 Apr 11., [PMID:16608919]

Abstract [show]
The ATP-binding cassette (ABC) transporter ABCG2 has been implicated to play a significant role in the response of patients to medication and/or the risk of diseases. To clarify the possible physiological or pathological relevance of ABCG2 polymorphisms, we have functionally validated single nucleotide polymorphisms (SNP) of ABCG2. In the present study, based on the currently available data on SNPs and acquired mutations, we have created a total of 18 variant forms of ABCG2 (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, F208S, S248P, E334stop, F431L, S441N, R482G, R482T, F489L, F571I, N590Y, and D620N) by site-directed mutagenesis and expressed them in insect cells. Because porphyrins are considered to be endogenous substrates for ABCG2, we have investigated the porphyrin transport activity of those variant forms in vitro. We herein provide evidence that the variants Q126stop, F208S, S248P, E334stop, and S441N are defective in porphyrin transport, whereas F489L exhibited impaired transport, approximately 10% of the activity observed for the wild type. Furthermore, Flp-In-293 cells expressing those variants were photosensitive. Thus, among those genetic polymorphisms of ABCG2, at least the hitherto validated alleles of Q126stop, S441N, and F489L are suggested to be of clinical importance related to the potential risk of porphyria.

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2 In the present study, based on the currently available data on SNPs and acquired mutations, we have created a total of 18 variant forms of ABCG2 (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, F208S, S248P, E334stop, F431L, S441N, R482G, R482T, F489L, F571I, N590Y, and D620N) by site-directed mutagenesis and expressed them in insect cells. Login to comment
82 GC indicates the percentage of guanine and cytosine contents in the PCR primer set. Tm shows the melting temperature (Tm) for each PCR primer set. Variant and Primers Primer Sequence (5Ј 3 3Ј) Primer Length GC Tm bases % °C V12M 33 39 55 Forward CGAAGTTTTTATCCCAATGTCACAAGGAAACAC Reverse GTGTTTCCTTGTGACATTGGGATAAAAACTTCG G51C 42 35 59 Forward ATCGAGTAAAACTGAAGAGTTGCTTTCTACCTTGTAGAAAAC Reverse GTTTTCGACAAGGTAGAAAGCAACTCTTCAGTTTTACTCGAT Q126stop 40 40 62 Forward GTAATTCAGGTTACGTGGTATAAGATGATGTTGTGATGGG Reverse CCCATCACAACATCATCTTATACCACGTAACCTGAATTAC Q141K 35 42 55 Forward CGGTGAGAGAAAACTTAAAGTTCTCAGCAGCTCTT Reverse AAGAGCTGCTGAGAACTTTAAGTTTTCTCTCACCG T153M 42 40 60 Forward CGGCTTGCAACAACTATGATGAATCATGAAAAAAACGAACGG Reverse CCGTTCGTTTTTTTCATGATTCATCATAGTTGTTGCAAGCCG Q166E 35 42 55 Forward GGATTAACAGGGTCATTGAAGAGTTAGGTCTGGAT Reverse ATCCAGACCTAACTCTTCAATGACCCTGTTAATCC I206L 36 44 59 Forward CTTATCACTGATCCTTCCCTCTTGTTCTTGGATGAG Reverse CTCATCCAAGAACAAGAGGGAAGGATCAGTGATAAG F208S 35 45 55 Forward TGATCCTTCCATCTTGTCCTTGGATGAGCCTACAA Reverse TTGTAGGCTCATCCAAGGACAAGATGGAAGGATCA S248P 35 40 55 Forward TTCATCAGCCTCGATATCCCATCTTCAAGTTGTTT Reverse AAACAACTTGAAGATGGGATATCGAGGCTGATGAA E334stop 35 31 55 Forward TCATAGAAAAATTAGCGTAGATTTATGTCAACTCC Reverse GGAGTTGACATAAATCTACGCTAATTTTTCTATGA F431L 28 60 62 Forward AGCTGGGGTTCTCCTCTTCCTGACGACC Reverse GGTCGTCAGGAAGAGGAGAACCCCAGCT S441N 34 47 59 Forward AACCAGTGTTTCAGCAATGTTTCAGCCGTGGAAC Reverse GTTCCACGGCTGAAACATTGCTGAAACACTGGTT F489L 46 34 62 Forward GAGGATGTTACCAAGTATTATACTTACCTGTATAGTGTACTTCATG Reverse CATGAAGTACACTATACAGGTAAGTATAATACTTGGTAACATCCTC F571I 36 47 61 Forward GTCATGGCTTCAGTACATCAGCATTCCACGATATGG Reverse CCATATCGTGGAATGCTGATGTACTGAAGCCATGAC N590Y 42 38 62 Forward CATAATGAATTTTTGGGACAATACTTCTGCCCAGGACTCAAT Reverse ATTGAGTCCTGGGCAGAAGTATTGTCCCAAAAATTCATTATG D620N 32 56 62 Forward GGTAAAGCAGGGCATCAATCTCTCACCCTGGG Reverse CCCAGGGTGAGAGATTGATGCCCTGCTTTACC veloped by using Western Lighting Chemiluminescent Reagent Plus (PerkinElmer Life and Analytical Sciences, Boston, MA) and detected by Lumino Imaging Analyzer FAS-1000 (Toyobo Engineering, Osaka, Japan). Login to comment
144 For this purpose, based on the currently available data on SNPs and acquired mutations, we generated variant forms (i.e., V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, F208S, S248P, E334stop, F431L, S441N, R482G, R482T, F489L, F571I, N590Y, and D620N) by site-directed mutagenesis. Login to comment
214 In the present study, based on the currently available data on SNPs and acquired mutations, we have created a total of 18 variant forms of ABCG2 (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, F208S, S248P, E334stop, F431L, S441N, R482G, R482T, F489L, F571I, N590Y, and D620N) by site-directed mutagenesis and expressed them in insect cells. Login to comment
219 The frequencies of the Q126stop, S441N, and F489L alleles are relatively low (less than 2%) compared with those of the V12M and Q141K alleles. Login to comment
224 Potential Risk Amino Acid Transport Allele Frequency cDNA Position Located on Exon Allele Data Sourcea Hemato MTX Wild-Type Allele % V12M ϩϩ ϩϩ 2.0-90.0 34 2 G A 1, 2, 4, 5, 7, 8 ૽૽ Q126stop - - 0.0-1.7 376 4 C T 1, 3, 5, 7 Q141K ϩϩ ϩϩ 0.0-35.5 421 5 C A 1, 2, 4, 5, 6, 7, 8 T153M ϩϩ ϩϩ 3.3 458 5 C T 5 R160Q N.D. N.D. 0.5 479 5 G A 8 Q166E ϩϩ ϩϩ N.D. 496 5 C G NCBI dbSNP rs1061017 I206L ϩϩ ϩϩ 10.0 616 6 A C 2 ૽૽ F208S - - N.D. 623 6 T C NCBI dbSNP rs1061018 ૽૽ S248P - - N.D. 742 7 T C NCBI dbSNP rs3116448 ૽૽ E334stop - - N.D. 1000 9 G T NCBI dbSNP rs3201997 F431L ϩϩ - 0.8 1291 11 T C 3 ૽૽ S441N - - 0.5 1322 11 G A 7 ૽ F489L ϩ - 0.5-0.8 1465 12 T C 3, 7 F571L ϩϩ ϩϩ 0.5 1711 14 T A NCBI dbSNP rs9282571 (૽૽) R575stop N.D. N.D. 0.5 1723 14 C T 8 N590Y ϩϩ ϩϩ 0.0-1.0 1768 15 A T 2, 5 D620N ϩϩ ϩϩ 0.5 1858 16 G A 8 Hemato, hematoporphyrin; NCBI, National Center for Biotechnology Information; N.D., not determined; ૽, risk of porphyria; (૽), potential risk is assumed as the lack of transport activity being as a result of a truncated protein. Login to comment

[hide] Genetic variation and haplotype structure of the A... Drug Metab Pharmacokinet. 2006 Apr;21(2):109-21. Maekawa K, Itoda M, Sai K, Saito Y, Kaniwa N, Shirao K, Hamaguchi T, Kunitoh H, Yamamoto N, Tamura T, Minami H, Kubota K, Ohtsu A, Yoshida T, Saijo N, Kamatani N, Ozawa S, Sawada J
Genetic variation and haplotype structure of the ABC transporter gene ABCG2 in a Japanese population.
Drug Metab Pharmacokinet. 2006 Apr;21(2):109-21., [PMID:16702730]

Abstract [show]
The ATP-binding cassette transporter, ABCG2, which is expressed at high levels in the intestine and liver, functions as an efflux transporter for many drugs, including clinically used anticancer agents such as topotecan and the active metabolite of irinotecan (SN-38). In this study, to elucidate the linkage disequilibrium (LD) profiles and haplotype structures of ABCG2, we have comprehensively searched for genetic variations in the putative promoter region, all the exons, and their flanking introns of ABCG2 from 177 Japanese cancer patients treated with irinotecan. Forty-three genetic variations, including 11 novel ones, were found: 5 in the 5'-flanking region, 13 in the coding exons, and 25 in the introns. In addition to 9 previously reported nonsynonymous single nucleotide polymorphisms (SNPs), 2 novel nonsynonymous SNPs, 38C>T (Ser13Leu) and 1060G>A (Gly354Arg), were found with minor allele frequencies of 0.3%. Based on the LD profiles between the SNPs and the estimated past recombination events, the region analyzed was divided into three blocks (Block -1, 1, and 2), each of which spans at least 0.2 kb, 46 kb, and 13 kb and contains 2, 24, and 17 variations, respectively. The two, eight, and five common haplotypes detected in 10 or more patients accounted for most (>90%) of the haplotypes inferred in Block -1, Block 1, and Block 2, respectively. The SNP and haplotype distributions in Japanese were different from those reported previously in Caucasians. This study provides fundamental information for the pharmacogenetic studies investigating the relationship between the genetic variations in ABCG2 and pharmacokinetic/pharmacodynamic parameters.

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17 In vitro studies have also indicated that a number of anticancer drugs are good substrates for ABCG2: e.g. topotecan, an irinotecan metabolite, 7-ethyl-10-hydroxycamptothecin (SN-38), and its glucuronide conjugate, SN-38G.810) Indeed, inhibition of the murine ABCG2 homologue, Bcrp 1, increases the bioavailability of topotecan when orally administered to mdr1aW1b- decient mice.11) In a clinical study, coadministration of topotecan with GF120918, a dual inhibitor for ABCG2 and P-glycoprotein, was shown to markedly increase the bioavailability and systemic exposure of topotecan.12) The cloning of ABCG2 from drug-selected cell lines revealed that acquired amino acid substitutions at residue 482 (Arg482Gly and Arg482Thr) of ABCG2 resulted in marked alterations in substrate recognition and transport ability.13) Thereafter, naturally occurring genetic variations in ABCG2 have been extensively examined in various ethnic populations1421) because they were expected to explain interindividual dierences in oral bioavailability and clearance of ABCG2 substrate drugs.22) Two nonsynonymous polymorphisms, 34GÀA (Val12Met) and 421CÀA (Gln141Lys), were detected at relatively high frequencies in most ethnic groups including Caucasians, Asians, and Africans.1416,1821,23) Both polymorphisms were reported to be associated with reduced protein expression in vitro andWor the increased sensitivity of the expressed cells toward several anticancer drugs although conicting data were also reported.16,2426) The expression of ABCG2 protein in placenta was signicantly lower in homozygotes with the 421A alleles than in those with the 421C alleles, while 34GÀA (Val12Met) did not aect ABCG2 protein expression.23) However, in intestinal samples, no association was found between the ABCG2 protein levels and the 421CÀA (Gln141Lys) genotype.18) A pharmacokinetic study showed that 421A (Gln141Lys) was unlikely to inuence the in vivo disposition of irinotecan in European Caucasian cancer patients.27) On the other hand, diomotecan pharmacokinetics were signicantly aected by the 421A genotype.28) To explain these inconsistencies, the elucidation of the haplotype structure of ABCG2 would be helpful; however, only limited information is available for the linkage disequilibrium (LD) prole and haplotype structure of this gene.20,21) Also, to facilitate future pharmacogenetic studies on ABCG2 genetic variations, haplotype analysis using its high-density SNPs found in a large number of samples is warranted. Login to comment
80 However, marked ethnic dierences in the allele frequencies were observed with |1203ä |1200delCTCA, 34GÀA (Val12Met), 421CÀA (Gln141Lys), and some intronic variations. Login to comment
85 115Haplotype Structure in Human ABCG2 (from |1836 to |1175 bp upstream of the translational start site) of the basal promoter,30) and was suggested to inuence irinotecan pharmacokinetics.31) The frequencies of two well-known nonsynonymous SNPs, 34GÀA (Val12Met) and 421CÀA (Gln141Lys), were 0.192 and 0.319 in our study, which were comparable to those in Chinese (0.204 and 0.2220.350, respectively).20,27) However, the frequencies were much higher than those in Caucasians (0.020.065 and 0.080.15), African-Americans (00.09 and 00.05), and a Swedish population (0.02 and 0.1).18,19,21,23,27) Of other relatively rare nonsynonymous SNPs, 376CÀT (Gln126X), 1291TÀC (Phe431Leu), 1322GÀA (Ser441Asn), 1465TÀC (Phe489Leu), and 1515delC (Phe506SerfsX4) were already detected in a Japanese population by Itoda et al.17) andWor Kobayashi et al.,23) but not found in other ethnic groups. Login to comment
101 Strong LDs (r2 À0.7) were observed between |262CÀT and 1098GÀA (Glu366Glu) (r2 0.798), between 421CÀA (Gln141Lys) and IVS12{49GÀT (r2 0.709), and among IVS11{20AÀG, IVS1321CÀT, and IVS15{110CÀT (r2 À0.720). Login to comment
115 No close associations were observed (r2 º0.70) between the variations across the blocks except for one pair of SNPs, 421CÀA (Gln141Lys) and IVS12{49GÀT, Fig. . Login to comment
130 In Block 1, seven haplotype groups (*1 to *7) were inferred, and the groups of *2 to *7 harbored non-synonymous SNPs, 421CÀA (Gln141Lys) (*2), 34GÀA (Val12Met) (*3), 376CÀT (Gln126X) (*4), 38CÀT (Ser13Leu) (*5), 479GÀA (Arg160Gln) (*6), and 1060GÀA (Gly354Arg) (*7). Login to comment
134 The haplotype-tagging SNPs (htSNPs) that were able to resolve the 8 common haplotypes were the following 7 variations: IVS199GÀA, 34GÀA (Val12Met), IVS293TÀC, 376CÀT (Gln126X), 421CÀA (Gln141Lys), IVS6217AÀG, and IVS6204CÀT. Login to comment
154 Mizuarai et al. showed that 34GÀA (Val12Met) was associated with reduced drug resistance in polarized LLC-PK1 cells, which might be caused by its impaired apical membrane localization.25) In contrast, several groups did not nd any signicant eects of Val12Met on the protein expression levels as well as drug resistance using stable and transient mammalian expression systems.16,24,26) According to Imai et al. and Kondo et al.,16,24) the Gln141Lys substitution resulted in decreased protein expression and reduced drug resistance. Login to comment
162 Except for Val12Met, Gln126X, and Gln141Lys, the allele frequencies of eight nonsynonymous SNPs were less than 0.01, and these low frequency variations do not largely contribute to the overall heterozygosity of ABCG2; however, they might have clinical importance. Login to comment
174 Nevertheless, the frequency (31.9z) of the haplotype *2a in Block 1 harboring 421CÀA (Gln141Lys) was comparable to their counterpart (20.4z). Login to comment
176 Therefore, the substitution itself of Gln141 to Lys is likely to be responsible for the reduced expression of ABCG2 protein in placenta as demonstrated by Kobayashi et al.23) Furthermore, Chinese and Japanese share several common Block 2 haplotypes, *1a, *1b, *1c, and *1dW *1e. Login to comment

[hide] Role of pharmacogenetics of ATP-binding cassette t... Pharmacol Ther. 2006 Nov;112(2):457-73. Cascorbi I
Role of pharmacogenetics of ATP-binding cassette transporters in the pharmacokinetics of drugs.
Pharmacol Ther. 2006 Nov;112(2):457-73., [PMID:16766035]

Abstract [show]
Interindividual differences of drug response are an important cause of treatment failures and adverse drug reactions. The identification of polymorphisms explaining distinct phenotypes of drug metabolizing enzymes contributed in part to the understanding of individual variations of drug plasma levels. However, bioavailability also depends on a major extent from the expression and activity of drug transport across biomembranes. In particular efflux transporters of the ATP-binding cassette (ABC) family such as ABCB1 (P-glycoprotein, P-gp), the ABCC (multidrug resistance-related protein, MRP) family and ABCG2 (breast cancer resistance protein, BCRP) have been identified as major determinants of chemoresistance in tumor cells. They are expressed in the apical membranes of many barrier tissue such as the intestine, liver, blood-brain barrier, kidney, placenta, testis and in lymphocytes, thus contributing to plasma, liquor, but also intracellular drug disposition. Since expression and function exhibit a broad variability, it was hypothesized that hereditary variances in the genes of membrane transporters could explain at least in part interindividual differences of pharmacokinetics and clinical outcome of a variety of drugs. This review focuses on the functional significance of single nucleotide polymorphisms (SNP) of ABCB1, ABCC1, ABCC2, and ABCG2 in in vitro systems, in vivo tissues and drug disposition, as well as on the clinical outcome of major indications.

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901 Both identified 34G>A (V12M) and 421C>A (Q141K). Login to comment
909 5.3. Association to activity and drug bioavailability HEK-293 cells, transfected with wild-type or V12N, ABCG2 showed apical staining with an ABCG2 antibody, but high intracellular staining in case of Q141K, suggesting impaired membrane trafficking or incorrect membrane insertion. Login to comment
917 0.005 Exon 4 c. 376 C>T Q126stop 0.01 Lack of function 0.00a 0.00b Exon 5 c. 421 C>A Q141K 0.35 Reduced activity (Mizuarai et al., 2004; Kobayashi et al., 2005; Sparreboom et al., 2005) 0.11a 0.02b IVS 5-16 A>G ? Login to comment

[hide] Role of BCRP 421C>A polymorphism on rosuvastatin p... Clin Chim Acta. 2006 Nov;373(1-2):99-103. Epub 2006 May 13. Zhang W, Yu BN, He YJ, Fan L, Li Q, Liu ZQ, Wang A, Liu YL, Tan ZR, Fen-Jiang, Huang YF, Zhou HH
Role of BCRP 421C>A polymorphism on rosuvastatin pharmacokinetics in healthy Chinese males.
Clin Chim Acta. 2006 Nov;373(1-2):99-103. Epub 2006 May 13., [PMID:16784736]

Abstract [show]
BACKGROUND: Rosuvastatin, a novel potent HMG-CoA reductase inhibitor, is excreted into bile mediated by breast cancer resistance protein (BCRP). Our objective was to determine the association between the most frequent single nucleotide polymorphisms (SNPs) of the BCRP (421C>A) and the pharmacokinetics of rosuvastatin. METHOD: Pre-screening of SLCO1B1 521TC and CYP2C9*1/*3 were performed before this pharmacokinetic study. Fourteen healthy volunteers who are SLCO1B1 521TT and CYP2C9*1/*1 wild-type homozygotes were selected to participate in this study. Seven were 421CC wild-type of BCRP, the others were carriers with at least one 421C>A mutant allele (five subjects had a genotype of 421CA and two were homozygotes of 421AA). Each was given a single oral dose of 20 mg rosuvastatin. The plasma concentrations of rosuvastatin were measured for up to 72 h by LC-MS. RESULTS: The pharmacokinetic parameters of rosuvastatin showed a significantly difference between the two genotyped groups. The AUC(0-72) and AUC(0-infinity) of rosuvastatin were lower in the 421CC group than in the 421CA+421AA group (33.8+/-11.4 vs. 59.6+/-22.2 ng.h/ml, P=0.018; 34.9+/-11.9 vs. 62.2+/-23.5 ng.h/ml, P=0.018), respectively. The C(max) value was higher in the 421CA+421AA group than that in the 421CC group (9.9+/-5.4 vs. 5.1+/-2.4 ng/ml, P=0.048). The CL/F value was lower in the 421CA+421AA group than that in the 421CC group (384.7+/-161.2 vs. 674.0+/-297.6 l/h, P=0.043). The T(1/2) and T(max) values showed no difference between these groups. CONCLUSIONS: The BCRP 421C>A polymorphism may play an important role in the pharmacokinetics of rosuvastatin in healthy Chinese males after the exclusion of impact of SLCO1B1 and CYP2C9 genetic polymorphism.

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91 In most of the ethnic populations, the 421C>A (Q141K) polymorphism occurred at high frequency. Login to comment
93 It is concluded that the functional impairment of 421C>A variant was due to reduced ATPase activity for Q141K [11]. Login to comment

[hide] The livestock photosensitizer, phytoporphyrin (phy... Res Vet Sci. 2006 Dec;81(3):345-9. Epub 2006 Jun 30. Robey RW, Fetsch PA, Polgar O, Dean M, Bates SE
The livestock photosensitizer, phytoporphyrin (phylloerythrin), is a substrate of the ATP-binding cassette transporter ABCG2.
Res Vet Sci. 2006 Dec;81(3):345-9. Epub 2006 Jun 30., [PMID:16808938]

Abstract [show]
Hepatogenous photosensitization occurs in livestock following damage to the liver or biliary apparatus that results in impaired excretion of phytoporphyrin (phylloerythrin), a photosensitizer. Based on earlier observations that porphyrin-based photosensitizers are substrates of the ATP-binding cassette transporter ABCG2, we examined the ability of the hepatic transporters ABCB1 (P-glycoprotein) and ABCG2 to transport phytoporphyrin. Transport of phytoporphyrin was blocked by the ABCG2-specific inhibitor fumitremorgin C (FTC) in human embryonic kidney cells transfected with full length human ABCG2, while no transport by cells transfected with human ABCB1 was noted. FTC-inhibited transport of phytoporphyrin was also demonstrated in ABCG2-expressing LLC-PK1 pig kidney cells, consistent with the idea that the pig orthologue, like human ABCG2, transports the photosensitizer. ABCG2 expression was confirmed by immunohistochemistry in the hepatocytes of cow, pig and sheep livers. We conclude that phytoporphyrin is a substrate for ABCG2 and that the transporter is likely responsible for its biliary excretion.

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54 A non-synonymous single nucleotide polymorphism in the human ABCG2 gene resulting in a glutamine to lysine change at amino acid 141 (Q141K) of the protein has been shown to result in impaired transport of substrate drugs and decreased ATPase activity compared to wild-type protein (Mizuarai et al., 2004; Morisaki et al., 2005). Login to comment

[hide] Multidrug resistance: retrospect and prospects in ... Curr Med Chem. 2006;13(16):1859-76. Perez-Tomas R
Multidrug resistance: retrospect and prospects in anti-cancer drug treatment.
Curr Med Chem. 2006;13(16):1859-76., [PMID:16842198]

Abstract [show]
Conventional cancer chemotherapy is seriously limited by the multidrug resistance (MDR) commonly exhibited by tumour cells. One mechanism by which a living cell can achieve multiple resistances is via the active efflux of a broad range of anticancer drugs through the cellular membrane by MDR proteins. Such drugs are exported in both ATP-dependent and -independent manners, and can occur despite considerable concentration gradients. To the ATP-dependent group belongs the ATP-binding cassette (ABC) transporter family, which includes P-gp, MRP, BCRP, etc. Another protein related to MDR, though not belonging to the ABC transporter family, is lung resistance-related protein (LRP). All of these proteins are involved in diverse physiological processes, and are responsible for the uptake and efflux of a multitude of substances from cancer cells. Many inhibitors of MDR transporters have been identified over the years. Firstly, MDR drugs were not specifically developed for inhibiting MDR; in fact, they had other pharmacological properties, as well as a relatively low affinity for MDR transporters. They included compounds of diverse structure and function, such as verapamil and cyclosporine, and caused side effects. Secondly, the new drugs were more inhibitor-specific, in terms of MDR transport, and were designed to reduce such side effects (e.g., R-verapamil, dexniguldipine, etc.). Unfortunately, they displayed poor response in clinical studies. Recently, new compounds obtained from drug development programs conducted by the pharmaceutical industry are characterized by a high affinity to MDR transporters and are efficient at nanomolar concentrations. Some of these compounds (e.g., MS-209) are currently under clinical trials for specific forms of advanced cancers. We aim to provide an overview of the properties associated with those mammalian MDR transporters known to mediate significant transport of relevant drugs in cancer treatments. We also summarize recent advances concerning resistance to cancer drug therapies with respect to the function and overexpression of ABC and LRP multidrug transporters.

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219 The results showed three BCRP-coding SNPs [G34A (V12M), C376T (Q126stop) and C421A (Q141K)] (Fig. 6). Login to comment
220 The V12M and the Q141K SNPs greatly Fig. (6). Login to comment
223 In addition, cells transfected with Q141K-BCRP cDNA expressed low amounts of this protein and showed low levels of drug resistance than did wild-type BCRP-transfected cells. Login to comment

[hide] Breast cancer resistance protein in pharmacokineti... Expert Opin Drug Metab Toxicol. 2005 Dec;1(4):595-611. Xia CQ, Yang JJ, Gan LS
Breast cancer resistance protein in pharmacokinetics and drug-drug interactions.
Expert Opin Drug Metab Toxicol. 2005 Dec;1(4):595-611., [PMID:16863427]

Abstract [show]
Breast cancer resistance protein (BCRP), also known as ABCG2, ABCP and MXR, is a member of the ATP-binding cassette transporter G family. BCRP functions as a biological barrier that extrudes xenobiotics out of cells. The broad substrate specificity and tissue distributions of BCRP in the body make this transporter one of the major efflux transporters in chemotherapy. Recent studies have demonstrated that BCRP exerts a great impact on drug absorption and disposition. This review focuses on the role of BCRP in pharmacokinetics as well as in vitro and in vivo strategies to evaluate hepatic/intestinal BCRP-mediated drug transports and drug-drug interactions. The impacts of polymorphism and gender difference of BCRP are also discussed.

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173 The mutation C421A at exon 5, with an amino acid change of Gln to Lys at codon 141, has been reported with a reduced BCRP protein level, but not mRNA level in PA-317 cells, and a decreased BCRP efflux function in transfected LLC-PK and HEK-283 cells [73,74,76]. Login to comment
511 Pharmacogenetics (2003) 13:19-28. 73. IMAI Y, NAKANE M, KAGE K et al.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Human ABC transporter ABCG2 in xenobiotic protecti... Drug Metab Rev. 2006;38(3):371-91. Wakabayashi K, Tamura A, Saito H, Onishi Y, Ishikawa T
Human ABC transporter ABCG2 in xenobiotic protection and redox biology.
Drug Metab Rev. 2006;38(3):371-91., [PMID:16877258]

Abstract [show]
Human ATP-binding cassette (ABC) transporter ABCG2 (BCRP/MXR/ABCP) is regarded as a member of the phase III system of xenobiotic metabolism. This efflux pump is suggested to be responsible for protecting the body from toxic xenobiotics and for removing toxic metabolites. The aim of this review article is to address new aspects of ABCG2 related to redox biology, namely the posttranslational modification (intra- and intermolecular disulfide bond formation) of ABCG2 protein and the transport of porphyrin and chlorophyll metabolites, as well as the high-speed screening and QSAR analysis method to evaluate ABCG2-drug interactions.

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176 Based on the currently available data on SNPs and acquired mutations, we have created a total of 18 variant forms of ABCG2 (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, F208S, S248P, E334stop, F431L, S441N, R482G, R482T, F489L, F571I, N590Y, and D620N) by site-directed mutagenesis and expressed them in Sf9 insect cells. Login to comment

[hide] Association of enzyme and transporter genotypes wi... Clin Pharmacol Ther. 2006 Aug;80(2):192-201. Gardner ER, Burger H, van Schaik RH, van Oosterom AT, de Bruijn EA, Guetens G, Prenen H, de Jong FA, Baker SD, Bates SE, Figg WD, Verweij J, Sparreboom A, Nooter K
Association of enzyme and transporter genotypes with the pharmacokinetics of imatinib.
Clin Pharmacol Ther. 2006 Aug;80(2):192-201., [PMID:16890580]

Abstract [show]
OBJECTIVE: Our objective was to explore the relationships between imatinib pharmacokinetics and 9 allelic variants in 7 genes coding for adenosine triphosphate-binding cassette transporters (ABCB1 and ABCG2) and enzymes (cytochrome P450 [CYP] 2C9, CYP2C19, CYP2D6, CYP3A4, and CYP3A5) of putative relevance for imatinib. METHODS: Imatinib transport in vitro was studied by use of human embryonic kidney 293 cells transfected with wild-type ABCG2 and an ABCG2 Q141K clone. Steady-state pharmacokinetics of imatinib was obtained in 82 patients with gastrointestinal stromal tumors treated with oral imatinib at doses ranging from 100 to 1000 mg/d. Genotyping was carried out via direct sequencing or restriction fragment length polymorphism-based techniques. RESULTS: Human embryonic kidney 293 cells transfected with ABCG2 Q141K exhibited greater drug accumulation in vitro in comparison with cells expressing wild-type ABCG2 (P = .028). However, pharmacokinetic parameters of imatinib in vivo were not statistically significantly different in 16 patients who were heterozygous for ABCG2 421C>A compared with 66 patients carrying the wild-type sequence (P = .479). The apparent oral clearance of imatinib was potentially reduced in individuals with at least 1 CYP2D6*4 allele (median, 7.78 versus 10.6 L/h; P = .0695). Pharmacokinetic parameters were not related to any of the other multiple-variant genotypes (P >or= .230), possibly because of the low allele frequencies. CONCLUSIONS: This study indicates that common genetic variants in the evaluated genes have only a limited impact on the pharmacokinetics of imatinib. Further investigation is required to quantitatively assess the clinical significance of homozygous variant ABCG2 and CYP2D6 genotypes in patients treated with imatinib.

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1 Methods: Imatinib transport in vitro was studied by use of human embryonic kidney 293 cells transfected with wild-type ABCG2 and an ABCG2 Q141K clone. Login to comment
4 Results: Human embryonic kidney 293 cells transfected with ABCG2 Q141K exhibited greater drug accumulation in vitro in comparison with cells expressing wild-type ABCG2 (P ‫؍‬ .028). Login to comment
26 In particular, a functional single-nucleotide polymorphism (SNP) has been identified in exon 5 of the ABCG2 gene, in which a CϾA nucleotide transition at position 421 (ABCG2 421CϾA) results in a nonsynonymous variant protein with a glutamine-to-lysine amino acid substitution at codon 141 (Q141K).5 In this study we tested the hypothesis that the extensive interindividual variability in the pharmacokinetics of imatinib is associated with the ABCG2 421CϾA polymorphism in a cohort of patients with gastrointestinal stromal tumors. Login to comment
29 Human embryonic kidney cells (HEK293) transfected with pcDNA3, wild-type ABCG2, or an ABCG2 Q141K clone were generated as described previously.8 Cells were maintained in N-[2-hydroxyethyl]piperazine-NЈ-[2-ethanesulfonic acid] (HEPES)-buffered RPMI-1640 medium (Gibco BRL, Paisley, United Kingdom) supplemented with 10% (vol/vol) fetal calf serum. Login to comment
82 To investigate the potential effect of the ABCG2 421CϾA variant on the protein`s ability to transport imatinib, human embryonic kidney cells (HEK293) transfected with pcDNA3, wild-type ABCG2, or an ABCG2 Q141K clone were exposed to 14 C-labeled imatinib for 2 hours, and the intracellular concentration was determined. Login to comment
84 The cells homozygous for the studied variant exhibited significantly greater drug accumulation than the cells expressing wild-type ABCG2 (P ϭ .028), suggesting impaired outward-directed transport of imatinib by the ABCG2 Q141K variant (Fig 1). Login to comment
102 Top, Comparative intracellular accumulation of 14 C-labeled imatinib in human embryonic kidney cells (HEK293) transfected with wild-type ABCG2 (HEK293/R) and ABCG2 Q141K clone (HEK293/K-5). Login to comment
117 The studied variant in ABCG2 is an SNP causing a nonsynonymous change in the protein sequence, in which a 421C-to-A transition in exon 5 leads to a glutamine-to-lysine amino acid substitution at codon 141 (Q141K). Login to comment
119 Although our own current in vitro transport studies in HEK293 cells transfected with the homozygous Q141K variant show Table I. Login to comment
120 Genotype and allele frequencies for studied variant genes Variant† Effect‡ Activity§ Genotype frequency [No. of patients (%)]࿣ Allele frequency¶ Wt Het Var p q Enzyme genotypes CYP2C9*2 (430CϾT) R144C (exon 3) Decreased 60 (85.7) 9 (12.9) 1 (1.43) 0.921 0.0786 CYP2C9*3 (1075AϾC) I359L (exon 7) Decreased 60 (85.7) 10 (14.3) 0 (0) 0.929 0.0714 CYP2C19*2 (681GϾA) Splice defect (exon 4) None 40 (57.1) 27 (38.6) 3 (4.29) 0.764 0.236 CYP2C19*3 (636GϾA) W212X (exon 5) None 70 (100) 0 (0) 0 (0) 1.000 0.000 CYP2D6*4 (1846GϾA) Splice defect (exon 4) None 42 (62.7) 24 (35.8) 1 (1.49) 0.806 0.194 CYP3A4*1B (-392AϾG) Promoter Normal/ increased 64 (91.4) 6 (8.57) 0 (0) 0.957 0.0429 CYP3A5*3C (6986AϾG) Splice defect (intron 3) Severely decreased 0 (0) 13 (18.6) 57 (81.4) 0.0929 0.907 Transporter genotypes ABCB1 3435CϾT I1145I (exon 26) Reduced 21 (25.6) 41 (50.0) 20 (24.4) 0.506 0.494 ABCG2 421CϾA Q141K (exon 5) Reduced 66 (80.5) 16 (19.5) 0 (0) 0.902 0.0976 Wt, Homozygous wild-type patient; Het, heterozygous patient; Var, homozygous variant patient; p, frequency for wild-type allele; q, frequency for variant allele. Login to comment

[hide] Expression and function of multidrug resistance tr... Expert Opin Drug Metab Toxicol. 2005 Aug;1(2):233-46. Scherrmann JM
Expression and function of multidrug resistance transporters at the blood-brain barriers.
Expert Opin Drug Metab Toxicol. 2005 Aug;1(2):233-46., [PMID:16922639]

Abstract [show]
The presence of active carrier-mediated transport of substrates from the brain to the blood is a major feature of the barrier properties of the blood-brain barrier (BBB). These proteins lie in the luminal or abluminal membranes of the endothelial cells that form the BBB. Some are ATP-binding cassette proteins (ABC) and many amphipathic cationic drugs are carried by P-glycoprotein (ABCB1) or ABCG2, which lie at the luminal pole of the BBB. Several multidrug resistance-associated proteins (MRPs, ABCCs) are also present on the membranes of brain microvessels; these are mainly involved in the efflux of anionic compounds. All these ABC proteins help to protect the brain and form a critical target for CNS pharmaceuticals, influencing the clinical variability of responses to, and the design of, these drugs.

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505 IMAI Y, NAKANE M, KAGE K et al.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Human multidrug resistance ABCB and ABCG transport... Physiol Rev. 2006 Oct;86(4):1179-236. Sarkadi B, Homolya L, Szakacs G, Varadi A
Human multidrug resistance ABCB and ABCG transporters: participation in a chemoimmunity defense system.
Physiol Rev. 2006 Oct;86(4):1179-236., [PMID:17015488]

Abstract [show]
In this review we give an overview of the physiological functions of a group of ATP binding cassette (ABC) transporter proteins, which were discovered, and still referred to, as multidrug resistance (MDR) transporters. Although they indeed play an important role in cancer drug resistance, their major physiological function is to provide general protection against hydrophobic xenobiotics. With a highly conserved structure, membrane topology, and mechanism of action, these essential transporters are preserved throughout all living systems, from bacteria to human. We describe the general structural and mechanistic features of the human MDR-ABC transporters and introduce some of the basic methods that can be applied for the analysis of their expression, function, regulation, and modulation. We treat in detail the biochemistry, cell biology, and physiology of the ABCB1 (MDR1/P-glycoprotein) and the ABCG2 (MXR/BCRP) proteins and describe emerging information related to additional ABCB- and ABCG-type transporters with a potential role in drug and xenobiotic resistance. Throughout this review we demonstrate and emphasize the general network characteristics of the MDR-ABC transporters, functioning at the cellular and physiological tissue barriers. In addition, we suggest that multidrug transporters are essential parts of an innate defense system, the "chemoimmunity" network, which has a number of features reminiscent of classical immunology.

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997 In healthy individuals or patients, altogether eight nonsynonymous (V12M, Q141K, I206L, F431L, S441N, F489L, N590Y, D620N), five synonymous (silent) (c. Login to comment
1006 From these numerous reported alterations, two protein variants, V12M, and Q141K, were found in relatively high frequencies, with significant differences in allele frequencies in different areas of the world (Fig. 11). Login to comment
1010 The Q141K polymorphism leads to the replacement of the uncharged glutamine residue with a positively charged lysine within the ATP-binding domain, between the Walker A motif (amino acids 83-89) and the signature region (amino acids 186-189). Login to comment
1011 The Q141K variant was also detected in all ethnic groups tested: the allele frequency ranged between 1 and 35% (the African and African-American subjects with low, while the Japanese and Chinese populations with high allele frequencies; see Refs. 45, 74, 185). Login to comment
1016 (251), by using stable mammalian expression systems, found that in PA317 or HEK-293 cells the expressed Q141K ABCG2 protein had a lower expression level than the wild-type ABCG2, or the V12M variant. Login to comment
1018 (251) demonstrated that both the V12M and Q141K ABCG2 could reach the plasma membrane in the HEK-293 cells, while a significant portion of Q141K remained intracellular. Login to comment
1019 Other studies found a 30-40% reduction in cell surface expression of the Q141K variant, despite a similar mRNA level than the wild-type ABCG2 (145, 188). Login to comment
1020 Recent investigation of 99 Japanese placenta samples revealed that individuals homozygous for the Q141K variant showed significantly lower expression levels of this transporter protein, while the heterozygous samples displayed an intermediate expression level (185). Login to comment
1023 (247) expressed ABCG2 in polarized LLC-PK1 cells, and by using confocal microscopy, the authors observed that the wild-type ABCG2 and Q141K showed mainly apical staining, while the V12M variant 1210 SARKADI, HOMOLYA, SZAKA´ CS, AND VA´ RADI Physiol Rev • VOL 86 • OCTOBER 2006 • www.prv.org showed mostly intracellular localization. Login to comment
1025 (188) also used LLC-PK1 cells to express the V12M and Q141K variants and found that all polymorphisms, including V12M and Q141K, had an apical localization. Login to comment
1028 (247), when the V12M or Q141K-transfected LLC-PK1 cells were challenged by mitoxantrone, topotecan, or an indolocarbazole topoisomerase I inhibitor. Login to comment
1030 (251) found that only the Q141K variant had a moderately lower level resistance against mitoxantrone, topotecan, or SN-38, compared with the wild-type ABCG2-transfected cells. Login to comment
1032 Q141K is mapped to a functionally important ABC region of ABCG2; therefore, it is possible that the ATPase activity of this variant is altered. Login to comment
1033 Two studies compared the vanadate-sensitive ATPase activity of ABCG2 V12M and Q141K variants, using Sf9 (Spodoptera frugiperda) cell membranes (247, 251). Login to comment
1034 A reduced basal ATPase activity was observed by both groups for the Q141K variant. Login to comment
1038 Clearly, more detailed studies are required to clarify the mechanism of a reduced protein expression for Q141K, and the altered cellular localization found for the V12M and Q141K variants under certain conditions. Login to comment
1047 A recent investigation performed an exploratory, retrospective evaluation of the functional consequence of the ABCG2 Q141K variant in 20 adult patients, treated with diflomotecan, a synthetic derivative of camptothecin (350). Login to comment
1050 This observation is in harmony with studies indicating a reduced protein expression and function for the Q141K variant, while it seems to contradict the results of De Jong et al. Login to comment
1052 As mentioned above, the allele frequency of Q141K varies in diverse populations, and in Japan and China, this polymorphism appears to be common, with an overall allele frequency of ϳ30%. Login to comment

[hide] Towards understanding the mechanism of action of t... J Mol Graph Model. 2007 Mar;25(6):837-51. Epub 2006 Aug 30. Li YF, Polgar O, Okada M, Esser L, Bates SE, Xia D
Towards understanding the mechanism of action of the multidrug resistance-linked half-ABC transporter ABCG2: a molecular modeling study.
J Mol Graph Model. 2007 Mar;25(6):837-51. Epub 2006 Aug 30., [PMID:17027309]

Abstract [show]
The ATP-binding cassette protein ABCG2 is a member of a broad family of ABC transporters with potential clinical importance as a mediator of multidrug resistance. We carried out a homology and knowledge-based, and mutationally improved molecular modeling study to establish a much needed structural framework for the protein, which could serve as guidance for further genetic, biochemical, and structural analyses. Based on homology with known structures of both full-length and nucleotide-binding domains (NBD) of ABC transporters and structural knowledge of integral membrane proteins, an initial model of ABCG2 was established. Subsequent refinement to conform to the lipophilic index distributions in the transmembrane domain (TMD) and to the results of site-directed mutagenesis experiments led to an improved model. The complete ABCG2 model consists of two identical subunits facing each other in a closed conformation. The dimeric interface in the nucleotide-binding domain (NBD) involves a characteristic nucleotide sandwich and the interface in the TMD consists of the TM helices 1-3 of one subunit and the helices 5 and 6 of the other. The interface between the NBD and the TMD is bridged by the conserved structural motif between TM2 and TM3, the intracellular domain 1 (ICD1), and the terminal beta-strand (S6) of the central beta-sheet in the NBD. The apparent flexibility of the ICD1 may play a role in transmitting conformational changes from the NBD to the TMD or from the TMD to the NBD.

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112 Q141K, a SNP [22,23], is located on the surface of the small sub-domain near the conserved H3 helix Y.-F Li et al. / Journal of Molecular Graphics and Modelling 25 (2007) 837-851 841 Table 1 Sequence identity and similarity in the NBD and TMD between ABCG2 and selected ABC transporters ABC transporter NBD domain (%)a ABC transporter TM domain (%)b Identity Similarityc Identity Similarity ABCG2 100.0 (234)d 100.0 (234) ABCG2 100.0 (295) 100.0 (295) EcMalK 26.5 (62) 42.7 (100) VcMsbA 9.5 (28) 19.5 (56) TlMalK 22.6 (53) 42.3 (99) EcMsbA 6.4 (19) 18.3 (54) His-P 18.4 (43) 40.6 (95) LcLmrA 8.5 (25) 21.0 (62) MJ1267 17.5 (41) 38.0 (89) Pgp-n 6.1 (18) 19.7 (58) LMR-A 20.9 (49) 40.2 (94) Pgp-c 6.1 (18) 15.6 (46) CFTR-N1 20.5 (48) 34.6 (81) Dro-Wht 22.0 (65) 40.3 (119) TAP1 17.1 (40) 42.7 (100) Pgp-n-reve (16) (30) a Sequence identities and similarities are based on alignments in Fig. 2. b Sequence identities and similarities are based on alignments in Fig. 4. c Similarity is defined for amino acid residues in the alignment, which share common physical or chemical properties such as size, hydrophobicity or charge. Login to comment
123 Patients carrying the Q141K SNP were shown to have elevated drug levels in plasma when treated with topotecan or diflomotecan [22,44-49]. Login to comment
124 From its location in the molecule, the Q141K polymorphism should not disrupt the folding of the subunit; rather it introduces an additional positive charge in an already very positively charged surface (residues R137, R147, K157, R160, and R163 are in close proximity). Login to comment
174 Subsequently, a symmetric dimer Y.-F Li et al. / Journal of Molecular Graphics and Modelling 25 (2007) 837-851844 Table 4 Locations of mutations as predicted by the ABCG2 model and functional correlation Mutation Position in ABCG2 Phenotype Reference V12M N-terminal Membrane localization, SNP, and somewhat lower expression and lower resistance [22] S25Pa N-terminal Low drug resistance for the cell line due to lower expression at cell surface [42] T82Aa NBD, Walker A Low drug resistance for the cell line due to lower expression at cell surface [42] K86M NBD, Walker A No expression at cell surface, retained in the Golgi [43] K86I NBD, Walker A No expressed at cell surface [43] Q141K NBD SNP with lower protein expression and low drug resistance [22,23] T237V NBD Fully functional b I239K,R NBD Loss of expression may be due to structural disruption b R309G Linkerc Low drug resistance [42] D315-6 Linker Deletion mutant for A315 and T316. Login to comment

[hide] Hoosier Oncology Group randomized phase II study o... Clin Cancer Res. 2006 Oct 15;12(20 Pt 1):6094-9. Hahn NM, Marsh S, Fisher W, Langdon R, Zon R, Browning M, Johnson CS, Scott-Horton TJ, Li L, McLeod HL, Sweeney CJ
Hoosier Oncology Group randomized phase II study of docetaxel, vinorelbine, and estramustine in combination in hormone-refractory prostate cancer with pharmacogenetic survival analysis.
Clin Cancer Res. 2006 Oct 15;12(20 Pt 1):6094-9., 2006-10-15 [PMID:17062685]

Abstract [show]
PURPOSE: To determine the safety and efficacy of two docetaxel doublets in hormone-refractory prostate cancer (HRPC) patients and to examine the prognostic role of polymorphisms in host genes important to docetaxel metabolism and transport. EXPERIMENTAL DESIGN: Sixty-four chemotherapy-naive patients with HRPC were randomized to docetaxel and vinorelbine (D, 20 mg/m2 i.v. days 1 and 8; V, 25 mg/m2 i.v. days 1 and 8) or docetaxel and estramustine phosphate (D, 60-70 mg/m2 i.v. day 1; E, 280 mg oral thrice daily days 1-5) administered q21d. Primary end point was clinically significant toxicity. A pharmacogenetic analysis of host genes was done in patients who received at least one cycle of docetaxel therapy. RESULTS: Grade 3/4 toxicity occurred in 15.6% of DV patients and in 28.6% DE patients. Neither arm exceeded the threshold of clinically significant toxicity. In the DV arm, objective response rate was 33%, prostate-specific antigen response rate was 20%, and median survival was 16.2 months. In the DE arm, objective response rate was 67%, prostate-specific antigen response rate was 43%, and median survival was 19.7 months. Pharmacogenetic analyses showed a significant association between survival beyond 15 months and the ABCG2 421 C > A (Q141K) polymorphism compared with the wild-type (C/C) genotype (66% versus 27%; P = 0.05). CONCLUSIONS: DV and DE doublets are active with a tolerable toxicity profile in patients with HRPC; however, efficacy does not seem superior to standard single-agent docetaxel. The ABCG2 421 C > A (Q141K) polymorphism may be an important predictor of response and survival in HRPC patients treated with docetaxel-based chemotherapy.

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11 Conclusions: DV and DE doublets are active with a tolerable toxicity profile in patients with HRPC; however, efficacy does not seem superior to standard single-agent docetaxel.The ABCG2 421C>A (Q141K) polymorphism may be an important predictor of response and survival in HRPC patients treated with docetaxel-based chemotherapy. Login to comment
121 In this pilot analysis, it was noted that 4 of 6 (66%) patients with the ABCG2 421 C>A (Q141K) polymorphism were alive past 15 months compared with only 12 of 44 (27%) patients with wild-type (C/C; P = 0.05). Login to comment

[hide] Involvement of breast cancer resistance protein (B... Drug Metab Dispos. 2007 Feb;35(2):209-14. Epub 2006 Nov 8. Enokizono J, Kusuhara H, Sugiyama Y
Involvement of breast cancer resistance protein (BCRP/ABCG2) in the biliary excretion and intestinal efflux of troglitazone sulfate, the major metabolite of troglitazone with a cholestatic effect.
Drug Metab Dispos. 2007 Feb;35(2):209-14. Epub 2006 Nov 8., [PMID:17093005]

Abstract [show]
Troglitazone sulfate (TGZS) is the major metabolite of troglitazone (TGZ), an antidiabetic agent, and thought to be a cause of the cholestasis induced by TGZ. The aim of the present study is to elucidate the involvement of breast cancer resistance protein (BCRP/ABCG2) in the hepatic disposition of TGZS. The basal-to-apical transport of TGZS was enhanced in organic anion transporting polypeptide 1B1-expressing Madin-Darby canine kidney II cells by infection of recombinant adenovirus harboring human BCRP and mouse Bcrp cDNA. TGZS was given to wild-type and Bcrp (-/-) mice by constant infusion. Biliary excretion is the predominant elimination pathway of TGZS in wild-type mice, and the biliary excretion clearance of TGZS with regard to the hepatic concentration was reduced to 30% of the control in Bcrp (-/-) mice. However, plasma and hepatic concentrations were unchanged, suggesting induction of compensatory mechanisms in Bcrp (-/-) mice for the elimination of TGZS. Involvement of BCRP in the intestinal efflux transport of TGZS was examined using everted sacs. The mucosal efflux clearance of TGZS showed only a slight reduction (15% reduction) in Bcrp (-/-) mice. Our results suggest that BCRP plays a major role in the biliary excretion but a minor role in the intestinal transport of TGZS.

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192 BCRP has some functional single nucleotide polymorphisms (SNP), such as C376T (Q126stop), C421A (Q141K), and G1322A (S441N). Login to comment

[hide] Pharmacogenetics of ABCG2 and adverse reactions to... J Natl Cancer Inst. 2006 Dec 6;98(23):1739-42. Cusatis G, Gregorc V, Li J, Spreafico A, Ingersoll RG, Verweij J, Ludovini V, Villa E, Hidalgo M, Sparreboom A, Baker SD
Pharmacogenetics of ABCG2 and adverse reactions to gefitinib.
J Natl Cancer Inst. 2006 Dec 6;98(23):1739-42., 2006-12-06 [PMID:17148776]

Abstract [show]
Gefitinib is an inhibitor of the epidermal growth factor receptor (EGFR) tyrosine kinase with activity in non-small-cell lung cancer. Diarrhea and skin toxicity are prominent gefitinib-related adverse events that potentially limit its use. Gefitinib is a substrate for ABCG2 (ABCP, BCRP, MXR), a polymorphic efflux transporter protein that is highly expressed in the intestines and liver. Here we investigated associations between allelic variants of EGFR, ABCG2, and the transporter protein ABCB1 with diarrhea and skin toxicity in gefitinib-treated patients. One variant, a common functional single-nucleotide polymorphism (SNP) in the ABCG2 gene, was associated with diarrhea in 124 patients treated with oral gefitinib 250 mg once daily; seven (44%) of 16 patients heterozygous for ABCG2 421C>A (Q141K) developed diarrhea, versus only 13 (12%) of 108 patients homozygous for the wild-type sequence (P = .0046). However, this SNP was not associated with skin toxicity (P = .99). The finding suggests that patients with reduced ABCG2 activity due to a common genetic variant are at increased risk for substrate drug-induced diarrhea, with implications for optimizing treatment with such agents.

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5 One variant, a common functional single-nucleotide polymorphism (SNP) in the ABCG2 gene, was associated with diarrhea in 124 patients treated with oral gefitinib 250 mg once daily; seven (44%) of 16 patients heterozygous for ABCG2 421C>A (Q141K) developed diarrhea, versus only 13 (12%) of 108 patients homozygous for the wild-type sequence (P = .0046). Login to comment
21 In particular, a functional single-nucleotide polymorphism (SNP) has been identified in exon 5 of the ABCG2 gene, in which a C → A nucleotide transition at position 421 (ABCG2 421C>A) results in a nonsynonymous variant protein in which a glutamine at position 141 is changed to lysine (Q141K) (15). Login to comment
64 The mechanism underlying the functional impact of the ABCG2 Q141K amino acid change is not entirely known, but it is most likely associated with reduced protein levels and altered ATPase activity and not with a change in localization of the protein (14). Login to comment

[hide] Toward individualized treatment: prediction of ant... Anticancer Drugs. 2007 Feb;18(2):111-26. Deeken JF, Figg WD, Bates SE, Sparreboom A
Toward individualized treatment: prediction of anticancer drug disposition and toxicity with pharmacogenetics.
Anticancer Drugs. 2007 Feb;18(2):111-26., [PMID:17159598]

Abstract [show]
A great deal of effort has been spent in defining the pharmacokinetics and pharmacodynamics of investigational and registered anticancer agents. Often, there is a marked variability in drug handling between individual patients, which contributes to variability in the pharmacodynamic effects of a given dose of a drug. A combination of physiological variables, genetic characteristics (pharmacogenetics) and environmental factors is known to alter the relationship between the absolute dose and the concentration-time profile in plasma. A variety of strategies are now being evaluated in patients with cancer to improve the therapeutic index of anticancer drugs by implementation of pharmacogenetic imprinting through genotyping or phenotyping individual patients. The efforts have mainly focused on variants in genes encoding the drug-metabolizing enzymes thiopurine S-methyltransferase, dihydropyrimidine dehydrogenase, members of the cytochrome P450 family, including the CYP2B, 2C, 2D and 3A subfamilies, members of the UDP glucuronosyltransferase family, as well as the ATP-binding cassette transporters ABCB1 (P-glycoprotein) and ABCG2 (breast cancer resistance protein). Several of these genotyping strategies have been shown to have substantial impact on therapeutic outcome and should eventually lead to improved anticancer chemotherapy.

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326 These SNPs occur at mRNA positions 34 (V12M; exon 2), 421 (Q141K, exon 16), 616 (I206L, exon 6) and 1768 (N590Y, exon 15). Login to comment
329 The nonsynonymous substitution C421A, in which a lysine is substituted for glutamine at codon 141 (Q141K), has been shown to have a functional significance. Login to comment
331 Table 12 Ethnic frequencies (%) of allelic variants in ABCG2 gene Allelic variant Caucasians African-Americans Asians Hispanics Africans Middle Easterns V12M 2 4 20-45 40 5 Q141K 11-14 2.3-5.0 15-35 10 1.0 13 I206L 0 0 0 10 0 N590Y 1 Sources: [150-153]. Login to comment
334 Plasma concentrations of orally administered drugs, including oral topotecan, are higher in patients with the Q141K SNP, which correlates with the protein`s role in drug excretion. Login to comment

[hide] Pharmacogenetic analysis of paclitaxel transport a... Pharmacogenomics J. 2007 Oct;7(5):362-5. Epub 2007 Jan 16. Marsh S, Somlo G, Li X, Frankel P, King CR, Shannon WD, McLeod HL, Synold TW
Pharmacogenetic analysis of paclitaxel transport and metabolism genes in breast cancer.
Pharmacogenomics J. 2007 Oct;7(5):362-5. Epub 2007 Jan 16., [PMID:17224914]

Abstract [show]
Paclitaxel is commonly used in the treatment of breast cancer. Variability in paclitaxel clearance may contribute to the unpredictability of clinical outcomes. We assessed genomic DNA from the plasma of 93 patients with high-risk primary or stage IV breast cancer, who received dose-intense paclitaxel, doxorubicin and cyclophosphamide. Eight polymorphisms in six genes associated with metabolism and transport of paclitaxel were analyzed using Pyrosequencing. We found no association between ABCB1, ABCG2, CYP1B1, CYP3A4, CYP3A5 and CYP2C8 genotypes and paclitaxel clearance. However, patients homozygous for the CYP1B1*3 allele had a significantly longer progression-free survival than patients with at least one Valine allele (P=0.037). This finding could reflect altered paclitaxel metabolism, however, the finding was independent of paclitaxel clearance. Alternatively, the role of CYP1B1 in estrogen metabolism may influence the risk of invasive or paclitaxel resistant breast cancer in patients carrying the CYP1B1*3 allele.

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16 ABCB1 appears to be the main paclitaxel transporter23,24 and variants in ABCB1 have been demonstrated to alter P-glycoprotein expression.25,26 A non-synonymous variant in ABCG2 (421C4A; Q141K) has been associated with drug resistance in vitro and in vivo,27-29 and with survival in prostate cancer patients receiving docetaxel chemotherapy (P ¼ 0.05),30 consequently, despite the lack of evidence associating taxane transport with ABCG2 the 421C4A variant was included in this study. Login to comment

[hide] Genetic polymorphisms of human ABC transporter ABC... J Exp Ther Oncol. 2006;6(1):1-11. Tamura A, Wakabayashi K, Onishi Y, Nakagawa H, Tsuji M, Matsuda Y, Ishikawa T
Genetic polymorphisms of human ABC transporter ABCG2: development of the standard method for functional validation of SNPs by using the Flp recombinase system.
J Exp Ther Oncol. 2006;6(1):1-11., [PMID:17228519]

Abstract [show]
The vector-mediated introduction of cDNA into mammalian cells by calcium phosphate co-precipitation or permeation with lipofectamine is widely used for the integration of cDNA into genomic DNA. However, integration of cDNA into the host's chromosomal DNA occurs randomly at unpredictable sites, and the number of integrated recombinant DNAs is not controllable. To investigate the effect of genetic polymorphisms of ABCG2 on the protein expression and the drug resistance profile, we developed the Flp-In method to integrate one single copy of ABCG2 variant-cDNA into FRT-tagged genomic DNA. More than 20 metaphase spreads were examined for both fluorescence in situ hybridization (FISH) mapping and multicolor-FISH analysis, and it has been revealed that ABCG2 cDNA was incorporated into the telomeric region of the short arm on one of chromosomes 12 in Flp-In-293 cells. Based on the currently available SNP data for human ABCG2, we have created a total of seven variants by site-directed mutagenesis and stably expressed them in Flp-In-293 cells. While mRNAs of those integrated ABCG2 variants and wild type were evenly expressed in Flp-In-293 cells, the protein expression levels of F208S and S441N variants were found to be markedly low. It is suggested that the protein instability due to enhanced degradation resulted in the low levels of their protein expression. Thus, the Flp recombinase system would provide a useful tool to validate the effect of nonsynonymous SNPs on the protein stability and post-translational modification of ABCG2.

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48 Standard method for functional validation of ABCG2 SNPs Journal of Experimental Therapeutics and Oncology Vol. 6 2006 3 Plasma Membrane inside outside S S S homodimer A B CH2N COOH V12M Q141K F208S S248P F431L S441N F489L R482G R482T Acquired mutation Figure 1. Login to comment
67 PCR primers and conditions for site-directed mutagenesis to create variants of ABCG2 Variant Forward/Reverse Primer sequence (5` →→ 3`) Primer length % GC Tm (ºC) (F/R) primers (bases) V12M F CGAAGTTTTTATCCCAATGTCACAAGGAAACAC 33 39 55 R GTGTTTCCTTGTGACATTGGGATAAAAACTTCG Q141K F CGGTGAGAGAAAACTTAAAGTTCTCAGCAGCTCTT 35 42 55 R AAGAGCTGCTGAGAACTTTAAGTTTTCTCTCACCG F208S F TGATCCTTCCATCTTGTCCTTGGATGAGCCTACAA 35 45 55 R TTGTAGGCTCATCCAAGGACAAGATGGAAGGATCA S248P F TTCATCAGCCTCGATATCCCATCTTCAAGTTGTTT 35 40 55 R AAACAACTTGAAGATGGGATATCGAGGCTGATGAA F431L F AGCTGGGGTTCTCCTCTTCCTGACGACC 28 60 62 R GGTCGTCAGGAAGAGGAGAACCCCAGCT S441N F AACCAGTGTTTCAGCAATGTTTCAGCCGTGGAAC 34 47 59 R GTTCCACGGCTGAAACATTGCTGAAACACTGGTT F489L F GAGGATGTTACCAAGTATTATACTTACCTGTATAGTGTACTTCATG 46 34 62 R CATGAAGTACACTATACAGGTAAGTATAATACTTGGTAACATCCTC Sites of mutagenesis are indicated by underbars. Login to comment
104 Standard method for functional validation of ABCG2 SNPs Journal of Experimental Therapeutics and Oncology Vol. 6 2006 0 1 2 RelativemRNAlevel Mock WT V12M Q141K mRNA A ABCG2 GAPDH Mock WT F208S S248P F431L S441N F489L ABCG2 GAPDH 0 1 2 RelativemRNAlevel mRNA B GAPDH ABCG2 Mock WT F208S S248P F431L S441N F489L Protein 0 1 2 Relativeproteinlevel * * * C DProtein GAPDH ABCG2 0 1 2 Relativeproteinlevel * * Mock WT V12M Q141K Figure 3. mRNA and protein expression levels of ABCG2 WT and variants expressed in Flp-In-293 cells. Login to comment
114 Characterization of V12M, Q141K, F208S, S248P, F431L, S441N, and F489L variants expressed in Flp-In-293 cells The mRNA levels of ABCG2 and GAPDH were measured by quantitative PCR, and the ratios of ABCG2 variants vs. GAPDH were plotted. Login to comment
119 Figure 3 demonstrates mRNA and protein levels of ABCG2 WT and V12M, Q141K, F208S, S248P, F431L, S441N, and F489L variants expressed in Flp-In-293 cells. Login to comment
121 The Q141K variant of ABCG2 stably expressed in Flp-In-293 cells had a lower expression level than did the wild-type ABCG2 or the V12M variant (Fig. 3C). Login to comment
124 The other variants, i.e., V12M, Q141K, S248P, F431L, and F489L, were expressed in plasma membrane as was ABCG2 WT. Login to comment
130 It is important to note that IC50 values of Q141K-expressing cells toward doxorubicin and daunorubicin were greater than that of WT-expressing cells, while Q141K-expressing cells were more sensitive to SN-38 and mitoxantron as compared with WT-expressing cells. Login to comment
132 Figure 4 summarizes the characteristics of those Tamura et al. 8 Journal of Experimental Therapeutics and Oncology Vol. 6 2006 Class Class Class Class WT V12M Q141K F431L S248P F489L F208S S441N R482G R482T Protein expression + + + + + + - - + + SN-38 resistance + + + + + / - - - - + + MX resistance + + + + / - - - - - + + Doxorubicin resistance - - - - - - - - + + Daunorubicin resistance - - - - - - - - + + Figure 4. Login to comment
138 WT, V12M, and Q141K form one group where protein expression and resistance to SN-38 and mitoxantrone are positive, but contribution to doxorubicin- and daurorubicin-resistance are negative. Login to comment
142 Finally, the acquired mutants R482G and R482T form another group, which is characteristic Standard method for functional validation of ABCG2 SNPs Journal of Experimental Therapeutics and Oncology Vol. 6 2006 9 Table 3 Remarks mRNA Protein Author Ref Host cell Vector Expression SNP expression expression Imai et al. (15) PA317 pHaL-IRES-DHFR bicistronic Stable V12M Similar to WT Similar to WT - - retrovirus vector plasmid - Q141K Similar to WT Lower than WT Mizuarai et al. (18) LLC-PK1 pcDNA3.1(+) Stable V12M Similar to WT N.D. - - - - Q141K Similar to WT N.D. Morisaki et al. (25) HEK293 pcDNA3.1 Stable V12M Vary among clones Vary among clones - - - - Q141K Vary among clones Vary among clones - - - - D620N Vary among clones Vary among clones Kondo et al. (26) LLC-PK1/ pcDNA3.1/ Stable/ V12M N.D. Similar to WT - HEK293 Adenovirus Transient Q141K N.D. 30 - 40% of WT - - - - A149P N.D. Similar to WT - - - - R163K N.D. Similar to WT - - - - Q166E N.D. Similar to WT - - - - P269S N.D. Similar to WT - - - - S441N N.D. Lower than WT Vethanayagam (27) HEK293 pcDNA3.1/myc-His(-) Stable I206L N.D. Vary among clones et al. - - - - N590Y N.D. Vary among clones - - - - D620N N.D. Vary among clones N.D.: No data Table 2. Login to comment
143 Resistance profile (IC50 ) of ABCG2 Compounds IC50 (nM) Mock WT V12M Q141K F208S S248P F431L S441N F489L SN-38 1.0 ± 0.2 49.9 ± 6.0 51.1 ± 13.8 17.7 ± 0.9 0.7 ± 0.0 3.6 ± 0.4 12.1 ± 1.5 0.8 ± 0.0 3.9 ± 0.4 (49.9)* (51.1)* (17.7)* (0.7) (3.6) (12.1)* (0.8) (3.9) Mitoxantorone 7.0 ± 1.1 108.0 ± 4.9 94.0 ± 18.6 46.7 ± 12.7 5.1 ± 1.0 13.4 ± 1.3 15.2 ± 1.4 5.7 ± 0.8 12.1 ± 6.2 (15.4)* (13.4)* (6.7)* (0.7) (1.9) (2.2)* (0.8) (1.7) Doxorubicin 38.8 ± 3.8 105.2 ± 24.9 123.6 ± 35.3 156.8 ± 27.5 19.9 ± 8.7 23.7 ± 6.7 43.5 ± 6.1 39.4 ± 4.1 47.6 ± 3.1 (2.7) (3.2) (4.0) (0.5) (0.6) (1.1) (1.0) (1.2) Daounorubicin 13.0 ± 0.6 32.3 ± 6.5 58.2 ± 5.0 57.7 ± 4.1 14.1 ± 2.3 22.1 ± 4.2 15.9 ± 1.2 13.3 ± 1.1 23.6 ± 1.6 (2.5) (4.5) (4.4) (1.1) (1.7) (1.2) (1.0) (1.8) Etoposide 117.1 ± 16.0 210.2 ± 18.4 297.3 ± 58.5 233.9 ± 54.2 122.9 ± 17.6 137.7 ± 14.8 139.1 ± 12.3 154.3 ± 8.5 186.9 ± 10.1 (1.8) (2.5) (2.0) (1.0) (1.2) (1.2) (1.3) (1.6) Vincristine 1.8 ± 0.2 4.3 ± 0.3 7.1 ± 1.4 5.6 ± 1.6 0.6 ± 0.0 4.3 ± 0.9 1.8 ± 0.3 0.9 ± 0.1 3.0 ± 0.7 (2.4) (3.0) (3.1) (0.3) (2.4) (1.0) (0.5) (1.7) The drug resistance profiles of ABCG2 WT and variants were obtained by incubating Flp-In-293/ABCG2 WT, V12M, Q141K, F208S, S248P, F431L, S441N, or F489L cells in the presence of SN-38, mitoxantrone, doxorubicin, daunorubicin, etoposide, or vincristine at different concentrations as described in Materials and Methods. Login to comment

[hide] Identification and functional assessment of BCRP p... Drug Metab Dispos. 2007 Apr;35(4):623-32. Epub 2007 Jan 19. Lee SS, Jeong HE, Yi JM, Jung HJ, Jang JE, Kim EY, Lee SJ, Shin JG
Identification and functional assessment of BCRP polymorphisms in a Korean population.
Drug Metab Dispos. 2007 Apr;35(4):623-32. Epub 2007 Jan 19., [PMID:17237154]

Abstract [show]
The breast cancer resistance protein (BCRP) is a member of the ATP-binding cassette transporters. The aim of the present study was to identify genetic variants of BCRP in Koreans and to assess the functional consequences of BCRP polymorphisms. Twenty single nucleotide polymorphisms (SNP), including four nonsynonymous SNP, were identified by DNA sequencing of the BCRP gene in 92 Korean subjects. BCRP V12M, Q141K, P269S, and Q126Stop were detected at frequencies of 23, 28, 0.2, and 1.9%, respectively. These four coding variants were also screened in Chinese and Vietnamese subjects; the allelic frequencies among the three populations were compared; and predictions were made as to the potential frequency of each variant. In vitro functional analyses of the P269S protein and the promoter SNP -19031C>T (mutated in the hypoxia-inducible factor-1alpha binding site) were performed and compared with those of the wild type. P269S exhibited a 35 to 40% decrease in vesicular uptake of [(3)H]estrone-3-sulfate and [(3)H]methotrexate compared with the wild type. The promoter SNP -19031C>T did not affect BCRP promoter activity in either the presence or absence of chemical-induced hypoxic stress. Our results suggest that the P269S variant could be a functionally altered variant. Genotyping of this variant in clinical studies is needed to address its phenotypic role. Genetic polymorphisms of BCRP were found to be very common in Koreans, as well as in other ethnic groups. Comparative analyses among three Asian populations revealed different frequencies for the four functional BCRP variants.

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3 BCRP V12M, Q141K, P269S, and Q126Stop were detected at frequencies of 23, 28, 0.2, and 1.9%, respectively. These four coding variants were also screened in Chinese and Vietnamese subjects; the allelic frequencies among the three populations were compared; and predictions were made as to the potential frequency of each variant. Login to comment
23 According to the current literature, the most frequent BCRP polymorphisms detected among different ethnic groups are 34GϾA, which codes for V12M, and 421CϾA, which codes for Q141K (Zamber et al., 2003). Login to comment
24 The BCRP Q141K SNP has been associated with decreased expression of the BCRP protein in that this genotype produces a reduced transporter activity phenotype (Imai et al., 2002; Kondo et al., 2004). Login to comment
34 The functional capability of the P269S variant to take up [3 H]estrone-3-sulfate (ES) and [3 H]MTX has been reported as being comparable with that of the wild-type protein TABLE 1 Primer sequences used for the amplification of the BCRP gene fragment and the annealing temperatures used in the PCR Name Region Primer Sequence (5Ј33Ј) Size PCR Condition base pair Tm; °C BCRP1P Promoter F: AACCCAGCTAGGTCAGACGA 557 60.0 R: TTTGAGTGGGCACAGCAC BCRP2P Promoter F: TTCCTAGGGTAGATGCAGCAG 509 60.0 R: CAGGGACAAGCCAAACACTC BCRP3P Promoter F: GTAGAGGCAGGGTTTCACCA 559 60.0 R: AAGTGATTGCGCATGTTCAG BCRP4P Promoter F: CGTGCCTGGCCTCTATGTAT 572 60.0 R: CTGACGCAGGCAGATCACT BCRP5P Promoter F: GCCACCACACCCAGTGTAAT 518 64.7 R: TGCAAAGTAAAAACAAATCAAAACC BCRP1E Exon 1 F: AGCTCGTCCCCTGGATGT 516 54.0 R: CCACCAACCTTTCCAGACAC BCRP2E Exon 2 F: CTGCTCATTGCCACACATTT 400 54.0 R: GCCAAAACCTGTGAGGTTCA BCRP3E Exon 3 F: GTCTCAAACTCCTGGCCTCA 403 54.0 R: GCGTTGCAAATGCTCAATAA BCRP4E Exon 4 F: TGGATTCAAAGTAGCCATGAGA 402 54.0 R: ATTCTCCCTGCCTTTTCACA BCRP5E Exon 5 F: GGTTCATCATTAGCTAGAACTTTACC 403 54.0 R: TGGAAAGCAACCATTTTTGA BCRP6E Exon 6 F: TCTTACAGGACTGGCACACG 426 54.0 R: CCTTCCCTACATTCTTACCTGCT BCRP7E Exon 7 F: TCAGGCTGAACTAGAGCAAACA 387 60.0 R: AGCACCAAATGGAACAAACA BCRP8E Exon 8 F: CATGGGAAGAAGAGAGAAAGAAA 412 60.0 R: CAAAAACACCAACAGCACTCA BCRP9E Exon 9 F: GGTGTTAGGGAAGCATCCAA 413 54.0 R: TGAAGCAGATGATAACAGAACCA BCRP10E Exon 10 F: GCCAAGCCATTGAGTGTTTA 386 60.0 R: TGGGCAACAGAGCATGAC BCRP11E Exon 11 F: CCACAACAATCCAAGACTGTG 423 60.0 R: GTAATCCTCCGGATCCCATC BCRP12E Exon 12 F: GGTCTAGCCCTGAGGATGTG 403 64.7 R: GAGTGCAAAATGGACAGGTG BCRP13E Exon 13 F: AGGGTGGTTGGAGAGTGGAT 412 60.0 R: AGCAGAGCCCCATTTACAGA BCRP14E Exon 14 F: TGAGTGTCTTGAGTAAGTGGAGAGA 420 54.0 R: GACTCCCCAGCCTTGTGTTA BCRP15E Exon 15 F: TCTTGATTGCCAGGGAAAAT 404 60.0 R: CGCGCACAACTCACTTTATG BCRP16E Exon 16 F: TGACGGATGCTAGGAATGAA 430 64.7 R: CCCATGGTTACTGTCTGAGGA TABLE 2 Primer sequences used for the pyrosequencing-based genotyping of functional BCRP variants SNPa Variant Primer Sequence (5Ј33Ј) Size PCR Condition base pair Tm; °C 34GϾA V12M 5Ј-Biotin-CTCTCCAGATGTCTTCCAGTAATG-3Ј 278 54.0 5Ј-GCCAAAACCTGTGAGGTTCA-3Ј For sequencing: 5Ј-AGTGTTCCTTTGTGGTTAC-3Ј 8191CϾT Q126Stop 5Ј-Biotin-ACTATCAGCCAAAGCACTTACCC-3Ј 174 54.5 5Ј-GTCTTAGCTGCAAGGAAAGATCCA-3Ј For sequencing: 5Ј-AATGTAATTCAGGTTACGTG-3Ј 8825CϾA Q141K 5Ј-Biotin-GTTGCAAGCCGAAGAGCTG-3Ј 69 54.0 5Ј-TGATGTTGTGATGGGCACTC-3Ј For sequencing: 5Ј-GACGGTGAGAGAAAACTT-3Ј 21850CϾT P269S 5Ј-Biotin-TAGCACCAAATGGAACAAACAC-3Ј 236 54.0 5Ј- TGTTTGATAGCCTCACCTTATTGG-3Ј For sequencing: 5Ј-GAAGACTTATGTTCCACG-3Ј a Position is indicated with respect to the start codon (ATG) of the BCRP gene; the A in the ATG triplet is designated as ϩ1, and the next base toward the 5Ј-end is designated as -1. Login to comment
37 Functional information on P269S is sparse, as compared with the amount of information that has been collected for other coding variants, such as V12M, Q141K, and the null allele Q126Stop. Login to comment
39 Therefore, we performed a functional study of the P269S variant among the four variants identified in the study (V12M, Q141K, P269S, and Q126Stop). Login to comment
65 All the subjects were screened for BCRP V12M, Q126Stop, Q141K, and P269S. Login to comment
105 SNP and Positionb Position Relative to Transcription Start Site Location Effect N Allelic Frequency % 1 -20296AϾG -1379 Promoter 92 13 2 -19855CϾT -938 Promoter 92 0.5 3 -19605AϾG -688 Promoter 92 0.5 4 -19031CϾT -114 Promoter 92 1.6 5 -18631CϾT ϩ286 5ЈUTR 92 2.2 6 34GϾA Exon 2 V12M 275 23 7 238AϾG Intron 2 92 25 8 7430AϾG Intron 3 92 9.8 9 8191CϾT Exon 4 Q126Stop 375 1.9 10 8825CϾA Exon 5 Q141K 275 28 11 21850CϾT Exon 7 P269S 674 0.2 12 26297GϾA Exon 9 92 1.1 13 38485AϾG Intron 11 92 24 14 40086insA Intron 12 92 0.5 15 40110GϾT Intron 12 92 22 16 42288CϾT Intron 13 92 67.4 17 42313TϾG Intron 13 92 2.2 18 44072CϾT Intron 13 92 23.4 19 44997AϾG Intron 14 92 49.5 20 45235CϾT Intron 15 92 20.1 a The reference sequence used has GenBank accession no. Login to comment
149 The four coding SNP were 34GϾA coding for V12M, 8191CϾT coding for Q126Stop, 8825CϾA coding for Q141K, and 21850CϾT coding for P269S. Login to comment
150 For more extensive evaluation of the allelic frequencies of the four BCRP variants found in the Korean population, the remaining 183 subjects were screened by pyrosequencing for the presence of V12M, Q126Stop, Q141K, and P269S. Login to comment
152 V12M and Q141K were found in 23 and 28% of Koreans, respectively. Login to comment
175 The most frequent allele was the wild type (26%), followed by the Q141K variant. Login to comment
176 The haplotype analysis suggests that none of the Q141K-containing haplotypes are linked to the V12M variant. Login to comment
177 To support this strong linkage, the V12M and Q141K variations were assigned to the same haplotype block among two discrete haplotype blocks of the BCRP gene (Fig. 1B). Login to comment
200 From the screening of four nonsynonymous variants in other ethnic groups, the allelic frequencies of V12M, Q126Stop, Q141K, and P269S were obtained (Table 4). Login to comment
201 The frequency of V12M was 10 to 13% higher in Chinese and Vietnamese subjects than in Koreans, whereas Q141K showed no significant differences among the three Asian ethnic groups. Login to comment
207 For example, the BCRP Q141K variant has been implicated in the altered pharmacokinetics of diflomotecan (Sparreboom et al., 2004). Login to comment
220 The Q141K variant was the most prevalent coding variant in Caucasians (14%), Japanese (35%), and Chinese (35%), as well as in Koreans (28%), whereas it was not detected in African-Americans or in subjects from Africa north of the Sahara (Zamber et al., 2003). Login to comment
231 In haplotypes 2, 7, and 13, Q141K (8825CϾA) variation is accompanied by 42288CϾT and 44997AϾG. Login to comment
234 Our results on BCRP haplotypes in the Korean population TABLE 4 Haplotype distribution of BCRP gene in Koreans PNS 69202- G>A 43 A>G 832 G>A 0347 G>A 5288 A>C 58483 G>A 01104 T>G 88224 T>C 27044 T>C 79944 G>A 53254 T>C ycneuqerF )%( egnahcAA K141QM21V 4.621 2 9.91 3 8.7 4 6.7 5 4.7 6 9.5 7 7.4 8 9.2 9 5.2 01 8.1 11 7.1 21 4.1 31 1.1 Haplotype 41 1.1 TABLE 5 Expected allelic frequencies of BCRP V12M, Q126Stop, Q141K, and P269S variants in different Asian populations Population No. Login to comment
235 of Subjects Allelic Frequency (95% CI) V12M Q126Stop Q141K P269S % % % % Korean 275-674a 23 (19.6-26.6) 1.9 (0.9-2.9) 28 (23.8-31.2) 0.2 (0-0.4) Chinese 191 33b (28.5-37.9) 0.5 (0-1.2) 29 (24.3-33.3) 0 (0-0.1) Vietnamese 140 36b (30.8-42.0) 0.4 (0-1.1) 31 (25.7-36.5) 0.7 (0-1.7) a The numbers of subjects genotyped for the V12M, Q126Stop, Q141K, and P269S variants were 275, 375, 275, and 674, respectively. Login to comment
238 In the case of BCRP, the 421CϾA variation (encodes Q141K) has been reported to play important roles in determining the expression level of the protein (Imai et al., 2002; Kobayashi et al., 2005), drug resistance, and ATPase activity (Mizuarai et al., 2004). Login to comment

[hide] Re-evaluation and functional classification of non... Cancer Sci. 2007 Feb;98(2):231-9. Tamura A, Wakabayashi K, Onishi Y, Takeda M, Ikegami Y, Sawada S, Tsuji M, Matsuda Y, Ishikawa T
Re-evaluation and functional classification of non-synonymous single nucleotide polymorphisms of the human ATP-binding cassette transporter ABCG2.
Cancer Sci. 2007 Feb;98(2):231-9., [PMID:17297656]

Abstract [show]
Impacts of genetic polymorphisms of the ATP-binding cassette (ABC) transporter BCRP/MXR1/ABCP (ABCG2) on drug response have been implicated; however, the hitherto reported data involve some inconsistencies. To re-evaluate the effect of single nucleotide polymorphisms (SNP) of ABCG2 in vitro, we created a total of seven variant cDNAs (V12M, Q141K, F208S, S248P, F431L, S441N and F489L) by site-directed mutagenesis and stably expressed each of them in Flp-In-293 cells using the Flp recombinase system. Multicolor fluorescence in situ hybridization mapping analysis revealed that one single copy of ABCG2 cDNA was incorporated into the telomeric region of chromosome 12p. It was proven that mRNAs of those integrated ABCG2 variants were expressed evenly in Flp-In-293 cells. However, the protein expression levels varied among those variants. In particular, expression of the F208S and S441N variants was markedly low, suggesting the instability of these variant proteins. Drug resistance profiles of Flp-In-293 cells expressing two major SNP variants (V12M and Q141K) toward the drug SN-38 demonstrated that the IC50 value (drug concentrations producing a 50% reduction of cell growth) for Q141K was approximately 50% of that for wild type. The contributions of the minor SNP variants (F208S, S248P, F431L, S441N and F489L) to drug resistance toward SN-38, mitoxantrone, doxorubicin, daunorubicin or etoposide were significantly lower than wild type. Based on our functional validation, the above-mentioned non-synonymous polymorphisms as well as acquired mutants (R482G and R482T) of ABCG2 were classified into four groups. Furthermore, new camptothecin analogs synthesized by our research group had potent effects in circumventing ABCG2-mediated drug resistance without any influence from major non-synonymous polymorphisms.

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3 To re-evaluate the effect of single nucleotide polymorphisms (SNP) of ABCG2 in vitro, we created a total of seven variant cDNAs (V12M, Q141K, F208S, S248P, F431L, S441N and F489L) by site-directed mutagenesis and stably expressed each of them in Flp-In-293 cells using the Flp recombinase system. Login to comment
7 Drug resistance profiles of Flp-In-293 cells expressing two major SNP variants (V12M and Q141K) toward the drug SN-38 demonstrated that the IC50 value (drug concentrations producing a 50% reduction of cell growth) for Q141K was approximately 50% of that for wild type. Login to comment
104 Characterization of WT ABCG2, V12M and Q141K expressed in Flp-In-293 cells. Login to comment
105 As shown in Fig. 2A, mRNA levels of WT ABCG2 as well as its major SNP variants (V12M and Q141K) were represented evenly in Flp-In cells. Login to comment
111 (16) As demonstrated in Fig. 2A, the protein expression level of the Q141K variant was approximately half that of the WT level. Login to comment
112 Figure 2B depicts the immunofluorescence images of Flp-In-293/Mock, Flp-In-293/ABCG2 (WT), Flp-In-293/ABCG2 (V12M) and Flp-In-293/ABCG2 (Q141K) cells. Login to comment
116 Drug resistance profiles of Flp-In-293 cells expressing WT ABCG2, V12M and Q141K variants toward camptothecin analogs. Login to comment
117 To examine the drug resistance profiles, we incubated Flp-In-293/ABCG2 (WT), Flp-In-293/ABCG2 (V12M) and Flp-In-293/ABCG2 (Q141K) cells with SN-38 and the new camptothecin analogs at different concentrations for 96 h, after which we observed the cell survival rates. Login to comment
118 Flp-In-293/ABCG2 (WT), Flp-In-293/ABCG2 (V12M) and Flp-In-293/ABCG2 (Q141K) cells exhibited strong resistance to SN-38, SN-355 and SN-398, as represented by large IC50 values (Fig. 3). Login to comment
120 In contrast, the Q141K variant appeared to render drug resistance at a level only half that of WT. Login to comment
135 (V12M) or Flp-In-293/ABCG2 (Q141K) cells exhibited resistance toward SN-22, SN-343, SN-348, SN-349, SN-351, SN-352, SN-353, SN-364, SN-397, SN-443 or SN-444. Login to comment
136 These results suggest that camptothecin analogs lacking the hydroxyl or amino group at position 10 or 11 are not substrates for WT ABCG2 or the V12M and Q141K variants. Login to comment
150 It is important to note that the IC50 values of Q141K-expressing cells toward doxorubicin and daunorubicin were greater than that of WT-expressing cells, whereas Q141K-expressing cells were more sensitive to SN-38 and mitoxantron compared with WT-expressing cells. Login to comment
151 These results suggest that the substrate specificity of the Q141K variant differs delicately from that of WT ABCG2. Login to comment
152 Considered overall, the minor SNP variants were less effective in drug resistance than the V12M and Q141K variants. Login to comment
155 For this purpose, we expressed WT ABCG2, V12M, Q141K, S248P, F431L, F489L, R482G and R482T in Sf9 insect cells and prepared plasma membranes as described previously,(16,35) as the plasma membrane of Sf9 cells has lower endogenous background ATPase activity than Flp-In-293 cells. Login to comment
159 Honjo et al. first identified non-synonymous SNP of V12M and Q141K. Login to comment
166 The Q141K polymorphism located in exon 5 (421C > A) leads to the replacement of the negatively charged glutamic acid residue with a positively charged lysine residue. Login to comment
168 The Q141K variant was also detected in all ethnic groups tested, with the allele frequency ranging from 0 to 35% (Africans North Fig. 3. Login to comment
169 The effect of camptothecin analogs on the growth of Flp-In-293/ABCG2 (wild type), Flp-In-293/ABCG2 (V12M) or Flp-In-293/ABCG2 (Q141K) cells. Login to comment
176 Resistance profile (IC50) of ABCG2 Compound IC50 (nM) Mock Wild type V12M Q141K F208S S248P F431L S441N F489L SN-38 0.9 40.0 (44.4) 40.0 (44.4) 17.0 (18.9) 0.6 (0.7) 3.0 (3.3) 10.0 (11.1) 0.7 (0.8) 3.1 (3.4) Mitoxantorone 5.2 >100 (>19) 92.0 (17.7) 45.0 (8.7) 4.5 (0.9) 11.0 (2.1) 21.0 (4.0) 4.6 (0.9) 11.0 (2.1) Doxorubicin 32.0 78.0 (2.4) 100.0 (3.1) 110.0 (3.4) 20.0 (0.6) 20.0 (0.6) 40.0 (1.3) 21.0 (0.7) 45.0 (1.4) Daunorubicin 12.0 30.0 (2.5) 50.0 (4.2) 50.0 (4.2) 12.0 (1.0) 21.0 (1.8) 14.0 (1.2) 12.0 (1.0) 19.0 (1.6) Etoposide 110.0 200.0 (1.8) 220.0 (2.0) 200.0 (1.8) 110.0 (1.0) 120.0 (1.1) 120.0 (1.1) 130.0 (1.2) 170.0 (1.5) Vincristine 1.4 4.0 (2.9) 5.0 (3.6) 4.5 (3.2) 0.6 (0.4) 4.0 (2.9) 1.4 (1.0) 0.8 (0.6) 2.8 (2.0) Relative resistances to mock cells are described in parentheses. Login to comment
179 (22,25,27,29) An investigation of the expression level of ABCG2 in 99 Japanese placenta samples revealed that individuals homozygous for the Q141K variant showed significantly lower expression levels of this transporter protein, whereas the heterozygous samples displayed an intermediate expression level. Login to comment
180 (27) It has been reported that the SNP Q141K may cause increased plasma concentrations of orally administered diflomotecan in cancer patients who express low levels of ABCG2 (Q141K) protein in the small intestine. Login to comment
182 By using an in-vitro system, Imai et al. showed that ABCG2 Q141K variant-transfected PA317 cells had a low-level of drug resistance associated with decreased protein expression. Login to comment
183 (25) This finding is apparently in accordance with a report by Kondo et al.(31) However, Morisaki et al.(30) reported that the protein expression level of ABCG2 Q141K varied among clones of HEK293 cells where ABCG2 Q141K cDNA was expressed stably with the conventional pcDNA3.1(+) vector. Login to comment
184 In contrast, Mizuarai et al.(28) described that approximately equal levels of ABCG2 were expressed in Q141K clones compared to the expression in WT, although they did not present any data in this respect. Login to comment
191 Based on the new method, our present study (Fig. 2A) supports the original report of Imai et al.,(25) suggesting that the protein stability of the Q141K variant is reduced without any detectable changes in its mRNA levels. Login to comment
202 As one of the specific aims of the present study, we functionally classified the non-synonymous polymorphisms (V12M, Q141K, F208S, S248P, F431L, S441N and F489L) in terms of their protein expression level, drug resistance profile and prazosin-stimulated ATPase activity. Login to comment
206 As shown in Fig. 5B, it is obvious that WT, V12M and Q141K form one group where protein expression, methotrexate and porphyrin transport, and resistance to SN-38 and mitoxantrone are positive, but contribution to doxorubicin- and daurorubicin- Fig. 4. Login to comment
221 (32,33) As demonstrated in Fig. 3, the new camptothecin analogs that were non-substrates for ABCG2 circumvented ABCG2-mediated drug resistance without any influence from major SNP (i.e. V12M and Q141K). Login to comment

[hide] Association of variant ABCG2 and the pharmacokinet... Cancer Biol Ther. 2007 Mar;6(3):432-8. Epub 2007 Mar 29. Li J, Cusatis G, Brahmer J, Sparreboom A, Robey RW, Bates SE, Hidalgo M, Baker SD
Association of variant ABCG2 and the pharmacokinetics of epidermal growth factor receptor tyrosine kinase inhibitors in cancer patients.
Cancer Biol Ther. 2007 Mar;6(3):432-8. Epub 2007 Mar 29., [PMID:17312388]

Abstract [show]
The purpose of the study was to determine if the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs), gefitinib and erlotinib, are substrates for the efflux transporter ABCG2, and to investigate the relevance of the ABCG2 421C>A (Q141K) polymorphism to the pharmacokinetics of gefitinib. Gefitinib and erlotinib transport in vitro was studied using HEK293 cells transfected with wild-type ABCG2 or a Q141K clone. Gefitinib pharmacokinetics was determined in 27 cancer patients. was. ABCG2 421C>A and ABCB1 3435C>T genotypes were determined using direct sequencing. Cells expressing wild-type ABCG2 exhibited lower intracellular accumulation of gefitinib and erlotinib at concentrations of 0.1 and 1 microM, and higher efflux at 1 microM than cells lacking ABCG2 (p < 0.05); no significant difference in cellular efflux and accumulation was observed in the variant cell line at lower concentrations nor in the three cell lines at 10 microM. In the presence of the ABCG2 inhibitor fumitremorgin C, cellular accumulation of gefitinib and erlotinib 1 microM was increased in wild-type (p < 0.05), but not in variant or null cells. Gefitinib accumulation during 28 days of treatment (C(ss,min)/C(1,min)) was higher in patients heterozygous at the ABCG2 421C>A locus than those with a wild-type genotype (median, 5.07 vs. 3.60, p = 0.004). No significant associations were observed between the ABCB1 3435C>T genotype and gefitinib pharmacokinetics. In conclusion, gefitinib and erlotinib are ABCG2 substrates, while they inhibit ABCG2 at higher concentrations. A functional variant of ABCG2 is associated with greater gefitinib accumulation at steady-state and may be relevant to toxicity and antitumor activity of EGFR TKIs.

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1 Jude Children's Research Hospital; 332 North Lauderdale Street, DTRC Room D1034, Mail Stop 314; Memphis, Tennessee USA 38105; Tel.: 901.495.3089; Fax: 901.495.3125; Email: sharyn.baker@stjude.org Original manuscript submitted: 12/21/06 Manuscript accepted: 12/29/06 Previously published online as a Cancer Biology & Therapy E-publication: http://www.landesbioscience.com/journals/cc/abstract.php?id=3763 Key words gefitinib, erlotinib, EGFR tyrosine kinase inhibitor, ABCG2, polymorphism, pharmacokinetics Abstract The purpose of the study was to determine if the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs), gefitinib and erlotinib, are substrates for the efflux transporter ABCG2, and to investigate the relevance of the ABCG2 421C>A (Q141K) polymorphism to the pharmacokinetics of gefitinib. Login to comment
2 Gefitinib and erlotinib transport in vitro was studied using HEK293 cells transfected with ��wild‑type ABCG2 or a Q141K clone. Login to comment
12 In particular, a functional single nucleotide polymorphism has been recently identified in exon 5 of the ABCG2 gene, in which a C>A nucleotide transition at position 421 (ABCG2 421C>A) results in a non-synonymous variant protein with a glutamine to lysine amino acid substitution in codon 141 (Q141K).19 The ABCG2 421C>A variant has been associated with low ABCG2 expression levels and altered substrate specificity,19 and has been found to alter the pharmacokinetics of diflomotecan and topotecan.13,24 Recently developed tyrosine kinase inhibitors, including imatinib and gefitinib, have been shown to inhibit ABCG2,25‑29 and imatinib has been shown to be a substrate for ABCG2.5 The aim of this study was to determine if ��ABCG2 transports the epidermal growth factor receptor tyrosine kinase inhibitors gefitinib and erlotinib, ��by examining cellular accumulation and efflux in human embryonic kidney cells (HEK293) transfected with empty vector, wild‑type ABCG2 and a 421C>A variant clone. Login to comment
71 Comparative intracellular accumulation of 3H-gefitinib (A) and erlotinib (B) in human embryonic kidney cells (HEK293) transfected with a pcDNA3 vector (pcDNA3), wild-type (R-5), and Q141K variant (Q141-K5) of ABCG2. Login to comment
74 *, ANOVA, significantly different from the control (pcDNA3), P<0.05; **, ANOVA, significantly different from the control (pcDNA3) and Q141 variant (Q141-K5), P<0.05 Table 1 Kinetic parameters a for cellular efflux of 3H‑gefitinib by �� HEK293 transfected with pcDNA3 vector (pcDNA), Q141K variant (Q141‑K5), and wild‑type (R‑5) of ABCG2, ��at the drug concentrations of 1 and 10 mM Gefitinib 1 mM Gefitinib 10 mM Emax (%) ET50 (h) Emax (%) ET50 (h) pcDNA 19.5 ± 4.19 1.40 ± 0.69 22.7 ± 10.6 1.86 ± 1.57 Q141‑K5 22.1 ± 2.23* 0.74 ± 0.09* 22.7 ± 2.30 1.00 ± 0.26 R‑5 27.3 ± 1.26*† 0.67 ± 0.09* 25.2 ± 4.04 1.13 ± 0.39 aIndividual cellular efflux curves were fitted with Michaelis‑Menten equation, where Emax is the maximum efflux fraction (%initial radioactivity), and ET50 is the time to achieve the half‑Emax. Login to comment
98 Relative intracellular accumulation of 3H-gefitinib (A) and erlotinib (B) in human embryonic kidney cells (HEK293) transfected with a pcDNA3 vector (pcDNA3), wild-type (R-5), and Q141K variant (Q141-K5) of ABCG2, in the absence and presence of 1 µM or 5 µM of the ABCG2 specific inhibi� tor fumitremorgin (FTC). Login to comment
101 *, ANOVA, significantly different from the control (in the absence of FTC), P<0.05 Table 2 Genotype and allele frequencies for the studied variant genes Genotype Frequenciesa Allele Frequenciesb Variantc Effectd Activitye Wt Het Var p q ABCB1 3435C > T I1145I (exon 26) Decreased 7 (26%) 12 (44%) 8 (30%) 0.48 0.52 ABCG2 421C > A Q141K (exon 5) Decreased 20 (74%) 7 (26%) 0 (0) 0.87 0.13 aNumber represents number of patients with percentage in parenthesis. Login to comment
110 Cellular efflux of 3H-gefitinib (A and B) and erlotinib (C and D) by human embryonic kidney cells (HEK293) transfected with a pcDNA3 vector (pcDNA3), wild-type (R-5), and Q141K variant (Q141-K5) of ABCG2, after cells were pre-incubated with 1 µM (A and C), and 10 µM (A and D) of the drug (i.e., gefitinib or erlotinib) for 1 h. Login to comment

[hide] Pharmacogenetics/genomics of membrane transporters... Cancer Metastasis Rev. 2007 Mar;26(1):183-201. Huang Y
Pharmacogenetics/genomics of membrane transporters in cancer chemotherapy.
Cancer Metastasis Rev. 2007 Mar;26(1):183-201., [PMID:17323126]

Abstract [show]
Inter-individual variability in drug response and the emergence of adverse drug reactions are main causes of treatment failure in cancer therapy. Recently, membrane transporters have been recognized as an important determinant of drug disposition, thereby affecting chemosensitivity and -resistance. Genetic factors contribute to inter-individual variability in drug transport and targeting. Therefore, pharmacogenetic studies of membrane transporters can lead to new approaches for optimizing cancer therapy. This review discusses genetic variations in efflux transporters of the ATP-binding cassette (ABC) family such as ABCB1 (MDR1, P-glycoprotein), ABCC1 (MRP1), ABCC2 (MRP2) and ABCG2 (BCRP), and uptake transporters of the solute carrier (SLC) family such as SLC19A1 (RFC1) and SLCO1B1 (SLC21A6), and their relevance to cancer chemotherapy. Furthermore, a pharmacogenomic approach is outlined, which using correlations between the growth inhibitory potency of anticancer drugs and transporter gene expression in multiple human cancer cell lines, has shown promise for determining the relevant transporters for any given drugs and predicting anticancer drug response.

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124 Moreover, Letourneau et al. examined 10 non-synonymous ABCC1 SNPs to determine Table 2 Summary of genetic variants in ABC transporters ABCB1, ABCC1, ABCC2 and ABCG2 involved in cancer chemotherapy Variants (location, effect) Phenotype Drug Sample Reference ABCB1 +103T>C (5'flanking, non-coding) Increased transcription Doxorubicin vincristine osteosarcoma Stein et al., 1994 [19] +8T>C (5'flanking, non-coding) Unknown Leukemia Rund et al., 1999 [21] 1236C>T (exon12, synonymous) Higher expression AML blasts Illmer et al., 2002 [47] Lower clearance Irinotecan Cancer patients Sai et al., 2003 [44] Higher exposure Irinotecan, SN-38 Cancer patients Mathijssen et al., 2003 [45] 2677G>T/A (exon21, A893S/T) Lower expression placenta Tanabe et al., 2001 [42] Lower expression placenta Hitzl et al., 2004 [37] Higher expression AML blasts Illmer et al., 2002 [47] Allele specific expression Cell lines, lymphoma Mickley et al., 1998 [22] Lower clearance Irinotecan Cancer patients Sai et al., 2003 [44] Survival leukemia Illmer et al., 2002 [47] Survival leukemia van den Heuvel-Eibrink et al., 2001 [48] Worse survival AML blasts Kim et al., 2006 [10] Higher efficacy Paclitaxel Ovarian cancer Green et al., 2006 [50] 2995G>A (exon24, A999T) None Cell lines, lymphoma Mickley et al., 1998 [22] 3435C>T (exon26, synonymous) Lower expression Duodenal protein Hoffmeyer et al., 2000 [26] Lower expression placenta Hitzl et al., 2004 [37] Higher expression Intestine mRNA Nakamura et al., 2002 [32] Higher expression AML blasts Illmer et al., 2002 [47] Lower clearance Irinotecan Cancer patients Sai et al., 2003 [44] Lower efflux Digoxin CD56+ NK cells Hitzl et al., 2001 [27] Higher plasma level Digoxin Healthy volunteers Hoffmeyer et al., 2000 [26] Higher AUC Cyclosporin transplant patients Bonhomme-Faivre et al., 2004 [36] Lower CNS relapse Cancer patients Stanulla et al., 2005 [46] Better survival leukemia Illmer et al., 2002 [47] Higher efficacy Breast cancer Kafka et al., 2003 [49] Higher activity, worse survival AML Kim et al., 2006 [10] Better survival Platinums Esophageal cancer Wu et al., 2006 [43] No difference Docetaxel patients Puisset et al., 2004 [41] No difference Irinotecan Cancer patients Mathijssen et al., 2004 [39] No difference Vincristine patients Plasschaert et al., 2004 [40] No difference colon Taniguchi et al., 2003 [24] ABCC1 -260G>C (5'flanking, non-coding) Higher activity Transfected cell line Wang et al., 2005 [62] Table 2 (Continued) Variants (location, effect) Phenotype Drug Sample Reference 128G>C (exon2, C43S) Reduced resistance Vincristine, arsenite Transfected cell line Leslie et al., 2003 [60] 1299G>T (exon10, R433S) Reduced transport of LTC4, increased resistance to doxorubicin Leukotriene C4, doxorubicin Transfected cell line Conrad et al., 2002 [59] 2012G>T (exon16, G671V) No change in activityLeukotriene C4 Transfected cell line Conrad et al., 2001 [58] Heart toxicity Doxorubicin nLon-Hodgkin lymphoma Wojnowski et al., 2005 [63] 2965G>A (exon22, A989T) Reduced transport Estradiol 17β-glucuronide Transfected cell line Letourneau et al., 2005 [61] ABCC2 1271A>G (exon10, R421G) Reduced drug elimination, increased nephrotoxicity Methotrexate One lymphoma patient Hulot et al., 2005 [79] 3972C>T (exon28, nonsynonymous) Reduced drug clearance Irinotecan Cancer patients Innocenti et al., 2004 [80] ABCG2 376C>T (exon4, Q126stop) Reduced transport Porphyrin Trensfected cell Tamura et al., 2006 [104] 421C>A (exon5, Q141K) Lower expression Transfected cell lines Imai et al., 2002 [94] Lower expression Transfected cell lines Kondo et al., 2004 [95] Lower expression Placenta Kobayashi et al., 2005 [98] Reduced ATPase activity Trensfected cell lines Mizuarai et al., 2004 [97] Higher plasma levels Diflomotecan patients Sparreboom et al., 2004 [100] Increased bioavailability Topotecan patients Sparreboom et al., 2005 [101] Increased bioavailability 9-Aminocamptothecin patients Zamboni et al., 2006 [81] Increased drug accumulation Imatinib Transfected cell lines Gardner et al., 2006 [96] Increased drug accumulation Topotecan Trensfected cell lines Imai et al., 2002 [94] No difference Imatinib patients Gardner et al., 2006 [96] No difference intestine Zamber et al., 2003 [99] No difference MTX Trensfected cell lines Kondo et al., 2004 [95] the effects on expression and function of this transporter in transfected HEK293T cells [61]. Login to comment
172 In vitro transfection experiments showed that 421C>A (substituting Lys for Gln-141, Q141K) resulted in decreased ABCG2 protein expression and a reduced level of drug resistance when compared with the wild type ABCG2 [94, 95]. Login to comment
175 ATPase activity of the Q141K variant was reduced ~1.3-fold compared to the activity of the wild type ABCG2 [97]. Login to comment

[hide] ABCG2: determining its relevance in clinical drug ... Cancer Metastasis Rev. 2007 Mar;26(1):39-57. Robey RW, Polgar O, Deeken J, To KW, Bates SE
ABCG2: determining its relevance in clinical drug resistance.
Cancer Metastasis Rev. 2007 Mar;26(1):39-57., [PMID:17323127]

Abstract [show]
Multidrug resistance is a major obstacle to successful cancer treatment. One mechanism by which cells can become resistant to chemotherapy is the expression of ABC transporters that use the energy of ATP hydrolysis to transport a wide variety of substrates across the cell membrane. There are three human ABC transporters primarily associated with the multidrug resistance phenomenon, namely Pgp, MRP1, and ABCG2. All three have broad and, to a certain extent, overlapping substrate specificities, transporting the major drugs currently used in cancer chemotherapy. ABCG2 is the most recently described of the three major multidrug-resistance pumps, and its substrates include mitoxantrone, topotecan, irinotecan, flavopiridol, and methotrexate. Despite several studies reporting ABCG2 expression in normal and malignant tissues, no trials have thus far addressed the role of ABCG2 in clinical drug resistance. This gives us an opportunity to critically review the disappointing results of past clinical trials targeting Pgp and to propose strategies for ABCG2. We need to know in which tumor types ABCG2 contributes to the resistance phenotype. We also need to develop standardized assays to detect ABCG2 expression in vivo and to carefully select the chemotherapeutic agents and clinical trial designs. This review focuses on our current knowledge about normal tissue distribution, tumor expression profiles, and substrates and inhibitors of ABCG2, together with lessons learned from clinical trials with Pgp inhibitors. Implications of SNPs in the ABCG2 gene affecting the pharmacokinetics of substrate drugs, including many non-chemotherapy agents and ABCG2 expression in the SP population of stem cells are also discussed.

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138 Expression of the Q141K SNP in cell lines has been shown to lead to significantly lower IC50 values for ABCG2 substrates, including mitoxantrone, irinotecan, and SN-38 [119, 122]. Login to comment
139 The Q141K SNP has been shown to influence the pharmacokinetics of orally administered drugs, including topotecan [125], diflomotecan [126] and 9-aminocamptothecin [127]. Login to comment
140 As noted earlier, the higher plasma drug levels due to the Q141K SNP may result in exquisite sensitivity to certain orally administered chemotherapy drugs. Login to comment

[hide] The identification of two germ-line mutations in t... Pharm Res. 2007 Jun;24(6):1108-17. Epub 2007 Mar 21. Yoshioka S, Katayama K, Okawa C, Takahashi S, Tsukahara S, Mitsuhashi J, Sugimoto Y
The identification of two germ-line mutations in the human breast cancer resistance protein gene that result in the expression of a low/non-functional protein.
Pharm Res. 2007 Jun;24(6):1108-17. Epub 2007 Mar 21., [PMID:17373578]

Abstract [show]
PURPOSE: We examined the effects of the nine nonsynonymous germ-line mutations/SNPs in the breast cancer resistance protein (BCRP/ABCG2) gene on the expression and function of the protein. MATERIALS AND METHODS: We generated cDNAs for each of these mutants (G151T, C458T, C496G, A616C, T623C, T742C, T1291C, A1768T, and G1858A BCRP) and compared the effects of their exogenous expression in PA317 cells with a wild-type control. RESULTS: PA/F208S cells (T623C BCRP-transfectants) expressed marginal levels of a BCRP protein species (65kDa), which is slightly smaller than wild-type (70kDa), but this mutant did not appear on the cell surface or confer drug resistance. PA/F431L cells (T1291C BCRP-transfectants) were found to express both 70 kDa and 65 kDa BCRP protein products. In addition, although PA/F431L cells expressed 70 kDa BCRP at comparable levels to PA/WT cells, they showed only marginal resistance to SN-38. PA/T153M cells (C458T BCRP-transfectants) and PA/D620N cells (G1858A BCRP-transfectants) expressed lower amounts of BCRP and showed lower levels of resistance to SN-38 compared with PA/WT cells. CONCLUSIONS: We have shown that T623C BCRP encodes a non-functional BCRP and that T1291C BCRP encodes a low-functional BCRP. Hence, these mutations may affect the pharmacokinetics of BCRP substrates in patients harboring these alleles.

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21 In our previous study, we identified three nonsynonymous SNPs within the BCRP gene, G34A substituting Met for Val-12 (V12M), C376T substituting a stop codon for Gln-126 (Q126Stop), and C421A substituting Lys for Gln141 (Q141K). Login to comment
42 The cells were selected with 120 ng/mL of methotrexate, and the resulting mixed populations of resistant cells were designated as PA/WT, PA/V12M, PA/ G51C, PA/Q141K, PA/T153M, PA/I206L, PA/F208S, PA/ S248P, PA/F431L, PA/N590Y and PA/D620N, respectively. The PA/F208S clones and PA/F431L clones were obtained by limiting dilution. Login to comment
43 Cell Growth Inhibition Assay Anticancer agent resistance levels in both the parental PA317 cells and in the various BCRP transfectants were Table I. Frequencies of Germ-line Mutations/SNPs Within The BCRP Gene Variation Frequency (%) Number Population Reference Nucleotide Amino acid G34A V12M 19 29 Japanese 17 G151T G51C 0.1a 350 Japanese C376T Q126Stop 1.2 124 Japanese 17 C421A Q141K 26.6 124 Japanese 17 C458T T153M 3.3 30 Cell line 32 C496G Q166E 0.3a 200 Japanese A616C I206L 20 10 Hispanic 33 T623C F208S 0.3a 200 Japanese T742C S248P 0.5a 200 Japanese T1291C F431L 0.6b 260 Japanese 34 A1768T N590Y 1.1 88 Caucasians 33 G1858A D620N 1.1 90 unknown 35 a Determined in this study. Login to comment
45 V12M Q141K D620N N590Y F431L S248P F208S I206L T153M G51C Q166E OUT MEMBRANE IN Fig. 1. Login to comment
75 SN-38 Resistance Levels of PA317 Transfectantsa Cell type IC50 (nmol/L) Degree of resistance PA317 11 T 0.2 1 PA/WT 550 T 16 50 PA/V12M 490 T 13 45 PA/Q141K 110 T 5.9 10 PA/T153M 260 T 15 24 PA/Q166E 680 T 40 62 PA/F208S 10 T 0.7 1 PA/F431L 34 T 0.9 3 PA/D620N 190 T 5.7 17 a Cells were cultured for 5 days with various concentrations of SN-38. Login to comment
85 Similar to previous findings (14), PA/V12M cells were observed to express similar amounts of BCRP compared with PA/WT cells, whereas PA/Q141K cells expressed significantly lower amounts of BCRP than PA/WT (Fig. 2a). Login to comment
88 PA/T153M and PA/D620N transfectants expressed lower amounts of BCRP than PA/WT cells, but these levels were higher than those in the PA/Q141K cells (Fig. 2a). Login to comment
94 PA/Q141K, PA/T153M and PA/D620N cells expressed lower amounts of BCRP on their cell surfaces than PA/WT cells (Fig. 2d). Login to comment
102 PA/Q141K, PA/ T153M, and PA/D620N cells showed 10Y24-fold higher resistance levels to SN-38 compared with the parental cells (Table II). Login to comment
130 The resulting mixed populations of cells were designated a PA/WT, PA/V12M, PA/G51C, PA/Q141K, PA/ T153M, PA/I206L, PA/F208S, PA/S248P, PA/F431L, PA/ N590Y and PA/D620N. Login to comment
168 Although PA/F431L cells express higher quantities of 70-kDa BCRP compared with PA/Q141K, PA/T153M, and PA/D620N cells (Fig. 2a) these cells in fact show a lower resistance to SN-38 than these other three transfectants (Table II). Login to comment
172 These results were similar to the data obtained for PA/Q141K cells and based upon these data, we hypothesize that the decrease in the resistance levels to SN-38 may not be due to functional alterations but to decreased protein expression. Login to comment

[hide] The emerging pharmacotherapeutic significance of t... Br J Pharmacol. 2007 May;151(2):163-74. Epub 2007 Mar 20. Hardwick LJ, Velamakanni S, van Veen HW
The emerging pharmacotherapeutic significance of the breast cancer resistance protein (ABCG2).
Br J Pharmacol. 2007 May;151(2):163-74. Epub 2007 Mar 20., [PMID:17375082]

Abstract [show]
The breast cancer resistance protein (also termed ABCG2) is an ATP-binding cassette transporter, which mediates the extrusion of toxic compounds from the cell, and which was originally identified in relation to the development of multidrug resistance of cancer cells. ABCG2 interacts with a range of substrates including clinical drugs but also substances such as sterols, porphyrins and a variety of dietary compounds. Physiological functions of ABCG2 at both cellular and systemic levels are reviewed. For example, ABCG2 expression in erythrocytes may function in porphyrin homeostasis. In addition, ABCG2 expression at apical membranes of cells such as hepatocytes, enterocytes, endothelial and syncytiotrophoblast cells may correlate to protective barrier or secretory functions against environmental or clinically administered substances. ABCG2 also appears influential in the inter-patient variation and generally poor oral bioavailability of certain chemotherapeutic drugs such as topotecan. As this often precludes an oral drug administration strategy, genotypic and environmental factors altering ABCG2 expression and activity are considered. Finally, clinical modulation of ABCG2 activity is discussed. Some of the more recent strategies include co-administered modulating agents, hammerhead ribozymes or antisense oligonucleotides, and with specificity in cell targeting, these may be used to reduce drug resistance and increase drug bioavailability to improve the profile of chemotherapeutic efficacy versus toxicity. While many such strategies remain in relative infancy at present, increased knowledge of modulators of ABCG2 could hold the key to novel approaches in medical treatment.

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145 Imai et al. (2002a) sequenced cDNA from 11 human tumours and identified SNPs G34A (V12M) and C421A (Q141K), a splice variant 944-949 deletion (A315- T316), and later, an additional C376T (G126Stop) polymorphism. Login to comment

[hide] Cloning, mapping and association studies of the ov... Anim Genet. 2007 Apr;38(2):126-31. Duncan EJ, Dodds KG, Henry HM, Thompson MP, Phua SH
Cloning, mapping and association studies of the ovine ABCG2 gene with facial eczema disease in sheep.
Anim Genet. 2007 Apr;38(2):126-31., [PMID:17403009]

Abstract [show]
Facial eczema (FE) is a hepatogenous mycotoxicosis in sheep caused by the fungal toxin sporidesmin. Resistance to FE is a multigenic trait. To identify QTL associated with this trait, a scan of ovine chromosomes was implemented. In addition, ABCG2 was investigated as a possible positional candidate gene because of its sequence homology to the yeast PDR5 protein and its functional role as a xenobiotic transporter. The sequence of ovine ABCG2 cDNA was obtained from liver mRNA by RT-PCR and 5' and 3' RACE. The predicted protein sequence shares >80% identity with other mammalian ABCG2 proteins. SNPs were identified within exon 6, exon 9 and intron 4. The intron 4 SNP was used to map ABCG2 to ovine chromosome 6 (OAR6), about 2 cM distal to microsatellite marker OarAE101. Interestingly, this chromosomal region contains weak evidence for a FE QTL detected in a previous genome-scan experiment. To further investigate the association of ABCG2 with FE, allele frequencies for the three SNPs plus three neighbouring microsatellite markers were tested for differences in sheep selected for and against FE. Significant differences were detected in the allele frequencies of the intronic SNP marker among the resistant, susceptible and control lines. No difference in the levels of ABCG2 expression between the resistant and susceptible animals was detected by Northern hybridisation of liver RNA samples. However, significantly higher expression was observed in sporidesmin-dosed sheep compared with naive animals. Our inference is that the ABCG2 gene may play a minor role in FE sensitivity in sheep, at least within these selection lines.

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95 The Q141K mutation affects the transport efficiency of the protein (Mizuarai et al. 2004), whereas the S441N mutation is known to alter the localisation of the mature protein from the cell membrane to an intracellular location. Login to comment

[hide] SLCO1B1 (OATP1B1, an uptake transporter) and ABCG2... Clin Pharmacol Ther. 2007 Nov;82(5):541-7. Epub 2007 Apr 25. Ieiri I, Suwannakul S, Maeda K, Uchimaru H, Hashimoto K, Kimura M, Fujino H, Hirano M, Kusuhara H, Irie S, Higuchi S, Sugiyama Y
SLCO1B1 (OATP1B1, an uptake transporter) and ABCG2 (BCRP, an efflux transporter) variant alleles and pharmacokinetics of pitavastatin in healthy volunteers.
Clin Pharmacol Ther. 2007 Nov;82(5):541-7. Epub 2007 Apr 25., [PMID:17460607]

Abstract [show]
To investigate the contribution of genetic polymorphisms of SLCO1B1 and ABCG2 to the pharmacokinetics of a dual substrate, pitavastatin, 2 mg of pitavastatin was administered to 38 healthy volunteers and pharmacokinetic parameters were compared among the following groups: 421C/C(*)1b/(*)1b (group 1), 421C/C(*)1b/(*)15 (group 2), 421C/C(*)15/(*)15 and 421C/A(*)15/(*)15 (group 3), 421C/A(*)1b/(*)1b (group 4), 421A/A(*)1b/(*)1b (group 5), and 421C/A(*)1b/(*)15 (group 6). In SLCO1B1, pitavastatin area under plasma concentration-time curve from 0 to 24 h (AUC(0-24)) for groups 1, 2, and 3 was 81.1+/-18.1, 144+/-32, and 250+/-57 ng h/ml, respectively, with significant differences among all three groups. In contrast to SLCO1B1, AUC(0-24) in groups 1, 4, and 5 was 81.1+/-18.1, 96.7+/-35.4, and 78.2+/-8.2 ng h/ml, respectively. Although the SLCO1B1 polymorphism was found to have a significant effect on the pharmacokinetics of pitavastatin, a nonsynonymous ABCG2 variant, 421C>A, did not appear to be associated with the altered pharmacokinetics of pitavastatin.

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230 Imai, Y. et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] C421 allele-specific ABCG2 gene amplification conf... Biochem Pharmacol. 2007 Jun 30;74(1):41-53. Epub 2007 Apr 1. Bram EE, Ifergan I, Grimberg M, Lemke K, Skladanowski A, Assaraf YG
C421 allele-specific ABCG2 gene amplification confers resistance to the antitumor triazoloacridone C-1305 in human lung cancer cells.
Biochem Pharmacol. 2007 Jun 30;74(1):41-53. Epub 2007 Apr 1., 2007-06-30 [PMID:17481587]

Abstract [show]
The A421 ABCG2 genotype is a frequent polymorphism encoding the K141 transporter, which is associated with a significant decrease in transporter expression and function when compared to the wild type (wt) C421 allele encoding the Q141 ABCG2. Here we show that during the acquisition of resistance to the novel triazoloacridone antitumor agent C-1305 in lung cancer cells harboring a heterozygous C421A genotype, a marked C421 allele-specific ABCG2 gene amplification occurred. This monoallelic C421 ABCG2 gene amplification brought about the overexpression of both C421 ABCG2 mRNA and the transporter at the plasma membrane. This resulted in the lack of cellular drug accumulation due to increased efflux of both C1305 and C-1311, a fluorescent imidazoacridone homologue of C-1305, as well as marked resistance to these antitumor agents and to established ABCG2 substrates including mitoxantrone and SN-38. Consistently, the accumulation and sensitivity to these drugs were restored upon incubation with the potent and specific ABCG2 transport inhibitors Ko143 and fumitremorgin C. Moreover, upon transfection into HEK293 cells, the wt Q141 ABCG2 allele displayed a significantly decreased accumulation of C-1311 and increased resistance to C-1305, C-1311 and mitoxantrone, when compared to the K141 ABCG2 transfectant. Hence, the current study provides the first evidence that during the exposure to anticancer drugs, an allele-specific Q141 ABCG2 gene amplification occurs that confers a drug resistance advantage when compared to the K141 ABCG2. These findings have important implications for the selection and expansion of malignant anticancer drug resistant clones during chemotherapy with ABCG2 drugs.

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110 C-1311 accumulation in HEK293 ABCG2 (Q141K) transfectant cells was carried out as above with the exceptions of using a C-1311 concentration range of 0.1-10 mM and the use of the alternative ABCG2 efflux inhibitor fumitremorgin C (5 mM) when co-incubated with C-1311. Login to comment
290 [17] Imai Y, Nakane M, Kage K, et al. C421 polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low level drug resistance. Login to comment

[hide] BCRP gene polymorphisms are associated with suscep... Carcinogenesis. 2007 Aug;28(8):1740-4. Epub 2007 May 10. Hu LL, Wang XX, Chen X, Chang J, Li C, Zhang Y, Yang J, Jiang W, Zhuang SM
BCRP gene polymorphisms are associated with susceptibility and survival of diffuse large B-cell lymphoma.
Carcinogenesis. 2007 Aug;28(8):1740-4. Epub 2007 May 10., [PMID:17494054]

Abstract [show]
To date, the biological significance of breast cancer resistance protein (BCRP) G34A and C421A polymorphisms is largely unknown. Analysis of these two polymorphisms in 156 diffuse large B-cell lymphoma (DLBCL) patients and 376 control subjects revealed an increased risk of DLBCL associated with variant BCRP 421 genotypes (CA and AA), when compared with the wild-type CC genotype [odds ratio = 1.49, 95% confidence interval (CI) 1.02-2.17, P = 0.042]. Moreover, the increased risk was more evident in younger patients (<or=50 years, odds ratio = 2.14, 95% CI 1.25-3.68, P = 0.006). Further evaluation for the association of these polymorphisms with overall survival of DLBCL showed that patients with 34AA alleles displayed worse survival compared with those carrying GG/GA genotypes [hazard ratio (HR) = 3.69, 95% CI 1.56-8.71, P = 0.001]. Significant association between 421CC genotypes and poorer survival of DLBCL was observed in patients younger at diagnosis (<or=50 years, HR = 5.80, 95% CI 1.16-28.90, P = 0.015) or with bulky tumor (HR = 4.36, 95% CI 1.04-18.31, P = 0.027). Furthermore, we found the combined effects of BCRP G34A and C421A on the overall survival. Compared with patients carrying BCRP 34(GG + GA)421(AA + CA) genotype, the individual with 34AA421CC displayed the worst survival (HR = 7.55, 95% CI 2.36-24.17, P = 0.001), while those with 34(GG + GA)421CC and 34AA421(AA + CA) combinations showed the intermediate survival. These results suggest that the BCRP G34A and C421A polymorphisms are associated with the risk and survival of DLBCL. Our finding warrants further investigations on the association of BCRP polymorphisms with susceptibility and clinical outcome of cancer.

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18 BCRP G34A (Val12Met) and C421A (Gln141Lys) polymorphisms occurred at high frequency in most ethnic populations and have been shown to be associated with the expression and activity of BCRP protein. Login to comment
21 Based on these evidences, we hypothesize that BCRP G34A (Val12Met) and C421A (Gln141Lys) polymorphisms should have potential effect on the susceptibility and prognosis of diseases. Login to comment

[hide] Evaluation of drug-transporter interactions using ... Curr Drug Metab. 2007 May;8(4):341-63. Xia CQ, Milton MN, Gan LS
Evaluation of drug-transporter interactions using in vitro and in vivo models.
Curr Drug Metab. 2007 May;8(4):341-63., [PMID:17504223]

Abstract [show]
Drug transporters, including efflux transporters (the ATP binding cassette (ABC) proteins) and uptake transporters (the solute carrier proteins (SLC)), have an important impact on drug disposition, efficacy, drug-drug interactions and toxicity. Identification of the interactions of chemical scaffolds with transporters at the early stages of drug development can assist in the optimization and selection of new drug candidates. In this review, we discuss current in vitro and in vivo models used to investigate the interactions between drugs and transporters such as P-gp, MRP, BCRP, BSEP, OAT, OATP, OCT, NTCP, PEPT1/2 and NT. In vitro models including cell-based, cell-free, and yeast systems as well as in vivo models such as genetic knockout, gene deficient and chemical knockout animals are discussed and compared. The applications, throughput, advantages and limitations of each model are also addressed in this review.

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119 The function of seven single nucleotide polymorphisms (SNPs) in BCRP (V12M, Q141K, A149P, R163K, Q166E, P269S, and S441N BCRP) was determined using membrane vesicles from HEK293 cells infected with the recombinant adenoviruses containing the corresponding BCRP cDNAs [45]. Login to comment
120 The expression levels of Q141K and S441N BCRP proteins were significantly lower compared to the wild type protein and the other five variants. Login to comment
121 Furthermore, the transport rate of estrone sulfate, dehydroepiandrosterone sulfate (DHEAS), methotrexate, and p-aminohippurate was almost the same for the wild type, V12M, Q141K, A149P, R163K, Q166E, and P269S BCRP variants when it is normalized by the expression levels of BCRP protein. Login to comment
122 These results suggest that Q141K SNPs may be associated with a lower expression level, and S441N SNPs may affect both the expression level and cellular localization. Login to comment

[hide] ATP-binding cassette transporter ABCG2 (BCRP) and ... Scand J Gastroenterol. 2007 Jun;42(6):726-33. Fischer S, Lakatos PL, Lakatos L, Kovacs A, Molnar T, Altorjay I, Papp M, Szilvasi A, Tulassay Z, Osztovits J, Papp J, Demeter P, Schwab R, Tordai A, Andrikovics H
ATP-binding cassette transporter ABCG2 (BCRP) and ABCB1 (MDR1) variants are not associated with disease susceptibility, disease phenotype response to medical therapy or need for surgeryin Hungarian patients with inflammatory bowel diseases.
Scand J Gastroenterol. 2007 Jun;42(6):726-33., [PMID:17505995]

Abstract [show]
OBJECTIVE: MDR1 (ABCB1), a member of the ATP-binding cassette (ABC) transporters, is an attractive candidate gene for the pathogenesis of inflammatory bowel diseases (IBD) and perhaps for response to therapy. Since limited data are available on MDR1 and ABCG2 polymorphisms in East European IBD patients, the aim of this study was to investigate ABCG2 and MDR1 variants and responses to medical therapy and/or disease phenotype in Hungarian patients. MATERIAL AND METHODS: A total of 414 unrelated IBD patients (Crohn's disease (CD): 265, age: 35.2+/-12.1 years, duration: 8.7+/-7.6 years and ulcerative colitis (UC): 149, age: 44.4+/-15.4 years, duration: 10.7+/-8.9 years) and 149 healthy subjects were investigated. ABCG2 G34A, C421A and MDR1 C3435T, G2677T/A single nucleotide polymorphisms (SNPs) were detected using real-time polymerase chain reaction (PCR). Detailed clinical phenotypes were determined by reviewing the medical charts. RESULTS: The frequency of the ABCG2 and MDR1 SNPs was not significantly different among IBD, CD, UC patients and controls. There was no difference in risk for steroid resistance in CD patients carrying variant ABCG2 (19.6% versus non-carriers 18.4%, p=NS) or MDR1 3435T (CC: 22.2% versus CT/TT: 17.6%) alleles. In addition, carriage of the variant allele was not associated with disease phenotype, presence of extra-intestinal manifestations, smoking, response to infliximab therapy or the need for surgery. In UC, the carriage of variant ABCG2 alleles seemed to be preventive for arthritis (15.5% versus 31.7%, OR: 0.39, 95% CI: 0.16-0.98). CONCLUSIONS: MDR1 and ABCG2 SNPs were not associated with disease susceptibility or disease phenotype in Hungarian patients, and variant alleles did not predict the response to medical therapy or the need for surgery. Further studies are needed to clarify the association between the presence of ABCG2 variants and arthritis in UC.

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41 For example, Sparreboom et al. [25] found that the ABCG2 C421A (Q141K) variant affected the pharmacokinetics of diflomotecan, a synthetic derivative of camptothecin. Login to comment
60 Detection of MDR1 and ABCG2 polymorphisms The sequence variants (SNPs) of the two ABC transporters studied were: ABCB1 (MDR1): c.2677G/T/A or G2677T/A (Ala893Ser/Thr, exon 21, SNP database ID: rs2032582) and c.3435C/T or C3435T (silent base-substitution I1145I, exon 26, SNP database ID: rs1045642); ABCG2 (BCRP/MXR): c.34G/A or G34A (Val12Met, exon 2, SNP database ID: rs2231137; c.421C/A or C421A (Gln141Lys, exon 5, SNP database ID: rs2231142). Login to comment

[hide] Concordance of pharmacogenetic polymorphisms in tu... Cancer Epidemiol Biomarkers Prev. 2007 May;16(5):1038-41. Weiss JR, Baer MR, Ambrosone CB, Blanco JG, Hutson A, Ford LA, Moysich KB
Concordance of pharmacogenetic polymorphisms in tumor and germ line DNA in adult patients with acute myeloid leukemia.
Cancer Epidemiol Biomarkers Prev. 2007 May;16(5):1038-41., [PMID:17507636]

Abstract [show]
Archived tumor tissue is a useful resource for retrospective studies addressing relationships between genetic polymorphisms and treatment outcomes. However, genotypes determined in tumor and somatic tissues may differ due to cytogenetic and molecular changes associated with malignant transformation and progression. Discordance between germ line and tumor genotypes may be particularly relevant in leukemia because cytogenetic abnormalities are frequent. We compared genotypes determined in DNA extracted from paired pretreatment bone marrow and buccal samples from 80 adult patients with acute myeloid leukemia (AML). Paired AML and buccal DNA samples were genotyped for polymorphisms (21 single nucleotide polymorphisms and 2 gene deletions) on genes encoding proteins involved in drug metabolism (CYP3A4, CYP2C8, CDA, and GSTP1), oxidative stress mechanisms (CAT, MnSOD, GSTT1, GSTM1, GSTA1, and GPX1), drug transport (MDR1, MRP1, and BCRP), and DNA repair (MGMT, XPD, and XRCC1). Genotypes were determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, except GSTM1 and GSTT1, for which deletion genotypes were determined using multiplex PCR. Concordance of genotypes was tested by kappa statistics. kappa statistics for paired AML and buccal DNA samples ranged between 0.94 and 1.00, indicating excellent agreement. The GSTT1 and GSTM1 genotypes were in perfect concordance for the paired samples. Agreement was also excellent for genes at AML chromosome deletion and translocation breakpoints, including MDR1 at 7q21.1 and MRP1 at 16p13.1. Based on these data, genotypes derived from archived AML bone marrow samples were not likely to differ from those from genomic DNA, and archived bone marrow samples may be useful for the conduct of retrospective pharmacogenetic studies.

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60 Gene information and K statistics for matched AML and buccal DNA samples (N = 80) Gene Single nucleotide polymorphism location Chromosome rs nos. j Asymptotic error Confidence interval No. of evaluable cases (%) No. of nonmatching genotype calls ABCB1-03 Ex27-55A>C>G>T; I1144M 7q21.1 rs1045642 1.00 77 (96.3) 0 ABCB1-05 Ex22-9A>C>G>T; A892S 7q21.1 rs2032582 0.96 0.031 0.89-1.02 76 (95.0) 2 ABCB1-24 Ex13+12 A>G; G412G 7q21.1 rs1128503 0.98 0.023 0.95-1.02 76 (95.0) 1 ABCC1 Ex8 T825C; V275V 16p13.1 rs246221 1.00 76 (95.0) 0 ABCC1 Ex28 G4002A; S1334S 16p13.1 rs2230671 1.00 78 (97.5) 0 ABCC1 Ex9 T1062C; L562L 16p13.1 rs35587 0.98 0.022 0.93-1.02 77 (96.3) 1 ABCG2 Ex5 C421A; Q141K 4q22 rs2231142 1.00 75 (93.8) 0 ABCG2 Ex2 G34A; V12M 4q22 rs2231137 1.00 75 (93.8) 0 CAT-01 À329T>C 11p13 rs1001179 1.00 78 (97.5) 0 GPX1-01 Ex1-226C>T; P200L 3p21.3 rs1050450 0.96 0.029 0.90-1.02 79 (98.8) 2 SOD2-01 Ex2+24C>T; V16A 6q25.3 rs1799725 0.98 0.020 0.94-1.02 77 (96.3) 1 GSTA1-01 À4621T>C 6p12.1 rs3957357 1.00 78 (97.5) 0 GSTP1-01 Ex5-24A>G; I105V 11q13 rs947894 0.96 0.030 0.90-1.02 78 (97.5) 2 GSTM1 Gene deletion 1p13.3 1.00 77 (96.3) 0 GSTT1 Gene deletion 22q11.23 1.00 77 (96.3) 0 CYP3A4-02 À391A>G 7q21.1 rs2740574 0.94 0.053 0.89-1.05 77 (96.3) 1 CYP2C8 A1196G; R399K 10q23.33 rs10509681 0.96 0.040 0.88-1.04 77 (96.3) 1 CYP2C8 C792G; M264I 10q23.33 rs1058930 0.96 0.041 0.88-1.04 77 (96.3) 1 MGMT-01 Ex4-13A>G; I143V 10q26 rs2308321 1.00 80 (100.0) 0 XPD312 Ex10-16G>A; D312N 19q13.3 rs1799793 1.00 73 (91.3) 0 XPD751 Ex23-61A>C; K751Q 19q13.3 rs13181 0.98 0.022 0.93-1.02 76 (95.0) 1 XRCC1 Ex10-4A>G; Q399R 19q13.2 rs25487 1.00 74 (92.5) 0 CDA A79C; K27Q 1p36.2-p35 rs2072671 1.00 77 (96.3) 0 NOTE: Values of n: j > 0.75 (excellent agreement beyond chance), j between 0.40 and 0.75 (fair to good agreement), j < 0.40 (poor agreement). Login to comment
62 j Asymptotic error Confidence interval No. of evaluable cases (%) No. of nonmatching genotype calls ABCB1-03 Ex27-55A>C>G>T; I1144M 7q21.1 rs1045642 1.00 77 (96.3) 0 ABCB1-05 Ex22-9A>C>G>T; A892S 7q21.1 rs2032582 0.96 0.031 0.89-1.02 76 (95.0) 2 ABCB1-24 Ex13+12 A>G; G412G 7q21.1 rs1128503 0.98 0.023 0.95-1.02 76 (95.0) 1 ABCC1 Ex8 T825C; V275V 16p13.1 rs246221 1.00 76 (95.0) 0 ABCC1 Ex28 G4002A; S1334S 16p13.1 rs2230671 1.00 78 (97.5) 0 ABCC1 Ex9 T1062C; L562L 16p13.1 rs35587 0.98 0.022 0.93-1.02 77 (96.3) 1 ABCG2 Ex5 C421A; Q141K 4q22 rs2231142 1.00 75 (93.8) 0 ABCG2 Ex2 G34A; V12M 4q22 rs2231137 1.00 75 (93.8) 0 CAT-01 À329T>C 11p13 rs1001179 1.00 78 (97.5) 0 GPX1-01 Ex1-226C>T; P200L 3p21.3 rs1050450 0.96 0.029 0.90-1.02 79 (98.8) 2 SOD2-01 Ex2+24C>T; V16A 6q25.3 rs1799725 0.98 0.020 0.94-1.02 77 (96.3) 1 GSTA1-01 À4621T>C 6p12.1 rs3957357 1.00 78 (97.5) 0 GSTP1-01 Ex5-24A>G; I105V 11q13 rs947894 0.96 0.030 0.90-1.02 78 (97.5) 2 GSTM1 Gene deletion 1p13.3 1.00 77 (96.3) 0 GSTT1 Gene deletion 22q11.23 1.00 77 (96.3) 0 CYP3A4-02 À391A>G 7q21.1 rs2740574 0.94 0.053 0.89-1.05 77 (96.3) 1 CYP2C8 A1196G; R399K 10q23.33 rs10509681 0.96 0.040 0.88-1.04 77 (96.3) 1 CYP2C8 C792G; M264I 10q23.33 rs1058930 0.96 0.041 0.88-1.04 77 (96.3) 1 MGMT-01 Ex4-13A>G; I143V 10q26 rs2308321 1.00 80 (100.0) 0 XPD312 Ex10-16G>A; D312N 19q13.3 rs1799793 1.00 73 (91.3) 0 XPD751 Ex23-61A>C; K751Q 19q13.3 rs13181 0.98 0.022 0.93-1.02 76 (95.0) 1 XRCC1 Ex10-4A>G; Q399R 19q13.2 rs25487 1.00 74 (92.5) 0 CDA A79C; K27Q 1p36.2-p35 rs2072671 1.00 77 (96.3) 0 NOTE: Values of n: j > 0.75 (excellent agreement beyond chance), j between 0.40 and 0.75 (fair to good agreement), j < 0.40 (poor agreement). Login to comment

[hide] The effect of ABCG2 V12M, Q141K and Q126X, known f... Br J Clin Pharmacol. 2007 Nov;64(5):645-54. Epub 2007 May 17. Kim HS, Sunwoo YE, Ryu JY, Kang HJ, Jung HE, Song IS, Kim EY, Shim JC, Shon JH, Shin JG
The effect of ABCG2 V12M, Q141K and Q126X, known functional variants in vitro, on the disposition of lamivudine.
Br J Clin Pharmacol. 2007 Nov;64(5):645-54. Epub 2007 May 17., [PMID:17509035]

Abstract [show]
AIMS: To evaluate the effects of three ABCG2 variants (Q141K, V12M and Q126X), which are known to have altered transport properties in vitro, on the disposition of lamivudine in healthy subjects. METHODS: To evaluate whether lamivudine is a substrate of ABCG2, intracellular accumulation and vectorial transport of 3H-lamivudine were determined in MDCK-ABCG2 cells. The pharmacokinetic parameters of lamivudine were compared among subjects with four different ABCG2 genotypes, including wild type (seven subjects), K141/K141 (six subjects), Q126/Stop126 (four subjects) and M12/M12 (five subjects) after a single oral dose of 100 mg lamivudine. RESULTS: The intracellular accumulation of lamivudine in MDCK-ABCG2 cells was significantly lower than that in MDCK-mock cells, but fumitremorgin C reversed the intracellular lamivudine concentration to that of MDCK-mock cells. The ABCG2-mediated transport of lamivudine was saturable and the values of Km and Vmax were 216.5 +/- 58 microm and 20.42 +/- 2.9 nmol h(-1) per 10(6) cells, respectively. After lamivudine administration to healthy subjects, the AUC of lamivudine showed no difference among subjects with different ABCG2 genotypes; 2480 +/- 502, 2207 +/- 1019, 2422 +/- 239, 2552 +/- 698 ng h(-1) ml(-1) for wild type, K141/K141, Q126/Stop126 and M12/M12 genotype, respectively (P = 0.85). The estimated 95% confidence intervals for the mean difference between K141/K141, Q126/Stop126, M12/M12 and wild as reference were (-1053, 507), (-555, 439) and (-552, 696), respectively. No other pharmacokinetic parameters were estimated to be significantly different among four different ABCG2 genotypes tested. CONCLUSIONS: Lamivudine appeared to be a substrate of ABCG2 in vitro, but the disposition of lamivudine was not significantly influenced by known in vitro functional variants of ABCG2, Q141K, V12M and Q126X in healthy subjects.

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0 The effect of ABCG2 V12M, Q141K and Q126X, known functional variants in vitro, on the disposition of lamivudine Ho-Sook Kim,1 Yu Eun Sunwoo,1 Ji Young Ryu,1 Ho-Jin Kang,1 Hye-Eun Jung,1 Im-Sook Song,1 Eun-Young Kim,1,2 Joo-Cheol Shim,1,3 Ji-Hong Shon1,2 & Jae-Gook Shin1,2 1 Department of Pharmacology and Pharmacogenomics Research Centre, Inje University College of Medicine, 2 Department of Clinical Pharmacology and 3 Department of Psychiatry, Inje University Busan Paik Hosptial, Busan, Korea Correspondence Jae-Gook Shin, MD, PhD, Department of Pharmacology and Clinical Pharmacology, Pharmacogenomics Research Centre, Inje University College of Medicine, 633-165 Gaegum-dong, Jin-gu, Busan 614-735, Korea. Login to comment
2 Received 19 September 2006 Accepted 15 March 2007 Published OnlineEarly 17 May 2007 Aims To evaluate the effects of three ABCG2 variants (Q141K, V12M and Q126X), which are known to have altered transport properties in vitro, on the disposition of lamivudine in healthy subjects. Login to comment
10 Conclusions Lamivudine appeared to be a substrate of ABCG2 in vitro, but the disposition of lamivudine was not significantly influenced by known in vitro functional variants of ABCG2, Q141K, V12M and Q126X in healthy subjects. Login to comment
21 The ABCG2 C421A allele, resulting in a Gln141Lys change, has been associated with low levels of ABCG2 expression and altered sensitivity to the anticancer drugs SN-38, mitoxantrone and topotecan in vitro, compared with the wild type [14]. Login to comment
25 Recently, it was reported that patients with the ABCG2 Gln141Lys variant had elevated plasma concentrations of diflomotecan, a substrate of ABCG2, compared with patients with two wild-type alleles, after intravenous drug administration [16]. Login to comment
26 Very recently, it has also been demonstrated that ABCG2 Gln141Lys polymorphism may play an important role in the pharmacokinetics of rosuvastatin in healthy Chinese men, after excluding the impact of OATP-C and CYP2C9 genetic polymorphisms [17]. Login to comment
53 Weighted nonlinear regression analysis was performed in the fitting using Sigma plot (version 9.0; Systat Software Inc., Richmond, CA, USA) Subjects Unrelated Korean subjects (n = 183) had been genotyped for theABCG2 variants G34A(Val12Met), C376T (Gln126stop) and C421A (Gln141Lys). Login to comment
62 ABCG2 gene to determine the presence of the Val12Met, Gln126Stop and Gln141Lys alleles. Login to comment
85 After washing Table 2 Sequences of primers used for the amplification and sequencing analysis of ABCG2 genotype and the denaturation temperatures used in the PCR Name Primer sequence (5Ј,3Ј) Size PCR ( Tm; °C) ABCG2 V12M F: Biotin-CTCTCCAGATGTCTTCCAGTAATG 278 54 R: GCCAAAACCTGTGAGGTTCA S: CATTGGTGTTTCCTTGTGA ABCG2 Q126X F: GTCTTAGCTGCAAGGAAAGATCCA 174 54.5 R: Biotin-ACTATCAGCCAAAGCACTTACCC S: AATGTAATTCAGGTTACGTG ABCG2 Q141K F: TGATGTTGTGATGGGCACTC 69 54 R: Biotin-GTTGCAAGCCGAAGAGCTG S: GACGGTGAGAGAAAACTT F, forward primer; R, reverse primer; S, sequencing primer; Tm, melting temperature; PCR, polymerase chain reaction. Login to comment
116 Effect of ABCG2 variants on the pharmacokinetics of lamivudine To study the influence of ABCG2 variants on lamivudine pharmacokinetics, we included subjects with the Val12Met, Gln126Stop or Gln141Lys ABCG2 variants. Login to comment
134 Among the known ABCG2 variants with altered functional properties in vitro, the ABCG2 Gln141Lys polymorphism has been reported to be involved in regulation of the level of protein expression [14, 23], drug resistance level and ATPase activity [15]. Login to comment
138 The ABCG2 Val12Met and Gln141Lys variants are common in Koreans, with allelic frequencies of 23 and 27.5%, respectively. Login to comment
145 There has been a previous demonstration that patients carrying the ABCG2 Gln141Lys variant have elevated plasma concentrations of diflomotecan, a substrate of ABCG2, compared with patients with two wild-type alleles, after intravenous drug administration [16]. Login to comment
165 However, the Val12Met, Gln126stop and Gln141Lys polymorphisms of ABCG2 studied here, which have been previously shown to cause functional differences in the protein, did not alter the disposition of a single oral dose of 100 mg lamivudine in healthy volunteers. Login to comment

[hide] Associations of ABCB1, ABCC2, and ABCG2 polymorphi... Cancer. 2007 Jul 1;110(1):138-47. Han JY, Lim HS, Yoo YK, Shin ES, Park YH, Lee SY, Lee JE, Lee DH, Kim HT, Lee JS
Associations of ABCB1, ABCC2, and ABCG2 polymorphisms with irinotecan-pharmacokinetics and clinical outcome in patients with advanced non-small cell lung cancer.
Cancer. 2007 Jul 1;110(1):138-47., 2007-07-01 [PMID:17534875]

Abstract [show]
BACKGROUND: The authors investigated whether ABCB1, ABCC2, and ABCG2 genetic polymorphisms affect pharmacokinetics (PK) of irinotecan and treatment outcome of patients with advanced nonsmall cell lung cancer (NSCLC). METHODS: Blood samples from 107 NSCLC patients treated with irinotecan and cisplatin chemotherapy were used for genotyping ABCB1 (1236C > T, 2677G > T/A, 3435C > T), ABCC2 (-24C > T, 1249G > A, 3972C > T), and ABCG2 (34G > A, 421C > A) polymorphisms. Genotypes were correlated with irinotecan-PK, toxicity, tumor response, and survival. RESULTS: Among 8 polymorphisms, 3435TT and 2677TT were associated with AUC(SN-38G) and CL(SN-38G). When haplotypes are assigned, 2677TT/3435TT carriers showed significantly lower AUC(SN-38G) (P = .006), whereas 2677GG/3435CC carriers showed significantly higher AUC(SN-38) (P = .039). These findings suggest that 2677TT and 3435TT variants are associated with higher efflux activity. In toxicity, the 2677G/T or A was associated with grade 4 neutropenia. The 2677GG carriers showed significantly lower absolute neutrophil count during the 1(st) cycle (P = .012) as well as entire course of chemotherapy (P = .042). The 3435TT was associated with higher frequency of grade 3 diarrhea (P = .047). In tumor response, ABCC2 -24TT and 3972TT genotypes were associated with higher response rates (P = .031 and P = .048, [corrected] respectively) and longer progression-free survival (P = .010 and P = .019, [corrected] respectively), which was sustained in haplotype analysis. CONCLUSIONS: Specific polymorphisms of ABCB1 and ABCC2 can influence disposition and tumor response to irinotecan by regulating transporter activity. These findings may help to individualize irinotecan-based chemotherapy in patients with advanced NSCLC.

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81 of patients Genotype frequencies{ Allele frequencies§ w/w w/m m/m w m ABCB1 1236C > T Synonymous 105 14 57 34 0.405 0.595 2677G > T Ala893Ser 105 22 37 (GT) 10 (TT) 0.457 0.338 (T) 2677G > A Ala893Thr 15 (GA) 14 (TA) 0.205 (A) 7 (AA) 3435C > T Synonymous 105 43 51 11 0.652 0.348 ABCC2 À24C > T - 107 57 47 3 0.752 0.248 1249G > A Val417Ile 107 86 19 2 0.893 0.107 3972C > T Synonymous 107 51 48 8 0.701 0.299 ABCG2 34G > A Val12Met 106 60 41 5 0.759 0.241 421C > A Gln141Lys 105 59 42 4 0.762 0.238 w indicates wild type allele; m, mutant type allele. Login to comment

[hide] Role of UGT1A1*6, UGT1A1*28 and ABCG2 c.421C>A pol... Cancer Sci. 2007 Sep;98(9):1461-7. Epub 2007 Jul 11. Jada SR, Lim R, Wong CI, Shu X, Lee SC, Zhou Q, Goh BC, Chowbay B
Role of UGT1A1*6, UGT1A1*28 and ABCG2 c.421C>A polymorphisms in irinotecan-induced neutropenia in Asian cancer patients.
Cancer Sci. 2007 Sep;98(9):1461-7. Epub 2007 Jul 11., [PMID:17627617]

Abstract [show]
The objectives of the present study were (i) to study the pharmacogenetics of UGT1A1*6, UGT1A1*28 and ABCG2 c.421C>A in three distinct healthy Asian populations (Chinese, Malays and Indians), and (ii) to investigate the polygenic influence of these polymorphic variants in irinotecan-induced neutropenia in Asian cancer patients. Pharmacokinetic and pharmacogenetic analyses were done after administration of irinotecan as a 90-min intravenous infusion of 375 mg/m(2) once every 3 weeks (n = 45). Genotypic-phenotypic correlates showed a non-significant influence of UGT1A1*28 and ABCG2 c.421C>A polymorphisms on the pharmacokinetics of SN-38 (P > 0.05), as well as severity of neutropenia (P > 0.05). Significantly higher exposure levels to SN-38 (P = 0.018), lower relative extent of glucuronidation (REG; P = 0.006) and higher biliary index (BI; P = 0.003) were found in cancer patients homozygous for the UGT1A1*6 allele compared with patients harboring the reference genotype. The mean absolute neutrophil count (ANC) was 85% lower and the prevalence of grade 4 neutropenia (ANC < or = 500/microL) was 27% in patients homozygous for UGT1A1*6 compared with the reference group. Furthermore, the presence of the UGT1A1*6 allele was associated with an approximately 3-fold increased risk of developing severe grade 4 neutropenia compared with patients harboring the reference genotype. These exploratory findings suggest that homozygosity for UGT1A1*6 allele may be associated with altered SN-38 disposition and may increase the risk of severe neutropenia in Asian cancer patients, particularly in the Chinese cancer patients who comprised 80% (n = 36) of the patient population in the present study.

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21 Apart from the influence of UGT1A1*6 and UGT1A1*28 on irinotecan-induced neutropenia, a recent in vitro study by Imai et al.(16) suggested that the non-synonymous ABCG2 c.421C>A (Q141K) polymorphism in exon 5 exhibited low-level drug resistance and increased the sensitivity of normal cells to SN-38 compared with cell lines transfected with ABCG2 gene harboring the reference genotype. Login to comment

[hide] Membrane cholesterol selectively modulates the act... Biochim Biophys Acta. 2007 Nov;1768(11):2698-713. Epub 2007 Jul 10. Telbisz A, Muller M, Ozvegy-Laczka C, Homolya L, Szente L, Varadi A, Sarkadi B
Membrane cholesterol selectively modulates the activity of the human ABCG2 multidrug transporter.
Biochim Biophys Acta. 2007 Nov;1768(11):2698-713. Epub 2007 Jul 10., [PMID:17662239]

Abstract [show]
The human ABCG2 multidrug transporter provides protection against numerous toxic compounds and causes multidrug resistance in cancer. Here we examined the effects of changes in membrane cholesterol on the function of this protein. Human ABCG2 was expressed in mammalian and in Sf9 insect cells, and membrane cholesterol depletion or enrichment was achieved by preincubation with beta cyclodextrin or its cholesterol-loaded form. We found that mild cholesterol depletion of intact mammalian cells inhibited ABCG2-dependent dye and drug extrusion in a reversible fashion, while the membrane localization of the transporter protein was unchanged. Cholesterol enrichment of cholesterol-poor Sf9 cell membrane vesicles greatly increased ABCG2-driven substrate uptake, substrate-stimulated ATPase activity, as well as the formation of a catalytic cycle intermediate (nucleotide trapping). Interestingly, modulation of membrane cholesterol did not significantly affect the function of the R482G or R482T substrate mutant ABCG2 variants, or that of the MDR1 transporter. The selective, major effect of membrane cholesterol on the wild-type ABCG2 suggests a regulation of the activity of this multidrug transporter during processing or in membrane micro-domain interactions. The experimental recognition of physiological and pharmacological substrates of ABCG2, as well as the fight against cancer multidrug resistance may be facilitated by demonstrating the key role of membrane cholesterol in this transport activity.

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30 There are several polymorphic variants of ABCG2 present in large percentage in the human population (e.g. V12M, Q141K), and the possible alterations in the transport capacity and substrate handling of these variants have been examined in numerous experimental systems [18-24]. Login to comment

[hide] Breast cancer resistance protein 1 limits fetal di... Drug Metab Dispos. 2007 Dec;35(12):2154-8. Epub 2007 Sep 4. Zhang Y, Wang H, Unadkat JD, Mao Q
Breast cancer resistance protein 1 limits fetal distribution of nitrofurantoin in the pregnant mouse.
Drug Metab Dispos. 2007 Dec;35(12):2154-8. Epub 2007 Sep 4., [PMID:17785426]

Abstract [show]
The efflux transporter, the breast cancer resistance protein (BCRP), is most abundantly expressed in the apical membrane of the placental syncytiotrophoblasts, indicating that it could play an important role in protecting the fetus by limiting xenobiotic/drug penetration across the placental barrier. In the present study, we examined whether Bcrp1, the murine homolog of human BCRP, limits fetal distribution of the model BCRP/Bcrp1 substrate, nitrofurantoin (NFT), in the pregnant mouse. NFT was administered i.v. to FVB wild-type and Bcrp1(-/-) pregnant mice. The maternal plasma samples and fetuses were collected at various times (5-60 min) after drug administration. The NFT concentrations in the maternal plasma samples and homogenates of fetal tissues were determined by a high-performance liquid chromatography/UV assay. Although the maternal plasma area under the concentration-time curve (AUC) of NFT in the Bcrp1(-/-) pregnant mice (97.4 +/- 10.0 microg . min/ml plasma) was only slightly (but significantly) higher than that in the wild-type pregnant mice (78.4 +/- 6.0 microg . min/ml plasma), the fetal AUC of NFT in the Bcrp1(-/-) pregnant mice (1493.0 +/- 235.3 ng . min/g of fetus) was approximately 5 times greater than that in the wild-type pregnant mice (298.6 +/- 77.4 ng . min/g of fetus). These results clearly suggest that Bcrp1 significantly limits fetal distribution of NFT in the pregnant mouse, but has only a minor effect on the systemic clearance of the drug.

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174 Thus, the fetuses of pregnant women with BCRP natural variants, such as Q141K, that display lower transport activity (Imai et al., 2002) may have an increased risk for potential NFT toxicity. Login to comment

[hide] Pharmacogenetic assessment of toxicity and outcome... J Clin Oncol. 2007 Oct 10;25(29):4528-35. Marsh S, Paul J, King CR, Gifford G, McLeod HL, Brown R
Pharmacogenetic assessment of toxicity and outcome after platinum plus taxane chemotherapy in ovarian cancer: the Scottish Randomised Trial in Ovarian Cancer.
J Clin Oncol. 2007 Oct 10;25(29):4528-35., 2007-10-10 [PMID:17925548]

Abstract [show]
PURPOSE: Standard therapy for advanced ovarian cancer consists of a platinum agent in combination with a taxane, which has a 5-year survival rate of approximately 45%. The large individual variability for ovarian cancer patients in both outcome and toxicity risk from chemotherapy makes the identification of pharmacogenetic markers that can be used to screen patients before therapy selection an attractive prospect. PATIENTS AND METHODS: We assessed 27 selected polymorphisms based on previously described associations or putative functional effects in 16 key genes from pathways that may influence cellular sensitivity to taxanes (ABCB1, ABCC1, ABCC2, ABCG2, CDKN1A, CYP1B1, CYP2C8, CYP3A4, CYP3A5, MAPT, and TP53) and platinum (ABCC2, ABCG2, ERCC1, ERCC2, GSTP1, MPO, and XRCC1) using polymerase chain reaction and Pyrosequencing in 914 ovarian cancer patients from the Scottish Randomised Trial in Ovarian Cancer phase III trial who were treated at presentation with carboplatin and taxane regimens after cytoreductive surgery. RESULTS: No reproducible significant associations between genotype and outcome or toxicity were found for any of the genes analyzed. Previously reported genotype associations could not be replicated in this large study of a well-defined patient population within one specific clinical trial. CONCLUSION: There are no clear candidates for taxane/platinum pharmacogenetic markers. This study highlights the need for validation of putative genetic markers in large, well-defined clinical sample sets.

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55 Homozygous Wild-Type Heterozygous Homozygous Variant p‫ء‬ q† ABCB1 1236CϾT Pac, Doc 896 287 448 161 0.57 0.43 ABCB1 2677GϾT‡§ Pac, Doc 900 269 416 162 0.55 0.42 ABCB1 3435CϾT Pac, Doc 898 207 445 246 0.48 0.52 ABCC1 S1334S Pac, Doc 890 469 360 61 0.73 0.27 ABCC1 IVS18-30CϾG Pac, Doc 886 649 215 22 0.85 0.15 ABCC2 24CϾT Pac, Doc, Carb 899 593 277 29 0.81 0.19 ABCC2 IVS12ϩ148AϾG Pac, Doc, Carb 892 338 422 132 0.62 0.38 ABCC2 V417I Pac, Doc, Carb 867 575 264 28 0.82 0.18 ABCG2 Q141K Pac, Doc, Carb 904 726 168 10 0.90 0.10 CDKN1A 10971CϾT Pac, Doc 873 749 124 0 0.93 0.07 CYP1B1‫ء‬ 3 Pac, Doc 823 228 421 174 0.53 0.47 CYP2C8 M264I Pac 892 683 191 18 0.94 0.06 CYP2C8 R139K Pac 905 799 104 2 0.87 0.13 CYP2C8 K399R Pac 841 650 174 17 0.88 0.12 CYP3A4‫ء‬ 1B Pac, Doc 875 838 37 0 0.98 0.02 CYP3A4‫ء‬ 3ʈ Pac, Doc 900 893 7 0 0.996 0.004 CYP3A5‫ء‬ 3C Pac, Doc 883 6 95 782 0.06 0.94 ERCC1 17677GϾT Carb 889 742 136 11 0.91 0.09 ERCC1 8092CϾA Carb 854 477 317 60 0.74 0.26 ERCC1 N118N Carb 869 347 396 126 0.63 0.37 ERCC2 K751Q Carb 901 367 398 136 0.63 0.37 GSTP1 A114V Carb 867 748 114 5 0.93 0.07 GSTP1 I105V Carb 881 394 367 120 0.66 0.34 MAPT P587P Pac, Doc 853 791 59 3 0.96 0.04 MPO -463GϾA Carb 888 565 277 46 0.79 0.21 TP53 R72P Pac, Doc 877 495 323 59 0.75 0.25 XRCC1 R399Q Carb 869 390 395 84 0.68 0.32 Abbreviations: SCOTROC1, Scottish Randomised Trial in Ovarian Cancer; Pac, paclitaxel; Doc, docetaxel; Carb, carboplatin. Login to comment
72 However, in a study Table 3. Uncorrected P Values From ␹2 Analysis of Development Set From SCOTROC1 Data Gene and Variant Neurotoxicity (paclitaxel) Neurotoxicity (docetaxel) Hematologic (docetaxel) Grade 3/4 GI (paclitaxel) Grade 3/4 GI (docetaxel) ABCB1 1236CϾT .790 .179 .864 .649 .559 ABCB1 2677GϾT/A .816 .783 .740 .382 .472 ABCB1 3435CϾT .362 .771 .345 .507 .919 ABCC1 S1334S .981 .745 .327 .880 .125 ABCC1 IVS18-30CϾG .943 1.000 .623 .053 .354 ABCC2 -24CϾT .975 .567 .348 .961 .537 ABCC2 IVS12ϩ148AϾG .655 .935 .227 .570 .017 ABCC2 V417I .059 .738 .351 .838 .182 ABCG2 Q141K .711 .318 .838 .086 .205 CDKN1A 10971CϾT .359 1.000 .094 .483 .002‫ء‬ CYP1B1‫ء‬ 3 .709 .587 .764 .175 .006‫ء‬ CYP2C8 M264I .601 .069 1.000 1.000 .148 CYP2C8 R139K .421 .315 .314 .020 .157 CYP2C8 K399R .232 .353 .285 .007 .147 CYP3A4‫ء‬ 1B .590 1.000 .714 .750 .743 CYP3A5‫ء‬ 3C .121 .243 1.000 .730 .272 ERCC1 17677GϾT .171 .079 .705 .159 .638 ERCC1 8092CϾA .010 .662 .069 .683 .975 ERCC1 N118N .261 .427 .171 .655 .981 ERCC2 K751Q .206 .209 .501 .062 .108 GSTP1 A114V .231 .400 .812 1.000 .072 GSTP1 I105V .205 .018 .467 .566 .277 MAPT P587P .797 .308 .590 .329 .361 MPO -463GϾA .236 .724 .829 .496 .138 TP53 R72P .298 .954 .945 .490 .157 XRCC1 R399Q .917 .467 .068 .793 .977 NOTE. Login to comment
94 For example, polymorphisms in SLCO1B3 were not associated with taxanepharmacokinetics.42 Inadditiontoidentifyingfunctionalpoly- morphisms in known candidate genes, ongoing genome-wide strategies may provide novel candidate genes/chromosome regions for future pharmacogenetics studies.43 Recent studies also suggest that Table 4. Uncorrected P Values From ␹2 (response) and Log-Rank (progression-free survival) Analysis for Genotype-Outcome Associations in the Development Set Gene and Variant Progression-Free Survival (567-594 patients) CA-125 Response (260-287 patients) Clinical/Radiologic Response (295- 341 patients) ABCB1 1236CϾT .719 .897 .541 ABCB1 2677GϾT/A‫ء‬ .023 .308 .138 ABCB1 3435CϾT .614 .500 .740 ABCC1 S1334S .671 1.000 .903 ABCC1 IVS18-30CϾG .335 .226 .305 ABCC2 -24CϾT .619 .616 .693 ABCC2 IVS12ϩ148AϾG .712 .689 .848 ABCC2 V417I .278 .582 .503 ABCG2 Q141K .427 1.000 .344 CDKN1A 10971CϾT .305 .014 1.000 CYP1B1‫ء‬ 3 .120 .865 .291 CYP2C8 M264I .481 .762 .295 CYP2C8 R139K .572 .080 .154 CYP2C8 K399R .617 .004 .319 CYP3A4‫ء‬ 1B .687 1.000 .780 CYP3A5‫ء‬ 3C .499 .570 .909 ERCC1 17677GϾT .517 .292 .635 ERCC1 8092CϾA .677 .029 .593 ERCC1 N118N .851 .214 .204 ERCC2 K751Q .562 .613 .198 GSTP1 A114V .178 .512 .243 GSTP1 I105V .693 .187 .939 MAPT P587P .253 1.000 .827 MPO -463GϾA .537 .349 .088 TP53 R72P .212 .878 .309 XRCC1 R399Q .254 .422 .256 NOTE. Login to comment

[hide] Synergistic interaction of ABCB1 and ABCG2 polymo... Pharmacogenomics J. 2008 Oct;8(5):321-7. Erdilyi DJ, Kamory E, Csokay B, Andrikovics H, Tordai A, Kiss C, Filni-Semsei A, Janszky I, Zalka A, Fekete G, Falus A, Kovacs GT, Szalai C
Synergistic interaction of ABCB1 and ABCG2 polymorphisms predicts the prevalence of toxic encephalopathy during anticancer chemotherapy.
Pharmacogenomics J. 2008 Oct;8(5):321-7., [PMID:17938643]

Abstract [show]
Polymorphisms of the ABCB1 (MDR1) and ABCG2 (BCRP) genes were reported to alter the expression and function of these drug transporters. Both proteins are present at the main pharmacokinetic barriers including the blood-brain barrier. Data from 291 children with acute lymphoblastic leukaemia were analysed in this retrospective study. ABCB1 3435T>C, 2677G>T/A, 1236C>T and ABCG2 421C>A, 34G>A genotypes were determined. Encephalopathy episodes were more frequent among those with ABCB1 3435TT genotype than in the 3435CC/CT group (odds ratio (OR) 3.5; P=0.03). Patients with the ABCG2 421A allele tended to have more complications than wild type homozygotes (OR=2.0; P=0.25). The rate of the adverse effect was similar in those harbouring no or only one of the predisposing genotypes, that is, either ABCB1 3435TT or ABCG2 421AA/AC. However, significantly more children suffered encephalopathy in the group with both predisposing genotypes (OR=12.3; P=0.005). In conclusion, these variations exert synergistic effect in predisposing patients to toxic neurological complications of chemotherapy.

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23 According to the first theory this SNP and other variations (2677G4T/A and 1236C4T) in linkage disequilibrium are linked to a putative functional polymorphism that causes the differences observed.10 On the other hand it was shown that the 3435T mRNA is characterized by decreased stability, while the linked alleles themselves have no effect on gene expression.11 The ABCG2 34G4A and 421C4A are rare non-synonymous polymorphisms resulting in V12M and Q141K variations. Login to comment

[hide] Erlotinib (Tarceva, OSI-774) antagonizes ATP-bindi... Cancer Res. 2007 Nov 15;67(22):11012-20. Shi Z, Peng XX, Kim IW, Shukla S, Si QS, Robey RW, Bates SE, Shen T, Ashby CR Jr, Fu LW, Ambudkar SV, Chen ZS
Erlotinib (Tarceva, OSI-774) antagonizes ATP-binding cassette subfamily B member 1 and ATP-binding cassette subfamily G member 2-mediated drug resistance.
Cancer Res. 2007 Nov 15;67(22):11012-20., 2007-11-15 [PMID:18006847]

Abstract [show]
It has been reported that gefitinib, an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), has the ability to modulate the function of certain ATP-binding cassette (ABC) transporters and to reverse ABC subfamily B member 1 (ABCB1; P-glycoprotein)- and ABC subfamily G member 2 (ABCG2; breast cancer resistance protein/mitoxantrone resistance protein)-mediated multidrug resistance (MDR) in cancer cells. However, it is unknown whether other EGFR TKIs have effects similar to that of gefitinib. In the present study, we have investigated the interaction of another EGFR TKI, erlotinib, with selected ABC drug transporters. Our findings show that erlotinib significantly potentiated the sensitivity of established ABCB1 or ABCG2 substrates and increased the accumulation of paclitaxel or mitoxantrone in ABCB1- or ABCG2-overexpressing cells. Furthermore, erlotinib did not significantly alter the sensitivity of non-ABCB1 or non-ABCG2 substrates in all cells and was unable to reverse MRP1-mediated MDR and had no effect on the parental cells. However, erlotinib remarkably inhibited the transport of E(2)17 beta G and methotrexate by ABCG2. In addition, the results of ATPase assays show that erlotinib stimulated the ATPase activity of both ABCB1 and ABCG2. Interestingly, erlotinib slightly inhibited the photolabeling of ABCB1 with [(125)I]iodoarylazidoprazosin (IAAP) at high concentration, but it did not inhibit the photolabeling of ABCG2 with IAAP. Overall, we conclude that erlotinib reverses ABCB1- and ABCG2-mediated MDR in cancer cells through direct inhibition of the drug efflux function of ABCB1 and ABCG2. These findings may be useful for cancer combinational therapy with erlotinib in the clinic.

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226 November 15, 2007 11018 www.aacrjournals.org American Association for Cancer ResearchCopyright (c) 2007 on May 3, Recently, Cusatis et al. (43) reported that one common functional single-nucleotide polymorphism in the ABCG2 gene, ABCG2 421C!A (Q141K), is associated with diarrhea, which is a gefitinib-induced adverse effect, and led to a high risk of diarrhea in patients treated with oral gefitinib. Login to comment

[hide] Determination of ancestral allele for possible hum... Cancer Genet Cytogenet. 2008 Jan 1;180(1):24-9. Maruta Y, Okayama N, Hiura M, Suehiro Y, Hirai H, Hinoda Y
Determination of ancestral allele for possible human cancer-associated polymorphisms.
Cancer Genet Cytogenet. 2008 Jan 1;180(1):24-9., 2008-01-01 [PMID:18068529]

Abstract [show]
To determine ancestral allele in possible cancer-associated polymorphisms, DNA samples from 10 chimpanzees (Pan troglodytes) were sequenced for alleles corresponding to 17 polymorphisms: 8 short tandem repeats [IL1RN (alias IL-1RA) variable number tandem repeat (VNTR); TYMS (previously TS) VNTR; AR CAG repeat; dinucleotide repeats of UGT1A1, IGF1, IFNG (alias IFN-gamma), ESR1 (alias ER-alpha), and EGFR] and 9 single nucleotide polymorphisms (MMP1-1607 1G/2G, MMP3-1171 5A/6A, OGG1 Ser326Cys, ALDH2 Gly487Lys, TP53 Arg72Pro, ABCG2 Gln141Lys, MGMT Leu84Phe, SOD2 Ala-9Val, and MTHFR Ala222Val). No chimpanzee polymorphism corresponded to human IL1RN VNTR; the ancestral allele was a repeat lost in humans. Dinucleotide repeat polymorphisms of IGF1, IFNG, ESR1, and EGFR were shared by chimpanzees, but the length of repeats tended to be longer in humans than in chimpanzees. This tendency was particularly evident for IGF1. All of the SNPs tested are human-specific nucleotide changes. The ancestral allele 7A was shown to be lost in MMP3-1171 5A/6A. Thus, all of the possible cancer-associated polymorphisms tested have human-specific alleles, and the ancestral allele is lost in three polymorphisms (IL1RN VNTR, UGT1A1 CA repeat, and MMP3-1171 5A/6A), suggesting a possible involvement of human-specific alleles in cancer susceptibility.

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0 Determination of ancestral allele for possible human cancer-associated polymorphisms Yuichi Marutaa,b , Naoko Okayamab , Mikako Hiuraa , Yutaka Suehiroa , Hirohisa Hiraic , Yuji Hinodaa,b,* a Department of Laboratory Medicine, Yamaguchi University Graduate School of Medicine, 1-1-1, Minami-Kogushi, Ube, Yamaguchi 755-8505 Japan b Division of Clinical Laboratory, Yamaguchi University Hospital, Ube, Japan c Department of Cellular and Molecular Biology, Primate Research Institute, Kyoto University, Inuyama, Japan Received 31 August 2007; accepted 19 September 2007 Abstract To determine ancestral allele in possible cancer-associated polymorphisms, DNA samples from 10 chimpanzees (Pan troglodytes) were sequenced for alleles corresponding to 17 polymorphisms: 8 short tandem repeats [IL1RN (alias IL-1RA) variable number tandem repeat (VNTR); TYMS (previously TS ) VNTR; AR CAG repeat; dinucleotide repeats of UGT1A1, IGF1, IFNG (alias IFN-g), ESR1 (alias ER-a), and EGFR] and 9 single nucleotide polymorphisms (MMP1-1607 1G/2G, MMP3-1171 5A/6A, OGG1 Ser326Cys, ALDH2 Gly487Lys, TP53 Arg72Pro, ABCG2 Gln141Lys, MGMT Leu84Phe, SOD2 Ala-9Val, and MTHFR Ala222Val). Login to comment
42 subfamily G2 ABCG2 Gln141Lys (rs2231142); O-6-methylguanine-DNA methyltransferase MGMT Leu84Phe (rs12917); mitochondrial superoxide dismutase SOD2 (alias MnSOD) Ala-9Val (rs4880); and methylenetetrahydrofolate reductase MTHFR Ala222Val (rs1801133). Login to comment

[hide] The breast cancer resistance protein (Bcrp1/Abcg2)... Mol Pharmacol. 2008 Mar;73(3):949-59. Epub 2007 Dec 13. Zhou L, Naraharisetti SB, Wang H, Unadkat JD, Hebert MF, Mao Q
The breast cancer resistance protein (Bcrp1/Abcg2) limits fetal distribution of glyburide in the pregnant mouse: an Obstetric-Fetal Pharmacology Research Unit Network and University of Washington Specialized Center of Research Study.
Mol Pharmacol. 2008 Mar;73(3):949-59. Epub 2007 Dec 13., [PMID:18079276]

Abstract [show]
Breast cancer resistance protein (BCRP) is most abundantly expressed in the apical membrane of placental syncytiotrophoblasts, suggesting that it may protect the fetus by impeding drug penetration across the placental barrier. Glyburide (GLB) is an antidiabetic drug routinely used to treat gestational diabetes. In this study, we first determined whether GLB is a BCRP/Bcrp1 substrate. The intracellular [(3)H]GLB concentrations in Madin-Darby canine kidney (MDCK)/BCRP cells were significantly lower than those in MDCK/vector cells. The addition of 10 muM fumitremorgin C, a specific BCRP inhibitor, significantly increased the intracellular [(3)H]GLB concentrations approximately 2-fold in MDCK/BCRP cells, but it had no effect in MDCK/vector cells. Similar results were obtained using MDCKII parent and MDCKII/Bcrp1 cells. GLB was also shown to be a BCRP/Bcrp1 substrate in transwell transport experiments. We then examined whether Bcrp1 limits fetal distribution of GLB in the pregnant mouse. GLB was administered by retro-orbital injection to the wild-type and Bcrp1(-/-) pregnant mice. The maternal plasma samples and fetuses were collected at various times (0.5-240 min) after drug administration. The GLB concentrations in the maternal plasma samples and homogenates of fetal tissues were determined by high-performance liquid chromatography/mass spectrometry. Although the maternal plasma area under the concentration-time curves (AUCs) of GLB in the wild-type and Bcrp1(-/-) pregnant mice were comparable, the fetal AUC of GLB in the Bcrp1(-/-) pregnant mice was approximately 2 times greater than that in the wild-type pregnant mice. These results suggest that GLB is a BCRP/Bcrp1 substrate, and Bcrp1 significantly limits fetal distribution of GLB in the pregnant mouse, but it has only a minor effect on the systemic clearance of the drug.

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323 For example, the fetuses of pregnant women carrying the BCRP natural variant Q141K, which displays lower transport activity (Imai et al., 2002), may be at increased risk for the GLB toxicity. Login to comment

[hide] ABC multidrug transporters: structure, function an... Pharmacogenomics. 2008 Jan;9(1):105-27. Sharom FJ
ABC multidrug transporters: structure, function and role in chemoresistance.
Pharmacogenomics. 2008 Jan;9(1):105-27., [PMID:18154452]

Abstract [show]
Three ATP-binding cassette (ABC)-superfamily multidrug efflux pumps are known to be responsible for chemoresistance; P-glycoprotein (ABCB1), MRP1 (ABCC1) and ABCG2 (BCRP). These transporters play an important role in normal physiology by protecting tissues from toxic xenobiotics and endogenous metabolites. Hydrophobic amphipathic compounds, including many clinically used drugs, interact with the substrate-binding pocket of these proteins via flexible hydrophobic and H-bonding interactions. These efflux pumps are expressed in many human tumors, where they likely contribute to resistance to chemotherapy treatment. However, the use of efflux-pump modulators in clinical cancer treatment has proved disappointing. Single nucleotide polymorphisms in ABC drug-efflux pumps may play a role in responses to drug therapy and disease susceptibility. The effect of various genotypes and haplotypes on the expression and function of these proteins is not yet clear, and their true impact remains controversial.

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355 Over 80 SNPs, missense, nonsense and frameshift mutations in the ABCG2 gene have been identified in different ethnic groups [23,170], including V12M (N-terminal cytosolic region), Q141K (NBD) and Q126stop (in which no active protein is produced). Login to comment
357 In a study of six different SNP variants, the C421A polymorphism (nonsynonymous, Q141K) was expressed at lower levels, and the S441N variant had both lower expression and altered localization [171]. Login to comment
358 The Q141K mutation is located between the Walker A and signature motifs, so altered ATPase activity is possible. Login to comment
359 Compared with wild-type ABCG2, the Q141K variant displayed lower ATPase activity and lower mitoxantrone efflux when expressed in HEK-293 cells, whereas the V12M and D620N proteins showed little change [172]. Login to comment
360 Somewhat different results were reported by another group for the V12M and Q141K variants [173]. Login to comment
361 A recent study examined seven ABCG2 variants in detail, and found that cells expressing both V12M and Q141K had reduced resistance towards the drug SN-38 [174]. Login to comment
362 Several different studies that examined the expression level of the Q141K variant, both in human tissues and in transfected cell lines, have yielded contradictory results [175]. Login to comment
363 The frequency of the Q141K polymorphism varies considerably among different ethnic populations, being commonly found in China and Japan, and was found to be part of a common haplotype [176,177]. Login to comment
365 The accumulation of the tyrosine kinase inhibitor gefitinib was higher in patients heterozygous at the C421A (Q141K) locus compared with those with a wild-type genotype, indicating that this polymorphism may indeed affect the outcome of anticancer drug treatment [181]. Login to comment

[hide] In vitro evaluation of photosensitivity risk relat... Drug Metab Pharmacokinet. 2007 Dec;22(6):428-40. Tamura A, Onishi Y, An R, Koshiba S, Wakabayashi K, Hoshijima K, Priebe W, Yoshida T, Kometani S, Matsubara T, Mikuriya K, Ishikawa T
In vitro evaluation of photosensitivity risk related to genetic polymorphisms of human ABC transporter ABCG2 and inhibition by drugs.
Drug Metab Pharmacokinet. 2007 Dec;22(6):428-40., [PMID:18159130]

Abstract [show]
Since porphyrins are regarded as endogenous substrates for the ATP-binding cassette (ABC) transporter ABCG2, it is hypothesized that functional impairment owing to genetic polymorphisms or inhibition of ABCG2 by drugs may result in a disruption of cellular porphyrin homeostasis. In the present study, we expressed ABCG2 genetic variants, i.e., V12M, Q141K, S441N, and F489L, as well as the wild type (WT) in Flp-In-293 cells to examine the hypothesis. Cells expressing S441N and F489L variants exhibited high levels of both cellularly accumulated pheophorbide a and photosensitivity, when those cells were incubated with pheophorbide a and irradiated with visible light. To further elucidate the significance of ABCG2 in cellular porphyrin homeostasis, we observed cellular accumulation and compartmentation of porphyrin and pheophorbide a by means of a new fluorescence microscopy technology, and found that accumulation of porphyrin and pheophorbide a in the cytoplasm compartment was maintained at low levels in Flp-In-293 cells expressing ABCG2 WT, V12M, or Q141K. When ABCG2 was inhibited by imatinib or novobiocin, however, those cells became sensitive to light. Based on these results, it is strongly suggested that certain genetic polymorphisms and/or inhibition of ABCG2 by drugs can enhance the potential risk of photosensitivity.

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8 In the present study, we expressed ABCG2 genetic variants, i.e., V12M, Q141K, S441N, and F489L, as well as the wild type (WT) in Flp-In-293 cells to examine the hypothesis. Login to comment
10 To further elucidate the signiˆcance of ABCG2 in cellular porphyrin homeostasis, we observed cellular accumulation and compartmentation of porphyrin and pheophorbide a by means of a new ‰uorescence microscopy technology, and found that accumulation of porphyrin and pheophorbide a in the cytoplasm compartment was maintained at low levels in Flp-In-293 cells expressing ABCG2 WT, V12M, or Q141K. Login to comment
98 Genetic polymorphisms of human ABCG2 and pheophorbide a-photosensitivity In vitro experiments SNP data IC50 (mM) Photosensitivity ratio (fold) Ethnic group N Allele frequency (z) Reference WT 3.0 1.0 - - - - V12M 4.1 0.7 Caucasian 546 5.6 22, 13, 21, 20, 14 Japanese 259 17.6 18, 22, 20 African 181 6.3 22, 20 Q141K 2.9 1.0 Caucasian 717 11.0 22, 13, 21, 15, 20, 14 Japanese 354 30.6 18, 22, 20 African 1213 1.4 22, 15, 14 S441N 0.5 6.0 Japanese 100 0.5 20 F489L 1.7 1.8 Japanese 160 0.6 19, 20 Pheophorbide a-photosensitivity ratios and IC50 values were determined from the data shown in Fig. 2B. 432 Ai TAMURA, et al. a Fluoroskan Ascent FL (Thermo Labsystems, Helsinki, Finland) (excitation at 405 nm; emission at 612 nm). Login to comment
99 Results Expression of ABCG2 WT and SNP variants in Flp-In-293 cells: In the present study, we aimed to examine the impact of hitherto reported major SNPs (V12M, Q141K, S441N, or F489L) on the photo-sensitivity. Login to comment
108 In addition, the protein level of the Q141K SNP variant was moderately lower than that of ABCG2 WT. Login to comment
110 Figure 1B depicts the immuno‰uorescence images of Flp-In-293 cells expressing ABCG2 WT and those SNP variants (i.e., V12M, Q141K, S441N, and F489L) as well as mock vector-transfected cells (Flp-In-293/ Mock). Login to comment
114 In contrast, strong green ‰uorescence was observed at the plasma membrane and within intracellular compartments in Flp-In-293 cells expressing ABCG2 WT as well as the SNP variants of V12M, Q141K, and F489L. Login to comment
121 In contrast, intracellular accumulations of pheophorbide a in both Flp-In-293/ABCG2 (V12M) and Flp-In-293/ABCG2 (Q141K) cells were signiˆcantly lower, being similar to the levels observed in Flp-In-293/ABCG2 (WT) cells. Login to comment
122 It is suggested that two variants, V12M and Q141K, actively extruded pheophorbide a out of cells as did ABCG2 WT, 1. Login to comment
123 Expression of human ABCG2 WT, V12M, Q141K, S441N, and F489L in Flp-In-293 cells. Login to comment
130 Photosensitivity of Flp-In-293 cells expressing ABCG2 WT and SNP variants: Figure 2B demonstrates the cellular photosensitivity proˆles of Flp-In-293 cells expressing ABCG2 WT, V12M, Q141K, S441N, and F489L, as well as that of Flp-In-293/Mock cells. Login to comment
136 Flp-In-293/Mock and Flp-In-293/ABCG2 (S441N) cells were very sensitive to light, whereas Flp-In-293/ABCG2 (V12M), Flp-In-293/ABCG2 (Q141K), and Flp-In-293/ABCG2 (WT) cells were signiˆcantly more resistant. Login to comment
141 A, Flp-In-293 cells expressing human ABCG2 WT and SNP variants (V12M, Q141K, S441N, and F489L) were incubated with pheophorbide a at diŠerent concentrations (0, 0.63, 1.25, and 2.5 mM) at 379C for 4 hours. Login to comment
201 While the protein expression level of Q141K was lower than that of WT in Flp-In-293 cells (Fig. 1A), its level is considered to be su‹cient to extrude pheophorbide a from Flp-In-293/ABCG2(Q141K) cells. Login to comment

[hide] Association of breast cancer resistance protein/AB... Drug Metab Dispos. 2008 Apr;36(4):780-95. Epub 2008 Jan 7. Poonkuzhali B, Lamba J, Strom S, Sparreboom A, Thummel K, Watkins P, Schuetz E
Association of breast cancer resistance protein/ABCG2 phenotypes and novel promoter and intron 1 single nucleotide polymorphisms.
Drug Metab Dispos. 2008 Apr;36(4):780-95. Epub 2008 Jan 7., [PMID:18180275]

Abstract [show]
The hypothesis was tested that sequence diversity in breast cancer resistance protein (BCRP)'s cis-regulatory region is a significant determinant of BCRP expression. The BCRP promoter and intron 1 were resequenced in lymphoblast DNA from the polymorphism discovery resource (PDR) 44 subset. BCRP single nucleotide polymorphisms (SNPs) were genotyped in donor human livers, intestines, and lymphoblasts quantitatively phenotyped for BCRP mRNA expression. Carriers of the -15622C>T SNP had lower BCRP expression in multiple tissues. The intron 1 SNP 16702C>T was associated with high expression in livers; 1143G>A was associated with low expression in intestine; 12283T>C was associated with higher expression in the PDR44 and White livers. The -15994C>T promoter SNP was significantly associated with higher BCRP expression in multiple tissues. Patients with the -15994C>T genotype had substantially higher clearance of p.o. imatinib. We next determined whether BCRP expression was related to polymorphic alternative splicing or alternative promoter use. Liver polymorphically expressed an alternatively spliced mRNA [splice variant (SV) 1] skipping exon 2. Although SV1+ livers did not uniformly carry the exon 2 G34A allele, 90% of G34A livers expressed SV1 (versus 4% of 34GG livers). BCRP mRNA was significantly lower among Hispanic livers with the G34A variant genotype and may be due, in part, to polymorphic exon 2 splicing. Analysis of allele expression imbalance (AEI) showed that PDR44 samples with AEI had lower BCRP mRNA expression; however, no linked cis-polymorphisms were identified. BCRP used multiple promoters, and livers differentially using alternative exon 1b had lower BCRP. In conclusion, BCRP expression in lymphoblasts, liver, and intestine is associated with novel promoter and intron 1 SNPs.

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32 Among the coding single nucleotide polymorphisms (SNPs), G34A (V12M) in exon 2 and C421A (Q141K) in exon 5 occur in most racial groups but with a higher allele frequency in Asians and Hispanics. Login to comment

[hide] BCRP/ABCG2 in the placenta: expression, function a... Pharm Res. 2008 Jun;25(6):1244-55. Mao Q
BCRP/ABCG2 in the placenta: expression, function and regulation.
Pharm Res. 2008 Jun;25(6):1244-55., [PMID:18202831]

Abstract [show]
Knowledge concerning transport of maternally administered drugs across the placental barrier is essential for determining potential toxicity of drugs to the fetus and the value of drug therapy during pregnancy. An important determinant for fetal drug exposure is the expression of efflux transporters in the placenta. Among human tissues, the ATP-binding cassette efflux transporter BCRP (gene symbol ABCG2) is most abundantly expressed in the apical membrane of placental syncytiotrophoblasts. Although the precise physiological role of BCRP in the placenta is still unclear, existing data strongly suggest that BCRP plays an important role in protecting the fetus against the potential toxicity of drugs, xenobiotics, and metabolites by expelling them across the placental barrier. In this review, we summarize the current knowledge with respect to the expression, function, and polymorphisms of BCRP, as well as transcriptional and posttranscriptional regulation of the transporter in the placenta. Finally, clinical significance of BCRP in the placenta for drug therapy in pregnant women is discussed.

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138 Notably, the single nucleotide polymorphisms (SNPs) G34A and C421A, resulting in alterations of BCRP protein at position 12 (V12M) and 141 (Q141K), respectively, occur at a relatively high frequency in most ethnic populations. Login to comment
149 The study by Imai et al. (79) showed that the C421A polymorphism was associated with significantly lower expression and activity of the Q141K variant compared with the wild-type protein. Login to comment

[hide] Ubiquitin-mediated proteasomal degradation of non-... Biochem J. 2008 May 1;411(3):623-31. Nakagawa H, Tamura A, Wakabayashi K, Hoshijima K, Komada M, Yoshida T, Kometani S, Matsubara T, Mikuriya K, Ishikawa T
Ubiquitin-mediated proteasomal degradation of non-synonymous SNP variants of human ABC transporter ABCG2.
Biochem J. 2008 May 1;411(3):623-31., 2008-05-01 [PMID:18237272]

Abstract [show]
Clinical relevance is implicated between the genetic polymorphisms of the ABC (ATP-binding cassette) transporter ABCG2 (ABC subfamily G, member 2) and the individual differences in drug response. We expressed a total of seven non-synonymous SNP (single nucleotide polymorphism) variants in Flp-In-293 cells by using the Flp (flippase) recombinase system. Of these, ABCG2 F208S and S441N variants were found to be expressed at markedly low levels, whereas their mRNA levels were equal to those of the other SNP variants and ABCG2 WT (wild-type). Interestingly, protein expression levels of the ABCG2 F208S and S441N variants increased 6- to 12-fold when Flp-In-293 cells were treated with MG132, a proteasome inhibitor. Immunoprecipitation followed by immunoblot analysis showed that the ABCG2 F208S and S441N variant proteins were endogenously ubiquitinated in Flp-In-293 cells, and treatment with MG132 significantly enhanced the level of these ubiquitinated variants. Immunofluorescence microscopy demonstrated that MG132 greatly affected the ABCG2 F208S and S441N variants in terms of both protein levels and intracellular distribution. Immunoblot analysis revealed that those variants were N-glycosylated; however, their oligosaccharides were immature compared with those present on ABCG2 WT. The ABCG2 F208S and S441N variant proteins do not appear to be processed in the Golgi apparatus, but undergo ubiquitin-mediated protein degradation in proteasomes, whereas ABCG2 WT is sorted to the plasma membrane and then degraded via the lysosomal pathway. The present study provides the first evidence that certain genetic polymorphisms can affect the protein stability of ABCG2. Control of proteasomal degradation of ABCG2 would provide a novel approach in cancer chemotherapy to circumvent multidrug resistance of human cancers.

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208 In a previous study using the Flp recombinase system [33], we functionally characterized the non-synonymous polymorphisms (V12M, Q141K, F208S, S248P, F431L, S441N and F489L) in terms of their protein expression level, drug resistance profile and prazosin-stimulated ATPase activity. Login to comment

[hide] Homology modeling of breast cancer resistance prot... J Struct Biol. 2008 Apr;162(1):63-74. Epub 2007 Dec 15. Hazai E, Bikadi Z
Homology modeling of breast cancer resistance protein (ABCG2).
J Struct Biol. 2008 Apr;162(1):63-74. Epub 2007 Dec 15., [PMID:18249138]

Abstract [show]
BCRP (also known as ABCG2, MXR, and ABC-P) is a member of the ABC family that transports a wide variety of substrates. BCRP is known to play a key role as a xenobiotic transporter. Since discovering its role in multidrug resistance, considerable efforts have been made in order to gain deeper understanding of BCRP structure and function. The recent study was aimed at predicting BCRP structure by creating a homology model. Based on sequence similarity with known structures of full-length, NB and TM domain of ABC transporters, TM, NB, and linker regions of BCRP were defined. The NB domain of BCRP was modeled using MalK as a template. Based on secondary structure prediction of BCRP and comparison of the transmembrane connecting regions of known structures of ABC transporters, the TM domain arrangement of BCRP was established and was found to resemble to that of the recently published crystal structure of Sav1866. Thus, an initial alignment of TM domain of BCRP was established using Sav1866 as a template. This alignment was subsequently refined using constrains derived from secondary structure and TM predictions and the final model was built. Finally, the complete homodimer ABCG2 model was generated using Sav1866 as template. Furthermore, known ligands of BCRP were docked to our model in order to define possible binding sites. The results of molecular dockings of known BCRP substrates to the BCRP model were in agreement with recently published experimental data indicating multiple binding sites in BCRP.

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245 However, in our model, R482 cannot form interaction with rhodamine, but L484 is in interacting distance Table 3 Mutations on BCRP and their effect on its function Mutation Effect/results Reference V12M Did not effect Hemato and MTX transport Tamura et al. (2006) G51C Did not effect Hemato and MTX transport Tamura et al. (2006) K86M Inactivates transporter (dominant negative effect on ATPase activity); alters subcellular distribution Henriksen et al. (2005a) K86M Transporter inactive, but still able to bind ATP Ozvegy et al. (2002) Q126stop Defective porphyrin transport Tamura et al. (2006) Q141K Did not effect Hemato and MTX transport Tamura et al. (2006) T153M Did not effect Hemato and MTX transport Tamura et al. (2006) Q166E Did not effect Hemato and MTX transport Tamura et al. (2006) I206L Did not effect Hemato and MTX transport Tamura et al. (2006) F208S Defective porphyrin transport Tamura et al. (2006) S248P Defective porphyrin transport Tamura et al. (2006) E334stop Defective porphyrin transport Tamura et al. (2006) F431L Effects MTX transport Tamura et al. (2006) S441N Defective porphyrin transport Tamura et al. (2006) E446-mutants No drug resistance Miwa et al. (2003) R482G, R482T Effects MTX transport Tamura et al. (2006) R482T Substrate drug transport and inhibitor efficiency is not mediated by changes in drug-binding Pozza et al. (2006) R482G, R482T Substitution influence the substrate specificity of the transporter Ozvegy et al. (2002) R482G, R482T Altered substrate specificity Honjo et al. (2001) R482G Methotrexate not transported Chen et al. (2003b) Mitomo et al. (2003) R482G Resistance to hydrophilic antifolates in vitro, G482-ABCG2 mutation confers high-level resistance to various hydrophilic antifolates Shafran et al., (2005) R482G Three distinct drug, binding sites Clark et al. (2006) R482G Altered substrate specificity, granulocyte maturation uneffected Ujhelly et al. (2003) R482 mutants Higher resistance to mitoxantrone and doxorubicin than wt Miwa et al. (2003) R482X Affects substrate transport and ATP hydrolysis but not substrate binding Ejendal et al. (2006) F489L Impaired porphyrin transport Tamura et al. (2006) G553L; G553E Impaired trafficing, expression, and N-linked glycosylation Polgar et al. (2006) L554P Dominant negative effect on drug sensitivity Kage et al. (2002) N557D Resistance to MTX, but decreased transport of SN-38; N557E no change in transport compared to wt Miwa et al. (2003) F571I Did not effect Hemato and MTX transport Tamura et al. (2006) N590Y Did not effect Hemato and MTX transport Tamura et al. (2006) C592A Impaired function and expression Henriksen et al. (2005b) C592A/C608A Restored plasma mb expression; MTX transport normal, BODIPY-prazosin impaired Henriksen et al. (2005b) C603A Disulfide bridge; no functional or membrane targeting change Henriksen et al. (2005b) C608A Impaired function and expression Henriksen et al. (2005b) D620N Did not effect Hemato and MTX transport Tamura et al. (2006) H630X No change in transport Miwa et al. (2003) Cand N-terminal truncated Impaired trafficing Takada et al. (2005) with the ligand. Login to comment

[hide] Drug transporters: recent advances concerning BCRP... Br J Cancer. 2008 Mar 11;98(5):857-62. Epub 2008 Feb 5. Lemos C, Jansen G, Peters GJ
Drug transporters: recent advances concerning BCRP and tyrosine kinase inhibitors.
Br J Cancer. 2008 Mar 11;98(5):857-62. Epub 2008 Feb 5., 2008-03-11 [PMID:18253130]

Abstract [show]
Multidrug resistance is often associated with the (over)expression of drug efflux transporters of the ATP-binding cassette (ABC) protein family. This minireview discusses the role of one selected ABC-transporter family member, the breast cancer resistance protein (BCRP/ABCG2), in the (pre)clinical efficacy of novel experimental anticancer drugs, in particular tyrosine kinase inhibitors.

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45 Breedveld et al (2005) showed that Table 1 BCRP substrates and polymorphisms Substrates Classical anticancer drugs Novel targeted drugs Mitoxantrone Canertinib (CI-1033)a Anthracyclinesb Imatiniba Camptothecins Nilotiniba Antifolatesb Gefitiniba Erlotinib Flavopiridol Polymorphismsc Variant Amino-acid change Effect G34A Val12Met (V12M) No change C376T Gln126stop (Q126T) No active BCRP protein C421A Gln141Lys (Q141K) Decreased protein levels and drug resistance Increased gefitinib-associated toxicity (diarrhoea) Increased imatinib accumulation in vitro, but no changes in the pharmacokinetic parameters of imatinib in vivo G1322A Ser441Asn (S441N) Decreased protein levels and different subcellular localisation BCRP ¼ breast cancer resistance protein. Login to comment
103 A nonsynonymous SNP C421A resulting in a glutamine to lysine amino-acid change at position 141 (Q141K) has been associated with markedly decreased levels of BCRP protein expression and also low levels of drug resistance. Login to comment
104 Nonetheless, cross-resistance patterns were similar for the wild type and the Q141K BCRP variant, suggesting that this polymorphism does not affect substrate recognition of BCRP (Imai et al, 2002). Login to comment

[hide] Pharmacogenomic and pharmacokinetic determinants o... J Clin Oncol. 2008 Mar 1;26(7):1119-27. Rudin CM, Liu W, Desai A, Karrison T, Jiang X, Janisch L, Das S, Ramirez J, Poonkuzhali B, Schuetz E, Fackenthal DL, Chen P, Armstrong DK, Brahmer JR, Fleming GF, Vokes EE, Carducci MA, Ratain MJ
Pharmacogenomic and pharmacokinetic determinants of erlotinib toxicity.
J Clin Oncol. 2008 Mar 1;26(7):1119-27., 2008-03-01 [PMID:18309947]

Abstract [show]
PURPOSE: To assess the pharmacogenomic and pharmacokinetic determinants of skin rash and diarrhea, the two primary dose-limiting toxicities of the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor erlotinib. PATIENTS AND METHODS: A prospective clinical study of 80 patients with non-small-cell lung cancer, head and neck cancer, and ovarian cancer was performed. Detailed pharmacokinetics and toxicity of erlotinib were assessed. Polymorphic loci in EGFR, ABCG2, CYP3A4, and CYP3A5 were genotyped, and their effects on pharmacokinetics and toxicities were evaluated. RESULTS: A novel diplotype of two polymorphic loci in the ABCG2 promoter involving -15622C/T and 1143C/T was identified, with alleles conferring lower ABCG2 levels associated with higher erlotinib pharmacokinetic parameters, including area under the curve (P = .019) and maximum concentration (P = .006). Variability in skin rash was best explained by a multivariate logistic regression model incorporating the trough erlotinib plasma concentration (P = .034) and the EGFR intron 1 polymorphism (P = .044). Variability in diarrhea was associated with the two linked polymorphisms in the EGFR promoter (P < .01), but not with erlotinib concentration. CONCLUSION: Although exploratory in nature, this combined pharmacogenomic and pharmacokinetic model helps to define and differentiate the primary determinants of skin and gastrointestinal toxicity of erlotinib. The findings may be of use both in designing trials targeting a particular severity of rash and in considering dose and schedule modifications in patients experiencing dose-limiting toxicities of erlotinib or similarly targeted agents. Further studies of the relationship between germline polymorphisms in EGFR and the toxicity and efficacy of EGFR inhibitors are warranted.

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28 Recent studies suggest that gefitinib and erlotinib are substrates of ABCG2.32-35 Two nonsynonymous ABCG2 SNPs, 421 CϾA (Q141K, rs2231142) and 34GϾA (V12M, rs2231137), are common.36-39 The 141K polymorphism has been associated with lower expression and activity of ABCG2 and with higher accumulation of both gefitinib and erlotinib.35,36,40 A recent clinical study showed an association between 141K and diarrhea in patients treated with gefitinib.41 We have recently identified four functional polymorphisms in the 5Ј-regulatory region of ABCG2 (Poonkuzhali et al, manuscript submitted for publication). Login to comment

[hide] Drug-induced phototoxicity evoked by inhibition of... Expert Opin Drug Metab Toxicol. 2008 Mar;4(3):255-72. Tamura A, An R, Hagiya Y, Hoshijima K, Yoshida T, Mikuriya K, Ishikawa T
Drug-induced phototoxicity evoked by inhibition of human ABC transporter ABCG2: development of in vitro high-speed screening systems.
Expert Opin Drug Metab Toxicol. 2008 Mar;4(3):255-72., [PMID:18363541]

Abstract [show]
BACKGROUND: Photosensitivity depends on both genetic and environmental factors. Pheophorbide a, present in various plant-derived foods and food supplements, can be absorbed by the small intestine. Accumulation of pheophorbide a and porphyrins in the systemic blood circulation can result in phototoxic lesions on light-exposed skin. OBJECTIVE: As the human ATP-binding cassette (ABC) transporter ABCG2 has been suggested to be critically involved in porphyrin-mediated photosensitivity, we aimed to develop in vitro screening systems for drug-induced phototoxicity. CONCLUSION: Functional impairment owing to inhibition of ABCG2 by drugs or its genetic polymorphisms can lead to the disruption of porphyrin homeostasis. This review article provides an overview on drug-induced photosensitivity, as well as our hypothesis on a potential role of ABCG2 in phototoxicity.

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230 Plasma membrane Outside Inside ATP-binding cassette H2 N COOH V12M G51C Q126stop Q141K T153M R160Q Q166E I206L F208S S248P E334stop F431L F489L S441N R482G R482T F571I R575stop N590Y D620N T542A A528T D296H P269S A. Login to comment
231 0.0 0.1 0.2 0.3 0.4 0.5 Mock WT V12M G51C Q126stop Q141K T153M Q166E I206L F208S S248P E334stop F431L S441N F489L F571I N590Y D620N R482G R482T ATP-dependenthematoporphyrintransport (nmol/min/mgprotein) B. interactions should also take into consideration the presence of multiple flavonoids. Login to comment
245 Based on the presently available data on SNPs and acquired mutations, we have created a total of 18 variant forms of ABCG2 (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, F208S, S248P, E334stop, F431L, S441N, R482G, R482T, F489L, F571I, N590Y, and D620N) by site-directed mutagenesis and expressed them in insect cells. Login to comment
249 To further elucidate the significance of ABCG2 in cellular porphyrin homeostasis, we observed the cellular accumulation and compartmentation of porphyrin and pheophorbide a by means of a new fluorescence microscopy technology, and found that the accumulation of porphyrin and pheophorbide a in the cytoplasmic compartment was significantly higher in Flp-In-293 cells expressing S441N and F489L variants, as compared with those expressing WT, V12M, or Q141K [88]. Login to comment
252 Amino acid Porphyrin transport* Allele frequency (%)‡ cDNA position Location Wild-type allele Variant alllele V12M ++ 2.0 - 90.0 34 Exon 2 G A Q126stop - 0.0 - 1.7 376 Exon 4 C T Q141K ++ 0.0 - 35.5 421 Exon 5 C A T153M ++ 3.3 458 Exon 5 C T Q166E ++ N.D. 496 Exon 5 C G I206L ++ 10.0 616 Exon 6 A C F208S - N.D. 623 Exon 6 T C S248P - N.D. 742 Exon 7 T C E334stop - N.D. 1000 Exon 9 G T F431L ++ 0.8 1291 Exon 11 T C S441N - 0.5 1322 Exon 11 G A F489L + 0.5 - 0.8 1465 Exon 12 T C F571L ++ 0.5 1711 Exon 14 T A N590Y ++ 0.0 - 1.0 1768 Exon 15 A T D620N ++ 0.5 1858 Exon 16 G A *Transport of hematoporphyrin is indicated by either '+` (positive) or '-' (negative). Login to comment

[hide] Pharmacogenomics of drug-metabolizing enzymes and ... Methods Mol Biol. 2008;448:63-76. Bosch TM
Pharmacogenomics of drug-metabolizing enzymes and drug transporters in chemotherapy.
Methods Mol Biol. 2008;448:63-76., [PMID:18370231]

Abstract [show]
There is wide variability in the response of individuals to standard doses of drug therapy. This is an important problem in clinical practice, where it can lead to therapeutic failures or adverse drug events. Polymorphisms in genes coding for metabolizing enzymes and drug transporters can affect drug efficacy and toxicity. Pharmacogenomics aims to identify individuals predisposed to high risk of toxicity and low response from standard doses of anticancer drugs. This chapter focuses on the clinical significance of polymorphisms in drug-metabolizing enzymes and drug transporters in influencing efficacy and toxicity of anticancer therapy. The most important examples to demonstrate the influence of pharmacogenomics on anticancer therapy are thiopurine methyltransferase (TPMT), UGT (uridine diphosphate glucuronosyltransferase) 1A1*28, and DPD (dihydropyrimidine dehydrogenase) *2A, respectively, for 6-mercaptopurine, irinotecan, and 5-fluorouracil therapy. However, in most other anticancer therapies no clear association has been found for polymorphisms in drug-metabolizing enzymes and drug transporters and pharmacokinetics or pharmacodynamics of anticancer drugs. Evaluation of different regimens and tumor types showed that polymorphisms can have different, sometimes even contradictory, pharmacokinetic and pharmacodynamic effects in different tumors in response to different drugs. The clinical application of pharmacogenomics in cancer treatment therefore requires more detailed information regarding the different polymorphisms in drug-metabolizing enzymes and drug transporters. A greater understanding of complexities in pharmacogenomics is needed before individualized therapy can be applied on a routine basis.

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139 Mizuarai et al. (77) analyzed the effect of the polymorphisms G34A and C8825A, leading to an amino acid change of V12M and Q141K, respectively, on the transporter function of the protein. Login to comment
140 Drug resistance to indolocarbazole, a topoisomerase I inhibitor, of cells expressing V12M or Q141K was less than 1/10 compared to wild-type ABCG2-transfected cells and was accompanied by increased drug accumulation and decreased drug efflux in the variant ABCG2-expressing cells. Login to comment
143 In a study by Imai et al. (78), Japanese volunteers with the mutant C8825A polymorphism (allele frequency of 46%) expressed a low amount of the Q141K ABCG2 protein. Login to comment

[hide] ABCG2: structure, function and role in drug respon... Expert Opin Drug Metab Toxicol. 2008 Jan;4(1):1-15. Polgar O, Robey RW, Bates SE
ABCG2: structure, function and role in drug response.
Expert Opin Drug Metab Toxicol. 2008 Jan;4(1):1-15., [PMID:18370855]

Abstract [show]
ABCG2 was discovered in multi-drug-resistant cancer cells, with the identification of chemotherapeutic agents, such as mitoxantrone, flavopiridol, methotrexate and irinotecan as substrates. Later, drugs from other therapeutic groups were also described as substrates, including antibiotics, antivirals, HMG-CoA reductase inhibitors and flavonoids. An expanding list of compounds inhibiting ABCG2 has also been generated. The wide variety of drugs transported by ABCG2 and its normal tissue distribution with highest levels in the placenta, intestine and liver, suggest a role in protection against xenobiotics. ABCG2 also has an important role in the pharmacokinetics of its substrates. Single nucleotide polymorphisms of the gene were shown to alter either plasma concentrations of substrate drugs or levels of resistance against chemotherapeutic agents in cell lines. ABCG2 was also described as the determinant of the side population of stem cells. All these aspects of the transporter warrant further research aimed at understanding ABCG2's structure, function and regulation of expression.

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82 Some of these are in the coding region of ABCG2 and alter the protein sequence, such as V12M, Q141K, I206L and N590Y. Login to comment
83 Of these, Q141K, the substitution of glutamine with lysine at amino acid 141 in the NBD region is by far the most extensively studied. Login to comment
84 The Q141K SNP has an especially high incidence rate in the Asian population (~ 30%) and is also frequently found in Caucasians (~ 10%) [68]. Login to comment
86 Some investigators have also reported reduced surface expression for the Q141K SNP [71,72]. Login to comment
87 In the authors` hands, the Q141K mutant when transfected into human embryonic kidney (HEK) cells was clearly detected on the cell surface by flow cytometry with the 5D3 antibody and in immunofluorescence studied with the BXP-21 antibody, although some intracellular staining was also observed [69]. Login to comment
88 Recently, studies aimed at evaluating the clinical relevance of the Q141K SNP in patients undergoing chemotherapy and in healthy volunteers have also been reported: elevated plasma concentrations of gefitinib [73] and diflomotecan [74] and increased bioavailability of oral topotecan [75] were found. Login to comment
89 In one study a higher incidence of diarrhea in patients carrying the Q141K SNP was observed following treatment with gefitinib [76]. Login to comment
175 Membrane Out In 200 100 300 400 500 600 ATP site N-terminus C-terminus V12M Q141K I206L GXXXG, G406/G410 R482 G553 C603 N596 N590Y protein synthesis. Login to comment
344 Pharm Res 2004;21(10):1895-903 72. Imai Y, Nakane M, Kage K, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment
348 Cancer Biol Ther 2007;6(3):432-8 • Potential relevance of the Q141K SNP in the pharmacokinetics of ABCG2 substrates. Login to comment
350 Clin Pharmacol Ther 2004;76(1):38-44 • Potential relevance of the Q141K SNP in the pharmacokinetics of ABCG2 substrates. Login to comment
352 Cancer Biol Ther 2005;4(6):650-8 • Potential relevance of the Q141K SNP in the pharmacokinetics of ABCG2 substrates. Login to comment
355 J Natl Cancer Inst 2006;98(23):1739-42 • Potential relevance of the Q141K SNP in the pharmacokinetics of ABCG2 substrates. Login to comment

[hide] Breast cancer resistance protein (ABCG2) and drug ... Pharmacogenet Genomics. 2008 May;18(5):439-48. Urquhart BL, Ware JA, Tirona RG, Ho RH, Leake BF, Schwarz UI, Zaher H, Palandra J, Gregor JC, Dresser GK, Kim RB
Breast cancer resistance protein (ABCG2) and drug disposition: intestinal expression, polymorphisms and sulfasalazine as an in vivo probe.
Pharmacogenet Genomics. 2008 May;18(5):439-48., [PMID:18408567]

Abstract [show]
Breast cancer resistance protein (BCRP) is an efflux transporter expressed in tissues that act as barriers to drug entry. Given that single nucleotide polymorphisms (SNPs) in the ABCG2 gene encoding BCRP are common, the possibility exists that these genetic variants may be a determinant of interindividual variability in drug response. The objective of this study is to confirm the human BCRP-mediated transport of sulfasalazine in vitro, evaluate interindividual variation in BCRP expression in human intestine and to determine the role of ABCG2 SNPs to drug disposition in healthy patients using sulfasalazine as a novel in vivo probe. To evaluate these objectives, pinch biopsies were obtained from 18 patients undergoing esophagogastro-duodenoscopy or colonoscopy for determination of BCRP expression in relation to genotype. Wild-type and variant BCRP were expressed in a heterologous expression system to evaluate the effect of SNPs on cell-surface trafficking. A total of 17 healthy individuals participated in a clinical investigation to determine the effect of BCRP SNPs on sulfasalazine pharmacokinetics. In vitro, the cell surface protein expression of the common BCRP 421 C>A variant was reduced in comparison with the wild-type control. Intestinal biopsy samples revealed that BCRP protein and mRNA expression did not significantly differ between patients with 34GG/421CC versus patients with 34GG/421CA genotypes. Remarkably, in subjects with 34GG/421CA genotype, sulfasalazine area under the concentration-time curve was 2.4-fold greater compared with 34GG/421CC subjects (P<0.05). This study links commonly occurring SNPs in BCRP with significantly increased oral sulfasalazine plasma exposure in humans. Accordingly, sulfasalazine may prove to have utility as in vivo probe for assessing the clinical impact of BCRP for the disposition and efficacy of drugs.

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21 In addition, several nonsynonymous single nucleotide polymorphisms (SNPs) of the ABCG2 gene have been described including ABCG2 34G > A (V12M) located in exon 2 and 421C > A (Q141K) in exon 5. Login to comment
62 Site-directed mutagenesis was utilized to create the nonsynonymous allelic variants, 34G > A (V12M) and 421C > A (Q141K). Login to comment

[hide] The ABCG2 C421A polymorphism does not affect oral ... Br J Clin Pharmacol. 2008 Aug;66(2):233-9. Epub 2008 Apr 22. Adkison KK, Vaidya SS, Lee DY, Koo SH, Li L, Mehta AA, Gross AS, Polli JW, Lou Y, Lee EJ
The ABCG2 C421A polymorphism does not affect oral nitrofurantoin pharmacokinetics in healthy Chinese male subjects.
Br J Clin Pharmacol. 2008 Aug;66(2):233-9. Epub 2008 Apr 22., [PMID:18429968]

Abstract [show]
AIMS: A number of drugs are substrates or inhibitors of the efflux transporter breast cancer resistance protein (BCRP; ABCG2), which can limit systemic exposure by reducing absorption and/or increasing biliary elimination. The identification of a BCRP-selective clinical probe drug would provide a useful tool to understand the effect of genetic polymorphisms and transporter-based drug interactions on drug pharmacokinetics. The aim of this study was to assess the utility of nitrofurantoin as a clinical probe substrate for BCRP activity by evaluating the impact of genetic variation on nitrofurantoin pharmacokinetics. METHODS: Nitrofurantoin pharmacokinetics were studied in an open-label, single-oral dose (100 mg) study in 36 male Chinese subjects who were pre-screened for ABCG2 421 CC, CA and AA genotypes (n = 12 each). Plasma and urine concentrations of nitrofurantoin were determined by LC/MS/MS and LC/UV respectively. anova was used to compare pharmacokinetic parameters among genotypes. RESULTS: There were no significant differences in nitrofurantoin pharmacokinetics among the genotypic cohorts. The geometric mean nitrofurantoin plasma AUC((0-infinity)) (95% confidence interval) values were 2.21 (2.00, 2.45), 2.42 (2.11, 2.78) and 2.32 (1.99, 2.70) microg h ml(-1) and half-life values were 0.79 (0.59, 1.0), 0.76 (0.64, 0.89) and 0.72 (0.62, 0.84) h for ABCG2 421 genotypes CC, CA and AA, respectively. The percentage of dose excreted unchanged in the urine was 43, 44 and 39%, respectively. CONCLUSIONS: The ABCG2 C421A polymorphism had no effect on nitrofurantoin plasma and urine pharmacokinetic parameters in healthy Chinese subjects. These results indicate that nitrofurantoin is not a suitable clinical probe substrate for assessing BCRP activity.

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19 C421A, which results in a substitution of lysine for glutamine (Q141K) in the BCRP protein, is one of the most studied polymorphisms. Login to comment

[hide] Sequential topoisomerase targeting and analysis of... Anticancer Drugs. 2008 Apr;19(4):411-20. Saraiya B, Gounder M, Dutta J, Saleem A, Collazo C, Zimmerman L, Nazar A, Gharibo M, Schaar D, Lin Y, Shih W, Aisner J, Strair RK, Rubin EH
Sequential topoisomerase targeting and analysis of mechanisms of resistance to topotecan in patients with acute myelogenous leukemia.
Anticancer Drugs. 2008 Apr;19(4):411-20., [PMID:18454051]

Abstract [show]
Resistance to topoisomerase I (TOP1)-targeting drugs such as topotecan often involves upregulation of topoisomerase II (TOP2), with accompanying increased sensitivity to TOP2-targeting drugs such as etoposide. This trial was designed to investigate sequential topoisomerase targeting in the treatment of patients with high-risk acute myelogenous leukemia. An initial cohort of patients received topotecan and cytosine arabinoside daily for 5 days. Serial samples of circulating mononuclear cells were examined to evaluate peak elevations of TOP2-alpha protein expression. In subsequent cohorts, etoposide was administered daily for 3 days, beginning 6 h after initiation of the topotecan infusion. The etoposide dose was escalated to determine a maximum-tolerated dose. Circulating mononuclear cells were analyzed for TOP1 mutations and ABCG2 protein expression. In addition, systemic and intracellular topotecan concentrations were measured. Thirty-one patients were enrolled. On the basis of TOP1-alpha protein levels in three patients with peripheral blast counts greater than 50%, etoposide administration began 6 h after initiation of the topotecan/cytosine arabinoside infusion. Using this schedule of administration, the maximum-tolerated dose of etoposide was 90 mg/m. No TOP1 mutations were identified, but increases in ABCG2 expression during the infusion were observed in mononuclear cells from two of four evaluable patients. Administration of etoposide 6 h after initiation of a topotecan/cytosine arabinoside infusion is feasible and is associated with clinical activity. Analysis of TOP2-alpha protein levels in this small number of patients indicated that peak increases occurred earlier than expected based on earlier publications. Upregulation of ABCG2 was detected in circulating cells and may represent an inducible form of drug resistance that should be investigated further.

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91 For ABCG2, primers were designed to amplify regions containing the following four SNPs: G34A (V12M, exon 1), C376T (stop, exon 3), A616C (I206L, exon 5), C421A (Q141K, exon 12), and A1768T (N590Y, exon 14). Login to comment

[hide] Pharmacogenomics of MRP transporters (ABCC1-5) and... Drug Metab Rev. 2008;40(2):317-54. Gradhand U, Kim RB
Pharmacogenomics of MRP transporters (ABCC1-5) and BCRP (ABCG2).
Drug Metab Rev. 2008;40(2):317-54., [PMID:18464048]

Abstract [show]
Elucidation of the key mechanisms that confer interindividual differences in drug response remains an important focus of drug disposition and clinical pharmacology research. We now know both environmental and host genetic factors contribute to the apparent variability in drug efficacy or in some cases, toxicity. In addition to the widely studied and recognized genes involved in the metabolism of drugs in clinical use today, we now recognize that membrane-bound proteins, broadly referred to as transporters, may be equally as important to the disposition of a substrate drug, and that genetic variation in drug transporter genes may be a major contributor of the apparent intersubject variation in drug response, both in terms of attained plasma and tissue drug level at target sites of action. Of particular relevance to drug disposition are members of the ATP Binding Cassette (ABC) superfamily of efflux transporters. In this review a comprehensive assessment and annotation of recent findings in relation to genetic variation in the Multidrug Resistance Proteins 1-5 (ABCC1-5) and Breast Cancer Resistance Protein (ABCG2) are described, with particular emphasis on the impact of such transporter genetic variation to drug disposition or efficacy.

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250 It should be noted that many xeno- and endobiotic BCRP Figure 5 Predicted membrance topology of BCRP (ABCG2) based on hydrophobicity analysis. Locations of the non-synonymous polymorphisms are indicated with arrows. See Table 5 for allele frequencies and description of funtional consequences. NH2 COOH NBD Val12Met Gly51Cys Gln126* Ala149Pro Gln141Lys Thr153Met Arg160Gln Arg163Lys Gln166Glu Phe506Ser Phe507Leu Val508Leu Met509* Phe489Leu Ser441Asn Phe431Leu Glu334* Ile206Leu Ala315del Thr316del Phe208Ser Asp296His Ser248Pro Pro269Ser Phe571Ile Arg575* Asn590Tyr Asp620Asn in out Membrane BCRP (ABCG2) NBD Val12Met NBDNBD Val12Met substrates are also transported by other efflux transporters, especially P-glycoprotein, thus extrapolating BCRP related in vitro data to the in vivo situation may be difficult. Login to comment
258 One of the most extensively studied amino acid changes in BCRP is Gln141Lys located within the nucleotide binding domain of the transporter (Fig. 5). Login to comment
266 Further in vitro studies investigated the functional relevance of this amino acid substitution and found that Gln141Lys negatively affects the transport efficiency of ABCG2 for several substrates (Morisaki et al., 2005; Sparreboom et al., 2005) and, therefore, leads to a loss of drug resistance (Mizuarai et al., 2004; Tamura et al., 2007). Login to comment
267 In addition, a few in vivo studies also indicate a significant impact of Gln141Lys on drug pharmacokinetics. Login to comment
318 Available data in relation to BCRP pharmacogenetics suggest at least one of the known non-synonymous polymorphisms (421C>A/Gln141Lys) and possibly a few others (34G>A/Val12Met and 1322G>T/Ser441Asn) may be of functional and clinical importance. Login to comment

[hide] A dose-ranging study of the pharmacokinetics and p... Cancer Chemother Pharmacol. 2009 Feb;63(3):477-89. Epub 2008 May 29. O'Bryant CL, Lieu CH, Leong S, Boinpally R, Basche M, Gore L, Leonardi K, Schultz MK, Hariharan S, Chow L, Diab S, Gibbs A, Eckhardt SG
A dose-ranging study of the pharmacokinetics and pharmacodynamics of the selective apoptotic antineoplastic drug (SAAND), OSI-461, in patients with advanced cancer, in the fasted and fed state.
Cancer Chemother Pharmacol. 2009 Feb;63(3):477-89. Epub 2008 May 29., [PMID:18509645]

Abstract [show]
PURPOSE: To evaluate the safety, pharmacokinetics and determine the recommended dose of the selective apoptotic antineoplastic drug, OSI-461 administered on a twice-daily regimen to patients with advanced solid malignancies. METHODS: In this phase I trial, 33 patients were treated with OSI-461 doses ranging from 400 to 1,200 mg given twice daily in 4-week cycles. Pharmacokinetic studies were performed to characterize the plasma disposition of OSI-461 and the effect of food intake on OSI-461 absorption. Secondary biomarker studies were performed to assess the biologic activity of OSI-461 including the measurement of pGSK-3beta, a PKG substrate, and pharmacogenetic studies to identify polymorphisms of CYP3A that influence drug metabolism and of ABCG2, involved in drug resistance. RESULTS: Thirty-three patients were treated with 86 courses of OSI-461. The dose-limiting toxicities were grade 3 abdominal pain, found in one patient at the 1,000 mg BID fed dose level and all patients at the 1,200 mg BID fed dose level. There was also one episode each of grade 3 fatigue and grade 3 constipation at the 1,000 and 1,200 mg BID fed dose levels, respectively. Other common toxicities included mild to moderate fatigue, nausea, anorexia and mild elevation in bilirubin. Pharmacokinetic studies of OSI-461 revealed approximately a twofold increase in AUC(0-24) when OSI-461 was administered with food. An increase in pGSK-3beta post-dose was seen in the majority of patients and was greater at higher dose levels. No patients exhibited CYP3A4 polymorphisms, while 100% of patients were found to have the CYP3A5*3/CYP3A5*3 polymorphism. Two known polymorphisms of the ABCG2 gene, G34 --> A34 and C421 --> A421, occurred at frequencies of 11.76 and 29%, respectively. CONCLUSIONS: Toxicity and pharmacodynamic data show that the recommended oral dose of OSI-461 is 800 mg twice daily administered with food. The drug appears to be well-tolerated, and overall bioavailability appears to be markedly increased when the drug is administered with food. These results support further disease-directed evaluations of OSI-461 at a dose of 800 mg BID in combination with other chemotherapeutic agents.

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273 A mutation results in markedly decreased protein expression, and low level drug resistance, as compared to wild-type BRCP c For ABCG-2 #34 the of the A mutation frequency in Caucasians is 4.7%; and for ABCG-2 #421 the A mutation frequency in Caucasians is 25.9% Pt # ABCG-2 #34a ABCG-2 #421b Racec Val12 Met mutation Sequencing result Gln141 Lys mutation Sequencing result 01015 No Wild-type: G/G No Wild-type: C/C White 01016 No Wild-type: G/G Yes Heterozygote: C/A White 01017 Yes Homozygote: A/A No Wild-type: C/C Asian 01018 No Wild-type: G/G Yes Heterozygote: C/A White 01019 No Wild-type: G/G No Wild-type: C/C White 01020 Yes Heterozygote: A/G No Wild-type: C/C White 01021 No Wild-type: G/G Yes Heterozygote: C/A White 01022 No Wild-type: G/G No Wild-type: C/C Black 01023 No Wild-type: G/G Yes Heterozygote: C/A White 01024 No Wild-type: G/G No Wild-type: C/C White 01026 No Wild-type: G/G No Wild-type: C/C White 01027 No Wild-type: G/G No Wild-type: C/C White 01028 No Wild-type: G/G No Wild-type: C/C White 01029 No Wild-type: G/G No Wild-type: C/C White 01031 No Wild-type: G/G Yes Homozygote: A/A White 01032 No Wild-type: G/G No Wild-type: C/C White 01033 No Wild-type: G/G No Wild-type: C/C White A prior study of OSI-461 demonstrated that Cmax and AUC values increased in an approximate linear fashion as the dose was increased from 100 to 400 mg BID with a signiWcant amount of interpatient variability [35]. Login to comment

[hide] Intestinal efflux transporters and drug absorption... Expert Opin Drug Metab Toxicol. 2008 Jul;4(7):923-39. Murakami T, Takano M
Intestinal efflux transporters and drug absorption.
Expert Opin Drug Metab Toxicol. 2008 Jul;4(7):923-39., [PMID:18624680]

Abstract [show]
BACKGROUND: The intestinal epithelial membrane expresses ATP-binding cassette (ABC) transporters such as P-glycoprotein (P-gp), multi-drug resistance-associated proteins (MRPs) and breast cancer resistance protein (BCRP), in addition to various solute carrier (SLC) transporters. These ABC transporters affect the oral bioavailability of their substrate drugs. OBJECTIVE: To review the contribution of ABC efflux transporters such as P-gp, MRP2, MRP3, and BCRP in the intestinal absorption of substrate drugs. METHODS: Discussion was made by focusing on the site-specific expression and function of these ABC transporters, and the solubility and permeability of their substrate compounds. RESULTS/CONCLUSION: The increase in the solubility and permeability of orally administered drugs could be the key to escape barrier function of ABC transporters, especially P-gp.

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183 A common single polymorphism (SNP) in• ABCG2 (421C > A; Q141K) in two patients showed a 1.34-fold increased oral bioavailability of irinotecan (topotecan) compared to the bioavailability in ten patients with the wild type allele (42.0% versus 31.4%; p = 0.037) [77]. Login to comment

[hide] Human ABC transporters ABCG2 (BCRP) and ABCG4. Xenobiotica. 2008 Jul;38(7-8):863-88. Koshiba S, An R, Saito H, Wakabayashi K, Tamura A, Ishikawa T
Human ABC transporters ABCG2 (BCRP) and ABCG4.
Xenobiotica. 2008 Jul;38(7-8):863-88., [PMID:18668433]

Abstract [show]
1. The human ABC transporter ABCG2 is regarded as a member of the phase III system for xenobiotic metabolism, and it has been suggested that this efflux pump is responsible for protecting the body from toxic xenobiotics and for removing metabolites. 2. This review paper will address the new aspects of ABCG2 in terms of post-translational modifications (i.e., disulfide bond formation, ubiquitination, and endoplasmic reticulum-associated degradation) of ABCG2 protein, high-speed screening, and quantitative structure-activity relationship (QSAR) analysis to evaluate ABCG2-drug interactions, and genetic polymorphisms potentially associated with photosensitivity. 3. In addition, new aspects of human ABCG4 and mouse Abcg4 are presented with respect to their molecular properties and potential physiological roles. Considering a high sequence similarity between ABCG1 and ABCG4, both Abcg4 and ABCG4 may be involved in the transport of cholesterol from neurons and astrocytes. Furthermore, high expression of the mouse Abcg4 protein in the testis implicates its involvement in transport of certain sex hormones.

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225 Based on the currently available data on SNPs and acquired mutations, a total of 18 variant forms of ABCG2 (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, F208S, S248P, E334stop, F431L, S441N, R482G, R482T, F489L, F571I, N590Y, and D620N) were created by site-directed mutagenesis and expressed in Sf9 insect cells (Tamura et al. 2006, 2007). Login to comment

[hide] Pharmacogenetics of intestinal absorption. Curr Drug Deliv. 2008 Jul;5(3):153-69. Nakamura T, Yamamori M, Sakaeda T
Pharmacogenetics of intestinal absorption.
Curr Drug Deliv. 2008 Jul;5(3):153-69., [PMID:18673259]

Abstract [show]
The small intestine is the primary site of absorption for many drugs administered orally and so is the target tissue for pharmacotherapeutic strategies to control the oral absorption of drugs. Drug transporters, including the ATP-binding cassette (ABC) superfamily and the solute carrier (SLC) superfamily, have been considered to play a physiological role in regulating the absorption of xenobiotics, and variations in their expression level and function in the small intestine cause intra- and inter-individual variation in the oral absorption of drugs. Recent advances in molecular biology have suggested that genetic polymorphisms are associated with the expression level and function, and thereby inter-individual variation. In this review, the pharmacogenetics of these transporters is summarized, and their future significance in the clinical setting is discussed.

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76 Among the SNPs identified so far, two nonsynonymous SNPs, 34G>A and 421C>A, in the coding region, resulting in Val12Met and Gln141Lys, respectively, are relatively frequently identified and well-studied. Login to comment
77 Honjo et al. sequenced 90 genomic DNA samples and identified Val12Met and Gln141Lys at a frequency of 12.2% and 6.1%, respectively [86]. Login to comment
78 The Val12Met and Gln141Lys polymorphisms were also identified in a sequence analysis of ABCG2 cDNA clones isolated from 11 human tumor cell lines and were confirmed to have allelic frequencies of 19.0% and 26.6%, respectively, by a genomic DNA analysis in 124 Japanese [85]. Login to comment
81 Kondo and coworkers showed that the expression level of Gln141Lys ABCG2 was approximately 30-40% that of wild-type ABCG2 in membrane vesicles prepared from HEK293 cells infected with a recombinant adenovirus [94]. Login to comment
85 Exon Polymorphism Effect dbSNP Cell Expression Function Reference mRNA ( ) Protein (n.s.) Membrane localization (n.s.) Drug sensitivity (n.s.) Mitoxantrone efflux (n.s.) Hoechst 33342 efflux (n.s.) Morisaki et al. [92] HEK293 Protein (n.s.) Transport activity (n.s.) Kondo et al. [94] Protein (n.s.) ATPase activity (n.s.) Mizuarai et al. [88] Sf9 Protein ( ) ATPase activity (n.s.) Hoechst 33342 efflux ( ) Morisaki et al. [92] Exon 2 114T>C synonymous rs12721640 Exon 4 369C>T synonymous rs2231139 PA317 mRNA (n.s.) Protein ( ) Drug sensitivity ( ) Intracellular uptake ( ) Imai et al. [85] mRNA (n.s.) Protein (n.s.) Apical localization (n.s.) Drug sensitivity ( ) Indolocarbazole uptake ( ) Indolocarbazole efflux ( ) Mizuarai et al. [88] LLC-PK1 Apical localization (n.s.) Kondo et al. [94] mRNA ( ) Protein (n.s.) Membrane localization (impaired) Drug sensitivity ( ) Mitoxantrone efflux ( ) Hoechst 33342 efflux (n.s.) Morisaki et al. [92] HEK293 Protein ( ) Transport activity (n.s.) Kondo et al. [94] Protein (n.s.) ATPase activity ( ) Mizuarai et al. [88] 421C>A Gln141Lys rs2231142 Sf9 Protein (n.s.) ATPase activity ( ) Hoechst 33342 efflux (n.s.) Morisaki et al. [92] LLC-PK1 Apical localization (n.s.) Exon 5 496C>G Gln166Glu rs1061017 HEK293 Protein (n.s.) Transport activity (n.s.) Kondo et al. [94] 564A>G synonymous rs3116439 616A>C Ile206Leu rs12721643 HEK293 Protein ( or n.s.) Membrane localization (n.s.) Efflux activity ( ) Drug sensitivity ( ) ATPase activity (n.s.) Vethanayagam et al. [95] 617T>G Ile206Ser 617T>C Ile206Thr 617T>A Ile206Asn rs28365037 Exon 6 623T>C Phe208Ser rs1061018 Exon 7 742T>C Ser248Pro rs3116448 Exon 9 1000G>T Glu334stop rs3201997 Exon 14 2204T>A Phe571Ile rs9282571 SLC15A1 CHO Cephalexin uptake (n.s.)61G>A Val21Ile rs8187818 Cos7 Cephalexin uptake (n.s.) Substrate selectivity (n.s.) CHO Cephalexin uptake ( ) Cos7 Cephalexin uptake ( ) Substrate selectivity (VAC inhibition?) Login to comment
94 Exon Polymorphism Effect dbSNP Subject Expression Function Reference 114T>C synonymous rs12721640 369C>T synonymous rs2231139 421C>A Gln141Lys rs2231142 Patient (Caucasian) 9-nitrocamptotecin PK (CC CA) 9-aminocamptotecin PK [AUC/Dose] (CC<CA) Zamboni et al. [55] Nasopharyngeal cancer patient Irinotecan PK (CC CA+AA) SN-38 PK (CC CA+AA) SN-38G PK (CC CA+AA) Zhou et al. [56] HIV patient (Caucasian) Nelfinavir intracellular AUC (CC CA AA) Colombo et al. [58] Cancer patient Irinotecan PK (CC CA+AA) SN-38 PK (CC CA+AA) SN-38G PK (CC CA+AA) de Jong et al. [90] Patient (Japanese) Placental mRNA (CC CA AA) Placental protein (CC>CA>AA) Kobayashi et al. [91] Cancer patient Diflomotecan PK [AUC, Cmax] (CC<CA), [F] (CC>CA) Sparreboom et al. [96] Healthy (Chinese) Rosuvastatin PK [AUC, Cmax] (CC<CA+AA), [CL/F] (CC>CA+AA), [T1/2, Tmax] (CC CA+AA) Zhang et al. [97] Exon 4 496C>G Gln166Glu rs1061017 564A>G synonymous rs3116439 616A>C Ile206Leu rs12721643 617T>G Ile206Ser 617T>C Ile206Thr 617T>A Ile206Asn rs28365037 Exon 6 623T>C Phe208Ser rs1061018 Exon 7 742T>C Ser248Pro rs3116448 Exon 9 1000G>T Glu334Stop rs3201997 Exon 14 1711T>A Phe571Ile rs9282571 SLC15A1 61G>A Val21Ile rs8187818Exon 3 83T>A Phe28Tyr rs8187817 258G>A synonymous rs8187823 330C>T synonymous rs8187822 350G>A Ser117Asn rs2297322 351C>A Ser117Arg rs8187821 Exon 5 364G>A Val122Met rs8187820 Exon 7 501C>T synonymous rs3737087 Exon 11 843G>A synonymous r8187812 Exon 15 1147G>A Asp383Asn rs1782674 1179C>T synonymous rs8187836Exon 16 1256G>C Gly419Ara rs4646227 1347T>C synonymous rs1339067 Allelic mRNA imbalance (2030%) Anderle et al. [101] 1348G>A Val450Ile rs2274828 1352C>A Thr451Asn rs8187838 Exon 17 1375C>T Arg459Cys rs2274827 Exon 18 1446A>G synonymous rs8187828 (Table 3) contd…. Login to comment
97 PK: pharmacokinetics DNT: dysembryoplastic neuroepithelial tumors SN-38, SN-38G: irinotecan metabolites HIV: human Immunodeficiency Virus AUC: area under time-drug concentration curve Cmax: maximum drug concentration F: bioavailability CL/F: oral clearance T1/2: blood elimination half-life Tmax: time to peak concetration pendent groups, Mizuarai et al. [88] and Morisaki et al. [92], have shown comparable ABCG2 protein expression in wild-type and variant Gln141Lys transfectants in an immunostaining analysis using LLC-PK1 cells and immunoblot analysis using HEK-293 cells, respectively. Login to comment
99 In the report of Mizuarai et al., Val12Met but not Gln141Lys affected apical membrane localization of ABCG2 in transfectant- LLC-PK1 cells, whereas Morisaki et al. reported impaired membrane trafficking or incorrect membrane insertion of Gln141Lys ABCG2 in HEK-293 cells. Login to comment
100 Both groups also demonstrated that variant Gln141Lys ABCG2-expressing cells exhibited altered ATPase activity in Sf9 insect cells with ABCG2-bearing baculovirus and reduced drug resistance to ABCG2 substrates such as mitoxantrone, topotecan, and an indolocarbazole topoisomerase I inhibitor in LLC-PK1 cells or HEK293 cells [88, 92]. Login to comment

[hide] Pharmacogenomic importance of ABCG2. Pharmacogenomics. 2008 Aug;9(8):1005-9. Cusatis G, Sparreboom A
Pharmacogenomic importance of ABCG2.
Pharmacogenomics. 2008 Aug;9(8):1005-9., [PMID:18681776]

Abstract [show]
The ATP-binding cassette transporter ABCG2 (BCRP, MXR and ABCP) is highly expressed in the gastrointestinal tract and liver, and governs absorption, distribution and excretion of a wide variety of clinically important drugs. Common germline polymorphisms in the ABCG2 gene have been described that can affect expression, cellular localization and/or substrate recognition of the encoded protein. Alteration of transporter function by either of these mechanisms contributes significantly to interindividual variability in drug disposition and treatment outcome with certain, but not all, substrates for ABCG2.

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24 In particular, a SNP in exon 5 of the ABCG2 gene has been described, in which a 421C>A transversion results in a lysine to glutamine amino acid change at codon 141 (Q141K) [12]. Login to comment
33 Substrates • 9-aminocamptothecin, abacavir, bisantrene, BNP1350, chlorin E6, CI1033, ciprofloxacin, cladribine, daunorubicin, diflomotecan (BN-80915), dipyridamole, doxorubicin, DX-8951f, edevarone (metabolites), epirubicin, erlotinib, etoposide, flavopiridol, gefitinib, gimatecan, glyburide, GW1843, heterocyclic amines, homocamptothecin, imatinib, J-107088, JNJ-7706621, leflunomide, methotrexate, mitoxantrone, NB-506, nelfinavir, nitrofurantoin, norfloxacin, ofloxacin, olmesartan, pheophorbide A, pitavastatin, posuvastatin, protoporphyrin IX, pyropheophorbide A, SN-38 (irinotecan metabolite), sulfasalazine, teniposide, tomudex, topotecan, triglutamate, UCN-01, zidovudine Inhibitors • 17-β-estradiol, 6-prenlychrysin, abacavir, acacetin, amprenavir, atazanavir, biricodar (VX-710), chrysin, CI1033, curcumin, cyclosporin A, delavirdine, diethylstilbestrol, dihydropyridines, dipyridamole, efavirenz, elacridar (GF120918), estrone, fumitremorgin C (FTC), gefitinib, genistein, ginsenosides, imatinib, Ko143 (FTC analog), lopinavir, naringenin, nelfinavir, nicardapine, nimodipine, nitrendipine, novobiocin, ortataxel, pantoprazole, quercetin, reserpine, ritonavir, rosuvastatin, saquinavir, silymarin, sirolimus (rapamycin), tacrolimus, tamoxifen, tariquidar (XR9576), techochrysin, tryprostatin A, WK-X-34 demonstrated that subjects with a reduced ABCG2 activity owing to the Q141K variant are at an increased risk for gefitinib-induced diarrhea [17] (Figure 1), and altered pharmacokinetics of 9-aminocamptothecin [18], diflomotecan [19], irinotecan [20], rosuvastatin [21], sulfasalazine [22,23] and topotecan [24]. Login to comment

[hide] Association of the ABCG2 C421A polymorphism with p... BJU Int. 2008 Dec;102(11):1694-9. Epub 2008 Aug 14. Gardner ER, Ahlers CM, Shukla S, Sissung TM, Ockers SB, Price DK, Hamada A, Robey RW, Steinberg SM, Ambudkar SV, Dahut WL, Figg WD
Association of the ABCG2 C421A polymorphism with prostate cancer risk and survival.
BJU Int. 2008 Dec;102(11):1694-9. Epub 2008 Aug 14., [PMID:18710444]

Abstract [show]
OBJECTIVE: To determine if the C421A single nucleotide polymorphism (SNP) in the ATP-binding cassette transporter ABCG2 increases prostate cancer risk or affects survival. PATIENTS, SUBJECTS AND METHODS: Numerous studies have suggested that dietary, hormonal and environmental factors all play a role in the initiation in prostate cancer; among these, the carcinogenic heterocyclic amine 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), a known substrate of the ABCG2. A SNP of ABCG2, C421A, resulting in a glutamine to lysine change at amino acid 141, has been shown to result in decreased function of the protein. Due to the expression of ABCG2 in the prostate, together with the purported role of dietary carcinogens and steroids in the development and progression of prostate cancer, 311 individuals were genotyped for the ABCG2 C421A SNP, 170 patients with androgen-independent prostate cancer (AIPC) and 141 'healthy' controls. We also evaluated the effect of this SNP on the intracellular accumulation of PhIP and testosterone in vitro. RESULTS: There were no significant differences in the prevalence of prostate cancer based on ABCG2 genetic variation in this population. However, survival was significantly longer for individuals with wild-type ABCG2, as compared with those hetero- or homozygous for the C421A SNP (7.4 years vs 5.3 years, P = 0.044). Intracellular accumulation of PhIP was 80% higher in HEK293 cells transfected with Q141K ABCG2 than in wild-type cells, confirming that this SNP decreases transport of PhIP. In contrast, testosterone was not transported by either wild-type or variant transfected cells, nor did it act as in inhibitor of ABCG2 in subsequent transport assays. CONCLUSION: Increased exposure to PhIP may decrease survival, but the ABCG2 C421A polymorphism does not appear to increase the risk of prostate cancer.

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5 Intracellular accumulation of PhIP was 80% higher in HEK293 cells transfected with Q141K ABCG2 than in wild-type cells, confirming that this SNP decreases transport of PhIP. Login to comment
10 A SNP of ABCG2, C421A, resulting in a glutamine to lysine change at amino acid 141, has been shown to result in decreased function of the protein. Login to comment
43 CELL LINES Human embryonic kidney cells (HEK293) stably transfected with empty pcDNA3.1 vector (pcDNA3.1-HEK) or pcDNA3.1 vector containing full-length wild-type (482R-HEK) or Q141K (Q141K-HEK) ABCG2 have been previously characterized and were maintained as described [18,27]. Login to comment
46 [14 C]-PhIP AND [3 H]-TESTOSTERONE ACCUMULATION ASSAYS pcDNA3.1-HEK, R482-HEK and Q141K-HEK cells (2.5 × 105 cells/well) were grown in monolayer in a 24-well tissue culture plate at 37 °C in the presence of 5% CO2 in Dulbecco`s Modified Eagle`s Medium supplemented with 10% FBS. Login to comment
49 DETERMINATION OF CELL SURFACE EXPRESSION OF ABCG2 The cell surface expression of ABCG2 in pcDNA3.1-HEK, R482-HEK and Q141K-HEK cells was determined as described previously [29]. Login to comment
69 EFFECT OF C421A SNP ON IN VITRO PhIP TRANSPORT Comparison of the pcDNA3.1-HEK cells that do not express ABCG2 with the R482-HEK cells that express wild-type ABCG2 showed a three-fold decrease in intracellular accumulation of [14 C]-PhIP, in the absence of FTC (P < 0.001) as shown in Fig. 2a. The cells expressing the C421A variant form of ABCG2 (Q141K-HEK) had a reduced ability to efflux [14 C]-PHIP, resulting in a 1.8-fold increase in intracellular accumulation of this substrate compared with the wild-type cells (P < 0.001). Login to comment
71 Comparative ABCG2 expression levels were confirmed in the R482-HEK and Q141K-HEK cell lines, using flow cytometry as shown in Fig. 2b. Login to comment
72 EFFECT OF C421A SNP ON IN VITRO TESTOSTERONE TRANSPORT Transport assays using transfected R482-HEK and Q141K-HEK cells showed that testosterone was not transported by either TABLE 1 ComparisonofgenotypicfrequenciesofABCG2C421ASNPinpatientsdiagnosedwithAIPCand healthy volunteers (control). Login to comment
89 Green and red bars represent testosterone accumulation with and without the presence of FTC, respectively. pcDNA3.1-HEK cells do not express ABCG2. R482-HEK cells express wild-type ABCG2 and Q141K-HEK cells are homozygous variant for the C421A SNP. Each bar represents the mean accumulation (n = 3) and the error bars indicate the SD. 50 40 30 20 10 0 pcDNA3.1-HEK R482-HEK Q141K-HEK FIG. 4. Login to comment
94 C421ASNPresultsindecreasedintracellularPhIPaccumulation in vitro.Greenandredebarsrepresent PhIP accumulation with and without the presence of FTC, respectively. pcDNA3.1-HEK cells do not express ABCG2. R482-HEK cells express wild-type ABCG2 and Q141K-HEK cells are homozygous variant for the C421A SNP. Each bar represents the mean accumulation (n = 3) and the error bars indicate the SD. Login to comment
96 (a) (b) 1400 1200 1000 800 600 400 200 0 pcDNA3.1-HEK R482-HEK Q141K-HEK 14C-Phipaccumulation, nmoles/millioncells pcDNA3.1-HEK R482-HEK FL2-Height 0 30 60 90 120 150 0 30 60 90 120 150 0 30 60 90 120 150 2000 400 600800 1000 2000 400 600 800 1000 2000 400 600 800 1000 CountsCountsCounts FL2-Height FL2-Height Q141K-HEK accumulation of docetaxel, leading to improved response. Login to comment

[hide] Impact of genetic polymorphisms of transporters on... Drug Metab Pharmacokinet. 2008;23(4):223-35. Maeda K, Sugiyama Y
Impact of genetic polymorphisms of transporters on the pharmacokinetic, pharmacodynamic and toxicological properties of anionic drugs.
Drug Metab Pharmacokinet. 2008;23(4):223-35., [PMID:18762709]

Abstract [show]
As the importance of drug transporters in the clinical pharmacokinetics of drugs is recognized, genetic polymorphisms of drug transporters have emerged as one of the determinant factors to produce the inter-individual variability of pharmacokinetics. Many clinical studies have shown the influence of genetic polymorphisms of drug transporters on the pharmacokinetics and subsequent pharmacological and toxicological effects of drugs. The functional change in a transporter in clearance organs such as liver and kidney affects the drug concentration in the blood circulation, while that in the pharmacological or toxicological target can alter the local concentration at the target sites without changing its plasma concentration. As for the transporters for organic anions, some single nucleotide polymorphisms (SNPs) or haplotypes occurring with high frequency in organic anion transporting polypeptide (OATP) 1B1, multidrug resistance 1 (MDR1), and breast cancer resistance protein (BCRP) have been extensively investigated in both human clinical studies and in vitro functional assays. We introduce some examples showing the relationship between haplotypes in transporters and pharmacokinetics and pharmacological effects of drugs. We also discuss how to predict the effect of functional changes in drug transporters caused by genetic polymorphisms on the pharmacokinetics of drugs from in vitro data.

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107 The most important SNP in BCRP is C421A (Gln141Lys), which is located at the N-terminus domain (Fig. 5). Login to comment

[hide] Natural allelic variants of bovine ATP-binding cas... Drug Metab Dispos. 2009 Jan;37(1):5-9. Epub 2008 Sep 29. Merino G, Real R, Baro MF, Gonzalez-Lobato L, Prieto JG, Alvarez AI, Marques MM
Natural allelic variants of bovine ATP-binding cassette transporter ABCG2: increased activity of the Ser581 variant and development of tools for the discovery of new ABCG2 inhibitors.
Drug Metab Dispos. 2009 Jan;37(1):5-9. Epub 2008 Sep 29., [PMID:18824523]

Abstract [show]
ATP-binding cassette transporter ABCG2 [breast cancer resistance protein (BCRP)] is a member of the ABC transporter superfamily that actively extrudes xenotoxins from cells and is a major determinant of the bioavailability of many compounds. ABCG2 expression is strongly induced during lactation in the mammary gland and is related to the active secretion of drugs into the milk. The presence of drug residues and environmental pollutants in milk is an outstanding problem for human milk consumption and milk industrial processes, involving important risks to public health and the dairy industry. In cows, a single nucleotide polymorphism (SNP) in this protein has been described previously (Tyr581) and is associated with higher fat and protein percentages and lower milk yield. However, whether this amino acid substitution affects ABCG2-mediated drug transport in cows, including milk secretion, required further exploration. We cloned the two variants of bovine ABCG2 and evaluated the effect of this SNP on mitoxantrone accumulation assays performed in ovine primary fibroblasts transiently expressing either of the variants. It is interesting to note that statistically significant differences in activity between both variants were observed, and the Ser581 variant was related with an increased efflux activity. In addition, we demonstrated that genistein is a very good inhibitor of bovine ABCG2 and identified new inhibitors of the transporter, such as the macrocyclic lactones, ivermectin, and selamectin. Moreover, the inhibitory effect of these compounds on human and murine ABCG2 homologs was confirmed using transduced Marbin-Dabin canine kidney II cells. These findings may have important implications regarding the presence of drug residues in milk and drug interactions affecting the pharmacological behavior of ABCG2 substrates.

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18 For instance, the significance of the Q141K SNP has been shown in vitro (Mizuarai et al., 2004) and in clinical studies (Sparreboom et al., 2004; Yamasaki et al., 2008). Login to comment

[hide] Phase II study of imatinib in patients with recurr... J Clin Oncol. 2008 Oct 1;26(28):4659-65. Raymond E, Brandes AA, Dittrich C, Fumoleau P, Coudert B, Clement PM, Frenay M, Rampling R, Stupp R, Kros JM, Heinrich MC, Gorlia T, Lacombe D, van den Bent MJ
Phase II study of imatinib in patients with recurrent gliomas of various histologies: a European Organisation for Research and Treatment of Cancer Brain Tumor Group Study.
J Clin Oncol. 2008 Oct 1;26(28):4659-65., 2008-10-01 [PMID:18824712]

Abstract [show]
PURPOSE: To evaluate the safety and the efficacy of imatinib in recurrent malignant gliomas. PATIENTS AND METHODS: This was a single-arm, phase II study. Eligible patients had recurrent glioma after prior radiotherapy with an enhancing lesion on magnetic resonance imaging. Three different histologic groups were studied: glioblastomas (GBM), pure/mixed (anaplastic) oligodendrogliomas (OD), and low-grade or anaplastic astrocytomas (A). Imatinib was started at a dose of 600 mg/d with dose escalation to 800 mg in case of no toxicity; during the trial this dose was increased to 800 mg/d with escalation to 1,000 mg/d. Trial design was one-stage Fleming; both an objective response and 6 months of progression-free survival (PFS) were considered a successful outcome to treatment. RESULTS: A total of 112 patients (51 patients with GBM, 25 patients with A, and 36 patients with OD) were enrolled. Imatinib was in general well tolerated. The median number of cycles was 2.0 (range, 1 to 43 cycles). Five patients had an objective partial response, including three patients with GBM; all had 6 months of PFS. The 6-month PFS rate was 16% (95% CI, 8.0% to 34.0%) in GBM, 4.0% (95% CI, 0.3% to 15.0%) in OD, and 9% (95% CI, 2.0% to 25.0%) in A. The exposure to imatinib was significantly lower in patients using enzyme-inducing antiepileptic drugs. The presence of ABCG2 point mutations were not correlated with pharmacokinetic findings. No somatic activating mutations of KIT or platelet-derived growth factor receptor-A or -B were found. CONCLUSION: In the dose range of 600 to 1,000 mg/d, single-agent imatinib is well tolerated but has limited antitumor activity in patients with recurrent gliomas.

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61 Raymond et al 4660 17) as previously described.17 Genotyping for the hypomorphic Q141K ABCG2 polymorphism was performed using tumor DNA and a Fluorescent Resonance Energy Transfer-based hybridization probes and melting curve analysis using a Roche LC480 instrument (Roche, Neuilly sur Seine, France; primer, probe, and polymerase chain reaction conditions available on request).18 Statistical Analysis Demographic, response rate, safety, laboratory, PK, and pharmacodynamic data were analyzed using descriptive statistics. Login to comment
100 Twelve patients had the Q141K exon 5 SNP. Login to comment

[hide] Lapatinib (Tykerb, GW572016) reverses multidrug re... Cancer Res. 2008 Oct 1;68(19):7905-14. Dai CL, Tiwari AK, Wu CP, Su XD, Wang SR, Liu DG, Ashby CR Jr, Huang Y, Robey RW, Liang YJ, Chen LM, Shi CJ, Ambudkar SV, Chen ZS, Fu LW
Lapatinib (Tykerb, GW572016) reverses multidrug resistance in cancer cells by inhibiting the activity of ATP-binding cassette subfamily B member 1 and G member 2.
Cancer Res. 2008 Oct 1;68(19):7905-14., 2008-10-01 [PMID:18829547]

Abstract [show]
Lapatinib is active at the ATP-binding site of tyrosine kinases that are associated with the human epidermal growth factor receptor (Her-1 or ErbB1) and Her-2. It is conceivable that lapatinib may inhibit the function of ATP-binding cassette (ABC) transporters by binding to their ATP-binding sites. The aim of this study was to investigate the ability of lapatinib to reverse tumor multidrug resistance (MDR) due to overexpression of ABC subfamily B member 1 (ABCB1) and ABC subfamily G member 2 (ABCG2) transporters. Our results showed that lapatinib significantly enhanced the sensitivity to ABCB1 or ABCG2 substrates in cells expressing these transporters, although a small synergetic effect was observed in combining lapatinib and conventional chemotherapeutic agents in parental sensitive MCF-7 or S1 cells. Lapatinib alone, however, did not significantly alter the sensitivity of non-ABCB1 or non-ABCG2 substrates in sensitive and resistant cells. Additionally, lapatinib significantly increased the accumulation of doxorubicin or mitoxantrone in ABCB1- or ABCG2-overexpressing cells and inhibited the transport of methotrexate and E(2)17betaG by ABCG2. Furthermore, lapatinib stimulated the ATPase activity of both ABCB1 and ABCG2 and inhibited the photolabeling of ABCB1 or ABCG2 with [(125)I]iodoarylazidoprazosin in a concentration-dependent manner. However, lapatinib did not affect the expression of these transporters at mRNA or protein levels. Importantly, lapatinib also strongly enhanced the effect of paclitaxel on the inhibition of growth of the ABCB1-overexpressing KBv200 cell xenografts in nude mice. Overall, we conclude that lapatinib reverses ABCB1- and ABCG2-mediated MDR by directly inhibiting their transport function. These findings may be useful for cancer combinational therapy with lapatinib in the clinic.

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307 Recently, Cusatis and colleagues (45) reported that one common functional single-nucleotide polymorphism in the ABCG2 gene, ABCG2 421C!A (Q141K), is associated with diarrhea, a gefitinib-induced adverse effect, and led to a high risk of diarrhea in patients treated with oral gefitinib. Login to comment

[hide] Association of three genetic loci with uric acid c... Lancet. 2008 Dec 6;372(9654):1953-61. Epub 2008 Oct 1. Dehghan A, Kottgen A, Yang Q, Hwang SJ, Kao WL, Rivadeneira F, Boerwinkle E, Levy D, Hofman A, Astor BC, Benjamin EJ, van Duijn CM, Witteman JC, Coresh J, Fox CS
Association of three genetic loci with uric acid concentration and risk of gout: a genome-wide association study.
Lancet. 2008 Dec 6;372(9654):1953-61. Epub 2008 Oct 1., 2008-12-06 [PMID:18834626]

Abstract [show]
BACKGROUND: Hyperuricaemia, a highly heritable trait, is a key risk factor for gout. We aimed to identify novel genes associated with serum uric acid concentration and gout. METHODS: Genome-wide association studies were done for serum uric acid in 7699 participants in the Framingham cohort and in 4148 participants in the Rotterdam cohort. Genome-wide significant single nucleotide polymorphisms (SNPs) were replicated in white (n=11 024) and black (n=3843) individuals who took part in the study of Atherosclerosis Risk in Communities (ARIC). The SNPs that reached genome-wide significant association with uric acid in either the Framingham cohort (p<5.0 x 10(-8)) or the Rotterdam cohort (p<1.0 x 10(-7)) were evaluated with gout. The results obtained in white participants were combined using meta-analysis. FINDINGS: Three loci in the Framingham cohort and two in the Rotterdam cohort showed genome-wide association with uric acid. Top SNPs in each locus were: missense rs16890979 in SLC2A9 (p=7.0 x 10(-168) and 2.9 x 10(-18) for white and black participants, respectively); missense rs2231142 in ABCG2 (p=2.5 x 10(-60) and 9.8 x 10(-4)), and rs1165205 in SLC17A3 (p=3.3 x 10(-26) and 0.33). All SNPs were direction-consistent with gout in white participants: rs16890979 (OR 0.59 per T allele, 95% CI 0.52-0.68, p=7.0 x 10(-14)), rs2231142 (1.74, 1.51-1.99, p=3.3 x 10(-15)), and rs1165205 (0.85, 0.77-0.94, p=0.002). In black participants of the ARIC study, rs2231142 was direction-consistent with gout (1.71, 1.06-2.77, p=0.028). An additive genetic risk score of high-risk alleles at the three loci showed graded associations with uric acid (272-351 mumol/L in the Framingham cohort, 269-386 mumol/L in the Rotterdam cohort, and 303-426 mumol/L in white participants of the ARIC study) and gout (frequency 2-13% in the Framingham cohort, 2-8% in the Rotterdam cohort, and 1-18% in white participants in the ARIC study). INTERPRETATION: We identified three genetic loci associated with uric acid concentration and gout. A score based on genes with a putative role in renal urate handling showed a substantial risk for gout.

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71 **Chromosome 4 (89271347) in ABCG2; alleles= G/T (Q141K). Login to comment
154 A missense SNP in ABCG2 (rs2231142; Q141K) was associated with uric acid concentration and gout in both white and black individuals and may be a causal candidate variant. Login to comment
165 This SNP in exon 5 leads to a glutamine-to-lysine aminoacid substitution (Q141K). Login to comment
168 Three other SNPs located downstream of and in disequilibrium with the Q141K variant were associated with uric acid, two of which are within the polycystic kidney disease 2 gene (PKD2). Login to comment
169 However, neither these nor other SNPs in the region were independently associated with uric acid when adjusted for the Q141K variant in either the Framingham or the Rotterdam cohort. Login to comment
170 This evidence together with the weak linkage disequilibrium pattern in the ABCG2 region in the HapMap Yoruban sample and thesignificantassociationofrs2231142inblackparticipants in the ARIC study, despite the low minor allele frequency of 3%, suggests that the ABCG2 Q141K variant (rs2231142) could be causally related to uric acid concentration. Login to comment

[hide] Clinical pharmacogenetics and potential applicatio... Curr Drug Metab. 2008 Oct;9(8):738-84. Zhou SF, Di YM, Chan E, Du YM, Chow VD, Xue CC, Lai X, Wang JC, Li CG, Tian M, Duan W
Clinical pharmacogenetics and potential application in personalized medicine.
Curr Drug Metab. 2008 Oct;9(8):738-84., [PMID:18855611]

Abstract [show]
The current 'fixed-dosage strategy' approach to medicine, means there is much inter-individual variation in drug response. Pharmacogenetics is the study of how inter-individual variations in the DNA sequence of specific genes affect drug responses. This article will highlight current pharmacogenetic knowledge on important drug metabolizing enzymes, drug transporters and drug targets to understand interindividual variability in drug clearance and responses in clinical practice and potential use in personalized medicine. Polymorphisms in the cytochrome P450 (CYP) family may have had the most impact on the fate of pharmaceutical drugs. CYP2D6, CYP2C19 and CYP2C9 gene polymorphisms and gene duplications account for the most frequent variations in phase I metabolism of drugs since nearly 80% of drugs in use today are metabolised by these enzymes. Approximately 5% of Europeans and 1% of Asians lack CYP2D6 activity, and these individuals are known as poor metabolizers. CYP2C9 is another clinically significant drug metabolising enzyme that demonstrates genetic variants. Studies into CYP2C9 polymorphism have highlighted the importance of the CYP2C9*2 and CYP2C9*3 alleles. Extensive polymorphism also occurs in a majority of Phase II drug metabolizing enzymes. One of the most important polymorphisms is thiopurine S-methyl transferases (TPMT) that catalyzes the S-methylation of thiopurine drugs. With respect to drug transport polymorphism, the most extensively studied drug transporter is P-glycoprotein (P-gp/MDR1), but the current data on the clinical impact is limited. Polymorphisms in drug transporters may change drug's distribution, excretion and response. Recent advances in molecular research have revealed many of the genes that encode drug targets demonstrate genetic polymorphism. These variations, in many cases, have altered the targets sensitivity to the specific drug molecule and thus have a profound effect on drug efficacy and toxicity. For example, the beta (2)-adrenoreceptor, which is encoded by the ADRB2 gene, illustrates a clinically significant genetic variation in drug targets. The variable number tandem repeat polymorphisms in serotonin transporter (SERT/SLC6A4) gene are associated with response to antidepressants. The distribution of the common variant alleles of genes that encode drug metabolizing enzymes, drug transporters and drug targets has been found to vary among different populations. The promise of pharmacogenetics lies in its potential to identify the right drug at the right dose for the right individual. Drugs with a narrow therapeutic index are thought to benefit more from pharmacogenetic studies. For example, warfarin serves as a good practical example of how pharmacogenetics can be utilized prior to commencement of therapy in order to achieve maximum efficacy and minimum toxicity. As such, pharmacogenetics has the potential to achieve optimal quality use of medicines, and to improve the efficacy and safety of both prospective and licensed drugs.

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618 Only a small portion of them are non-synonymous (V12M, Q141K, Q166E, I206L, F208S, S248P, D296H, L525R, A528T, F571I, and Y590N) and there is one frameshift (1515delC) mutation observed in the coding region of ABCG2. Login to comment
619 Among the above variations, 34G>A (V12M) AND 421C>A (Q141K) have been testified to be polymorphic in numerous populations [267, 271]. Login to comment
624 The Q141K polymorphism located in exon 5 leads to the replacement of the negatively charged glutamic acid residue with a positively charged lysine residue. Login to comment
626 The Q141K variant was detected in all ethnic groups tested; it was found that Q141K occurs frequently in Asian populations ranging from (~30-60%) and relatively low allele frequency in Caucasians and African American subjects (~5-10%) [273]. Login to comment
627 For example, 60% are heterozygous for Q141 K in a Chinese population; 39-50% heterozygous for a Japanese population and 7% homozygous for the variant Q141K. Login to comment
630 Studies have discovered that the expression levels of Q141K ABCG2 protein is lower than the wild-type, or the V12M variant when expressed in PA317 or HEK-293 cells [273]. Login to comment
631 It was also found that a portion of Q141K remained intracellular despite having a low level of expression, as both V12M and Q141K BCRP could reach the plasma membrane in the HEK-293 cells [273]. Login to comment
632 Other reports quoted that there is a 30-40% reduction in expression of cell surface Q141K variant, despite having a similar mRNA level compared to the wild-type. Login to comment
633 Individuals homozygous for the Q141K variant had significantly lower expression levels of BCRP in the placenta while the heterozygous samples displayed an intermediate expression level [274]. Login to comment
635 ABCG2 is expressed in polarized LLC-PKI cells, and a study has demonstrated that the V12M variant has an intracellular localization whereas the wild-type ABCG2 and Q141K show mainly apical staining [275]. Login to comment
637 All polymorphisms, including V12M and Q141K, had an apical localization, and only the S441N variant displayed an intracellular staining [275]. Login to comment
639 Further studies are required to clarify the mechanism of a reduced protein expression for Q141K, and the change of cellular localization for the V12M and Q141K variants found under specific conditions. Login to comment
641 There was a 10-fold decrease in drug resistance compared with the wild-type ABCG2 when the V12M or Q141K-transfected LLC-PKI cells were challenged by mitoxantrone or topotecan [275]. Login to comment
642 In contrast, when compared to wild-type ABCG2-transfected cells, Q141K variant alone had a fairly lower level resistance against mitoxantrone, topotecan or SN-38. Login to comment
644 In the ATP-binding cassette region of ABCG2, Q141K is between the Walker A and the signature motifs; hence variant ATPase activity alteration is possible. Login to comment
645 Experiments were undertaken to observe between the vanadate-sensitive ATPase activity of ABCG2 V12M and Q141K variants, using Sf9 (Spodoptera frugiperda) cell membranes [276]. Login to comment
646 There was a 1.3 and 1.8-fold lower basal ATPase activity, respectively, for V12M and Q141K compared to wild-type [276]. Login to comment

[hide] Major SNP (Q141K) variant of human ABC transporter... Pharm Res. 2009 Feb;26(2):469-79. Epub 2008 Oct 29. Furukawa T, Wakabayashi K, Tamura A, Nakagawa H, Morishima Y, Osawa Y, Ishikawa T
Major SNP (Q141K) variant of human ABC transporter ABCG2 undergoes lysosomal and proteasomal degradations.
Pharm Res. 2009 Feb;26(2):469-79. Epub 2008 Oct 29., [PMID:18958403]

Abstract [show]
PURPOSE: Single nucleotide polymorphisms (SNPs) of the ATP-binding cassette (ABC) transporter ABCG2 gene have been suggested to be a significant factor in patients' responses to medication and/or the risk of diseases. We aimed to evaluate the impact of the major non-synonymous SNP Q141K on lysosomal and proteasomal degradations. METHODS: ABCG2 WT and the Q141K variant were expressed in Flp-In-293 cells by using the Flp recombinase system. Their expression levels and cellular localization was measured by immunoblotting and immunofluorescence microscopy, respectively. RESULTS: The protein level of the Q141K variant expressed in Flp-In-293 cells was about half that of ABCG2 WT, while their mRNA levels were equal. The protein expression level of the Q141K variant increased about two-fold when Flp-In-293 cells were treated with MG132. In contrast, the protein level of ABCG2 WT was little affected by the same treatment. After treatment with bafilomycin A1, the protein levels of ABCG2 WT and Q141K increased 5- and 2-fold in Flp-In-293 cells, respectively. CONCLUSIONS: The results strongly suggest that the major non-synonymous SNP Q141K affects the stability of the ABCG2 protein in the endoplasmic reticulum and enhances its susceptibility to ubiquitin-mediated proteasomal degradation.

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0 Research Paper Major SNP (Q141K) Variant of Human ABC Transporter ABCG2 Undergoes Lysosomal and Proteasomal Degradations Tomoka Furukawa,1 Kanako Wakabayashi,1,2 Ai Tamura,1,2 Hiroshi Nakagawa,1 Yoshihiro Morishima,3 Yoichi Osawa,3 and Toshihisa Ishikawa1,4 Received July 30, 2008; accepted October 8, 2008; published online October 29, 2008 Purpose. Login to comment
2 We aimed to evaluate the impact of the major non-synonymous SNP Q141K on lysosomal and proteasomal degradations. Login to comment
4 ABCG2 WT and the Q141K variant were expressed in Flp-In-293 cells by using the Flp recombinase system. Login to comment
7 The protein level of the Q141K variant expressed in Flp-In-293 cells was about half that of ABCG2 WT, while their mRNA levels were equal. Login to comment
8 The protein expression level of the Q141K variant increased about two-fold when Flp-In-293 cells were treated with MG132. Login to comment
10 After treatment with bafilomycin A1, the protein levels of ABCG2 WT and Q141K increased 5-and 2-fold in Flp-In-293 cells, respectively. Login to comment
12 The results strongly suggest that the major non-synonymous SNP Q141K affects the stability of the ABCG2 protein in the endoplasmic reticulum and enhances its susceptibility to ubiquitin-mediated proteasomal degradation. Login to comment
20 The most extensively studied of those SNPs with potential clinical relevance is 421 C>A, which results in a glutamine to lysine substitution (Q141K) in the ABCG2 protein. Login to comment
21 The Q141K SNP was identified with varying frequencies in different ethnic groups and was found to be the most prevalent in Japanese and Chinese populations (approximately 30% in allele frequency) (2). Login to comment
22 Q141K has been associated with lower levels of protein expression and impaired transport in vitro; however, some controversies exist in the publications characterizing this SNP (12-18). Login to comment
30 Furthermore, the Q141K SNP was reportedly associated with a higher incidence of diarrhea in non-small cell lung cancer patients treated with gefitinib (22). Login to comment
36 In this context, it has been hypothesized that the reduced expression levels of the Q141K variant may be due to the ubiquitin-mediated proteasomal degradation. Login to comment
43 We here provide evidence that the major nonsynonymous SNP variant Q141K undergoes both ubiquitin-mediated protein degradation in proteasomes and lysosomal proteolysis and that the protein level of the Q141K variant is thereby significantly reduced. Login to comment
47 Preparation of Plasmids Carrying ABCG2 Q141K Variant cDNA. Login to comment
48 Wild-type (WT) ABCG2 cDNA inserted into the pcDNA5/FRT plasmid was used as the template, and the nonsynonymous SNP Q141K variant was generated by using the QuikChange® Site-Directed Mutagenesis Kit (Stratagene, La Jolla, CA, USA). Login to comment
53 Expression of ABCG2 WTand Q141K Variant in Flp-In-293 Cells. Login to comment
59 Flp-In-293 cells expressing ABCG2 WT or Q141K, named Flp-In-293/ABCG2 (WT) or Flp-In-293/ABCG2 (Q141K), were maintained in high-glucose Dulbecco`s modified Eagle`s medium (D-MEM) supplemented with 10% (v/v) heat-inactivated fetal calf serum (ICN Biomedicals, Inc., Aurora, OH, USA), 2 mM L-glutamine, 100 U/ml penicillin, 100 μg/ml streptomycin, 250 ng/ml amphotericin B, and Fig. 1. Schematic illustration of the protein structure of human ABCG2. Login to comment
60 The sites of three non-synonymous SNPs, Q141K, F208S and S441N, are indicated. Login to comment
98 RESULTS AND DISCUSSION Protein Expression Levels of ABCG2 WT and Q141K in Flp-In-293 Cells. Login to comment
101 The Q141K variant is reportedly associated with lower levels of protein expression; however, little is known about the underlying molecular mechanism. Login to comment
102 In the present study, ABCG2 WT and the Q141K variant were individually expressed in Flp-In-293 cells by using the Flp recombinase system, where one single copy of the cDNA encoding either the WT or the variant was integrated into the genomic DNA at the designated FRT site prepared in chromosome 12 (Fig. 2A). Login to comment
103 As shown in Fig. 2B, mRNA levels of ABCG2 WT as well as Q141K were evenly represented in Flp-In-293 cells, where the mRNA levels of ABCG2 and GAPDH were measured by RT-PCR. Login to comment
104 Both WT and Q141K proteins were found as glycosylated homodimers. Login to comment
105 To quantify the levels of ABCG2 WT and Q141K proteins, we treated the samples with PNGase F and mercaptoethanol (ME) to remove the glycomoieties and to break the cysteinyl disulfide bond forming the homodimers (Fig. 2B). Login to comment
107 As demonstrated in Fig. 2B, the protein level of the Q141K variant was about 45% of the ABCG2 WT protein level. Login to comment
109 Flp-mediated integration of the ABCG2 cDNA into FRT-tagged genomic DNA (A), the mRNA and protein levels of ABCG2 WT and Q141K (B) and the relationship between the relative intensity of ABCG2 immunoreactivity and the amount of cellular proteins (C). Login to comment
113 B The mRNA level was analyzed by RT-PCR with total RNA extracted from Flp-In-293 cells expressing ABCG2 WT or Q141K. Login to comment
115 The level of the Q141K variant protein was normalized to the WT level. Login to comment
120 Effect of MG132 (A) or bafilomycin A1 (B) on the protein levels of ABCG2 WT and Q141K variant. Login to comment
121 A Flp-In-293 cells expressing ABCG2 WT or Q141K were incubated with 2 μ;M MG132 for 0 and 24 h. ABCG2 protein levels were analyzed by immunoblotting after PNGase F treatment. Login to comment
122 B Flp-In-293 cells expressing ABCG2 WT or Q141K were incubated with 10 nM bafilomycin A1 for 0, 12, and 24 h. ABCG2 protein levels were analyzed as described above. Data are expressed as mean values ± SD in triplicate experiments. Login to comment
124 Effect of MG132 and Bafilomycin A1 on the Protein Expression Levels of ABCG2 WT and Q141K Variant. Login to comment
126 Flp-In-293 cells expressing WT or Q141K were incubated in the presence of MG132 at a concentration of 2.0 μM for 24 h, and then cell lysate samples were immediately prepared. Login to comment
129 Protein expression levels of the WT and the Q141K variant were determined by immunoblotting after PNGase F treatment in the same way as described above. Login to comment
130 As shown in Fig. 3A, the protein level of the Q141K variant was approximately two-fold enhanced by treatment with the proteasome inhibitor MG132. Login to comment
133 These results suggest that the Q141K variant undergoes proteasomal proteolysis but the WT does not. Login to comment
135 The Q141K variant protein level was only two-fold enhanced by Fig. 4. Login to comment
136 Immunocytochemical staining of Flp-In-293 cells expressing ABCG2 WT or Q141K proteins (A) and statistical data to analyze the effects of MG132 on the cellular localization of ABCG2 WT or Q141K proteins (B, C). Login to comment
144 It is suggested that the WT is degraded substantially in lysosomes and that the Q141K variant protein undergoes degradation in both proteasomes and lysosomes. Login to comment
145 Effect of MG132 on the Cellular Localization of ABCG2 WT and Q141K Variant. Login to comment
146 Fig. 4A depicts the immunofluorescence images of Flp-In-293 cells expressing ABCG2 WT or Q141K that were incubated with or without 2 μM MG132 for 24 h. The ABCG2 protein was probed with either BXP-21 or 5D3 antibody. Login to comment
152 In contrast with the WT, immunofluorescence of the Q141K variant was relatively weak at the plasma membrane as well as within intracellular compartments. Login to comment
154 The 5D3 antibody-immunofluorescence image demonstrated even more clearly that MG132 treatment enhanced the localization of the Q141K variant protein at the plasma membrane (Fig. 4A). Login to comment
155 Fig. 4B and C depict the statistical data on the cellular localization of ABCG2 WT and Q141K. Login to comment
160 Marked differences were observed after the MG132 treatment in terms of the cellular localization and/or amount of the Q141K variant protein. Login to comment
161 It is noteworthy that the protein level of the Q141K variant increased in the plasma membrane after the MG132 treatment (Fig. 4B). Login to comment
162 Effect of MG132 on the Drug Resistance of Flp-In-293 Cells Expressing ABCG2 Q141K Variant. Login to comment
165 incubation conditions, Flp-In-293 cells expressing the Q141K variant exhibited a cellular resistance to SN-38, but their resistance profile was relatively weaker than that of cells expressing the WT (Fig. 5). Login to comment
166 This may be due to the lower expression level of the Q141K variant protein as compared with the ABCG2 WT in Flp-In-293 cells. Login to comment
167 It was of great interest to know how the inhibition of proteasomal protein degradation by MG132 affects the drug resistance of cells expressing the Q141K variant protein. Login to comment
168 As demonstrated in Fig. 5B, the MG132 treatment enhanced cellular resistance to SN-38 in Flp-In-293 cells expressing the Q141K variant, whereas little was changed in cells expressing ABCG2 WT, even after the MG132 treatment (data not shown). Login to comment
169 Thus, it is suggested that the Q141K variant sorted to the plasma membrane domain after the MG132 treatment was functionally active to extrude SN-38 out of the cells. Login to comment
177 We recently found that such reduced protein levels were associated with the instabil- Fig. 6. Schematic illustration of plausible pathways for protein folding and degradation of ABCG2 WT and Q141K variant protein. Login to comment
190 In contrast, Q141K undergoes both lysosomal proteolysis and ubiquitination-mediated proteasomal degradation. Login to comment
193 Drug resistance profiles of Flp-In-293 cells expressing ABCG2 WT or Q141K (A) and the effect of MG132 on the SN-38 resistance of Flp-In-293 cells expressing Q141K (B). Login to comment
198 *P<0.01 as compared with Flp-In-293 cells expressing ABCG2 Q141K treated with MG132. Login to comment
200 In our functional validation, the Q141K variant and the WT showed very similar substrate specificities and drug resistance profiles when they were expressed in Sf9 insect cells (42) and Flp-In-293 cells (18). Login to comment
201 The level of the Q141K variant protein was significantly lower than that of the WT, however when it was expressed in Flp-In-293 cells (Fig. 2B) and in other mammalian cell lines (12-18). Login to comment
202 Based on those recent findings, we have examined the contribution of proteasomal degradation to the lower-level expression of the Q141K variant by testing the effect of MG132. Login to comment
203 In fact, MG132 enhanced both the protein level of the Q141K variant (Figs. Login to comment
204 3A and 4A) and the drug resistance profile of the Q141K-expressing Flp-In-293 cells (Fig. 5B). Login to comment
205 It is strongly suggested that the lower protein expression level of the Q141K variant is due to both ubiquitin-mediated proteasomal degradation and the lysosomal proteolysis. Login to comment
221 The clinical impact of the Q141K SNP has been most extensively studied. Login to comment
222 The Q141K SNP was identified with varying frequencies in different ethnic groups and was found to be the most prevalent in Japanese and Chinese populations (2). Login to comment
223 Q141K is reportedly associated with lower levels of protein expression and impaired transport (12-18), whereas some controversies exist in the publications characterizing this SNP (15,16). Login to comment
224 Zamber et al. (15) and Urquhart et al. (16) reported that the levels of ABCG2 in the intestine were not significantly affected by the Q141K SNP. Login to comment
225 It is important to note, however, that ABCG2 protein levels in the intestinal biopsy samples were measured by immunoblotting and compared between homozygous WT/WT and heterozygous WT/Q141K human subjects in those reports (15,16). Login to comment
226 No data was demonstrated for the ABCG2 protein level in homozygous Q141K/Q141K subjects. Login to comment
227 Since ABCG2 forms homo- and hetero-dimers linked through a cysteinyl disulfide bond, it is plausible that the heterodimer consisting of the WT and the Q141K variant is preferentially preserved in the ER as is the WT homodimer. Login to comment
228 The present in vitro study provides clear evidence that this major non-synonymous SNP Q141K affects the protein stability of ABCG2 in the ER and enhances its susceptibility to proteasomal degradation, thus affecting the plasma membrane localization of this non-synonymous SNP. Login to comment
229 Further studies will be needed to verify the impact of the homozygous Q141K SNP on ABCG2 protein degradation in the intestine in vivo. Login to comment
1 We aimed to evaluate the impact of the major non-synonymous SNP Q141K on lysosomal and proteasomal degradations. Login to comment
5 The protein expression level of the Q141K variant increased about two-fold when Flp-In-293 cells were treated with MG132. Login to comment
23 Q141K has been associated with lower levels of protein expression and impaired transport in vitro; however, some controversies exist in the publications characterizing this SNP (12-18). Login to comment
25 Furthermore, the Q141K SNP was reportedly associated with a higher incidence of diarrhea in non-small cell lung cancer patients treated with gefitinib (22). Login to comment
31 In this context, it has been hypothesized that the reduced expression levels of the Q141K variant may be due to the ubiquitin-mediated proteasomal degradation. Login to comment
38 We here provide evidence that the major nonsynonymous SNP variant Q141K undergoes both ubiquitin-mediated protein degradation in proteasomes and lysosomal proteolysis and that the protein level of the Q141K variant is thereby significantly reduced. Login to comment
41 Preparation of plasmids carrying ABCG2 Q141K variant cDNA Wild-type (WT) ABCG2 cDNA inserted into the pcDNA5/FRT plasmid was used as the template, and the nonsynonymous SNP Q141K variant was generated by using the QuikChange® Site-Directed Mutagenesis Kit (Stratagene, La Jolla, CA, USA). Login to comment
46 Expression of ABCG2 WT and Q141K variant in Flp-In-293 cells Flp-In-293 cells were transfected with the ABCG2-pcDNA5/FRT plasmid, the Flp recombinase expression plasmid pOG44, and LipofectAmine™-2000 (Invitrogen, Carlsbad, CA, USA) according to the manufacturer`s instructions (see Fig. 2A). Login to comment
51 Flp-In-293 cells expressing ABCG2 WT or Q141K, named Flp-In-293/ABCG2 (WT) or Flp-In-293/ABCG2 (Q141K), were maintained in high-glucose Dulbecco`s modified Eagle`s medium (D-MEM) supplemented with 10% (v/v) heat-inactivated fetal calf serum (ICN Biomedicals, Inc., Aurora, OH, USA), 2 mM L-glutamine, 100 U/ml penicillin, 100 μg/ml streptomycin, 250 ng/ml amphotericin B, and 100 μg/ml hygromycin B at 37°C in a in air. Login to comment
83 Results and Discussion Protein expression levels of ABCG2 WT and Q141K in Flp-In-293 cells We previously found that protein expression levels varied among those SNP variants (18). Login to comment
85 The Q141K variant is reportedly associated with lower levels of protein expression; however, little is known about the underlying molecular mechanism. Login to comment
86 In the present study, ABCG2 WT and the Q141K variant were individually expressed in Flp-In-293 cells by using the Flp recombinase system, where one single copy of the cDNA encoding either the WT or the variant was integrated into the genomic DNA at the designated FRT site prepared in chromosome 12 (Fig. 2A). Login to comment
87 As shown in Figure 2B, mRNA levels of ABCG2 WT as well as Q141K were evenly represented in Flp-In-293 cells, where the mRNA levels of ABCG2 and GAPDH were measured by RT-PCR. Login to comment
88 Both WT and Q141K proteins were found as glycosylated homodimers. Login to comment
89 To quantify the levels of ABCG2 WT and Q141K proteins, we treated the samples with PNGase F and mercaptoethanol (ME) to remove the glycomoieties and to break the cysteinyl disulfide bond forming the homodimers (Fig. 2B). Login to comment
91 As demonstrated in Figure 2B, the protein level of the Q141K variant was about 45% of the ABCG2 WT protein level. Login to comment
92 Effect of MG132 and bafilomycin A1 on the protein expression levels of ABCG2 WT and Q141K variant We investigated the effect of MG132, an inhibitor of proteasomal proteolysis, on the protein expression levels of those SNP variants. Login to comment
93 Flp-In-293 cells expressing WT or Q141K were incubated in the presence of MG132 at a concentration of 2.0 μM for 24 h, and then cell lysate samples were immediately prepared. Login to comment
96 Protein expression levels of the WT and the Q141K variant were determined by immunoblotting after PNGase F treatment in the same way as described above. Login to comment
97 As shown in Figure 3A, the protein level of the Q141K variant was approximately two-fold enhanced by treatment with the proteasome inhibitor MG132. Login to comment
100 These results suggest that the Q141K variant undergoes proteasomal proteolysis but the WT does not. Login to comment
111 In contrast with the WT, immunofluorescence of the Q141K variant was relatively weak at the plasma membrane as well as within intracellular compartments. Login to comment
114 Figures 4B and 4C depict the statistical data on the cellular localization of ABCG2 WT and Q141K. Login to comment
119 Marked differences were observed after the MG132 treatment in terms of the cellular localization and/or amount of the Q141K variant protein. Login to comment
123 It was of great interest to know how the inhibition of proteasomal protein degradation by MG132 affects the drug resistance of cells expressing the Q141K variant protein. Login to comment
125 Thus, it is suggested that the Q141K variant sorted to the plasma membrane domain after the MG132 treatment was functionally active to extrude SN-38 out of the cells. Login to comment
134 The level of the Q141K variant protein was significantly lower than that of the WT, however when it was expressed in Flp-In-293 cells (Fig. 2B) and in other mammalian cell lines (12-18). Login to comment
137 3A and 4A) and the drug resistance profile of the Q141K-expressing Flp-In-293 cells (Fig. 5B). Login to comment
138 It is strongly suggested that the lower protein expression level of the Q141K variant is due to both ubiquitin-mediated proteasomal degradation and the lysosomal proteolysis. Login to comment
153 The Q141K SNP was identified with varying frequencies in different ethnic groups and was found to be the most prevalent in Japanese and Chinese populations (2). Login to comment
156 It is important to note, however, that ABCG2 protein levels in the intestinal biopsy samples were measured by immunoblotting and compared between homozygous WT/ WT and heterozygous WT/Q141K human subjects in those reports (15,16). Login to comment
157 No data was demonstrated for the ABCG2 protein level in homozygous Q141K/Q141K subjects. Login to comment
158 Since ABCG2 forms homo- and hetero-dimmers linked through a cysteinyl disulfide bond, it is plausible that the heterodimer consisting of the WT and the Q141K variant is preferentially preserved in the ER as is the WT homodimer. Login to comment
159 The present in-vitro study provides clear evidence that this major non-synonymous SNP Q141K affects the protein stability of ABCG2 in the ER and enhances its susceptibility to proteasomal degradation, thus affecting the plasma membrane localization of this nonsynonymous SNP. Login to comment
344 The sites of three nonsynonymous SNPs, Q141K, F208S and S441N, are indicated. Login to comment
349 Flp-mediated integration of the ABCG2 cDNA into FRT-tagged genomic DNA (A), the mRNA and protein levels of ABCG2 WT and Q141K (B) and the relationship between the relative intensity of ABCG2 immunoreactivity and the amount of cellular proteins (C). Login to comment
353 B: The mRNA level was analyzed by RT-PCR with total RNA extracted from Flp-In-293 cells expressing ABCG2 WT or Q141K. Login to comment
355 The level of the Q141K variant protein was normalized to the WT level. Login to comment
360 Effect of MG132 (A) or bafilomycin A1 (B) on the protein levels of ABCG2 WT and Q141K variant. Login to comment
361 A: Flp-In-293 cells expressing ABCG2 WT or Q141K were incubated with 2 μM MG132 for 0 and 24 h. ABCG2 protein levels were analyzed by immunoblotting after PNGase F treatment. Login to comment
362 B: Flp-In-293 cells expressing ABCG2 WT or Q141K were incubated with 10 nM bafilomycin A1 for 0, 12, and 24 h. ABCG2 protein levels were analyzed as described above. Data are expressed as mean values ± S.D. in triplicate experiments. Login to comment
365 Immunocytochemical staining of Flp-In-293 cells expressing ABCG2 WT or Q141K proteins (A) and statistical data to analyze the effects of MG132 on the cellular localization of ABCG2 WT or Q141K proteins (B, C). Login to comment
373 Drug resistance profiles of Flp-In-293 cells expressing ABCG2 WT or Q141K (A) and the effect of MG132 on the SN-38 resistance of Flp-In-293 cells expressing Q141K (B). Login to comment
378 *, P<0.01 as compared with Flp-In-293 cells expressing ABCG2 Q141K treated with MG132. Login to comment
379 Figure 6. Schematic illustration of plausible pathways for protein folding and degradation of ABCG2 WT and Q141K variant protein. Login to comment
392 In contrast, Q141K undergoes both lysosomal proteolysis and ubiquitination-mediated proteasomal degradation. Login to comment

[hide] The 315-316 deletion determines the BXP-21 antibod... Mol Cell Biochem. 2009 Feb;322(1-2):63-71. Epub 2008 Nov 11. Polgar O, Deeken JF, Ediriwickrema LS, Tamaki A, Steinberg SM, Robey RW, Bates SE
The 315-316 deletion determines the BXP-21 antibody epitope but has no effect on the function of wild type ABCG2 or the Q141K variant.
Mol Cell Biochem. 2009 Feb;322(1-2):63-71. Epub 2008 Nov 11., [PMID:19002564]

Abstract [show]
ABCG2 is a half-transporter initially described in multidrug-resistant cancer cells and lately identified as an important factor in the pharmacokinetics of its substrates. Q141K is by far the most intensively studied single nucleotide polymorphism of ABCG2 with potential clinical relevance. Here we used stably transfected HEK cells to study the Q141K polymorphism together with the deletion of amino acids 315-316, which were recently reported to coexist in two cancer cell lines (A549 and SK-OV-3). Functional studies confirmed our previous report that when normalized to surface expression, Q141K has impaired transport of mitoxantrone. This result was extended to include the ABCG2-specific substrate pheophorbide a. While we found no functional consequence of deleting amino acids 315 and 316, we did find that the deletion mutant is no longer recognized by the BXP-21 antibody. We conclude that amino acids 315 and 316 form part of the epitope for the BXP-21 antibody.

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0 The 315-316 deletion determines the BXP-21 antibody epitope but has no effect on the function of wild type ABCG2 or the Q141K variant Orsolya Polgar Æ John F. Deeken Æ Lilangi S. Ediriwickrema Æ Akina Tamaki Æ Seth M. Steinberg Æ Robert W. Robey Æ Susan E. Bates Received: 30 May 2008 / Accepted: 22 October 2008 / Published online: 11 November 2008 Ó Springer Science+Business Media, LLC. 2008 Abstract ABCG2 is a half-transporter initially described in multidrug-resistant cancer cells and lately identified as an important factor in the pharmacokinetics of its substrates. Login to comment
1 Q141K is by far the most intensively studied single nucleotide polymorphism of ABCG2 with potential clinical relevance. Login to comment
2 Here we used stably transfected HEK cells to study the Q141K polymorphism together with the deletion of amino acids 315-316, which were recently reported to coexist in two cancer cell lines (A549 and SK-OV-3). Login to comment
3 Functional studies confirmed our previous report that when normalized to surface expression, Q141K has impaired transport of mitoxantrone. Login to comment
18 The most extensively studied of these SNPs with potential clinical relevance is 421 C [ A resulting in a glutamic acid to lysine substitution (Q141K) in the protein. Login to comment
19 The Q141K SNP was identified with varying frequencies in different ethnic groups and was found to be the most prevalent in the Japanese population (*30%). Login to comment
20 Q141K has been associated with lower levels of protein expression and impaired transport in vitro, though some controversies exist in the publications characterizing this SNP [13-17]. Login to comment
22 Further, Q141K was associated with a higher incidence of diarrhea in non-small cell lung cancer patients treated with gefitinib [21]. Login to comment
23 Sequencing the ABCG2 gene in the 60 cancer cell lines maintained and used for screening in the National Cancer Institute by various groups revealed that ten of the cell lines carried the Q141K polymorphism. Login to comment
26 In the present study, we expanded our previous functional studies on the Q141K SNP [13] using pheophorbide a, a fluorescent compound that was recently reported by our group as an ABCG2-specific substrate [22]. Login to comment
27 In addition, we examined the effect of the D315-316 variant with or without the Q141K polymorphism on ABCG2 function. Login to comment
30 Mutagenesis and transfection The D315-316 and the Q141K/D315-316 mutants were generated by site-directed mutagenesis in the pcDNA3.1/ Myc-HisA(-) vector (Invitrogen) as previously described [23]. Login to comment
31 The wild type (R482), Q141K, R482G, and empty vector-transfected (pcDNA) cells were previously reported [13]. Login to comment
51 Once strong inverse correlations were ruled out, then a Kruskal-Wallis test was used to determine if the three clones of particular mutant, or a Wilcoxon rank sum test if 315-316-3 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 #Cells 315-316-6 315-316-8 Q141K/ 315-316-5 Q141K/ 315-316-15 Q141K/ 315-316-17 wild type Fig. 1 ABCG2 surface expression in the mutants. Login to comment
52 Flow cytometry with the 5D3 monoclonal antibody recognizing an extracellular epitope of ABCG2 is shown for the wild type and three clones of both mutants (D315-316 and Q141K/D315-316, with clone numbers indicated). Login to comment
62 We have previously used the same expression system to study non-synonymous SNPs, such as Q141K, V12M, and D620N, and found that Q141K results in impaired function [13]. Login to comment
63 Given that Q141K and D315-316 were found to coexist in the A549 and SK-OV-3 carcinoma cell lines, we also generated mutants carrying both the deletion and the Q141K SNP (Q141K/D315-316). Login to comment
65 We selected three clones of both the D315-316 and the Q141K/D315-316 mutants that displayed significant levels of surface expression (Fig. 1) and expanded these clones for further studies. Login to comment
66 Crude membranes extracted from the D315-316 and the Q141K/D315-316 clones and from the wild type and Q141K transfectants previously generated in our lab were subjected to immunoblotting with the commercially available BXP-21 monoclonal antibody. Login to comment
67 Surprisingly, despite sufficient surface expression levels seen on flow cytometry, none of the D315-316 and Q141K/D315-316 clones were detectable on immunoblot with the BXP-21 antibody (Fig. 2), while the wild type and the Q141K mutant clones were readily detected. Login to comment
70 To prove this, we used the 405 polyclonal antibody created in our laboratory [24] on the immunoblot previously probed with the BXP-21 antibody and found that with this antibody the D315-316 and the Q141K/D315-316 clones were also detectable, though the 405 antibody gives a weaker signal when compared to BXP-21 (Fig. 2). Login to comment
72 72 kDa 72 kDa BXP-21 ab 405 ab wt ∆ 315-316-3,6,8 Q141K/ ∆ 315-316 Q141K-5,8 -5,15,17 Fig. 2 The D315-316 mutant is not detected on immunoblot with the BXP-21 antibody. Login to comment
73 Membrane proteins extracted from HEK 293 cells stably transfected with the wild type (482R) and the mutants (Q141K, D315-316, and Q141K/D315-316) were separated on SDS/PAGE (18 lg/lane), transferred onto a PVDF membrane, and incubated overnight with the mouse monoclonal anti-ABCG2 antibody BXP-21 (upper panel) or the 405 rabbit polyclonal anti-ABCG2 antibody (lower panel). Login to comment
74 Two clones are shown for the Q141K mutant and three clones for the D315-316 and Q141K/D315-316 mutants, with clone numbers indicated. Login to comment
79 Efflux is represented by the shift between the histograms with (dashed line) and without (solid line) FTC c wildtype 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 #Cells 10 0 10 1 10 2 10 3 10 4 FL2-H 0 100 200 300 #Cells 5D3 mitoxantrone pheophorbide a 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 500 #Cells 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 500 #Cells 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 500 #Cells 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 500 #Cells 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 #Cells 10 0 10 1 10 2 10 3 10 4 FL4-H 0 100 200 300 400 #Cells Q141K/315-316-5315-316-6Q141K-8R482G-2 We next utilized a flow cytometry-based assay measuring the efflux of mitoxantrone and pheophorbide a to determine the activity of the mutants. Login to comment
83 For the Q141K variant, where a reduced surface expression has been confirmed [13, 16, 17], our previous studies also demonstrated impaired transport when mitoxantrone efflux was normalized to surface expression [13]. Login to comment
89 Using a Wilcoxon rank sum test, differences with unadjusted P-values \0.05 were found when the wild type was compared to the Q141K and Q141K/D315-316 mutants for both mitoxantrone and pheophorbide a. Login to comment
90 In agreement with previous reports, the transport efficacy of the R482G mutant was found to be significantly higher than that of the wild type for mitoxantrone [13]. Login to comment
91 The average ratio calculated for pheophorbide a for the D315-316 mutant was significantly higher than that for the Q141K and the Q141K/D315-316 mutants. Login to comment
92 In summary, the mutant carrying the deletion was similar in transport efficacy to the wild type for both substrates, while any mutant carrying the Q141K was impaired. Login to comment
94 Of the numerous SNPs identified so far in the ABCG2 gene, 421 C [ A, resulting in a mutant protein with a glutamine to lysine substitution at residue 141 (Q141K), is the most extensively studied and is suggested to have impaired transport function. Login to comment
95 Interestingly, some cell lines of the NCI-60 panel were reported to have the Q141K SNP together with a splicing variant resulting in the deletion of amino acids 315 and 316. Login to comment
96 Here, we investigated the functional consequences of the 315-316 deletion with/or without the Q141K mutation in stably transfected HEK 293 cells using a flow cytometry-based assay to analyze mitoxantrone and pheophorbide a efflux. Login to comment
97 After normalizing the measured efflux values to ABCG2 expression by the 5D3 monoclonal antibody, we have found that the D315-316 mutant has equivalent efflux 0.331 0.289 0.261 0.256 0.520 0.000 0.100 0.200 0.300 0.400 0.500 0.600 0.700 482R Q141K 316 R482G p=0.028 p<0.0001 p=0.0099 Mitoxantroneefflux/5D3antibody n=14 n=15n=23n=23n=22 0.445 0.410 0.263 0.313 0.504 0.000 0.100 0.200 0.300 0.400 0.500 0.600 0.700 482R Q141K 316 R482G p=0.00075 p=0.0000052 p=0.013 p=0.039 Pheophorbideaefflux/5D3antibody n=6 n=10n=14n=15 n=12 A B ∆315-316 Q141K/∆315- ∆315-316 Q141K/∆315- Fig. 4 Efflux normalized to cell surface expression. Login to comment
98 The mean FTC-inhibitable mitoxantrone (panel A) and pheophorbide a (panel B) efflux was plotted for the wild type (482R) and each mutant (Q141K, D315-316, aQ141K/D315-316, and R482G). Login to comment
102 The numbers above the bars represent the mean values plotted capacity to the wild type, while the double mutant (Q141K/ D315-316) was similar to the Q141K SNP. Login to comment
103 Additionally, the experiments reconfirmed that the Q141K SNP has significantly impaired function compared to the wild type protein when normalized by surface expression. Login to comment
125 In conclusion, the results presented here confirm our previous finding that the clinically relevant, frequently occurring Q141K SNP impairs transport of the chemotherapeutic agent mitoxantrone by ABCG2 [13]. Login to comment
127 We believe the Q141K mutation results in decreased surface expression due to impaired trafficking of ABCG2 [13], which, combined with impaired transporter function, effects a clinically detectable reduction in transporter function [19]. Login to comment
24 The Q141K SNP was identified with varying frequencies in different ethnic groups and was found to be the most prevalent in the Japanese population (~30%). Login to comment
25 Q141K has been associated with lower levels of protein expression and impaired transport in vitro, though some controversies exist in the publications characterizing this SNP [13-17]. Login to comment
28 Sequencing the ABCG2 gene in the 60 cancer cell lines maintained and used for screening in the National Cancer Institute by various groups revealed that ten of the cell lines carried the Q141K polymorphism. Login to comment
32 In addition, we examined the effect of the Δ315-316 variant with or without the Q141K polymorphism on ABCG2 function. Login to comment
35 Mutagenesis and transfection The Δ315-316 and the Q141K/D315-316 mutants were generated by site-directed mutagenesis in the pcDNA3.1/Myc-HisA(-) vector (Invitrogen) as previously described [23]. Login to comment
36 The wild type (R482), Q141K, R482G, and empty vector-transfected (pcDNA) cells were previously reported [13]. Login to comment
64 We have previously used the same expression system to study non-synonymous SNPs, such as Q141K, V12M, and D620N, and found that Q141K results in impaired function [13]. Login to comment
68 Crude membranes extracted from the Δ315-316 and the Q141K/Δ315-316 clones and from the wild type and Q141K transfectants previously generated in our lab were subjected to immunoblotting with the commercially available BXP-21 monoclonal antibody. Login to comment
69 Surprisingly, despite sufficient surface expression levels seen on flow cytometry, none of the Δ315-316 and Q141K/Δ315-316 clones were detectable on immunoblot with the BXP-21 antibody (Fig. 2), while the wild type and the Q141K mutant clones were readily detected. Login to comment
78 For the Q141K variant, where a reduced surface expression has been confirmed [13,16,17], our previous studies also demonstrated impaired transport when mitoxantrone efflux was normalized to surface expression [13]. Login to comment
84 Using a Wilcoxon rank sum test, differences with unadjusted P-values <0.05 were found when the wild type was compared to the Q141K and Q141K/Δ315-316 mutants for both mitoxantrone and pheophorbide a. Login to comment
86 The average ratio calculated for pheophorbide a for the Δ315-316 mutant was significantly higher than that for the Q141K and the Q141K/Δ315-316 mutants. Login to comment
87 In summary, the mutant carrying the deletion was similar in transport efficacy to the wild type for both substrates, while any mutant carrying the Q141K was impaired. Login to comment
93 Additionally, the experiments reconfirmed that the Q141K SNP has significantly impaired function compared to the wild type protein when normalized by surface expression. Login to comment
115 In conclusion, the results presented here confirm our previous finding that the clinically relevant, frequently occurring Q141K SNP impairs transport of the chemotherapeutic agent mitoxantrone by ABCG2 [13]. Login to comment
117 We believe the Q141K mutation results in decreased surface expression due to impaired trafficking of ABCG2 [13], which, combined with impaired transporter function, effects a clinically detectable reduction in transporter function [19]. Login to comment

[hide] ABCG2 Q141K polymorphism is associated with chemot... Cancer Sci. 2008 Dec;99(12):2496-501. Epub 2008 Nov 20. Kim IS, Kim HG, Kim DC, Eom HS, Kong SY, Shin HJ, Hwang SH, Lee EY, Lee GW
ABCG2 Q141K polymorphism is associated with chemotherapy-induced diarrhea in patients with diffuse large B-cell lymphoma who received frontline rituximab plus cyclophosphamide/doxorubicin/vincristine/prednisone chemotherapy.
Cancer Sci. 2008 Dec;99(12):2496-501. Epub 2008 Nov 20., [PMID:19032367]

Abstract [show]
ATP-binding cassette transporter G2 (ABCG2) is the most recently described transporter of the multidrug-resistance pump and it promotes resistance to anticancer drugs such as doxorubicin, mitoxantrone, topotecan, and SN-38. Of the ABCG2 polymorphisms, V12M and Q141K alter the functional activity of the ABCG2 transporter and influence the drug response and various toxicities to chemotherapeutic agents. We therefore evaluated the impact of the ABCG2 V12M and Q141K polymorphisms on the therapeutic outcomes and toxicities of primary rituximab plus cyclophosphamide/doxorubicin/vincristine/prednisone (R-CHOP) therapy in 145 Korean patients with diffuse large B-cell lymphoma (DLBCL). ABCG2 V12M and Q141K genotyping was carried out by pyrosequencing of polymerase chain reaction products. The clinical characteristics, treatment outcomes, toxicities of the patients, and the predictive value of the polymorphisms on response, survival, and adverse events to R-CHOP for 145 patients were analyzed according to the ABCG2 V12M and Q141K polymorphisms. No differences were observed according to ABCG2 Q141K and V12M genotype in patient characteristics, disease characteristics, response, survival, or hematology toxicity profiles in patients with DLBCL who received frontline R-CHOP chemotherapy. On multivariate analysis, grade I-IV diarrhea was statistically significant according to ABCG2 Q141K polymorphism (the QQ genotype vs the QK or KK genotypes; hazard ratio 2.835; 95% confidence interval 1.432-5.613; P = 0.003). This study demonstrates that the ABCG2 Q141K polymorphism may correlate with chemotherapy-induced diarrhea in patients with DLBCL who have received frontline R-CHOP chemotherapy, and this has implications for optimizing treatment with such agents.

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1 12 | 2496-2501 doi: 10.1111/j.1349-7006.2008.00985.x (c) 2008 Japanese Cancer Association Blackwell Publishing Asia ABCG2 Q141K polymorphism is associated with chemotherapy-induced diarrhea in patients with diffuse large B-cell lymphoma who received frontline rituximab plus cyclophosphamide/doxorubicin/ vincristine/prednisone chemotherapy In-Suk Kim,1,8 Hoon-Gu Kim,2,8 Dong Chul Kim,3 Hyeon-Seok Eom,4 Sun-Young Kong,5 Ho-Jin Shin,6 Sang-Hyun Hwang,7 Eun-Yup Lee7 and Gyeong-Won Lee2,9 1 Department of Laboratory Medicine; 2 Division of Hematology-Oncology, Department of Internal Medicine and 3 Department of Pathology, Gyeongsang National University Hospital, 90 Chilam-dong, Jinju 660-702; 4 Division of Hematology-Oncology, Department of Internal Medicine and 5 Department of Laboratory Medicine, Research Institute and Hospital, National Center, 809 Madu-dong, Ilsan-gu, Goyang-si, Gyeonggi-do 410-769; 6 Division of Hematology-Oncology, Department of Internal Medicine, and 7 Department of Laboratory Medicine, School of Medicine Pusan National University, 1-ga 10, Ami-dong, Seo-gu, Busan 602-739, Korea (Received June 27, 2008/Revised July 30, 2008; August 12, 2008/Accepted August 14, 2008/Online publication November 20, 2008) ATP-binding cassette transporter G2 (ABCG2) is the most recently described transporter of the multidrug-resistance pump and it promotes resistance to anticancer drugs such as doxorubicin, mitoxantrone, topotecan, and SN-38. Login to comment
2 Of the ABCG2 polymorphisms, V12M and Q141K alter the functional activity of the ABCG2 transporter and influence the drug response and various toxicities to chemotherapeutic agents. Login to comment
3 We therefore evaluated the impact of the ABCG2 V12M and Q141K polymorphisms on the therapeutic outcomes and toxicities of primary rituximab plus cyclophosphamide/doxorubicin/vincristine/prednisone (R-CHOP) therapy in 145 Korean patients with diffuse large B-cell lymphoma (DLBCL). Login to comment
4 ABCG2 V12M and Q141K genotyping was carried out by pyrosequencing of polymerase chain reaction products. Login to comment
5 The clinical characteristics, treatment outcomes, toxicities of the patients, and the predictive value of the polymorphisms on response, survival, and adverse events to R-CHOP for 145 patients were analyzed according to the ABCG2 V12M and Q141K polymorphisms. No differences were observed according to ABCG2 Q141K and V12M genotype in patient characteristics, disease characteristics, response, survival, or hematology toxicity profiles in patients with DLBCL who received frontline R-CHOP chemotherapy. Login to comment
6 On multivariate analysis, grade I-IV diarrhea was statistically significant according to ABCG2 Q141K polymorphism (the QQ genotype vs the QK or KK genotypes; hazard ratio 2.835; 95% confidence interval 1.432-5.613; P = 0.003). Login to comment
7 This study demonstrates that the ABCG2 Q141K polymorphism may correlate with chemotherapy-induced diarrhea in patients with DLBCL who have received frontline R-CHOP chemotherapy, and this has implications for optimizing treatment with such agents. Login to comment
15 (7-11) In particular, two non-synonymous polymorphisms, c.34G>A (p.Val12Met, V12M) and c.421C>A (p.Gln141Lys, Q141K), have been detected at relatively high frequencies in most ethnic groups, including Asians, Caucasians, and Africans. Login to comment
17 (15) A recently published study found that the ABCG2 V12M and Q141K polymorphisms are associated with susceptibility to and survival from DLBCL. Login to comment
18 (16) In the present study, we evaluated the impact of the ABCG2 Q141K and V12M polymorphisms on the therapeutic outcomes and adverse reactions of primary R-CHOP therapy in 145 patients with DLBCL, including treatment response, overall survival (OS) and event-free survival (EFS), hematological toxicities, and non-hematological toxicities. Login to comment
35 ABCG2 Q141K and V12M genotyping was carried out by pyrosequencing of the polymerase chain reaction (PCR) products from the ABCG2 gene to determine the presence of the Q141K and V12M polymorphisms. Login to comment
39 Patient characteristics and treatment outcomes according to the ABCG2 Q141K and V12M polymorphisms ABCG2 Q141K P-value ABCG2 V12M P-value QQ QK KK VV VM MM No. patients (%) 67 (46.2%) 69 (47.6%) 9 (6.2%) 71 (49.0%) 63 (43.4%) 11 (7.6%) Sex (no. Login to comment
43 Sequences and information of primers used for pyrosequencing Name Primer sequence (5' to 3') Size (bp) Polymerase chain reaction (Tm; °C) ABCG2 V12M F: CTCTCCAGATGTCTTCCAGTAATG 110 59 R: Biotin-TCCTTCAGTAAATGCCTTCAGGT S: TCGAAGTTTTTATCCCA ABCG2 Q141K F: Biotin-ACTGCAGGTTCATCATTAGCTAGA 238 60 R: CCGTTCGTTTTTTTCATGATTC S: CGAAGAGCTGCTGAGAA F, forward; R, reverse; S, sequencing; Tm, melting temperature. Login to comment
55 The primary objective of the current study was to correlate the ABCG2 Q141K and V12M polymorphisms with the response and survival outcomes, including OS and EFS to R-CHOP therapy. Login to comment
56 The secondary objectives were to correlate the ABCG2 Q141K and V12M polymorphisms with the hematological and non- hemtological toxicities of R-CHOP therapy. Login to comment
57 The clinical characteristics, treatment outcomes, and toxicities of the patients were compared using χ2 -tests, Fisher`s exact tests, or Mann-Whitney U-tests according to the ABCG2 Q141K and V12M polymorphisms. Login to comment
58 Logistic regression analysis was conducted to determine the predictive value of the polymorphisms on response and adverse reaction to R-CHOP for the 145 patients who had available both the ABCG2 Q141K and V12M polymorphism data. Login to comment
59 The variables included stage (stages 1 and 2 vs 3 and 4), IPI score (0-2 vs 3-5), age (<60 vs ≥60 years), performance status (Eastern Cooperative Oncology Group [ECOG] 0 and 1 vs ≥2), lactate dehydrogenase level (normal vs beyond normal range), extranodal involvement (≤1 vs ≥2 sites), B symptoms (absence vs presence), hematological toxicity (grade 0-II vs III-IV or grade 0 vs grade I-IV), non-hematological toxicity (grade 0-II vs III-IV or grade 0 vs grade I-IV), and ABCG2 Q141K (QQ, QK, and KK) and ABCG2 V12M (VV, VM, and MM) genotypes. Login to comment
66 Results Frequency of ABCG2 Q141K and V12M polymorphisms. Login to comment
67 In the frontline R-CHOP therapy group, the distribution of the QQ, QK, and KK genotypes of the ABCG2 Q141K polymorphism was 46.2, 47.6, and 6.2%, respectively (Table 1). Login to comment
70 Patient characteristics according to the ABCG2 Q141K and V12M polymorphisms. Login to comment
72 In brief, no differences in the patient and disease characteristics were observed according to the ABCG2 Q141K and V12M polymorphisms. Login to comment
73 Response to frontline R-CHOP therapy according to the ABCG2 Q141K and V12M polymorphisms. Login to comment
75 As shown in Table 1, no significant difference in the response rate or the ORR was observed according to the ABCG2 Q141K and V12M polymorphisms. No statistically significant difference in the response rate or ORR was observed according to the ABCG2 Q141K and V12M polymorphisms. Login to comment
76 Survival analysis according to the ABCG2 Q141K and V12M polymorphisms. Login to comment
79 When comparing OS and EFS according to the ABCG2 Q141K and V12M polymorphisms, neither the Q141K nor V12M polymorphisms had any impact on OS or EFS (Fig. 1). Login to comment
80 Side effects according to the ABCG2 Q141K and V12M polymorphisms. Login to comment
82 No significant differences of such hematological toxicities as anemia, leukocytopenia, neutropenia, and thrombocytopenia were observed according to the ABCG2 Q141K and V12M alleles. Login to comment
83 Among the severe non-hematological side effects (grade III-IV), fever, mucositis, infection, and diarrhea were observed more frequently in the patients with the non-QQ alleles of the ABCG Q141K polymorphisms. Login to comment
84 Of them, fever and infection were statistically significant according to the ABCG2 Q141K polymorphism (QQ vs QK or KK genotype; P = 0.037 and 0.046, respectively). Login to comment
85 When considering the presence of toxicity (grade I-IV), regardless of severity, differences for fever (P = 0.007) and diarrhea (P = 0.002) in the non-hematological toxicity profiles were noted and these were statistically significant according to the ABCG2 Q141K polymorphism (Table 4). Login to comment
86 On multivariate analysis, grade I-IV diarrhea was the statistically significant factor according to the ABCG2 Q141K polymorphism (HR 2.835; 95% CI 1.432-5.613; P = 0.003). Login to comment
87 Discussion Among several naturally occurring ABCG2 polymorphisms, V12M in exon 2 and Q141K in exon 5 occur in most racial groups, but they occur with a higher allellic frequency in Asians. Login to comment
88 In the present study, the frequencies of two polymorphisms, V12M and Q141K, were 0.293 and 0.300, respectively, and these values were comparable to those reported in the literature for Asians (0.230-0.241 and 0.150-0.360, respectively). Login to comment
89 (8,10,16,19-21) When the frequencies of the present study were compared with those reported in the literature for Asian populations, the frequencies of Q141K in these Korean DLBCL patients and the Asian healthy controls, including Korean, Chinese, and Malaysian, were statistically insignificant using the χ2 -test (Table 5). Login to comment
90 (8,10,16,19) Unlike a previous study that reported that ABCG2 Q141K is a candidate susceptibility gene in Chinese DLBCL patients,(16) our findings suggest the ABCG2 Q141K polymorphism may not be associated with an increased risk of DLBCL in Koreans. Login to comment
92 The present study demonstrated that the ABCG2 Q141K and V12M polymorphisms were not predictive of the response, OS, or EFS to R-CHOP chemotherapy in patients with DLBCL. Login to comment
93 No association of the ABCG2 Q141K and V12M polymorphisms with response and survival was noted in the present study for the following reasons: (1) the relatively small number of patients; (2) the relatively short period of follow up may not have been enough to see a significant difference in survival; and (3) other unknown chemoresistance mechanisms are probably important in the mechanism of R-CHOP action, and this would be expected to affect the response and survival of DLBCL patients. Login to comment
95 Several groups have reported that the Q141K genotype is associated with altered pharmacokinetic parameters of some ABCG2 substrates. Login to comment
96 (8,15,22) Notably, the Q141K genotype has the signature of recent positive selection and it is the strongest in the Asian population,(23) suggesting that there is some advantageous property for this genotype. Login to comment
97 The Q141K polymorphism has been associated with lower expression and activity of ABCG2 and with higher accumulation of ABCG2 substrates. Login to comment
98 (24) On univariate analysis, the present study showed statistically significant results for non-hematological toxicity such as grade I-IV diarrhea, grade I-IV fever, grade III-IV fever, and grade III-IV infection, according to ABCG2 Q141K polymorphism (QQ genotype vs non-QQ genotype, P = 0.002, 0.007, 0.037, and 0.046, respectively). Login to comment
99 On multivariate analysis, the present study demonstrated that the ABCG2 Q141K polymorphism was associated with grade I-IV diarrhea (P = 0.003), just like a previous report for which the ABCG2 Q141K polymorphism was associated with diarrhea in gefitinib-treated patients with non-small cell cancer. Login to comment
101 Survival curve after frontline rituximab plus cyclophosphamide-doxorubicin-vincristine-prednisone (R-CHOP) therapy according to the ABCG2 Q141K genotype. Login to comment
103 Hematological and non-hematological toxicity outcomes according to the ABCG2 Q141K and V12M polymorphisms (grade 0-II vs grade III-IV) ABCG2 Q141K P-value ABCG2 V12M P-value QQ QK or KK VV VM or MM No. patients (%) 67 (46.2%) 78 (53.8%) 71 (49.0%) 74 (51.0%) Grade III-IV, n (%) Anemia 9 (13.4%) 9 (11.5%) 0.730 10 (14.1%) 8 (10.8%) 0.550 Leukocytopenia 29 (43.3%) 32 (41.0%) 0.784 33 (46.5%) 28 (37.8%) 0.292 Neutropenia 38 (56.7%) 39 (50.0%) 0.419 38 (53.5%) 39 (52.7%) 0.921 Thrombocytopenia 4 (6.0%) 11 (14.1%) 0.109 9 (12.7%) 6 (8.1%) 0.367 Grade III-IV, n (%) Fever 8 (11.9%) 20 (25.6%) 0.037 16 (22.5%) 12 (16.2%) 0.335 Mucostis 11 (16.4%) 22 (28.2%) 0.091 19 (26.8%) 14 (18.9%) 0.260 Infection 9 (13.4%) 21 (27.5%) 0.046 13 (18.3%) 17 (23.0%) 0.488 Nausea and vomiting 2 (3.0%) 5 (6.4%) 0.337 3 (4.2%) 4 (5.4%) 0.740 Diarrhea 2 (3.0%) 9 (11.5%) 0.052 4 (5.6%) 7 (9.5%) 0.384 Alopecia 27 (40.3%) 38 (48.7%) 0.309 32 (45.1%) 33 (44.6%) 0.954 Neurotoxicity 2 (3.0%) 4 (5.1%) 0.518 3 (4.2%) 3 (4.1%) 0.959 Bold and italic values significant at P < 0.05. Login to comment
105 Non-hematological toxicity outcomes according to ABCG2 Q141K genotype (grade 0 vs grade I-IV) ABCG2 Q141K P-value QQ QK or KK No. patients, n (%) 67 (46.2%) 78 (53.8%) Grade I-IV, n (%) Fever 18 (26.9%) 38 (48.7%) 0.007 Mucositis 43 (64.2%) 50 (64.1%) 0.992 Infection 24 (35.8%) 40 (51.3%) 0.062 Diarrhea 21 (31.3%) 44 (56.4%) 0.002 Bold and italic values significant at P < 0.05. is one of the most common side effects of chemotherapy. Login to comment
110 Therefore, the present study demonstrates the necessity of study for determining the association between ABCG2 Q141K polymorphism and adverse reactions to chemotherapy. Login to comment
114 The mechanism underlying the functional impact of the ABCG2 Q141K amino acid change is not entirely known, but it is most likely associated with reduced protein levels and altered ATPase activity. Login to comment
115 (15) The association we observed between ABCG2 Q141K genotype status and observed clinical adverse reactions such as diarrhea, infection, and fever may reflect a role for this transporter in the metabolism and elimination pathways of R-CHOP, especially doxorubicin. Login to comment
116 In summary, the present study demonstrates that the ABCG2 Q141K polymorphism is associated with chemotherapy-induced diarrhea in patients with DLBCL who have received frontline R-CHOP chemotherapy. Login to comment

[hide] Inhibiting the function of ABCB1 and ABCG2 by the ... Biochem Pharmacol. 2009 Mar 1;77(5):781-93. Epub 2008 Nov 18. Shi Z, Tiwari AK, Shukla S, Robey RW, Kim IW, Parmar S, Bates SE, Si QS, Goldblatt CS, Abraham I, Fu LW, Ambudkar SV, Chen ZS
Inhibiting the function of ABCB1 and ABCG2 by the EGFR tyrosine kinase inhibitor AG1478.
Biochem Pharmacol. 2009 Mar 1;77(5):781-93. Epub 2008 Nov 18., 2009-03-01 [PMID:19059384]

Abstract [show]
The tyrphostin 4-(3-chloroanilino)-6,7-dimethoxyquinazoline (AG1478) is a potent and specific EGFR tyrosine kinase inhibitor (TKI); its promising pre-clinical results have led to clinical trials. Overexpression of ATP-binding cassette (ABC) transporters such as ABCB1, ABCC1 and ABCG2 is one of the main causes of multidrug resistance (MDR) and usually results in the failure of cancer chemotherapy. However, the interaction of AG1478 with these ABC transporters is still unclear. In the present study, we have investigated this interaction and found that AG1478 has differential effects on these transporters. In ABCB1-overexpressing cells, non-toxic doses of AG1478 were found to partially inhibit resistance to ABCB1 substrate anticancer drugs as well as increase intracellular accumulation of [3H]-paclitaxel. Similarly, in ABCG2-overexpressing cells, AG1478 significantly reversed resistance to ABCG2 substrate anticancer drugs and increased intracellular accumulation of [3H]-mitoxantrone as well as fluorescent compound BODIPY-prazosin. AG1478 also profoundly inhibited the transport of [3H]-E(2)17betaG and [3H]-methotrexate by ABCG2. We also found that AG1478 slightly stimulated ABCB1 ATPase activity and significantly stimulated ABCG2 ATPase activity. Interestingly, AG1478 did not inhibit the photolabeling of ABCB1 or ABCG2 with [125I]-iodoarylazidoprazosin. Additionally, AG1478 did not alter the sensitivity of parental, ABCB1- or ABCG2-overexpressing cells to non-ABCB1 and non-ABCG2 substrate drug and had no effect on the function of ABCC1. Overall, we conclude that AG1478 is able to inhibit the function of ABCB1 and ABCG2, with a more pronounced effect on ABCG2. Our findings provide valuable contributions to the development of safer and more effective EGFR TKIs for use as anticancer agents in the clinic.

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261 Recently, one common functional single-nucleotide polymorphism of ABCG2 Q141K (421C ! Login to comment

[hide] Knocking down breast cancer resistance protein (Bc... Mol Pharm. 2009 Jan-Feb;6(1):134-43. Yue W, Abe K, Brouwer KL
Knocking down breast cancer resistance protein (Bcrp) by adenoviral vector-mediated RNA interference (RNAi) in sandwich-cultured rat hepatocytes: a novel tool to assess the contribution of Bcrp to drug biliary excretion.
Mol Pharm. 2009 Jan-Feb;6(1):134-43., [PMID:19105722]

Abstract [show]
BCRP transports numerous drugs/derived metabolites and toxins, and exhibits overlapping substrate specificity with P-glycoprotein (P-gp) and multidrug resistance-associated protein 2 (MRP2). Assessing the contribution of BCRP to drug/metabolite biliary excretion in intact hepatocytes remains a challenge. Current studies were designed to develop a novel in vitro tool to specifically assess the contribution of Bcrp to drug biliary excretion. Adenoviral vectors expressing short hairpin (sh) RNA targeting Bcrp (Ad-si01Bcrp) or a nontarget control (Ad-siNT) were packaged and infected into sandwich-cultured rat hepatocytes (SCRH). Protein levels were detected by immunoblot. Biliary excretion index (BEI) and in vitro biliary clearance (Cl(biliary)) of nitrofurantoin (BCRP substrate) and digoxin (P-gp substrate) were compared among noninfected, Ad-siNT- and Ad-si01Bcrp-infected SCRH. shRNA targeting Bcrp efficiently knocked down Bcrp in SCRH, while levels of other transport proteins (P-gp, Mrp2, Bsep, Mrp4 and Oatp1a1) were unaffected. In SCRH exhibiting Bcrp knockdown, cellular accumulation of nitrofurantoin was increased markedly and nitrofurantoin BEI and in vitro Cl(biliary) were decreased to 11% and 14% of control, respectively. Digoxin values were unaffected by knockdown of Bcrp. Results indicated that Bcrp in SCRH contributed predominantly to nitrofurantoin biliary excretion, but played a negligible role in digoxin biliary excretion. In summary, Bcrp knockdown in SCRH is the first in vitro model utilizing intact hepatocytes to assess the contribution of Bcrp to the biliary excretion of drugs. This approach may be useful in predicting drug-drug interactions in biliary excretion and the consequence of impaired BCRP function on the hepatic exposure of drugs/derived metabolites.

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14 For example, the single-nucleotide polymorphism in ABCG2 C421A (Q141K) is associated with * Corresponding author. Login to comment
40 (4) Imai, Y.; Nakane, M.; Kage, K.; Tsukahara, S.; Ishikawa, E.; Tsuruo, T.; Miki, Y.; Sugimoto, Y. C421A Polymorphism in the Human Breast Cancer Resistance Protein Gene Is Associated with Low Expression of Q141K Protein and Low-Level Drug Resistance. Login to comment

[hide] Functions of the breast cancer resistance protein ... Adv Drug Deliv Rev. 2009 Jan 31;61(1):26-33. Epub 2008 Dec 3. Noguchi K, Katayama K, Mitsuhashi J, Sugimoto Y
Functions of the breast cancer resistance protein (BCRP/ABCG2) in chemotherapy.
Adv Drug Deliv Rev. 2009 Jan 31;61(1):26-33. Epub 2008 Dec 3., 2009-01-31 [PMID:19111841]

Abstract [show]
The breast cancer resistance protein, BCRP/ABCG2, is a half-molecule ATP-binding cassette transporter that facilitates the efflux of various anticancer agents from the cell, including 7-ethyl-10-hydroxycamptothecin, topotecan and mitoxantrone. The expression of BCRP can thus confer a multidrug resistance phenotype in cancer cells, and its transporter activity is involved in the in vivo efficacy of chemotherapeutic agents. Thus, the elucidation of the substrate preferences and structural relationships of BCRP is essential to understanding its in vivo functions during chemotherapeutic treatments. Single nucleotide polymorphisms (SNPs) have also been found to be key factors in determining the efficacy of chemotherapeutics, and those therapeutics that inhibit BCRP activity, such as the SNP that results in a C421A mutant, may result in unexpected side effects of the BCRP- anticancer drugs interaction even at normal dosages. In order to modulate the BCRP activity during chemotherapy, various compounds have been tested as inhibitors of this protein. Estrogenic compounds including estrone, several tamoxifen derivatives in addition to phytoestrogens and flavonoids have been shown to reverse BCRP-mediated drug resistance. Intriguingly, recently developed molecular targeted cancer drugs, such as the tyrosine kinase inhibitors imatinib mesylate, gefitinib and others, can also interact with BCRP. Since both functional SNPs and inhibitory agents of BCRP modulate the in vivo pharmacokinetics and pharmacodynamics of its substrate drugs, BCRP activity is an important consideration in the development of molecular targeted chemotherapeutics.

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173 C421A (Q141K) BCRP SNP . Login to comment
847 C421A (Q141K) BCRP SNP In a previous study, we screened for BCRP SNPs among a population of Japanese individuals and in human cancer cell lines where we identified three variant BCRP cDNAs harboring the following substitutions: G34A (V12M), C421A (Q141K) and an amino acid deletion of residues 944-949 that lacks Ala-315 and Thr-316 (Δ315-6) [54]. Login to comment
848 The G34A and C421A variations were determined to be SNPs, and we have subsequently determined that the C421A BCRP-transfected murine fibroblast PA317 (PA/Q141K) cells show lower exogenous BCRP protein levels than the wild-type BCRP-transfected cells [54]. Login to comment
849 The intracellular topotecan accumulation in PA/Q141K cells was also found to be higher compared with other BCRP transfectants, indicating that the C421A (Q141K) SNP influences BCRP function. Login to comment
851 Regarding the Q141K BCRP SNP, there are conflicting reports on its effects upon expression levels, localization, and functionality [38,54,56-58]. Login to comment
852 Additional studies will be required to clarify the mechanism by which the Q141K mutation reduces the protein expression levels of BCRP. Login to comment
874 Among these SNPs, with the exception of C376T and C421A, only a few have been studied Table 1 Identified SNPs within the BCRP gene Variation Effect Domain A-1379G - Δ-654/-651 - G-286C - T-476C - Δ-235A - A-113G - A-29G - G34A V12M N-terminal T114C No change N-terminal G151T G51C N-terminal C369T No change NBD C376T Q126stop NBD C421A Q141K NBD C458T T153M NBD C474T No change NBD C496G Q166E NBD A564G No change NBD A616C I206L NBD T623C F208S NBD T742C S248P Linker G1000T E334stop Linker G1098A No change Linker T1291C F431L TMD A1425G No change TMD T1465C F489L TMD A1768T N590Y TMD G1858A D620N TMD G2237T - G2393T - NBD, nucleotide-binding domain; TMD, transmembrane domain. Login to comment
920 BCRP is also expressed in the intestinal epithelial cells, and Cusatis et al. [92] demonstrated that diarrhea, an adverse event related to oral gefitinib administration, is linked to genetic polymorphisms of BCRP, most notably C421A (Q141K). Login to comment

[hide] Quality control of human ABCG2 protein in the endo... Adv Drug Deliv Rev. 2009 Jan 31;61(1):66-72. Epub 2008 Dec 11. Wakabayashi-Nakao K, Tamura A, Furukawa T, Nakagawa H, Ishikawa T
Quality control of human ABCG2 protein in the endoplasmic reticulum: ubiquitination and proteasomal degradation.
Adv Drug Deliv Rev. 2009 Jan 31;61(1):66-72. Epub 2008 Dec 11., 2009-01-31 [PMID:19111842]

Abstract [show]
Human ATP-binding cassette (ABC) transporter ABCG2 (BCRP/MXR/ABCP) is a plasma membrane protein carrying intra- and inter-molecular disulfide bonds and an N-linked glycan. Both disulfide bond formation and N-glycosylation are critical check points determining the stability and degradation fate of ABCG2 protein in the endoplasmic reticulum (ER). Misfolded ABCG2 protein without those post-translational modifications is removed from the ER by retrotranslocation to the cytosol compartment, ubiquitination by ubiquitin ligase, and finally degradation by proteasomes. Certain non-synonymous SNP variants of ABCG2 undergo such ER-associated degradation (ERAD).

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813 Furthermore, certain non-synonymous single nucleotide polymorphisms (SNPs), such as Q141K, F208S, and S441N, were also found to greatly affect the stability of ABCG2 in the endoplasmic reticulum (ER) and to enhance the protein degradation rate via ubiquitination and proteasomal proteolysis [9a,b]. Login to comment
938 The most extensively studied among those SNPs with potential clinical relevance is 421 CNA resulting in a glutamic acid to lysine substitution (Q141K) in the ABCG2 protein. Login to comment
939 The Q141K SNP has been identified with varying frequencies in different ethnic groups and was found to be the most relevant in Japanese and Chinese populations (approximately 30% in the allele frequency) [37]. Login to comment
940 Q141K has been associated with lower levels of protein expression and impaired transport in vitro [34,43,46-49]. Login to comment
942 Furthermore, the Q141K SNP was reportedly Fig. 3. Schematic illustration of the stable expression of ABCG2 in Flp-In-293 cells. Login to comment
948 It has been hypothesized that the reduced expression levels of the Q141K variant may be due to its ubiquitin-mediated proteasomal degradation. Login to comment

[hide] Ins and outs of the ABCG2 multidrug transporter: a... Adv Drug Deliv Rev. 2009 Jan 31;61(1):47-56. Epub 2008 Dec 24. Hegedus C, Szakacs G, Homolya L, Orban TI, Telbisz A, Jani M, Sarkadi B
Ins and outs of the ABCG2 multidrug transporter: an update on in vitro functional assays.
Adv Drug Deliv Rev. 2009 Jan 31;61(1):47-56. Epub 2008 Dec 24., 2009-01-31 [PMID:19135105]

Abstract [show]
The major aim of this chapter is to provide a critical overview of the in vitro methods available for studying the function of the ABCG2 multidrug transporter protein. When describing the most applicable assay systems, in each case we present a short overview relevant to ABC multidrug transporters in general, and then we concentrate on the tools applicable to analysis of substrate-drug interactions, the effects of potential activators and inhibitors, and the role of polymorphisms of the ABCG2 transporter. Throughout this chapter we focus on recently developed assay systems, which may provide new possibilities for analyzing the pharmacological aspects of this medically important protein.

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983 C421A, resulting in the Q141K amino acid change, has been examined most extensively, due to its high frequency in Asian and Caucasian populations [27,28]. Login to comment

[hide] ABCG2: a perspective. Adv Drug Deliv Rev. 2009 Jan 31;61(1):3-13. Epub 2008 Dec 16. Robey RW, To KK, Polgar O, Dohse M, Fetsch P, Dean M, Bates SE
ABCG2: a perspective.
Adv Drug Deliv Rev. 2009 Jan 31;61(1):3-13. Epub 2008 Dec 16., 2009-01-31 [PMID:19135109]

Abstract [show]
ABCG2, or breast cancer resistance protein (BCRP), is an ABC transporter that has been the subject of intense study since its discovery a decade ago. With high normal tissue expression in the brain endothelium, gastrointestinal tract, and placenta, ABCG2 is believed to be important in the protection from xenobiotics, regulating oral bioavailability, forming part of the blood-brain barrier, the blood-testis barrier, and the maternal-fetal barrier. Notably, ABCG2 is often expressed in stem cell populations, where it likely plays a role in xenobiotic protection. However, clues to its epigenetic regulation in various cell populations are only beginning to emerge. While ABCG2 overexpression has been demonstrated in cancer cells after in vitro drug treatment, endogenous ABCG2 expression in certain cancers is likely a reflection of the differentiated phenotype of the cell of origin and likely contributes to intrinsic drug resistance. Notably, research into the transporter's role in cancer drug resistance and its development as a therapeutic target in cancer has lagged. Substrates and inhibitors of the transporter have been described, among them chemotherapy drugs, tyrosine kinase inhibitors, antivirals, HMG-CoA reductase inhibitors, carcinogens, and flavonoids. This broad range of substrates complements the efficiency of ABCG2 as a transporter in laboratory studies and suggests that, while there are redundant mechanisms of xenobiotic protection, the protein is important in normal physiology. Indeed, emerging studies in pharmacology and toxicology assessing polymorphic variants in man, in combination with murine knockout models have confirmed its dynamic role. Work in pharmacology may eventually lead us to a greater understanding of the physiologic role of ABCG2.

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1282 Of these, the nonsynonymous 421CNA single nucleotide polymorphism (SNP) that results in a glycine to lysine (Q141K) amino acid change has been studied most extensively. Login to comment
1283 The Q141K SNP has been linked to decreased plasma membrane expression of ABCG2, decreased drug transport or reduced ATPase activity [138-141]. Login to comment
1284 Some small studies have shown that the Q141K SNP alters the pharmacokinetics of chemotherapeutic agents such as topotecan, diflomotecan, and 9-aminocamptothecin [142-144]. Login to comment
1286 Thus, the Q141K SNP maylead to higher drug toxicity in some patient populations. Login to comment
1336 The Q141K SNP offers the possibility to do similar studies with ABCG2. Login to comment

[hide] Intracellular trafficking of MDR transporters and ... Curr Top Med Chem. 2009;9(2):197-208. Porcelli L, Lemos C, Peters GJ, Paradiso A, Azzariti A
Intracellular trafficking of MDR transporters and relevance of SNPs.
Curr Top Med Chem. 2009;9(2):197-208., [PMID:19200005]

Abstract [show]
Multi-drug resistance (MDR) frequently contributes to the failure of chemotherapeutic treatments in cancer patients. Mechanisms underlying the development of MDR have been extensively studied and are considered multifactorial. Among them, the ATP-Binding Cassette (ABC) family of proteins plays a pivotal role. Processes of cellular distribution and subcellular localization of MDR-ABC proteins are not yet well explored and to enlighten these topics could be crucial to understand cellular drug uptake and retention. In this review, we analysed literature data concerning i) intracellular trafficking of MDR-ABC proteins (BCRP, P-gp and MRP1) and ii) mechanisms altering their cellular localization and trafficking. Moreover, we describe single nucleotide polymorphisms (SNP) that have been reported for some multidrug resistance (MDR) transporters, such as BCRP and P-gp, emphasizing their ability to affect the expression, function and localization of the transporters, with implications on drug resistance phenotypes.

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139 The most studied ABCG2 polymorphism is the nonsynonymous SNP C421A, which results in a glutamine to lysine amino acid change at position 141 (Q141K). Login to comment
142 PA317 cells transfected with the ABCG2 C421A variant (PA/Q141K) had markedly decreased ABCG2 protein expression, compared with the wild-type ABCG2-transfected cells (PA/WT), despite similar levels of ABCG2 mRNA. Login to comment
143 Additionally, PA/Q141K cells were 2-3 fold more sensitive to SN-38, mitoxantrone and topotecan compared with PA/WT. Login to comment
144 Nonetheless, cross-resistance patterns were similar for the wild-type and the Q141K ABCG2 variant, suggesting that this polymorphism does not affect substrate recognition of ABCG2 [96]. Login to comment
148 [98] reported that the protein expression level of the Q141K variant was approximately half that of the wild-type ABCG2 in Flp-In- Table 1. Login to comment
149 Polymorphisms and Mutations Affecting the Cellular Localization of MDR Transporters Transporter Variant Amino-acid Change Localization References Intracellular + plasma membrane [99] C421A Q141K Plasma membrane [97, 98, 101] Intracellular + plasma membrane [98, 101] G34A V12M Plasma membrane [97, 99] ABCG2 G1322A S441N Intracellular [97, 98] ABCC1 G128C C43S Intracellular + plasma membrane [116] 4175-4180del RM1392-1393del Intracellular (ER) [118] C2302T R768W Intracellular (ER) [119] A3517T I1173F Intracellular (ER) [120, 121] C2366T S789F Intracellular + plasma membrane [122] ABCC2 G4348A A1459T Intracellular + plasma membrane [122] 293 transfected cells. Login to comment
150 Additionally, Q141K-expressing cells were more sensitive to SN-38 and mitoxantrone compared with wild-type-expressing cells, but they were more resistant to doxorubicin and daunorubicin. Login to comment
152 [96], suggesting that the substrate specificity of the Q141K variant slightly differs from that of wild-type ABCG2. Login to comment
154 [99] did not observe decreased expression of ABCG2 in HEK-293 cells transfected with Q141K ABCG2 compared with cells transfected with wild-type ABCG2. Login to comment
156 [100] did not find a correlation between the Q141K SNP and the level of expression of ABCG2. Login to comment
157 Despite similar levels of ABCG2 protein expression, Q141K-expressing HEK-293 cells were 1.2 to 5-fold more sensitive to mitoxantrone, topotecan, SN-38 and diflomotecan than cells transfected with wild-type ABCG2 [99]. Login to comment
158 Of note, basal ATPase activity was 1.8-fold lower in membrane protein isolated from Sf9 insect cells infected with recombinant baculoviruses containing full-length Q141K ABCG2 compared to protein from cells expressing wild-type ABCG2 [99]. Login to comment
160 [101] who reported that the ATPase activity of Q141K ABCG2 was 1.3-fold lower than wild-type ABCG2 in the Sf9 system. Login to comment
163 [99] reported that HEK-293 cells expressing the Q141K variant displayed high intra-cellular ABCG2 staining, as well as cell surface staining. Login to comment
165 These results suggested an impaired membrane trafficking or incorrect membrane insertion of Q141K ABCG2 [99], but they were not confirmed by others [97,98,101] who showed that both the wild-type and Q141K ABCG2 were localized in the cell surface. Login to comment
195 Additionally, the expression levels of S441N ABCG2 were much lower than the wild-type ABCG2 and also lower than the Q141K [97]. Login to comment
217 From these studies it is clear that only the S441N and, possibly, the V12M and Q141K variants might affect ABCG2 localization within the cell. Login to comment

[hide] ABCG2 polymorphism markedly affects the pharmacoki... Clin Pharmacol Ther. 2009 Aug;86(2):197-203. Epub 2009 May 27. Keskitalo JE, Zolk O, Fromm MF, Kurkinen KJ, Neuvonen PJ, Niemi M
ABCG2 polymorphism markedly affects the pharmacokinetics of atorvastatin and rosuvastatin.
Clin Pharmacol Ther. 2009 Aug;86(2):197-203. Epub 2009 May 27., [PMID:19474787]

Abstract [show]
The ABCG2 c.421C>A single-nucleotide polymorphism (SNP) was determined in 660 healthy Finnish volunteers, of whom 32 participated in a pharmacokinetic crossover study involving the administration of 20 mg atorvastatin and rosuvastatin. The frequency of the c.421A variant allele was 9.5% (95% confidence interval 8.1-11.3%). Subjects with the c.421AA genotype (n = 4) had a 72% larger mean area under the plasma atorvastatin concentration-time curve from time 0 to infinity (AUC(0-infinity)) than individuals with the c.421CC genotype had (n = 16; P = 0.049). In participants with the c.421AA genotype, the rosuvastatin AUC(0-infinity) was 100% greater than in those with c.421CA (n = 12) and 144% greater than in those with the c.421CC genotype. Also, those with the c.421AA genotype showed peak plasma rosuvastatin concentrations 108% higher than those in the c.421CA genotype group and 131% higher than those in the c.421CC genotype group (P < or = 0.01). In MDCKII-ABCG2 cells, atorvastatin transport was increased in the apical direction as compared with vector control cells (transport ratio 1.9 +/- 0.1 vs. 1.1 +/- 0.1). These results indicate that the ABCG2 polymorphism markedly affects the pharmacokinetics of atorvastatin and, even more so, of rosuvastatin-potentially affecting the efficacy and toxicity of statin therapy.

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187 et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low‑level drug resistance. Login to comment
18 The c.421C>A SNP (rs2231142), resulting in an amino acid change p.Gln141Lys, is one of the best-characterized sequence variations in the ABCG2 gene. Login to comment
22 this SNP affects ABCG2 function; some studies have suggested that it reduces ABCG2 expression,11,14-16 and other studies have suggested that it reduces the ATPase activity of ABCG2.12,17 However, the p.Gln141Lys variant is located within the cell apart from the ATP-binding region. Login to comment
186 et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low‑level drug resistance. Login to comment

[hide] Identification of a urate transporter, ABCG2, with... Proc Natl Acad Sci U S A. 2009 Jun 23;106(25):10338-42. Epub 2009 Jun 8. Woodward OM, Kottgen A, Coresh J, Boerwinkle E, Guggino WB, Kottgen M
Identification of a urate transporter, ABCG2, with a common functional polymorphism causing gout.
Proc Natl Acad Sci U S A. 2009 Jun 23;106(25):10338-42. Epub 2009 Jun 8., 2009-06-23 [PMID:19506252]

Abstract [show]
Genome-wide association studies (GWAS) have successfully identified common single nucleotide polymorphisms (SNPs) associated with a wide variety of complex diseases, but do not address gene function or establish causality of disease-associated SNPs. We recently used GWAS to identify SNPs in a genomic region on chromosome 4 that associate with serum urate levels and gout, a consequence of elevated urate levels. Here we show using functional assays that human ATP-binding cassette, subfamily G, 2 (ABCG2), encoded by the ABCG2 gene contained in this region, is a hitherto unknown urate efflux transporter. We further show that native ABCG2 is located in the brush border membrane of kidney proximal tubule cells, where it mediates renal urate secretion. Introduction of the mutation Q141K encoded by the common SNP rs2231142 by site-directed mutagenesis resulted in 53% reduced urate transport rates compared to wild-type ABCG2 (P < 0.001). Data from a population-based study of 14,783 individuals support rs2231142 as the causal variant in the region and show highly significant associations with urate levels [whites: P = 10(-30), minor allele frequency (MAF) 0.11; blacks P = 10(-4), MAF 0.03] and gout (adjusted odds ratio 1.68 per risk allele, both races). Our data indicate that at least 10% of all gout cases in whites are attributable to this causal variant. With approximately 3 million US individuals suffering from often insufficiently treated gout, ABCG2 represents an attractive drug target. Our study completes the chain of evidence from association to causation and supports the common disease-common variant hypothesis in the etiology of gout.

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4 Introduction of the mutation Q141K encoded by the common SNP rs2231142 by site-directed mutagenesis resulted in 53% reduced urate transport rates compared to wild-type ABCG2 (P < 0.001). Login to comment
35 Impaired Urate Transport of ABCG2 Q141K. Login to comment
47 To test whether this SNP is not only statistically associated but causally related to elevated urate levels, we introduced the mutation Q141K encoded by the rs2231142 T allele by site-directed mutagenesis. Login to comment
48 Wild-type and Q141K mutant ABCG2 showed similar expression levels at the cell surface when expressed in Xenopus oocytes (Fig. S1). Login to comment
49 Notably, Q141K-expressing oocytes showed 54% reduced urate transport rates compared with those expressing wild-type ABCG2 at similar levels (Fig. 2 B and C), and decreased urate efflux across a range of intracellular urate concentrations (Fig. 2D). Login to comment
50 Our evidence for Q141K as a causal loss-of-function variant is supported by data showing reduced transport of chemotherapeutic agents by this mutation (6). Login to comment
53 Information on the SNP rs2231442, encoding the Q141K variant, was available as an imputed SNP from genome-wide Affymetrix 6.0 data along with 601 other SNPs in a 600-kb region containing ABCG2, and also genotyped directly using the TaqMan assay. Login to comment
64 Among the white study participants, population-attributable risk calculations yielded that a conservatively estimated 10% of gout cases could be attributed to the Q141K mutation. Login to comment
65 Our findings are of substantial clinical interest because of the high prevalence of the Q141K mutation in individuals of European and Asian ancestry and the substantial population attributable risk. Login to comment
82 We show that ABCG2 is a urate efflux transporter and rs2231142 (Q141K) a loss-of-function mutation that causes hyperuricemia and gout. Login to comment
89 Oocytes after injection were cultured in a modified L-15 media (OR-3) and kept at 15-20 °C. mRNA was prepared using the SP6 mMessage mMachine (Ambion Inc.) according to the manufactures protocol. The Q141K mutant was created using site directed mutagenesis with the primers: 5Ј-GGT GAG AGA AAA CTT AAA GTT CTC AGC AGC TC 3-Ј and 5Ј-GAG CTG CTG AGA ACT TTA AGT TTT CTC TCA CC-3Ј and completed using the Quick-Change Lightning kit (Stratagene) according to the manufacturer`s protocol. Login to comment
97 For the 1-2-h accumulation studies, after incubation in the C-14 urate solution, 2 oocytes were placed in each scintillation tube (N) with lysis buffer of 1 N NaOH; a total Q141 *CFTR F508 ABCG2 CFTR 1 2 3 4 5 0.02 0.04 0.06 0.08 0.10 Internal oocyte concentration ( µM) nim/etycoo/lomp:xulffE 0.0 0.3 0.6 0.9 noitalumuccAetarU nim021/etycoo/lomp WT Q141K C D** 150 76 52 H2OWTQ141K α-BCRP α-actin B A Fig. 2. Login to comment
98 The ABCG2 Q141K mutation results in reduced urate transport. Login to comment
102 (B) Western blot of oocytes expressing ABCG2 WT or Q141K (monomers and dimers of ABCG2 are detected). Login to comment
104 (C) Accumulation rates in oocytes expressing ABCG2 WT or Q141K normalized to ABCG2 expression level. Login to comment
105 (D) Efflux rates for ABCG2 WT and Q141K. Login to comment
106 (WT [red triangles]: slope ϭ 0.01, N ϭ 55; Q141K [black squares]: slope ϭ 0.02, N ϭ 55). Login to comment

[hide] Oral sulfasalazine as a clinical BCRP probe substr... J Pharm Sci. 2010 Feb;99(2):1046-62. Adkison KK, Vaidya SS, Lee DY, Koo SH, Li L, Mehta AA, Gross AS, Polli JW, Humphreys JE, Lou Y, Lee EJ
Oral sulfasalazine as a clinical BCRP probe substrate: pharmacokinetic effects of genetic variation (C421A) and pantoprazole coadministration.
J Pharm Sci. 2010 Feb;99(2):1046-62., [PMID:19569219]

Abstract [show]
This study evaluated the utility of oral sulfasalazine as a probe substrate for Breast Cancer Resistance Protein (BCRP; ABCG2) activity by assessing the impact of genetic variation or coadministration of an inhibitor (pantoprazole) on plasma and urine pharmacokinetics of sulfasalazine and metabolites. Thirty-six healthy male subjects prescreened for ABCG2 421CC (reference activity), CA, and AA (lower activity) genotypes (N = 12 each) received a single 500 mg oral dose of enteric coated sulfasalazine alone, with 40 mg pantoprazole, or with 40 mg famotidine (gastrointestinal pH control) in a 3-period, single fixed sequence, crossover design. No significant difference in sulfasalazine or metabolite pharmacokinetics in 421AA or CA compared to 421CC subjects was found; however, high inter-subject variability was observed. Geometric mean (95% CI) sulfasalazine plasma AUC((0-infinity)) values were 32.1 (13.2, 78.1), 16.8 (7.15, 39.6) and 62.7 (33.4, 118) microg h/mL, and C(max) were 4.01 (1.62, 9.92), 1.70 (0.66, 4.40), and 6.86 (3.61, 13.0) microg/mL for CC, CA, and AA subjects, respectively. Pantoprazole and famotidine did not affect sulfasalazine pharmacokinetics in any genotypic cohort. These results suggest that the pharmacokinetics of oral, enteric-coated 500 mg sulfasalazine are not sufficiently sensitive to ABCG2 genetic variation or inhibitors to be useful as a clinical probe substrate of BCRP activity.

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14 The combined result of intestinal and liver BCRP activity is lower systemic exposures of substrate drugs.6 Several nonsynonymous single nucleotide polymorphisms (SNPs) in the human ABCG2 gene have been associated with reduced BCRP transport activity, including C421A, C376T, G34A, T1291C, and T623C.5 C421A, which results in a substitution of lysine for glutamine (Q141K) in the BCRP protein has been associated with increased drug exposure in vivo in man. Login to comment

[hide] Clinical relevance of a pharmacogenetic approach u... Clin Cancer Res. 2009 Jul 15;15(14):4750-8. Epub 2009 Jul 7. Kim DH, Sriharsha L, Xu W, Kamel-Reid S, Liu X, Siminovitch K, Messner HA, Lipton JH
Clinical relevance of a pharmacogenetic approach using multiple candidate genes to predict response and resistance to imatinib therapy in chronic myeloid leukemia.
Clin Cancer Res. 2009 Jul 15;15(14):4750-8. Epub 2009 Jul 7., 2009-07-15 [PMID:19584153]

Abstract [show]
PURPOSE: Imatinib resistance is major cause of imatinib mesylate (IM) treatment failure in chronic myeloid leukemia (CML) patients. Several cellular and genetic mechanisms of imatinib resistance have been proposed, including amplification and overexpression of the BCR/ABL gene, the tyrosine kinase domain point mutations, and MDR1 gene overexpression. EXPERIMENTAL DESIGN: We investigated the impact of 16 single nucleotide polymorphisms (SNP) in five genes potentially associated with pharmacogenetics of IM, namely ABCB1, multidrug resistance 1; ABCG2, breast-cancer resistance protein; CYP3A5, cytochrome P450-3A5; SLC22A1, human organic cation transporter 1; and AGP, alpha1-acid glycoprotein. The DNAs from peripheral blood samples in 229 patients were genotyped. RESULTS: The GG genotype in ABCG2 (rs2231137), AA genotype in CYP3A5 (rs776746), and advanced stage were significantly associated with poor response to IM especially for major or complete cytogenetic response, whereas the GG genotype at SLC22A1 (rs683369) and advanced stage correlated with high rate of loss of response or treatment failure to IM therapy. CONCLUSIONS: We showed that the treatment outcomes of imatinib therapy could be predicted using a novel, multiple candidate gene approach based on the pharmacogenetics of IM.

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201 Gardner et al. reported that the ABCG2 SNP Q141K (rs2231142 in the current article) is associated with the intracellular level of imatinib in vitro model (6). Login to comment
204 Similar to a previous result using an in vitro model (6), the ABCG2 genotype Q141K (rs2231142) was associated with molecular response to imatinib therapy in univariate analyses although it was not statistically significant in multivariate analyses. Login to comment

[hide] ABC-transporter gene-polymorphisms are potential p... Brain. 2009 Sep;132(Pt 9):2517-30. Epub 2009 Jul 15. Cotte S, von Ahsen N, Kruse N, Huber B, Winkelmann A, Zettl UK, Starck M, Konig N, Tellez N, Dorr J, Paul F, Zipp F, Luhder F, Koepsell H, Pannek H, Montalban X, Gold R, Chan A
ABC-transporter gene-polymorphisms are potential pharmacogenetic markers for mitoxantrone response in multiple sclerosis.
Brain. 2009 Sep;132(Pt 9):2517-30. Epub 2009 Jul 15., [PMID:19605531]

Abstract [show]
Escalation therapy with mitoxantrone (MX) in highly active multiple sclerosis is limited by partially dose-dependent side-effects. Predictors of therapeutic response may result in individualized risk stratification and MX dosing. ATP-binding cassette-transporters ABCB1 and ABCG2 represent multi-drug resistance mechanisms involved in active cellular MX efflux. Here, we investigated the role of ABC-gene single nucleotide polymorphisms (SNPs) for clinical MX response, corroborated by experimental in vitro and in vivo data. Frequencies of ABCB1 2677G>T, 3435C>T and five ABCG2-SNPs were analysed in 832 multiple sclerosis patients (Germany, Spain) and 264 healthy donors. Using a flow-cytometry-based in vitro assay, MX efflux in leukocytes from individuals with variant alleles in both ABC-genes (designated genotype ABCB1/ABCG2-L(ow), 22.2% of patients) was 37.7% lower than from individuals homozygous for common alleles (ABCB1/ABCG2-H(igh), P < 0.05, 14.8% of patients), resulting in genotype-dependent MX accumulation and cell death. Addition of glucocorticosteroids (GCs) inhibited MX efflux in vitro. ABC-transporters were highly expressed in leukocyte subsets, glial and neuronal cells as well as myocardium, i.e. cells/tissues potentially affected by MX therapy. In vivo significance was further corroborated in experimental autoimmune encephalomyelitis in Abcg2(-/-) animals. Using a MX dose titrated to be ineffective in wild-type animals, disease course and histopathology in Abcg2(-/-) mice were strongly ameliorated. Retrospective clinical analysis in MX monotherapy patients (n = 155) used expanded disability status scale, relapse rate and multiple sclerosis functional composite as major outcome parameters. The clinical response rate [overall 121 of 155 patients (78.1%)] increased significantly with genotypes associated with decreasing ABCB1/ABCG2-function [ABCB1/ABCG2-H 15/24 (62.5%) responders, ABCB1/ABCG2-I(ntermediate) 78/98 (79.6%), ABCB1/ABCG2-L 28/33 (84.8%), exact Cochran-Armitage test P = 0.039]. The odds ratio for response was 1.9 (95% CI 1.0-3.5) with each increase in ABCB1/ABCG2 score (from ABCB1/ABCG2-H to -I-, and -I to -L). In 36 patients with severe cardiac or haematological side effects no statistically relevant difference in genotype frequency was observed. However, one patient with biopsy proven cardiomyopathy only after 24 mg/m2 MX exhibited a rare genotype with variant, partly homozygous alleles in 3 ABC-transporter genes. In conclusion, SNPs in ABC-transporter genes may serve as pharmacogenetic markers associated with clinical response to MX therapy in multiple sclerosis. Combined MX/GC-treatment warrants further investigation.

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42 TaqManTM PCR was performed for ABCG2 V12M (reference SNP rs2231137) and Q141K (rs2231142) using Platinum qPCR SuperMix-UDG (Invitrogen, Karlsruhe, Germany) on a 7500 Real Time PCR system (Applied Biosystems, Darmstadt, Germany). Login to comment
70 Retrospective clinical correlation of genotype and MX response Patient samples were genotyped for ABCB1 2677 G4T, 3435C4T, ABCG2 V12M and Q141K and retrospectively correlated with clinical MX response. Login to comment
102 Of five different ABCG2 SNPs with potential functional significance investigated, only reference SNP (rs) 2231137 (G4A) leading to a V12M substitution and rs2231142 (C4A) resulting in a Q141K substitution were observed. Login to comment
110 A high degree of linkage disequilibrium was found between ABCG2 V12M and Q141K [linkage D` = 0.854, (Gaunt et al., 2007)]. Login to comment
207 For ABCG2, only V12M and Q141K polymorphisms could be detected, in vitro leading to disruption of apical membrane localization (V12M) or decreased ATPase function (Q141K) Table 3 Association of ABC-transporter genotype with therapeutic response to MX monotherapy, exact Cochran-Armitage test (P = 0.039) (panel A), or MX/GC combination therapy (P = 0.348) (panel B) Total, n (%) Responder, n (%) Non-Responder, n (%) A: MX monotherapy ABCB1/ABCG2-H 24 (15.5) 15 (62.5) 9 (37.5) ABCB1/ABCG2-I 98 (63.2) 78 (79.6) 20 (20.4) ABCB1/ABCG2-L 33 (21.3) 28 (84.8) 5 (15.2) Total 155 121 (78.1) 34 (21.9) B: MX/GC combination therapy ABCB1/ABCG2-H 21 (13.6) 12 (57.1) 9 (42.9) ABCB1/ABCG2-I 100 (64.9) 58 (58.0) 42 (42.0) ABCB1/ABCG2-L 33 (21.4) 21 (63.6) 12 (36.4) Total 154 91 (59.1) 63 (40.9) Retrospective analysis using EDSS, relapse rate and MSFC as main outcome parameters to define responders and non-responders, respectively. Login to comment

[hide] Identification of compounds that correlate with AB... Mol Pharmacol. 2009 Nov;76(5):946-56. Epub 2009 Jul 24. Deeken JF, Robey RW, Shukla S, Steadman K, Chakraborty AR, Poonkuzhali B, Schuetz EG, Holbeck S, Ambudkar SV, Bates SE
Identification of compounds that correlate with ABCG2 transporter function in the National Cancer Institute Anticancer Drug Screen.
Mol Pharmacol. 2009 Nov;76(5):946-56. Epub 2009 Jul 24., [PMID:19633067]

Abstract [show]
ABCG2 is an ATP-binding cassette transporter that counts multiple anticancer compounds among its substrates and is believed to regulate oral bioavailability as well as serve a protective role in the blood-brain barrier, the maternal-fetal barrier, and hematopoietic stem cells. We sought to determine whether novel compounds that interact with the transporter could be identified through analysis of cytotoxicity profiles recorded in the NCI Anticancer Drug Screen database. A flow cytometric assay was used to measure ABCG2 function in the 60 cell lines and generate a molecular profile for COMPARE analysis. This strategy identified >70 compounds with Pearson correlation coefficients (PCCs) >0.4, where reduced drug sensitivity correlated with ABCG2 expression, as well as >120 compounds with PCCs < -0.4, indicating compounds to which ABCG2 expression conferred greater sensitivity. Despite identification of known single nucleotide polymorphisms in the ABCG2 gene in a number of the cell lines, omission of these lines from the COMPARE analysis did not affect PCCs. Available compounds were subjected to validation studies to confirm interaction with the transporter, including flow cytometry, [(125)I]IAAP binding, and cytotoxicity assays, and interaction was documented in 20 of the 27 compounds studied. Although known substrates of ABCG2 such as mitoxantrone or topotecan were not identified, we characterized three novel substrates-5-hydroxypicolinaldehyde thiosemicarbazone (NSC107392), (E)-N-(1-decylsulfanyl-3-hydroxypropan-2-yl)-3-(6-methyl-2,4-dioxo-1H-pyri midin-5-yl)prop-2-enamide (NSC265473), and 1,2,3,4,7-pentahydroxy-1,3,4,4a,5,11b-hexahydro[1,3]dioxolo[4,5-j]phenanth ridin-6(2H)-one [NSC349156 (pancratistatin)]-and four compounds that inhibited transporter function-2-[methyl(2-pyridin-2-ylethyl)-amino]fluoren-9-one hydroiodide (NSC24048), 5-amino-6-(7-amino-5,8-dihydro-6-methoxy-5,8-dioxo-2-quinolinyl)-4-(2-hydr oxy-3,4-dimethoxyphenyl)-3-methyl-2-pyridinecarboxylic acid, methyl ester (NSC45384), (17beta)-2,4-dibromo-estra-1,3,5(10)-triene-3,17-diol (NSC103054), and methyl N-(pyridine-4-carbonylamino)carbamodithioate (NSC636795). In summary, COMPARE analysis of the NCI drug screen database using the ABCG2 functional profile was able to identify novel substrates and transporter-interacting compounds.

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23 One nonsynonymous substitution, 421CϾA (dbSNP 914CϾA, rs2231142), leads to an amino acid substitution of lysine for glutamine at position 141 and has been shown to result in lower plasma membrane expression, reduced drug efflux, and reduced ATPase activity (Imai et al., 2002; Mizuarai et al., 2004; Morisaki et al., 2005). Login to comment
135 DNA from 59 of the NCI-60 cell lines were genotyped to identify the presence of the Q141K allelic variant, previously shown to have diminished function compared with wild-type ABCG2 (Imai et al., 2002; Morisaki et al., 2005), as well as a SNP variant in the first intron of the gene recently found to correlate with chemotherapy toxicity in vivo (Rudin et al., 2008). Login to comment
137 For the Q141K SNP, 12 cell lines were found to contain a variant allele. Login to comment
139 The other 10 lines were heterozygous, containing one Q141K variant allele (A549, COLO205, HCT116, SF295, MALME-3M, SK-OV-3, CAKI-1, HOP62, HOP92, and MDA-MB-231). Login to comment
143 To determine whether these variants affected the correlation between gene expression and transporter function, the scatter plot analysis comparing expression with function was repeated after excluding those cell lines with the Q141K SNP or the intron 1 SNP. Login to comment
165 Nucleotide Change Amino Acid Reference Sequence Cell Lines Heterozygote Variants Homozygote Variants Intron 1 rs2622604 MOLT4, HOP-92, HCC2998, SF539, SNB19, SNB75, U251, SKMEL5, OVCAR3, OVCAR8, RXF393, TK10, NCI ADR-RES, MDA-MB-231, HS578T, 786-0 SW620, OVCAR 5, BT549, T47D 914CϾA Q141K rs2231142 A549, COLO205, HCT116, SF295, MALME-3M, SKOV-3, CAKI-1, HOP62, HOP92, MDA-MB-231 LOX IMVI, A498 862CϾT Y123Y rs2231139 None None 989CϾG Q166E rs1061017 None None 1057AϾG G188G rs3116439 None None 1116TϾC F208S rs1061018 None None 1235TϾC S248P rs3116448 None None 1493GϾT E334* rs3201997 None None levels of P-gp or MRP1 did not improve PCCs (data not shown). Login to comment

[hide] Single nucleotide polymorphism in ABCG2 is associa... J Hum Genet. 2009 Oct;54(10):572-80. Epub 2009 Aug 21. Cha PC, Mushiroda T, Zembutsu H, Harada H, Shinoda N, Kawamoto S, Shimoyama R, Nishidate T, Furuhata T, Sasaki K, Hirata K, Nakamura Y
Single nucleotide polymorphism in ABCG2 is associated with irinotecan-induced severe myelosuppression.
J Hum Genet. 2009 Oct;54(10):572-80. Epub 2009 Aug 21., [PMID:19696792]

Abstract [show]
Irinotecan is an anti-neoplastic agent that is widely used for treating colorectal and lung cancers, but often causes toxicities such as severe myelosuppression and diarrhea. In this study, we performed a two-stage case-control association study for irinotecan-induced severe myelosuppression (grades 3 and 4). In the first stage, 23 patients who developed severe myelosuppression and 58 patients who did not develop any toxicity were examined for 170 single nucleotide polymorphisms (SNPs) in 14 genes involved in the metabolism and transport of irinotecan. A total of five SNPs were identified to show the possible association with severe myelosuppression (P(Fisher)<0.01) and were further examined in 7 cases and 20 controls in the second stage of the study. An intronic SNP, rs2622604, in ABCG2 showed P(Fisher)=0.0419 in the second stage and indicated a significant association with severe myelosuppression in the combined study (P(Fisher)=0.000237; P(Corrected)=0.036). Although only limited subjects were investigated, our results suggested that a genetic polymorphism in ABCG2 might alter the transport activity for the drug and elevate the systemic circulation level of irinotecan, leading to severe myelosuppression.

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48 In addition to this SNP, rs7977213 in SLCO1B3 as well as the UGT1A7*3 variant Table 1 Demographical characteristic of patients First study Second study Combined study Case (N¼23) Control (N¼58) Case (N¼7) Control (N¼20) Case (N¼30) Control (N¼78) Age 62.04 (37-77) 60.29 (34-77) 60.29 (40-81) 67.60 (50-84) 61.63 (37-81) 62.21 (34-84) % of men 61 71 57 42 60 65 Types of cancers Lung 12 52% 10 17% 2 29% 4 17% 14 47% 14 18% Cervical 6 26% 2 3% 0 0% 0 0% 6 20% 2 3% Colorectal 3 13% 35 60% 3 43% 13 57% 6 20% 48 62% Gastric 1 4% 4 7% 1 14% 2 9% 2 7% 6 8% Ovarian 0 0% 2 3% 1 14% 1 4% 1 3% 3 4% Breast+cervical 1 4% 0 0% 0 0% 0 0% 1 3% 0 0% Pancreatic 0 0% 1 2% 0 0% 0 0% 0 0% 1 1% Esophageal 0 0% 1 2% 0 0% 0 0% 0 0% 1 1% Breast 0 0% 1 2% 0 0% 0 0% 0 0% 1 1% No information 0 0% 2 0% 0 0% 0 0% 0 0% 2 3% Types of regimens CPT-11 1 4% 11 19% 1 14% 2 9% 2 6% 13 17% Other drug combinations 22 96% 47 81% 6 86% 18 91% 28 94% 65 83% Associations of 170 SNPs with severe myelosuppression in subjects who received irinotecan therapy Allele Frequency of allele 1 Fisher test`s P-values Gene name SNP ID Position of SNP/functional SNP 1 2 dCase eControl f1vs2 g11vs h22vs Odds ratio UGT1A9 rs3832043 UGT1A9*1b or *22 9T 10T 0.63 0.63 1.00E+00 8.05EÀ01 7.33EÀ01 0.76 UGT1A7 rs17868323 Exon 1 (Lys 129 Asn) T G 0.54 0.63 3.72EÀ01 1.00E+00 1.16EÀ01 2.68 UGT1A7 1A7_R131Kaa Exon 1 (R131K) (UGT1A7*2) C A 0.54 0.63 3.72EÀ01 1.00E+00 1.16EÀ01 2.68 UGT1A7 1A7_R131Kba Exon 1 (R131K) (UGT1A7*2) G A 0.54 0.63 3.72EÀ01 1.00E+00 1.16EÀ01 2.68 UGT1A7 rs11692021 Exon 1 (Arg 208 Trp) (UGT1A7*3) T C 0.65 0.76 1.71EÀ01 1.00E+00 6.72EÀ03 15.56 UGT1A1 rs887829 Intron (tagged UGT1A1*28, promoter indel) G A 0.96 0.90 3.53EÀ01 1.64EÀ01 1.00E+00 5.15 UGT1A1 rs4148323 Exon 1 (Arg 71 Gly) (UGT1A1*6) G A 0.76 0.78 8.34EÀ01 4.51EÀ01 2.14EÀ02 12.00 UGT1A1 rs35350960 Exon 1 (Gln 229 Pro) (UGT1A1*27) C A 1.00 0.99 1.00E+00 1.00E+00 1.00E+00 NA BCHE rs697356 3' near gene G C 0.30 0.40 2.74EÀ01 4.95EÀ01 4.46EÀ01 1.64 BCHE rs1007845 3' near gene G A 0.83 0.78 6.67EÀ01 6.13EÀ01 1.00E+00 1.40 BCHE rs2048493 Intron 2 C G 0.78 0.62 6.41EÀ02 5.08EÀ02 4.29EÀ01 2.74 BCHE rs4639017 Intron 1 C G 0.72 0.60 2.07EÀ01 2.07EÀ01 7.17EÀ01 2.07 ABCC5 rs6810123 3' near gene A G 0.33 0.41 3.72EÀ01 1.00E+00 2.07EÀ01 2.07 ABCC5 rs12638772 3' near gene A G 0.36 0.32 7.07EÀ01 2.50EÀ01 1.00E+00 2.36 ABCC5 rs7613247 3' near gene A G 0.93 0.94 1.00E+00 1.00E+00 1.00E+00 1.15 ABCC5 rs2176825 3' near gene A G 0.25 0.37 1.81EÀ01 5.07EÀ01 2.07EÀ01 1.98 ABCC5 rs13066518 3' near gene T A 0.33 0.39 4.78EÀ01 1.00E+00 4.50EÀ01 1.62 ABCC5 rs3749443 3' near gene A G 0.20 0.18 1.00E+00 1.00E+00 7.90EÀ01 1.29 ABCC5 rs1402001 3' near gene A G 0.52 0.57 6.03EÀ01 7.95EÀ01 2.30EÀ01 2.10 ABCC5 rs6790814 3' near gene C G 0.66 0.76 2.33EÀ01 4.63EÀ01 1.25EÀ01 4.42 ABCC5 rs9838667 3' near gene T G 0.34 0.46 2.12EÀ01 5.64EÀ01 3.05EÀ01 1.85 ABCC5 rs2280392 3' near gene G A 0.25 0.23 8.35EÀ01 3.40EÀ01 8.02EÀ01 2.84 ABCC5 rs1879257 3' near gene A G 0.43 0.32 2.02EÀ01 3.07EÀ01 3.27EÀ01 2.02 ABCC5 rs3817403 3' near gene A G 0.87 0.90 7.82EÀ01 1.00E+00 1.00E+00 1.19 ABCC5 rs3805111 3' near gene T C 0.09 0.10 7.85EÀ01 1.00E+00 7.72EÀ01 1.24 ABCC5 rs3805108b 3' near gene A G 0.11 0.19 2.50EÀ01 6.69EÀ01 4.00EÀ01 1.97 ABCC5 rs2872247 3' near gene T G 0.76 0.70 4.49EÀ01 6.30EÀ01 6.67EÀ01 1.30 ABCC5 rs2293001 3' near gene T C 0.48 0.54 6.00EÀ01 7.79EÀ01 5.56EÀ01 1.44 ABCC5 rs4148572 Intron 2 C G 0.33 0.22 1.60EÀ01 1.36EÀ01 3.30EÀ01 4.20 ABCC5 rs4148568 Intron 2 A G 0.13 0.18 6.37EÀ01 5.85EÀ01 7.89EÀ01 NA ABCC5 rs4148564 Intron 2 A G 0.89 0.82 3.44EÀ01 3.00EÀ01 1.00E+00 1.89 ABCC5 rs4148560 Intron 2 A T 0.78 0.80 8.30EÀ01 7.96EÀ01 1.36EÀ01 4.20 ABCC5 rs7624838 Intron 2 T C 0.50 0.52 8.63EÀ01 7.82EÀ01 1.00E+00 1.26 ABCG2 rs2231164 Intron 14 C T 0.36 0.39 8.56EÀ01 5.57EÀ02 4.46EÀ01 NA ABCG2 rs2622611 Intron 10 T G 0.16 0.18 8.20EÀ01 3.29EÀ01 1.00E+00 NA ABCG2 rs1871744 Intron 6 T C 0.55 0.69 9.76EÀ02 3.22EÀ01 1.71EÀ01 2.73 ABCG2 rs2231142 Exon 5 (Gln 141 Lys) C A 0.78 0.78 1.00E+00 1.00E+00 1.00E+00 0.79 ABCG2 rs2231137 Exon 2 (Val 12 Met) G A 0.67 0.79 1.53EÀ01 3.19EÀ01 1.36EÀ01 4.20 ABCG2 rs1564481 Intron 1 C T 0.72 0.62 2.77EÀ01 3.15EÀ01 6.67EÀ01 1.74 ABCG2 rs2622624 Intron 1 A G 0.41 0.29 1.93EÀ01 2.78EÀ01 3.35EÀ01 2.41 ABCG2 rs2622604 Intron 1 T C 0.28 0.09 2.35EÀ03 7.81EÀ02 6.66EÀ03 4.40 ABCB1 rs1882478 Intron 27 C T 0.59 0.55 7.27EÀ01 4.31EÀ01 7.57EÀ01 1.57 ABCB1 rs6979885 Intron 27 G A 0.89 0.91 1.00E+00 1.00E+00 1.00E+00 1.19 ABCB1 rs2235047 Intron 27 T G 0.50 0.52 8.63EÀ01 7.82EÀ01 1.00E+00 1.26 ABCB1 rs1045642 Exon 27 (Ile 3 Ile) T C 0.37 0.44 4.80EÀ01 1.00E+00 4.34EÀ01 1.67 ABCB1 rs1002205 Intron 26 C G 0.52 0.60 3.75EÀ01 4.21EÀ01 7.14EÀ01 1.70 ABCB1 rs6949448 Intron 26 T C 0.35 0.40 5.91EÀ01 1.00E+00 6.09EÀ01 1.42 ABCB1 rs2235067 Intron 23 A G 0.07 0.11 5.60EÀ01 1.00E+00 5.36EÀ01 1.83 ABCB1 rs2373588 Intron 22 T C 0.43 0.39 5.98EÀ01 7.20EÀ01 7.99EÀ01 1.53 ABCB1 rs7787082 Intron 22 A G 0.50 0.50 1.00E+00 7.72EÀ01 7.72EÀ01 NA ABCB1 rs3789246 Intron 20 A G 0.09 0.19 1.50EÀ01 5.51EÀ01 2.69EÀ01 2.31 ABCB1 rs1922242b Intron 17 A T 0.78 0.68 2.45EÀ01 3.24EÀ01 4.90EÀ01 1.74 ABCB1 rs2235046 Intron 17 A G 0.72 0.59 1.51EÀ01 4.38EÀ02 1.00E+00 2.89 ABCB1 rs868755 Intron 9 C A 0.59 0.61 1.00E+00 1.00E+00 7.29EÀ01 1.36 ABCB1 rs4148734 Intron 8 C T 0.87 0.90 7.79EÀ01 5.42EÀ01 1.00E+00 1.55 ABCB1 rs2235018 Intron 6 A G 0.67 0.81 9.65EÀ02 2.09EÀ01 1.40EÀ01 4.13 ABCB1 rs10256836a Intron 5 C G 0.13 0.09 5.68EÀ01 1.00E+00 5.31EÀ01 1.73 ABCB1 rs10259849a Intron 5 C T 0.14 0.10 5.75EÀ01 1.00E+00 5.36EÀ01 1.65 ABCB1 rs1202172 Intron 5 T G 0.96 0.86 1.01EÀ01 8.01EÀ02 1.00E+00 4.00 Table 2 Continued Allele Frequency of allele 1 Fisher test`s P-values Gene name SNP ID Position of SNP/functional SNP 1 2 dCase eControl f1vs2 g11vs h22vs Odds ratio ABCB1 rs17327442 Intron 5 T A 0.89 0.94 5.12EÀ01 4.93EÀ01 1.00E+00 1.87 ABCB1 rs1202184 Intron 5 G A 0.24 0.33 3.44EÀ01 1.00E+00 2.25EÀ01 1.91 ABCB1 rs3789243 Intron 4 T C 0.50 0.31 2.81EÀ02 5.05EÀ02 1.38EÀ01 3.50 ABCB1 rs3213619 Exon 2 (5' UTR) C T 0.02 0.10 1.81EÀ01 1.00E+00 1.63EÀ01 5.02 ABCB1 rs4148732 Intron 1 A G 0.87 0.96 7.89EÀ02 6.87EÀ02 1.00E+00 3.67 ABCB1 rs13233308 Intron 1 T C 0.43 0.44 1.00E+00 7.60EÀ01 7.95EÀ01 0.82 ABCB1 rs1978095 Intron 1 T C 0.67 0.73 5.59EÀ01 6.21EÀ01 6.89EÀ01 1.36 ABCB1 rs2157929 Intron 1 T C 0.93 0.79 3.32EÀ02 5.60EÀ02 5.51EÀ01 3.81 ABCB1 rs10278483 Intron 1 T C 0.96 0.86 1.01EÀ01 1.36EÀ01 5.88EÀ01 3.34 CYP3A5 rs776746 Intron 3 (CYP3A5*3) A G 0.22 0.32 2.50EÀ01 6.67EÀ01 3.26EÀ01 1.79 CYP3A4 rs28371759c Exon 10 (Pro 293 Leu) (CYP3A4*18) 1.00 1.00 1.00E+00 1.00E+00 1.00E+00 NA CYP3A4 rs12721627b Exon 7 (Ser 185 Thr) (CYP3A4*16) C G 1.00 0.96 3.23EÀ01 3.22EÀ01 1.00E+00 NA ABCC2 rs12268782 5' near gene A G 0.22 0.13 2.26EÀ01 1.00E+00 1.81EÀ01 2.21 ABCC2 rs2804398 Intron 7 T A 0.83 0.84 8.14EÀ01 7.90EÀ01 1.00E+00 1.29 ABCC2 rs2756109 Intron 7 G T 0.63 0.65 8.55EÀ01 1.00E+00 1.00E+00 1.12 ABCC2 rs2273697 Exon 10 (Ile 417 Val) G A 0.83 0.92 8.91EÀ02 5.96EÀ02 1.00E+00 3.33 ABCC2 rs11190291a Intron 11 T C 0.17 0.08 8.91EÀ02 1.00E+00 5.96EÀ02 3.33 ABCC2 rs2002042 Intron 19 T C 0.33 0.39 4.78EÀ01 2.78EÀ01 1.00E+00 3.52 ABCC2 rs17222723c Exon 25 (Glu 1188 Val) T A 1.00 1.00 1.00E+00 1.00E+00 1.00E+00 NA ABCC2 rs3740065b Intron 29 T C 0.65 0.60 5.96EÀ01 6.18EÀ01 9.90EÀ02 6.36 ABCC2 rs12762549 3' near gene C G 0.48 0.36 2.13EÀ01 1.00E+00 6.81EÀ02 3.35 ABCC2 rs2862691 3' near gene T C 0.19 0.25 5.26EÀ01 6.05EÀ01 6.11EÀ01 NA ABCC2 rs11598781 3' near gene T C 0.17 0.27 2.30EÀ01 1.00E+00 1.40EÀ01 2.29 ABCC2 rs11190303 3' near gene T C 0.24 0.33 2.62EÀ01 5.33EÀ02 1.00E+00 NA SLCO1B3 rs12228798 Intron 1 C T 0.82 0.83 1.00E+00 7.77EÀ01 2.27EÀ01 NA SLCO1B3 rs7977213 Intron 2 G C 0.43 0.18 1.29EÀ03 1.31EÀ03 2.55EÀ02 NA SLCO1B3 rs10841661 Intron 2 T C 0.48 0.23 2.73EÀ03 5.65EÀ03 4.79EÀ02 9.88 SLCO1B3 rs4149118 Intron 4 A G 0.59 0.48 3.31EÀ01 3.49EÀ01 5.40EÀ01 1.85 SLCO1B3 rs3764009a Intron 4 A G 0.68 0.69 1.00E+00 1.00E+00 1.00E+00 1.16 SLCO1B3 rs7311358 Exon 7 (Ile 233 Met) A G 0.68 0.69 1.00E+00 1.00E+00 1.00E+00 1.16 SLCO1B3 rs11045585b Intron 12 G A 0.13 0.21 2.76EÀ01 3.14EÀ01 6.06EÀ01 NA SLCO1B3 rs980084 Intron 12 G C 0.28 0.45 7.48EÀ02 3.80EÀ01 7.66EÀ02 2.67 SLCO1B3 rs3764006 Exon 14 (Gly 611 Gly) T C 0.87 0.70 2.74EÀ02 1.32EÀ01 9.75EÀ02 NA SLCO1B3 rs919840 3' near gene C G 0.97 0.90 2.99EÀ01 2.77EÀ01 1.00E+00 3.74 SLCO1B3 rs2117032 3' near gene T C 0.52 0.47 6.05EÀ01 7.68EÀ01 7.82EÀ01 1.35 SLCO1B3 rs7312051 3' near gene C T 1.00 0.91 1.26EÀ01 1.04EÀ01 1.00E+00 NA SLCO1B3 rs10841714 3' near gene C T 0.15 0.17 8.01EÀ01 1.00E+00 1.00E+00 NA SLCO1B3 rs2174012 3' near gene T C 0.62 0.63 1.00E+00 5.72EÀ01 4.55EÀ01 0.39 SLCO1B3 rs11045639a 3' near gene G A 0.65 0.72 4.46EÀ01 2.17EÀ01 7.25EÀ01 2.05 SLCO1B3 rs2900459 3' near gene G A 0.46 0.56 2.95EÀ01 1.43EÀ02 5.70EÀ01 5.96 SLCO1B3 rs4762693 3' near gene G A 0.65 0.72 4.46EÀ01 2.17EÀ01 7.25EÀ01 2.05 SLCO1B3 rs10734711 3' near gene A G 0.28 0.35 4.61EÀ01 2.78EÀ01 8.05EÀ01 3.52 SLCO1B1 rs12228427 5' near gene (LST-3TM12 Intron 9) A G 0.96 0.88 1.59EÀ01 1.36EÀ01 1.00E+00 3.34 SLCO1B1 rs1910163 5' near gene (LST-3TM12 Intron 12) A T 0.28 0.34 4.66EÀ01 7.25EÀ01 6.21EÀ01 1.44 SLCO1B1 rs6487207 5' near gene (LST-3TM12 Intron 11) T G 0.78 0.73 5.54EÀ01 8.11EÀ01 3.22EÀ01 NA SLCO1B1 rs1604539 5' near gene (LST-3TM12 Intron 12) T G 0.80 0.75 5.41EÀ01 6.18EÀ01 1.00E+00 1.42 SLCO1B1 rs7973691 5' near gene T C 0.80 0.83 8.21EÀ01 7.98EÀ01 1.00E+00 1.22 SLCO1B1 rs10743408 Intron 2 C G 0.24 0.18 4.97EÀ01 1.00E+00 3.06EÀ01 1.73 SLCO1B1 rs976754 Intron 2 T C 0.83 0.69 1.15EÀ01 8.67EÀ02 6.67EÀ01 2.54 SLCO1B1 rs2291073 Intron 3 T G 0.78 0.72 4.35EÀ01 8.11EÀ01 1.76EÀ01 NA SLCO1B1 rs4149037 Intron 4 A G 0.70 0.80 2.12EÀ01 2.07EÀ01 6.19EÀ01 2.07 SLCO1B1 rs4149056 Exon 5 (Ala 174 Val) T C 0.72 0.84 8.02EÀ02 2.04EÀ01 8.01EÀ02 NA SLCO1B1 rs2417967 Intron 11 A G 0.33 0.21 1.54EÀ01 1.00E+00 8.68EÀ02 2.41 SLCO1B1 rs7969341b Intron 14 T C 0.50 0.46 7.24EÀ01 1.58EÀ01 5.60EÀ02 3.80 SLCO1B1 rs4149085 3' UTR T C 0.74 0.66 4.53EÀ01 8.05EÀ01 1.76EÀ01 NA SLCO1B1 rs12372067 3' near gene C A 0.21 0.33 2.06EÀ01 1.00E+00 1.68EÀ01 2.42 SLCO1B1 rs12310063 3' near gene A C 0.67 0.76 3.21EÀ01 2.13EÀ01 1.00E+00 2.07 ABCC1 rs8050881 5' near gene A G 0.33 0.23 2.37EÀ01 1.00E+00 1.44EÀ01 2.13 ABCC1 rs4148330 5' near gene G A 0.48 0.34 1.46EÀ01 2.95EÀ01 2.13EÀ01 2.02 Continued Allele Frequency of allele 1 Fisher test`s P-values Gene name SNP ID Position of SNP/functional SNP 1 2 dCase eControl f1vs2 g11vs h22vs Odds ratio ABCC1 rs7190484 Intron 1 T C 0.43 0.34 3.67EÀ01 5.03EÀ01 4.62EÀ01 1.52 ABCC1 rs215098 Intron 1 C A 0.41 0.34 4.63EÀ01 1.96EÀ01 1.00E+00 2.30 ABCC1 rs215096 Intron 1 T C 0.89 0.87 1.00E+00 5.63EÀ01 2.89EÀ01 0.00 ABCC1 rs152023 Intron 1 G A 0.48 0.41 4.75EÀ01 3.79EÀ01 8.05EÀ01 1.79 ABCC1 rs6498595 Intron 1 G C 0.41 0.31 2.70EÀ01 2.73EÀ02 1.00E+00 4.76 ABCC1 rs7196970 Intron 1 C G 0.70 0.63 4.71EÀ01 3.25EÀ01 1.00E+00 1.73 ABCC1 rs12935283 Intron 1 A G 0.67 0.61 4.78EÀ01 3.19EÀ01 1.00E+00 1.79 ABCC1 rs246240 Intron 5 A G 0.60 0.58 1.00E+00 1.00E+00 1.00E+00 1.06 ABCC1 rs924138b Intron 5 T C 0.68 0.63 5.84EÀ01 6.17EÀ01 1.00E+00 1.38 ABCC1 rs2062541 Intron 6 C T 0.71 0.64 4.46EÀ01 4.45EÀ01 7.51EÀ01 1.60 ABCC1 rs11647513 Intron 6 C T 0.78 0.70 3.34EÀ01 2.14EÀ01 1.00E+00 2.13 ABCC1 rs35593 Intron 11 T C 0.71 0.75 8.37EÀ01 1.00E+00 1.00E+00 2.70 ABCC1 rs3765129 Intron 11 C T 0.98 0.84 2.69EÀ02 1.75EÀ02 1.00E+00 9.90 ABCC1 rs17287570 Intron 12 A C 0.73 0.91 6.32EÀ03 1.03EÀ02 2.75EÀ01 4.27 ABCC1 rs35597 Intron 12 A G 0.59 0.55 7.19EÀ01 5.85EÀ01 1.00E+00 1.45 ABCC1 rs35598a Intron 12 A G 0.82 0.86 6.21EÀ01 7.86EÀ01 2.78EÀ01 NA ABCC1 rs9932506 Intron 12 G A 0.72 0.75 6.94EÀ01 6.21EÀ01 1.00E+00 1.44 ABCC1 rs35604 Intron 12 G A 0.24 0.25 1.00E+00 1.00E+00 1.00E+00 1.63 ABCC1 rs35606 Intron 13 C T 0.82 0.86 6.17EÀ01 7.82EÀ01 2.86EÀ01 NA ABCC1 rs35620 Intron 14 G C 0.26 0.23 8.39EÀ01 1.00E+00 6.19EÀ01 1.39 ABCC1 rs35625 Intron 14 C T 0.41 0.34 4.70EÀ01 7.52EÀ01 4.72EÀ01 1.45 ABCC1 rs35626 Intron 15 T G 0.43 0.40 7.21EÀ01 5.08EÀ01 2.96EÀ01 1.93 ABCC1 rs4148353 Intron 15 T G 0.20 0.07 2.42EÀ02 1.00E+00 1.69EÀ02 4.02 ABCC1 rs35629 Intron 15 C T 0.80 0.76 5.46EÀ01 6.24EÀ01 1.00E+00 1.32 ABCC1 rs2269800 Intron 20 A G 0.74 0.66 4.53EÀ01 8.06EÀ01 2.78EÀ01 3.52 ABCC1 rs11864374 Intron 21 G A 0.78 0.77 8.41EÀ01 1.00E+00 1.00E+00 1.63 ABCC1 rs3887893 Intron 22 A G 0.48 0.46 8.62EÀ01 2.30EÀ01 5.85EÀ01 2.10 ABCC1 rs4148376 Intron 23 A G 0.93 0.95 1.00E+00 1.00E+00 1.00E+00 1.30 ABCC1 rs2238475 Intron 23 C A 0.11 0.18 3.45EÀ01 1.00E+00 4.21EÀ01 1.80 ABCC1 rs212079 Intron 26 G A 0.76 0.83 3.77EÀ01 1.00E+00 6.71EÀ02 8.55 ABCC1 rs2283512 Intron 26 G T 0.48 0.46 1.00E+00 7.68EÀ01 1.00E+00 1.32 ABCC1 rs212081 Intron 27 T C 0.20 0.22 8.34EÀ01 5.73EÀ01 4.61EÀ01 1.50 ABCC1 rs212084 Intron 28 G A 0.68 0.77 2.90EÀ01 3.02EÀ01 6.00EÀ01 1.81 ABCC1 rs212087 Intron 28 C T 0.57 0.70 1.41EÀ01 1.38EÀ01 3.45EÀ01 2.30 ABCC1 rs4148380 3' UTR G A 0.87 0.89 7.88EÀ01 7.75EÀ01 1.00E+00 1.22 ABCC1 rs212091 3' UTR G A 0.22 0.34 1.84EÀ01 2.68EÀ01 3.35EÀ01 4.04 ABCC1 rs12448760 3' near gene G A 0.82 0.89 2.95EÀ01 3.79EÀ01 1.00E+00 1.79 ABCC1 rs9932935 3' near gene (ABCC6 Intron 29) A T 0.96 0.91 5.12EÀ01 5.00EÀ01 1.00E+00 1.93 ABCC1 rs2066738 3' near gene (ABCC6 Intron 28) C T 0.74 0.90 1.50EÀ02 5.28EÀ02 7.81EÀ02 2.95 ABCC1 rs169845 3' near gene (ABCC6 Intron 27) G C 0.26 0.40 1.04EÀ01 3.28EÀ01 2.13EÀ01 2.07 ABCC1 rs2238471 3' near gene (ABCC6 Intron 26) A C 0.93 0.81 5.49EÀ02 5.79EÀ02 1.00E+00 3.78 ABCC1 rs3213471 3' near gene (ABCC6 Intron 24) A G 0.91 0.92 1.00E+00 7.33EÀ01 1.00E+00 1.32 ABCC1 rs3213473 3' near gene (ABCC6 Intron 24) T G 0.16 0.13 7.98EÀ01 1.00E+00 7.71EÀ01 1.27 CES2 rs3843712 5' near gene G A 0.18 0.12 4.41EÀ01 1.00E+00 4.00EÀ01 1.80 CES2 rs8062110 5' near gene G C 0.61 0.65 7.16EÀ01 1.00E+00 7.28EÀ01 1.39 CES2 rs4783744 5' near gene G A 0.66 0.62 8.37EÀ01 7.76EÀ01 1.00E+00 1.24 CES2 rs7194513 5' near gene G A 0.26 0.27 1.00E+00 3.14EÀ01 6.29EÀ01 NA CES2 rs28382812c Exon 1 (Ile 37 Ile) C T 1.00 1.00 1.00E+00 1.00E+00 1.00E+00 NA CES2 rs2241410 Intron 2 T G 0.10 0.11 7.85EÀ01 1.00E+00 7.71EÀ01 1.26 CES2 rs2303218 Intron 2 G A 0.09 0.21 1.03EÀ01 5.55EÀ01 1.15EÀ01 2.70 CES2 rs8192924c Exon 5 (His 270 Arg) G A 1.00 1.00 1.00E+00 1.00E+00 1.00E+00 NA CES2 rs28382827c Exon 12 (Leu 613 Leu) C T 1.00 1.00 1.00E+00 1.00E+00 1.00E+00 NA Abbreviations: Chrom, chromosome; HWE, Hardy-Weinberg equilibrium; NA, not available; SNP, single nucleotide polymorphism. 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[hide] Additive effects of drug transporter genetic polym... Cancer Chemother Pharmacol. 2010 May;66(1):95-105. Epub 2009 Sep 22. Sai K, Saito Y, Maekawa K, Kim SR, Kaniwa N, Nishimaki-Mogami T, Sawada J, Shirao K, Hamaguchi T, Yamamoto N, Kunitoh H, Ohe Y, Yamada Y, Tamura T, Yoshida T, Matsumura Y, Ohtsu A, Saijo N, Minami H
Additive effects of drug transporter genetic polymorphisms on irinotecan pharmacokinetics/pharmacodynamics in Japanese cancer patients.
Cancer Chemother Pharmacol. 2010 May;66(1):95-105. Epub 2009 Sep 22., [PMID:19771428]

Abstract [show]
PURPOSE: Effects of genetic polymorphisms/variations of ABCB1, ABCC2, ABCG2 and SLCO1B1 in addition to "UGT1A1*28 or *6" on irinotecan pharmacokinetics/pharmacodynamics in Japanese cancer patients were investigated. METHODS: Associations between transporter haplotypes/variations along with UGT1A1*28 or *6 and SN-38 area under the time-concentration curve (AUC) or neutropenia were examined in irinotecan monotherapy (55 patients) and irinotecan-cisplatin-combination therapy (62 patients). RESULTS: Higher SN-38 AUC values were observed in ABCB1 2677G>T (A893S) (*2 group) for both regimens. Associations of grade 3/4 neutropenia were observed with ABCC2 -1774delG (*1A), ABCG2 421C>A (Q141K) and IVS12 + 49G>T ((#) IIB) and SLCO1B1 521T>C (V174A) (*15 x 17) in the irinotecan monotherapy, while they were evident only in homozygotes of ABCB1*2, ABCG2 (#) IIB, SLCO1B1*15 x 17 in the cisplatin-combination therapy. With combinations of haplotypes/variations of two or more genes, neutropenia incidence increased, but their prediction power for grade 3/4 neutropenia is still unsatisfactory. CONCLUSIONS: Certain transporter genotypes additively increased irinotecan-induced neutropenia, but their clinical importance should be further elucidated.

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3 Associations of grade 3/4 neutropenia were observed with ABCC2 -1774delG (*1A), ABCG2 421C[A (Q141K) and IVS12 ? Login to comment
40 Since the *9 haplotype with 1236C[T, 2677G[T (A893S) without 3435C[T [16] showed the same trend for PK/PD as *2 (data not shown), the current study classified the Table 1 List of major transporter haplotypes and their markers analyzed for Japanese cancer patients Gene Haplotype Tagging SNP Abbreviation used in this paper Haplotype frequency Monotherapy (N = 110)a With cisplatin (N = 124)a ABCB1 BJLb (block 1) -1789G[A 0.182 0.210 *2 groupc (block 2) 2677G[T(A893S) B 0.382 0.379 *10 groupd (block 2) 2677G[A(A893T) 0.182 0.169 *1b (block 3) IVS27-182G[T 0.200 0.169 ABCC2 *1A -1774delG C 0.373 0.371 *1C/G 3972C[T(I1324I) 0.218 0.266 ABCG2 # IIB [*1a-*2-*1b]e 421C[A(Q141K), IVS12 ? Login to comment
48 Major combinations in 177 patients were the wild type # IA (frequency = 0.291), # IIB [containing 421C[A (Q141K) and IVS12 ? Login to comment
50 Note that # IIB and # IIIC are subgroups of block 1 *2 [421C[A (Q141K)] and block 1*3 [34G[A (V12M)], respectively [28]. Login to comment
86 = UGT1A1*6 or *28. a BJL contains -1789G[A, *2 (block 1) = 325G[A (E109K), *3 (block 1) = 304G[A (G102R); b *2 (block 2) contains 2677G[T (A893S); c *1b (block 3) = IVS27-182G[T, *2 (block 3) = 3751G[ A (V1251I); d *1A contains -1774delG; e IIB contains 421C[A (Q141K) and IVS12 ? Login to comment
150 ABCG2# IIB contains the non-synonymous SNP 421C[A (Q141K), which was detected at higher frequencies in Asians and was reported to cause reduced expression of BCRP in vitro [36, 39-41]. Login to comment
151 In clinical studies, the association of 421C[A (Q141K) with higher plasma levels of diflomotecan was shown in Caucasians [42]. Login to comment
153 An association of 421C[A (Q141K) alone with irinotecan PK/PD was not significant in our hands (data not shown), but # IIB containing both 421C[A (Q141K) and IVS12 ? Login to comment

[hide] Drug transporters in the human blood-placental bar... Br J Pharmacol. 2009 Oct;158(3):665-78. Epub 2009 Sep 25. Vahakangas K, Myllynen P
Drug transporters in the human blood-placental barrier.
Br J Pharmacol. 2009 Oct;158(3):665-78. Epub 2009 Sep 25., [PMID:19788499]

Abstract [show]
Studies on the increasing number of transporters found in the placental barrier are gaining momentum, because of their tissue-specific expression, significance in physiology and disease, and the possible utilization of the emerging knowledge in pharmacology. In the placenta, both syncytiotrophoblast and fetal capillary endothelium express transporters. Fetal exposure is determined by the net effect of combination of transporters, their nature and localization in relation to placental cells and their substrate specificity. Although the significance of placental transporters on human fetal drug exposure is almost an unstudied field so far, their potential use to design drugs that do not cross the placenta is already being pursued. It is thus of interest to review the existing knowledge of human placental transporters. Transporters in all groups which take part in drug transport are found in human placenta. Especially, ATP-binding cassette transporters ABCG2/breast cancer resistance protein, ABCB1/P-glycoprotein and ABCC2/MRP2 are all expressed at the apical surface of syncytiotrophoblast facing maternal blood and are putatively important protective proteins both for placental tissue and the fetus, because they are efflux transporters and their substrates include many drugs and also environmental chemicals. Such protective effect has been shown in animals, but these results cannot be directly extrapolated to humans due to interspecies differences in placental structure and function. Experimental models utilizing human placental tissue, especially human placental perfusion, offer valuable possibilities, which have been insufficiently studied so far.

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99 Table 4 Significance of drug transporter polymorphisms in human placenta Protein Gene polymorphisms Type of study Significance in placenta Reference P-gp/MDR1 G2677A/T (Ala toThr/Ser) T-129C 100 human placentas Less P-gp protein Less P-gp protein 1 P-gp/MDR1 G2677T/A C3435T 73 human placentas from Caucasians No effect on MDR1 mRNA Homozygosity of 3435T and 2677T lead to lower protein levels 2 P-gp/MDR1 C3435T 44 human placentas T allele associated with a higher expression 3 P-gp/MDR1 C3435T and G2677A/T Human placental perfusion No effect on saquinavir transfer 3, 4 P-gp/MDR1 C3435T Human placental perfusion 3435T associated with increased transfer of quetiapine 5 MRP2 G1249A 58 human placentas Reduced expression of MRP2 mRNA in preterm placentas only 6 BCRP G34A (Val12Met) C421A (Gln141Lys) 99 human placentas No effect on protein level Protein decreased 7 References: (1) Tanabe et al. (2001), (2) Hitzl et al. (2004), (3) Rahi et al. (2008), (4) Mölsä et al. (2005), (5) Rahi et al. (2007), (6) Meyer zu Schwabedissen et al. (2005b), (7) Kobayashi et al. (2005). Login to comment
159 At least one non-synonymous polymorphism (C421A; Gln141Lys) seems to lead to a lower expression of ABCG2/BCRP protein both in vitro (Imai et al., 2002) and in human placenta (Kobayashi et al., 2005). Login to comment

[hide] Human ABC transporter ABCG2 in cancer chemotherapy... J Exp Ther Oncol. 2009;8(1):5-24. Ishikawa T, Nakagawa H
Human ABC transporter ABCG2 in cancer chemotherapy and pharmacogenomics.
J Exp Ther Oncol. 2009;8(1):5-24., [PMID:19827267]

Abstract [show]
The ability of cancer cells to acquire resistance to multiple anticancer agents, termed multidrug resistance, is often mediated by overexpression of ATP-binding cassette (ABC) transporters that remove drugs out of the cell against a concentration gradient. ABCG2, or breast cancer resistance protein (BCRP), is an ABC transporter that has been the subject of intense study since its discovery a decade ago. While ABCG2 overexpression has been demonstrated in cancer cells after in vitro drug treatment, endogenous ABCG2 expression in certain cancers is considered as a reflection of the differentiated phenotype of the cell of origin and likely contributes to intrinsic drug resistance. Notably, ABCG2 is often expressed in stem cell populations, where it plays a critical role in cellular protection. ABCG2 exhibits a broad range of substrate specificity. New technologies of high-speed screening and quantitative structure-activity-relationship (QSAR) analysis have been developed to analyze the interactions of drugs with ABCG2. As ABCG2 reportedly transports porphyrins, its contribution to photodynamic therapy of human cancer is also implicated. Protein expression levels of ABCG2 in cancer cells are regulated by both transcriptional activation and protein degradation. The ABCG2 protein undergoes endosomal and/or ubiquitin-mediated proteasomal degradations. Furthermore, genetic polymorphisms in the ABCG2 gene are important factors in cancer chemotherapy to circumvent adverse effects and/or to enhance the efficacy of anticancer drugs. The present review article addresses recent advances in molecular pharmacology and pharmacogenomics of ABCG2 and provides novelideas to improve cancer chemotherapy.

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219 The most extensively studied among those SNPs with potential clinical relevance is 421 C>A resulting in a glutamic acid to lysine substitution (Q141K) in the ABCG2 protein. Login to comment
220 The Q141K SNP has been identified with varying frequencies in different ethnic groups and was found to be the most relevant in Japanese and Chinese populations (approximately 30% in the allele frequency). Login to comment
222 COOH H2N N590Y V12M G51C Q126stop Q141K T153M Q166E I206L F208S S248P E334stop F431L F489L D620N R482G R482T S441N F571I OUT IN R160Q R575stop ATP-binding site Figure 7. Continued A 005-024 pp JETO-0900616-TI (Review).indd 8/7/2009 3:59:50 19 Q141K has been associated with lower levels of protein expression and impaired transport in vitro (Imai et al., 2002; Kobayashi et al., 2005; Misuarai et al., 2004; Zamber et al., 2003; Morisaki et al., 2008; Kondo et al., 2004). Login to comment
224 Furthermore, the Q141K SNP was reportedly associated with a higher incidence of diarrhea in non-small cell lung cancer patients treated with gefitinib (Cusatis et al., 2006). Login to comment
225 It has been demonstrated that the reduced expression levels of the Q141K variant may be due to its ubiquitin-mediated proteasomal degradation (Furukawa et al., 2009). Login to comment
228 Furthermore, the F208S, S248P, F431L, S441N, and F489L variants exhibited greatly altered protein expression levels and drug resistance profiles Figure 7. Continued WT V12M Q141K F208S S248P F431L S441N F489L R482G R482T Protein expression + + + - + + - + + + MTX transport + + + - - - - +/ - - Porphyrin transport + + + - - + - +/ + + SN-38 resistance + + + - +/ + - - + + MX resistance + + + - - - - - -- - - - - - - - +/ - - - - - - - - + + Doxorubicin resistance + + Daunorubicin resistance + + ATPase activity (Prazosin) + + WTV12M Q141K F431L F489L S248P F208S S441L R482G R482T ∆1.5 ∆3 ∆3.5 ∆5 ∆4 - - - - - - -- - - B 005-024 pp JETO-0900616-TI (Review).indd 8/7/2009 3:59:51 20 Journal of Experimental Therapeutics and Oncology Vol. 8 2009 (Tamura et al., 2007b). Login to comment
232 It is known that, in the ER, the N-linked glycans play pivotal roles in protein fold- 0.0 0.5 1.0 1.5 Mock WT V12M G51C Q126stop Q141K T153M Q166E I206L F208S S248P E334stop F431L S441N F489L F571I N590Y D620N R482G R482T Methotrexatetransport (nmol/min/mgprotein) Methotrexate 0.0 0.5 1.0 1.5 0.0 0.5 1.0 1.5 Mock WT V12M G51C Q126stop Q141K T153M Q166E I206L F208S S248P E334stop F431L S441N F489L F571I N590Y D620N R482G R482T Methotrexatetransport (nmol/min/mgprotein) MethotrexateMethotrexate Porphyrintransport (nmol/min/mgprotein) 0.0 0.1 0.2 0.3 0.4 0.5 0.0 0.1 0.2 0.3 0.4 0.5 Porphyrin Figure 7. Login to comment
251 The new camptothecin analogues that were non-substrates for ABCG2 circumvented ABCG2-mediated drug resistance without any influence from major SNPs, i.e., V12M and Q141K (Tamura et al. 2007b). Login to comment

[hide] Pharmacogenetics of drug transporters. Curr Pharm Des. 2010;16(2):220-30. Franke RM, Gardner ER, Sparreboom A
Pharmacogenetics of drug transporters.
Curr Pharm Des. 2010;16(2):220-30., [PMID:19835554]

Abstract [show]
During the last decade, a greater focus has been given to impact of genetic variation in membrane transporters on the pharmacokinetics and toxicity of numerous therapeutic drugs. While the majority of transporter-related pharmacogenetic research has been in regards to classic genes encoding the outward-directed ATP-binding cassette (ABC) transporters, such as ABCB1 (P-glycoprotein), ABCC2 (MRP2), and ABCG2 (BCRP), more studies have been conducted in recent years evaluating genes encoding solute carriers (SLC) that mediate the cellular uptake of drugs, such as SLCO1B1 (OATP1B1) and SLC22A1 (OCT1). The distribution of ABC and SLC transporters in tissues key to pharmacokinetics, such as intestine (absorption), blood-brain-barrier (distribution), liver (metabolism), and kidneys (excretion), strongly suggests that genetic variation associated with changes in protein expression or function of these transporters may have a substantial impact on systemic drug exposure and toxicity. In this current article, we will review recent advances in understanding the contribution of critical ABC and SLC transporters to interindividual pharmacokinetic and dynamic variability of substrate drugs.

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102 In particular, a SNP in exon 5 of the ABCG2 gene has been described, in which a 421C>A transversion results in a lysine to glutamine amino acid change at codon 141 (Q141K) [68]. Login to comment
108 Specifically, recent studies have demonstrated that subjects with a reduced ABCG2 activity due to the Q141K variant are at an increased risk for gefitinib-induced diarrhea [72], and altered pharmacokinetics of 9-aminocamptothecin, diflomotecan, irinotecan, rosuvastatin, sulfasalazine, and topotecan [64]. Login to comment

[hide] Different effects of the ABCG2 c.421C>A SNP on the... Pharmacogenomics. 2009 Oct;10(10):1617-24. Keskitalo JE, Pasanen MK, Neuvonen PJ, Niemi M
Different effects of the ABCG2 c.421C>A SNP on the pharmacokinetics of fluvastatin, pravastatin and simvastatin.
Pharmacogenomics. 2009 Oct;10(10):1617-24., [PMID:19842935]

Abstract [show]
AIMS: This study aimed to investigate possible effects of the ABCG2 c.421C>A (p.Gln141Lys; rs2231142) genotype on fluvastatin, pravastatin and simvastatin pharmacokinetics. MATERIALS & METHODS: In a crossover study, five healthy volunteers with the ABCG2 c.421A/A genotype, four with the c.421C/A genotype and 23 with the c.421C/C genotype ingested a single 40-mg dose of fluvastatin, pravastatin and simvastatin, with a washout period of 1 week. Plasma statin concentrations were measured up to 12 h. RESULTS: The estimated marginal mean area under the plasma concentration-time curve from 0 h to infinity (AUC(0-infinity)) of fluvastatin was 97% (p = 0.015) or 72% (p = 0.009) larger in participants with the A/A genotype than in those with the C/A or C/C genotype. The AUC(0-infinity) of simvastatin lactone was 111% (p = 0.005) larger in participants with the A/A genotype than in participants with the C/C genotype. The simvastatin acid:lactone AUC(0-infinity) ratio was 46% (p = 0.017) smaller in individuals with the A/A genotype than in those with the C/C genotype. The ABCG2 genotype had no significant effect on simvastatin acid or pravastatin pharmacokinetics. CONCLUSIONS: Genetic variability in ABCG2 markedly affects the pharmacokinetics of fluvastatin and simvastatin lactone, but has no significant effect on pravastatin or active simvastatin acid. Genotyping for ABCG2 in addition to SLCO1B1 and ABCB1 polymorphisms could help in predicting statin pharmacokinetics when selecting a statin and its dose for an individual patient.

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160 7 Imai Y, Nakane M, Kage K et al.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment
9 One of the most studied variations, c.421C>A (p.Gln141Lys; rs2231142), has been associated with reduced transport activity of ABCG2 in vitro [4-10]. Login to comment
17 The participants Aims: This study aimed to investigate possible effects of the ABCG2 c.421C>A (p.Gln141Lys; rs2231142) genotype on fluvastatin, pravastatin and simvastatin pharmacokinetics. Login to comment
161 7 Imai Y, Nakane M, Kage K etal.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Common polymorphisms influencing serum uric acid l... PLoS One. 2009 Nov 5;4(11):e7729. Stark K, Reinhard W, Grassl M, Erdmann J, Schunkert H, Illig T, Hengstenberg C
Common polymorphisms influencing serum uric acid levels contribute to susceptibility to gout, but not to coronary artery disease.
PLoS One. 2009 Nov 5;4(11):e7729., [PMID:19890391]

Abstract [show]
BACKGROUND: Recently, a large meta-analysis including over 28,000 participants identified nine different loci with association to serum uric acid (UA) levels. Since elevated serum UA levels potentially cause gout and are a possible risk factor for coronary artery disease (CAD) and myocardial infarction (MI), we performed two large case-control association analyses with participants from the German MI Family Study. In the first study, we assessed the association of the qualitative trait gout and ten single nucleotide polymorphisms (SNP) markers that showed association to UA serum levels. In the second study, the same genetic polymorphisms were analyzed for association with CAD. METHODS AND FINDINGS: A total of 683 patients suffering from gout and 1,563 healthy controls from the German MI Family Study were genotyped. Nine SNPs were identified from a recently performed genome-wide meta-analysis on serum UA levels (rs12129861, rs780094, rs734553, rs2231142, rs742132, rs1183201, rs12356193, rs17300741 and rs505802). Additionally, the marker rs6855911 was included which has been associated with gout in our cohort in a previous study. SNPs rs734553 and rs6855911, located in SLC2A9, and SNP rs2231142, known to be a missense polymorphism in ABCG2, were associated with gout (p=5.6*10(-7), p=1.1*10(-7), and p=1.3*10(-3), respectively). Other SNPs in the genes PDZK1, GCKR, LRRC16A, SLC17A1-SLC17A3, SLC16A9, SLC22A11 and SLC22A12 failed the significance level. None of the ten markers were associated with risk to CAD in our study sample of 1,473 CAD cases and 1,241 CAD-free controls. CONCLUSION: SNP markers in SLC2A9 and ABCG2 genes were found to be strongly associated with the phenotype gout. However, not all SNP markers influencing serum UA levels were also directly associated with the clinical manifestation of gout in our study sample. In addition, none of these SNPs showed association with the risk to CAD in the German MI Family Study.

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115 SNP Position a Major allele (1) Minor allele (2) Gene name(s) Function Call rate b rs12129861 Chr1: 144,437,046 G A PDZK1 59 Intergenic 98.4% rs780094 Chr2: 27,594,741 C T GCKR Intron 16 99.2% rs734553 Chr4: 9,532,102 T G SLC2A9 GLUT9 Intron 7 96.0% rs6855911 Chr4: 9,545,008 A G SLC2A9 GLUT9 Intron 7 99.0% rs2231142 Chr4: 89,271,347 G T ABCG2 Exon 5 Q141K 99.2% rs742132 Chr6: 25,715,550 A G LRRC16A Intron 34 99.4% rs1183201 Chr6: 25,931,423 T A SLC17A1 Intron 3 98.2% rs12356193 Chr10: 61,083,359 A G SLC16A9 Intron 5 98.7% rs17300741 Chr11: 64,088,038 G A SLC22A11 Intron 4 98.2% rs505802 Chr11: 64,113,648 T C SLC22A12 URAT1 59 Intergenic 99.3% a on human genome build 18. b in total sample (n = 4,960). Login to comment
120 This is the only marker examined that leads to a missense mutation with an amino acid exchange from glutamine to lysine at position 141 in ABCG2 transporter and therefore could have a direct and causal influence on development of the disease [6]. Login to comment

[hide] Disruption of N-linked glycosylation enhances ubiq... FEBS J. 2009 Dec;276(24):7237-52. Epub . Nakagawa H, Wakabayashi-Nakao K, Tamura A, Toyoda Y, Koshiba S, Ishikawa T
Disruption of N-linked glycosylation enhances ubiquitin-mediated proteasomal degradation of the human ATP-binding cassette transporter ABCG2.
FEBS J. 2009 Dec;276(24):7237-52. Epub ., [PMID:19909340]

Abstract [show]
The human ATP-binding cassette (ABC) transporter, ABCG2 (BCRP/MXR/ABCP), is a plasma membrane protein containing intramolecular and intermolecular disulfide bonds and an N-linked glycan at Asn596. We have recently reported that the intramolecular disulfide bond is a critical checkpoint for determining the degradation fates of ABCG2. In the present study, we aimed to analyze quantitatively the impact of the N-linked glycan on the protein stability of ABCG2. For this purpose, we incorporated one single copy of ABCG2 cDNA into a designated site of genomic DNA in Flp-In-293 cells to stably express ABCG2 or its variant proteins. When ABCG2 wild type-expressing cells were incubated with various N-linked glycosylation inhibitors, tunicamycin profoundly suppressed the protein expression level of ABCG2 and, accordingly, reduced the ABCG2-mediated cellular resistance to the cancer chemotherapeutic SN-38. When Asn596 was converted to Gln596, the resulting variant protein was not glycosylated, and its protein level was about one-third of the wild type level in Flp-In-293 cells. Treatment with MG132, a proteasome inhibitor, increased the level of the variant protein. Immunoblotting with anti-ubiquitin IgG1k after immunoprecipitation of ABCG2 revealed that the N596Q protein was ubiquitinated at levels that were significantly enhanced by treatment with MG132. Immunofluorescence microscopy demonstrated that treatment with MG132 increased the level of ABCG2 N596Q protein both in intracellular compartments and in the plasma membrane. In conclusion, we propose that the N-linked glycan at Asn596 is important for stabilizing de novo-synthesized ABCG2 and that disruption of this linkage results in protein destabilization and enhanced ubiquitin-mediated proteasomal degradation.

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22 In fact, the protein expression levels of ABCG2 SNP variants (Q141K, F208S and S441N) were significantly lower than that of the wild-type (WT) ABCG2 [23]. Login to comment

[hide] Genetic analysis of ABCG2 gene C421A polymorphism ... Hum Genet. 2010 Feb;127(2):245-6. Epub 2009 Nov 17. Wang B, Miao Z, Liu S, Wang J, Zhou S, Han L, Meng D, Wang Y, Li C, Ma X
Genetic analysis of ABCG2 gene C421A polymorphism with gout disease in Chinese Han male population.
Hum Genet. 2010 Feb;127(2):245-6. Epub 2009 Nov 17., [PMID:19921266]

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23 In the present study, gout in Chinese individuals was strongly associated with the ABCG2 missense SNP rs2231142 which leads to a glutamine-to-lysine amino acid substitution (Q141K) in exon 5. Login to comment

[hide] Polymorphisms in the xenobiotic transporter Multid... BMC Cancer. 2009 Nov 21;9:407. Andersen V, Ostergaard M, Christensen J, Overvad K, Tjonneland A, Vogel U
Polymorphisms in the xenobiotic transporter Multidrug Resistance 1 (MDR1) and interaction with meat intake in relation to risk of colorectal cancer in a Danish prospective case-cohort study.
BMC Cancer. 2009 Nov 21;9:407., [PMID:19930591]

Abstract [show]
BACKGROUND: The xenobiotic transporters, Multidrug Resistance 1 (MDR1/ABCB1) and Breast Cancer Resistance Protein (BCRP/ABCG2) may restrict intestinal absorption of various carcinogens, including heterocyclic amines (HCA) and polycyclic aromatic hydrocarbons (PAH). Cyclooxygenase-2 (COX-2) derived prostaglandins promote gastrointestinal carcinogenesis, affecting angiogenesis, apoptosis, and invasiveness.The aim of this study was to investigate if polymorphisms in these genes were associated with risk of colorectal cancer (CRC), and to investigate possible interactions with lifestyle factors such as smoking, meat consumption, and NSAID use. METHODS: The following polymorphisms were analyzed; a synonymous MDR1 C3435T (rs1045642) in exon26, G-rs3789243-A in intron3, the functional BCRP C421A (rs2231142), the two COX-2 A-1195G (rs689466) and G-765C (rs20417) in the promoter region, and the COX-2 T8473C (rs5275) polymorphisms in the 3'-untranslated region. The polymorphisms were assessed together with lifestyle factors in a nested case-cohort study of 359 cases and a random cohort sample of 765 participants from the Danish prospective Diet, Cancer and Health study. RESULTS: Carriers of the variant allele of MDR1 intron 3 polymorphism were at 1.52-fold higher risk of CRC than homozygous wild type allele carriers (Incidence rate ratio (IRR) = 1.52, 95% Confidence Interval (CI): 1.12-2.06). Carriers of the variant allele of MDR1 C3435T exon 26 had a lower risk of CRC than homozygous C-allele carriers (IRR = 0.71 (CI:0.50-1.00)). There was interaction between these MDR1 polymorphisms and intake of red and processed meat in relation to CRC risk. Homozygous MDR1 C3435T C-allele carriers were at 8% increased risk pr 25 gram meat per day (CI: 1.00-1.16) whereas variant allele carriers were not at increased risk (p for interaction = 0.02). COX-2 and BCRP polymorphisms were not associated with CRC risk. There was interaction between NSAID use and MDR1 C3435T and COX-2 T8473C (p-values for interaction 0.001 and 0.04, respectively). CONCLUSION: Two polymorphisms in MDR1 were associated with CRC risk and there was interaction between these polymorphisms and meat intake in relation to CRC risk. Our results suggest that MDR1 polymorphisms affect the relationship between meat and CRC risk.

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36 The variant allele of the non-synonymous BCRP C421A (Q141K) polymorphism has been associated with lower protein levels and lowered transport activity both in vitro [34,35] and in vivo [27,35], but no associations were found between either the intestinal levels of protein and mRNA and BRCP polymorphisms [36] or between BCRP polymorphisms and risk of CRC [4]. Login to comment

[hide] Pharmacogenetics of ATP-binding cassette transport... Methods Mol Biol. 2010;596:95-121. Cascorbi I, Haenisch S
Pharmacogenetics of ATP-binding cassette transporters and clinical implications.
Methods Mol Biol. 2010;596:95-121., [PMID:19949922]

Abstract [show]
Drug resistance is a severe limitation of chemotherapy of various malignancies. In particular efflux transporters of the ATP-binding cassette family such as ABCB1 (P-glycoprotein), the ABCC (multidrug resistance-associated protein) family, and ABCG2 (breast cancer resistance protein) have been identified as major determinants of chemoresistance in tumor cells. Bioavailability depends not only on the activity of drug metabolizing enzymes but also to a major extent on the activity of drug transport across biomembranes. They are expressed in the apical membranes of many barrier tissues such as the intestine, liver, blood-brain barrier, kidney, placenta, testis, and in lymphocytes, thus contributing to plasma, liquor, but also intracellular drug disposition. Since expression and function exhibit a broad variability, it was hypothesized that hereditary variances in the genes of membrane transporters could explain at least in part interindividual differences of pharmacokinetics of a variety of anticancer drugs and many others contributing to the clinical outcome of certain leukemias and further malignancies.

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224 Both identified 34G>A (V12M) and 421C>A (Q141K). Login to comment
231 HEK-293 cells, transfected with wild-type or V12N, ABCG2 showed apical staining with an ABCG2 antibody, but high intracellular staining in case of Q141K, suggesting impaired membrane trafficking or incorrect membrane insertion. Login to comment
242 0.005 c. 376 C>T Q126stop 0.01 0.00a 0.00b Lack of function c. 421 C>A Q141K 0.35 0.11a 0.02b Reduced activity [120, 124, 137] IVS 5 -16 A>G ? Login to comment

[hide] Flow cytometric evaluation of multidrug resistance... Methods Mol Biol. 2010;596:123-39. Aszalos A, Taylor BJ
Flow cytometric evaluation of multidrug resistance proteins.
Methods Mol Biol. 2010;596:123-39., [PMID:19949923]

Abstract [show]
There are several ways to detect proteins on cells. One quite frequently used method is flow cytometry. This method needs fluorescently labeled antibodies that can attach selectively to the protein to be investigated for flow cytometric detection. Flow cytometry scans individual cells, virtually without their surrounding liquid, and can scan many cells in a very short time. Because of this advantage of flow cytometry, it was adapted to investigate transport proteins on normal and cancerous human cells and cell lines. These transport proteins play important roles in human metabolism. Absorption in the intestine, excretion at the kidney, protection of the CNS compartment and the fetus from xenobiotics, and other vital functions depend on these transporters. However, several transporters are overexpressed in cancer cells. These overexpressed transporters pump out anticancer drugs from the cells and prevent their curative effects. The detection and quantitation of these types of transporters in cancer cells is important for this reason. Here, we review literature on flow cytometric detection of the three most studied transporters: P-glycoprotein, multidrug resistance-associated proteins, and breast cancer resistance protein.

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258 The V12M and Q141K variants were described by Zamber et al. Login to comment

[hide] Impact of breast cancer resistance protein on canc... Methods Mol Biol. 2010;596:251-90. Ross DD, Nakanishi T
Impact of breast cancer resistance protein on cancer treatment outcomes.
Methods Mol Biol. 2010;596:251-90., [PMID:19949928]

Abstract [show]
Breast cancer resistance protein (BCRP/ABCG2) was discovered in multidrug resistant breast cancer cells having an ATP-dependent transport-based resistance phenotype. This ABC transporter functions (at least in part) as a xenobiotic protective mechanism for the organism: in the gut and biliary tract, it prevents absorption and enhances elimination of potentially toxic substances. As a placental barrier, it protects the fetus; similarly, it serves as a component of blood-brain and blood-testis barrier; BCRP is expressed in stem cells and may protect them from potentially harmful agents. Therefore, BCRP could influence cancer outcomes by (a) endogenous BCRP affecting the absorption, distribution, metabolism, and elimination of anticancer drugs; (b) BCRP expression in cancer cells may directly cause resistance by active efflux of anticancer drugs; (c) BCRP expression in cancer cells could be a manifestation of the activity of metabolic and signaling pathways that impart multiple mechanisms of drug resistance, self-renewal (stemness), and invasiveness (aggressiveness)--i.e. impart a poor prognosis--to cancers. This chapter presents a synopsis of translational clinical studies relating BCRP expression in leukemias, lymphomas, and a variety of solid tumors with clinical outcome. Data are emerging that expression of BCRP, like P-glycoprotein/ABCB1, is associated with adverse outcomes in a variety of human cancers. Whether this adverse prognostic effect results from resistance imparted to the cancer cells as the direct result of BCRP efflux of anticancer drugs, or whether BCRP expression (and also Pgp expression - coexpression of these transporters is common among poor risk cancers) serves as indicators of the activity of signaling pathways that enhance cancer cellular proliferation, metastases, genomic instability, enhance drug resistance, and oppose programmed cell death mechanisms is yet unknown.

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70 A number of single nucleotide polymorphisms (SNPs) have been observed in the BCRP gene (77), and of those nonsynonymous SNPs observed in the coding region, the most common and most extensively studied are the G34A (exon 2) and C421A (exon 5) alleles, which cause alterations of amino acids 12 (V12M) and 141 (Q141K), respectively (78-81). Login to comment
79 In work done by Mizurai et al., the enforced expression of the G34A or C421A allele in porcine renal LLC-PK1 cells resulted in reduced transporter function, with a reduction in BCRP apical membrane localization for the G34A variant protein and reduced transporter function for the Q141K allele (82). Login to comment
84 transfected seven BCRP coding region SNPs into LLC-PK1 cells, and found lower BCRP -protein expression for the Q141K and S441N alleles (84). Login to comment
90 found four nonsynonymous coding region SNPs among 92 Korean subjects (V12M, Q141K, P269S, Q126Stop), and four SNPs in the BCRP promoter region, one of which was in the HIF-1 response element (C-19031T) (86). Login to comment
93 Tamura et al. used multicolor fluorescence in situ hybridization to assure uniform mRNA expression of cDNAs of seven BCRP SNPs (V12M, Q141K, F208S, S248P, F431L, S441N and F489L) transduced into Flp-In-293 cells (87, 88). Login to comment
94 Protein expression from the F208S and S441N variants was found to be low; the V12M and Q141K alleles had IC50 s for SN-38 that were approximately half that of the wild-type; all the other alleles examined had significantly lower IC50 values for SN-38, mitoxantrone, doxorubicin, daunorubicin and etoposide when compared with wild type alleles (88). Login to comment
104 The first report associating a BCRP polymorphism with altered drug exposure found that cancer patients (N = 5) heterozygous for the C421A (Q141K) allele had plasma levels of IV diflomotecan that were almost threefold higher than those of 15 patients harboring the wild-type allele (92). Login to comment
119 found the Q141K polymorphism is 3.1.2. Login to comment
124 Korenaga et al. investigated the C421A (Q141K) allele and found that individuals with the wild-type C/C genotype were more at risk for developing nonpapillary renal cell carcinoma (RCC) in a study of 200 Japanese patients with nonpapillary RCC and age-and sex-matched control subjects (108). Login to comment

[hide] Pharmacogenetics of membrane transporters: an upda... Mol Biotechnol. 2010 Feb;44(2):152-67. Sissung TM, Baum CE, Kirkland CT, Gao R, Gardner ER, Figg WD
Pharmacogenetics of membrane transporters: an update on current approaches.
Mol Biotechnol. 2010 Feb;44(2):152-67., [PMID:19950006]

Abstract [show]
This review provides an overview of the pharmacogenetics of membrane transporters including selected ABC transporters (ABCB1, ABCC1, ABCC2, and ABCG2) and OATPs (OATP1B1 and OATP1B3). Membrane transporters are heavily involved in drug clearance and alters drug disposition by actively transporting substrate drugs between organs and tissues. As such, polymorphisms in the genes encoding these proteins may have significant effects on the absorption, distribution, metabolism and excretion of compounds, and may alter pharmacodynamics of many agents. This review discusses the techniques used to identify substrates and inhibitors of these proteins and subsequently to assess the effect of genetic mutation on transport, both in vitro and in vivo. A comprehensive list of substrates for the major drug transporters is included. Finally, studies linking transporter genotype with clinical outcomes are discussed.

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57 This SNP results in an amino acid change of Gln to Lys at codon 141 and has been shown in Flp-In-293 cells to have half the protein expression of the wildtype [34]. Login to comment
123 [70, 71] Docetaxel AUC and overall liver with 3435TT genotype [100-102] Docetaxel is transported by ABCB1 [103] ABCG2 Topotecan Increased accumulation of drug in cells transfected with 421A (Q141K) [104] Increased bioavailability in heterozygous C421A patients [104] Imatinib Increased accumulation of drug in cells transfected with 421A (Q141K) [105] No significant difference in AUC or Cmax in heterozygous C421 patients [105] ABCC1 N/A None to date ABCC2 N/A None to date OATP1B1 Pravastatin Reduced membrane expression of OATP1B1*5 variant [45] Increased AUC in OATP1B1*5 [94, 106] OATP1B3 Leuprolide and goserelin Reduced transport of testosterone with the 334G/699A haplotype [24] Increased progression-free survival in patients with 334G/699A haplotype [97] consequences of the 2677G[T/A polymorphism may be explained by expression alterations alone and not necessarily by altered substrate binding or transport efficiency of the protein. Login to comment

[hide] Sex and age interaction with genetic association o... Atherosclerosis. 2010 Jun;210(2):474-8. Epub 2009 Dec 16. Brandstatter A, Lamina C, Kiechl S, Hunt SC, Coassin S, Paulweber B, Kramer F, Summerer M, Willeit J, Kedenko L, Adams TD, Kronenberg F
Sex and age interaction with genetic association of atherogenic uric acid concentrations.
Atherosclerosis. 2010 Jun;210(2):474-8. Epub 2009 Dec 16., [PMID:20053405]

Abstract [show]
BACKGROUND: High serum uric acid levels are associated with gout, atherosclerosis and cardiovascular disease. Three genes (SLC2A9, ABCG2, and SLC17A3) were reported to be involved in the regulation of uric acid levels. RESEARCH: Design and Methods: SNPs rs2231142 (ABCG2) and rs1165205 (SLC17A3) were genotyped in three cohorts (n=4492) and combined with previously genotyped SNPs within SLC2A9 (rs6855911, rs7442295, rs6449213, rs12510549). RESULTS: Each copy of the minor allele decreased uric acid levels by 0.30-0.38 mg/dL for SLC2A9 (p values: 10(-20)-10(-36)) and increased levels by 0.34 mg/dL for ABCG2 (p=1.1x10(-16)). SLC17A3 influenced uric acid levels only modestly. Together the SNPs showed graded associations with uric acid levels of 0.111 mg/dL per risk allele (p=3.8x10(-42)). In addition, we observed a sex-specific interaction of age with the association of SLC2A9 SNPs with uric acid levels, where increasing age strengthened the association of SNPs in women and decreased the association in men. CONCLUSIONS: Genetic variants within SLC2A9,ABCG2 and SLC17A3 show highly significant associations with uric acid levels, and for SNPs within SLC2A9 this association is strongly modified by age and sex.

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69 Bioinformatic analysis The ABCG2 SNP rs2231142 (Q141K) was recently shown to be a loss-of-function mutation that causes hyperuricemia and gout [9]. Login to comment

[hide] What lies behind serum urate concentration? Insigh... Genome Med. 2009 Dec 29;1(12):118. Ichida K
What lies behind serum urate concentration? Insights from genetic and genomic studies.
Genome Med. 2009 Dec 29;1(12):118., [PMID:20090896]

Abstract [show]
Many factors, including genetic components and acquired factors such as obesity and alcohol consumption, influence serum uric acid (urate) concentrations. Since serum urate concentrations are determined by the balance between renal urate excretion and the volume of urate produced via purine metabolism, urate transporter genes as well as genes coding for enzymes involved in purine metabolism affect serum urate concentrations. URAT1 was the first transporter affecting serum urate concentrations to be identified. Using the characterization of this transporter as an indicator, several transporters have been shown to transport urate, allowing the construction of a synoptic renal urate transport model. Notable re-absorptive urate transporters are URAT1 at apical membranes and GLUT9 at basolateral membranes, while ABCG2, MRP4 (multidrug resistance protein 4) and NPT1 are secretive transporters at apical membranes. Recent genome-wide association studies have led to validation of the in vitro model constructed from each functional analysis of urate transporters, and identification of novel candidate genes related to urate metabolism and transport proteins, such as glucokinase regulatory protein (GKRP), PDZK1 and MCT9. However, the function and physiologic roles of several candidates, as well as the influence of acquired factors such as obesity, foods, or alcoholic beverages, remain unclear.

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89 The frequency of the mutation Q141K, encoded by the common SNP rs2231142, is highly variable in the human population; the frequency of the A allele ranges from 1 to 5% in Africans, to approximately 30% in Asians. Login to comment
90 Urate transport by the ABCG2 mutant Q141K is about half that of the wild type [43]. Login to comment
91 Q126X shows stronger effects on gout development than Q141K, conferring an odds ratio of 5.97 [44]. Login to comment

[hide] Genetics of gout. Curr Opin Rheumatol. 2010 Mar;22(2):144-51. Choi HK, Zhu Y, Mount DB
Genetics of gout.
Curr Opin Rheumatol. 2010 Mar;22(2):144-51., [PMID:20110790]

Abstract [show]
PURPOSE OF REVIEW: This review provides an update on recent findings with regards to the genetics of hyperuricemia and gout, including recent data from genome-wide association studies (GWAS). RECENT FINDINGS: Five GWAS around the same time reported that genetic variants of SLC2A9/GLUT9 were associated with lower serum uric acid (SUA) levels and the effects were stronger among women (e.g. SUA level difference per copy of a minor allele, -0.46 mg/dl in women vs. -0.22 mg/dl in men). One study involving four cohorts and one meta-analysis of 14 genome-wide scans found that genetic variants of ABCG2 were associated with higher SUA concentrations and these effects were stronger among men (e.g. uric acid level difference per copy of the minor allele, 0.32 mg/dl in men vs. 0.18 mg/dl in women). Limited data indicate that these associations likely translate into those with the risk of gout. Functional determination that GLUT9 and ABCG2 can transport urate at the apical border of proximal tubules implicates them as substantial players in the renal excretion of urate. Furthermore, five novel genetic loci have been reported in the meta-analysis of 14 genome-wide scans. SUMMARY: Combined with their activities as urate transporters and their strong associations with serum uric acid concentrations, GLUT9 and ABCG2 appeared to be important modulators of uric acid levels and likely of the risk of gout. Together with a growing list of environmental risk factors, these genetic data add considerably to our understanding of the pathogenesis of hyperuricemia and gout.

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87 Furthermore, the mutation Q141K encoded by the common SNP rs2231142 resulted in a 53% reduction in urate transport rates, compared with wild-type ABCG2 [34 ]. Login to comment

[hide] ABCG2 polymorphism is associated with the low-dens... Clin Pharmacol Ther. 2010 May;87(5):558-62. Epub 2010 Feb 3. Tomlinson B, Hu M, Lee VW, Lui SS, Chu TT, Poon EW, Ko GT, Baum L, Tam LS, Li EK
ABCG2 polymorphism is associated with the low-density lipoprotein cholesterol response to rosuvastatin.
Clin Pharmacol Ther. 2010 May;87(5):558-62. Epub 2010 Feb 3., [PMID:20130569]

Abstract [show]
The ATP-binding cassette G2 (ABCG2) c.421C>A (rs2231142) polymorphism influences the pharmacokinetics of rosuvastatin. We examined whether this polymorphism influences the low-density lipoprotein cholesterol (LDL-C)-lowering efficacy of the drug. In 305 Chinese patients with hypercholesterolemia who were treated with rosuvastatin at a dosage of 10 mg daily, the c.421A variant was found to be significantly associated with greater reduction in LDL-C level, in a gene-dose-dependent manner. As compared with subjects with the c.421CC genotype, those with the c.421AA genotype showed a 6.9% greater reduction in LDL-C level, which would be equivalent to the effect obtained by doubling the dose of rosuvastatin.

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5 Rosuvastatin is one of the few drugs for which regulatory authorities, including the US Food and Drug Administration, have recommended starting with lower doses (5 mg instead of 10 mg) in Asian patients.3 This recommendation was based on data from single-dose pharmacokinetic studies that showed that systemic exposure to rosuvastatin was approximately twice as high in Asians as in non-Asians.4 These -differences are unlikely to be due to ethnicity-related variations in drug metabolizing enzymes, given that -rosuvastatin -undergoes relatively little enzymic modification and is a substrate for a number of drug transporters that influence its disposition.5,6 The efflux transporter ATP-binding cassette G2 (ABCG2) plays a significant role in the disposition of rosuvastatin in -vitro.7 The c.421C>A (rs2231142, Gln141Lys) single-nucleotide polymorphism of ABCG2 influences the pharmacokinetics of rosuvastatin in Chinese and Caucasian subjects.8,9 We examined whether this ABCG2 single-nucleotide polymorphism influences the reduction of LDL-C levels when rosuvastatin is administered to Chinese patients with hypercholesterolemia, including some with familial hypercholesterolemia (FH). Login to comment

[hide] Two novel SNPs of the ABCG2 gene and its associati... Mol Biol Rep. 2011 Jun;38(5):2927-32. Epub 2010 Feb 7. Yue W, Fang X, Zhang C, Pang Y, Xu H, Gu C, Shao R, Lei C, Chen H
Two novel SNPs of the ABCG2 gene and its associations with milk traits in Chinese Holsteins.
Mol Biol Rep. 2011 Jun;38(5):2927-32. Epub 2010 Feb 7., [PMID:20140710]

Abstract [show]
The ATP-binding cassette transporter ABCG2 (also known as breast cancer resistance protein, BCRP) belongs to the ATP-binding cassette (ABC) family of transmembrane drug transporters, playing a crucial role in the protection of various cells and tissues against xenotoxins and/or endotoxins. Recently, several studies have proposed it as the potential gene underlying the QTL on bovine chromosome 6. Hence, in this study, the PCR-SSCP method was applied to detect two polymorphisms (A --> C and A --> G) in the target sequence coding nucleotide-binding domain (NBD) region of ABCG2 and evaluate its associations with milk production traits and mastitis-related traits among Chinese Holsteins. In the analyzed population, the allelic frequencies for the A and B alleles were 0.5990 and 0.4010, respectively and the genotypic frequencies were in Hardy-Weinberg disequilibrium (P < 0.01). Moreover, significant statistical relationships between the polymorphisms of ABCG2 gene and following traits, including milk yields, milk protein percentage and somatic cell scores (SCS), were found (P < 0.05). When compared with AA genotype, BB genotype was associated with higher milk yields during 1st and 2nd lactations, as well as lower milk protein percentage and SCS. Thus, BB genotype is suggested to be a molecular marker for superior milk performance.

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18 For example, the significance of the Q141K SNP has been shown in vitro [14] and in clinical studies [15, 16]. Login to comment

[hide] Breast cancer resistance protein (BCRP)-mediated g... Drug Metab Dispos. 2010 May;38(5):740-4. Epub 2010 Feb 16. Pollex EK, Anger G, Hutson J, Koren G, Piquette-Miller M
Breast cancer resistance protein (BCRP)-mediated glyburide transport: effect of the C421A/Q141K BCRP single-nucleotide polymorphism.
Drug Metab Dispos. 2010 May;38(5):740-4. Epub 2010 Feb 16., [PMID:20159988]

Abstract [show]
The antidiabetic agent glyburide (glibenclamide) is frequently used for the treatment of type II diabetes and is increasingly being used for the treatment of gestational diabetes. Evidence suggests that breast cancer resistance protein/ATP-binding cassette, subfamily G, member 2 (ABCG2) expressed in the placenta protects the fetus against the accumulation of glyburide. A number of studies have investigated the significance of several single-nucleotide polymorphisms (SNPs) in the ABCG2 gene. Associations between the Q141K (C421A) SNP and ABCG2 protein expression, membrane surface translocation, efflux activity, or ATPase activity have been shown. Therefore, alterations in glyburide transport across the placenta, resulting in increased fetal glyburide exposure, may be seen in individuals carrying the C421A allele. The purpose of this study is to investigate whether the Q141K SNP causes alterations in ABCG2-mediated glyburide transport. Glyburide accumulation assays were carried out with stably transfected human embryonic kidney (HEK)-293 cells expressing wild-type ABCG2 (Arg482) and polymorphic ABCG2 (Q141K). Glyburide kinetic parameters were determined for comparison of wild-type and SNP ABCG2 activity by simultaneously fitting data for ABCG2-expressing cells (saturable transport) and empty vector-expressing cells (nonsaturable transport) by nonlinear regression analysis. The apparent K(t) and V(max) values for the transfected HEK-293 cells expressing the polymorphic variant (Q141K) of ABCG2 were significantly higher than those values determined for the wild-type ABCG2-expressing cells (p < 0.05). Our results indicate that the Q141K variant of ABCG2 may have the potential to alter the placental pharmacokinetics of glyburide used in pregnancy.

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0 Breast Cancer Resistance Protein (BCRP)-Mediated Glyburide Transport: Effect of the C421A/Q141K BCRP Single-Nucleotide Polymorphism Erika K. Pollex, Gregory Anger, Janine Hutson, Gideon Koren, and Micheline Piquette-Miller Division of Clinical Pharmacology and Toxicology (E.K.P., J.H., G.K.) and Motherisk Program (E.K.P., J.H., G.K.), Hospital for Sick Children, Toronto, Ontario, Canada; and Department of Pharmaceutical Sciences (E.K.P., G.A., G.K., M.P.-M.) and Institute of Medical Science (J.H., G.K.), University of Toronto, Toronto, Ontario, Canada Received October 14, 2009; accepted February 16, 2010 ABSTRACT: The antidiabetic agent glyburide (glibenclamide) is frequently used for the treatment of type II diabetes and is increasingly being used for the treatment of gestational diabetes. Login to comment
3 Associations between the Q141K (C421A) SNP and ABCG2 protein expression, membrane surface translocation, efflux activity, or ATPase activity have been shown. Login to comment
5 The purpose of this study is to investigate whether the Q141K SNP causes alterations in ABCG2-mediated glyburide transport. Login to comment
6 Glyburide accumulation assays were carried out with stably transfected human embryonic kidney (HEK)-293 cells expressing wild-type ABCG2 (Arg482) and polymorphic ABCG2 (Q141K). Login to comment
8 The apparent Kt and Vmax values for the transfected HEK-293 cells expressing the polymorphic variant (Q141K) of ABCG2 were significantly higher than those values determined for the wild-type ABCG2-expressing cells (p < 0.05). Login to comment
9 Our results indicate that the Q141K variant of ABCG2 may have the potential to alter the placental pharmacokinetics of glyburide used in pregnancy. Login to comment
28 The results of these studies vary, showing inconsistencies in the effect of the C421A (Q141K) polymorphism on ABCG2 protein activity. Login to comment
29 It has been suggested that the Q141K SNP may affect ABCG2 protein expression, membrane surface translocation, efflux activity, or ATPase activity. Login to comment
30 Of particular concern, in vivo studies have shown patients expressing the Q141K variant who were exposed to the chemotherapeutic agents topotecan and diflomotecan achieved higher plasma drug levels, suggesting a significant reduction in ABCG2-mediated excretion (Sparreboom et al., 2004). Login to comment
34 Because BCRP/ABCG2 is believed to protect the fetus against the accumulation of its substrates, it is important to determine the potential effect of the Q141K SNP on the placental transfer of drugs used in pregnancy. Login to comment
37 Therefore, the purpose of this study is to investigate whether the Q141K SNP causes alterations in ABCG2-mediated glyburide transport. Login to comment
38 Materials and Methods Stably transfected human embryonic kidney (HEK)-293 cells containing full-length ABCG2 encoding wild-type (Arg482) or the SNP variant (Q141K) of ABCG2, or an empty vector to serve as a control were obtained (Dr. Robert W. Robey, National Cancer Institute, Bethesda, MD), and expression was enforced by selection in G418 (Invitrogen, Carlsbad, CA). Login to comment
41 Flow cytometry assays were performed to confirm surface expression of ABCG2 in the wild-type (Arg482) and polymorphic (Q141K) clones. Login to comment
51 Transport activity was examined by measuring glyburide accumulation in stably transfected HEK-293 cells expressing wild-type ABCG2 (Arg482), polymorphic ABCG2 (Q141K), and an empty vector. Login to comment
72 Cells were incubated for 30 min with either phycoerythrin-labeled negative control antibody or the phycoerythrin-labeled anti-ABCG2 antibody 5D3 (red, negative control; green, Arg482; blue, Q141K). Login to comment
77 Data from the HEK-293 empty vector-transfected cell line and each of the ABCG2-transfected HEK-293 cell lines (Arg482, Q141K) were used simultaneously to fit the model parameters because the assumption was made that the permeability of the empty vector-transfected cell line and each of the ABCG2-transfected HEK-293 cell lines was the same. Login to comment
84 The surface expression of ABCG2 in HEK-293 cells transfected with empty vector, wild-type (Arg482), or SNP variant (Q141K) of FIG. 2. Login to comment
85 A, glyburide accumulation in the presence and absence of FTC (10 ␮M) in HEK293 cells transfected with wild-type ABCG2 (Arg482), ABCG2 polymorphic variant (Q141K), and control (empty vector) after 60 min of exposure to 20 ␮M glyburide. Login to comment
87 B, accumulation of glyburide in HEK-293 cells transfected with wild-type ABCG2 (Arg482), ABCG2 polymorphic variant (Q141K), and empty vector control after 20 min of exposure to various concentrations of glyburide in the presence and absence of FTC (10 ␮M). Login to comment
90 As shown in Fig. 1, surface expression of ABCG2 was detected at similar levels in the ABCG2-transfected HEK-293 cell lines expressing wild-type (Arg482) and the polymorphic variant (Q141K) of ABCG2 (Fig. 1). Login to comment
94 Inhibition of ABCG2/BCRP by fumitremorgin C (FTC) (10 ␮M) resulted in marked increases in intracellular accumulation of [3 H]glyburide in HEK-293 cells expressing the wild-type (Arg482) ABCG2 or SNP variant (Q141K) of ABCG2 after 60 min of exposure to 20 ␮M glyburide (Fig. 2A). Login to comment
96 [3 H]Glyburide accumulation experiments carried out for 20 min at various concentrations of glyburide (0.2-100 ␮M) show inhibitable transport in both Arg482- and Q141K-transfected HEK-293 cells exposed to up to 20 ␮M glyburide (Fig. 2B). Login to comment
98 Data obtained suggest that full FTC-mediated inhibition of BCRP/ ABCG2 is seen in the wild-type transfected cells at higher drug concentrations than in the SNP-transfected cells (Q141K) (Fig. 2B). Login to comment
101 In the transfected HEK-293 cells expressing the polymorphic variant (Q141K) of ABCG2, the apparent Kt and Vmax values were 80.37 Ϯ 22.96 ␮M and 384.79 Ϯ 69.71 pmol/min ⅐ mg protein, respectively. Login to comment
102 Both the apparent Kt and Vmax values for the transfected HEK-293 cells expressing the polymorphic variant (Q141K) of ABCG2 were found to be significantly higher than those values determined for the wild-type ABCG2-expressing cells (p Ͻ 0.05). Login to comment
106 The recent identification of the nonsynonymous SNP, Q141K (Honjo et al., 2002; Imai et al., 2002), in the coding region of ABCG2 and the potential functional consequences on drug disposition has led to an increasingly important area of research. Login to comment
107 To elucidate the potential effects of the Q141K SNP on BCRP-mediated glyburide transport, we examined and compared the accumulation of glyburide in HEK293 cells stably transfected with full-length ABCG2 encoding wild-type or the SNP variant (Q141K) of ABCG2. Login to comment
110 Regarding BCRP transport kinetics, the values obtained for apparent affinity (Kt) of glyburide for BCRP suggest that glyburide has greater affinity for the wild-type ABCG2 protein compared with the affinity determined for the Q141K ABCG2 variant. Login to comment
114 Rate of glyburide transport in wild-type (Arg482), SNP (Q141K), and empty vector-transfected HEK-293 cells after exposure to various concentrations of glyburide (0.2-100 ␮M) for 20 min. Login to comment
119 Metabolism of glyburide in this system is also thought to be minimal and similar in both wild-type and Q141K ABCG2-transfected cells; however, intracellular binding and glyburide metabolism could also confound the estimation of Kt values. Login to comment
120 It has been previously reported that the Q141K variant is associated with increased sensitivity to chemotherapeutic agents as a result of a decrease in protein expression in transfected cells (Imai et al., 2002). Login to comment
121 Data obtained in our study did not show reduced surface expression of Q141K ABCG2. Login to comment
122 Thus, the impaired ability of the ABCG2 protein to efflux glyburide seen in cells transfected with Q141K ABCG2 was likely the result of SNP-induced alterations in the function of the protein itself. Login to comment
123 Similar to our findings, Morisaki et al. (2005) and Zamber et al. (2003) did not observe a correlation between decreased expression of ABCG2 and the Q141K SNP. Login to comment
124 However, Morisaki et al. (2005) noted that a larger proportion of the surface-localized Q141K ABCG2 protein is intracellular, implying less efficient post-translational processing in the SNP variant and thus providing a potential mechanism for reduced efficiency. Login to comment
125 In summary, data from our accumulation studies indicate that the HEK-293 cell line expressing the Q141K polymorphic variant of the ABCG2/BCRP gene exhibits a reduced affinity for BCRP-mediated glyburide efflux. Login to comment
126 Therefore, the Q141K variant of ABCG2 may have the potential to alter the placental pharmacokinetics of glyburide and other clinically used BCRP substrate drugs in pregnancy. Login to comment
128 However, the results of this study provide sufficient reason to further investigate the impact of SNPs on ABCG2-mediated glyburide transport because the potential exists for increased fetal exposure to glyburide in pregnancy among patients carrying the Q141K polymorphic variant allele of ABCG2. Login to comment

[hide] Hepatic metabolism and transporter gene variants e... Circ Cardiovasc Genet. 2010 Jun;3(3):276-85. Epub 2010 Mar 5. Bailey KM, Romaine SP, Jackson BM, Farrin AJ, Efthymiou M, Barth JH, Copeland J, McCormack T, Whitehead A, Flather MD, Samani NJ, Nixon J, Hall AS, Balmforth AJ
Hepatic metabolism and transporter gene variants enhance response to rosuvastatin in patients with acute myocardial infarction: the GEOSTAT-1 Study.
Circ Cardiovasc Genet. 2010 Jun;3(3):276-85. Epub 2010 Mar 5., [PMID:20207952]

Abstract [show]
BACKGROUND: Pharmacogenetics aims to maximize benefits and minimize risks of drug treatment. Our objectives were to examine the influence of common variants of hepatic metabolism and transporter genes on the lipid-lowering response to statin therapy. METHODS AND RESULTS: The Genetic Effects On STATins (GEOSTAT-1) Study was a genetic substudy of Secondary Prevention of Acute Coronary Events-Reduction of Cholesterol to Key European Targets (SPACE ROCKET) (a randomized, controlled trial comparing 40 mg of simvastatin and 10 mg of rosuvastatin) that recruited 601 patients after myocardial infarction. We genotyped the following functional single nucleotide polymorphisms in the genes coding for the cytochrome P450 (CYP) metabolic enzymes, CYP2C9*2 (430C>T), CYP2C9*3 (1075A>C), CYP2C19*2 (681G>A), CYP3A5*1 (6986A>G), and hepatic influx and efflux transporters SLCO1B1 (521T>C) and breast cancer resistance protein (BCRP; 421C>A). We assessed 3-month LDL cholesterol levels and the proportion of patients reaching the current LDL cholesterol target of <70 mg/dL (<1.81 mmol/L). An enhanced response to rosuvastatin was seen for patients with variant genotypes of either CYP3A5 (P=0.006) or BCRP (P=0.010). Furthermore, multivariate logistic-regression analysis revealed that patients with at least 1 variant CYP3A5 and/or BCRP allele (n=186) were more likely to achieve the LDL cholesterol target (odds ratio: 2.289; 95% CI: 1.157, 4.527; P=0.017; rosuvastatin 54.0% to target vs simvastatin 33.7%). There were no differences for patients with variants of CYP2C9, CYP2C19, or SLCO1B1 in comparison with their respective wild types, nor were differential effects on statin response seen for patients with the most common genotypes for CYP3A5 and BCRP (n=415; odds ratio: 1.207; 95% CI: 0.768, 1.899; P=0.415). CONCLUSION: The LDL cholesterol target was achieved more frequently for the 1 in 3 patients with CYP3A5 and/or BCRP variant genotypes when prescribed rosuvastatin 10 mg, compared with simvastatin 40 mg. Clinical Trial Registration- URL: http://isrctn.org. Unique identifier: ISRCTN 89508434.

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28 In vitro pharmacokinetic evidence strongly suggests that common nonsynonymous SNPs in the SLCO1B1 (521TϾC; Val174 Ala) and BCRP (421CϾA; Gln141 Lys) genes both result in markedly decreased transport function and thus could potentially play a significant role in determining rosuvastatin concentration within the hepatocyte.23-29 It is a continual challenge to the medical profession to seek to match individual treatments to individual patients (personalized medicine). Login to comment

[hide] Pharmacological interaction with sunitinib is abol... Cancer Sci. 2010 Jun;101(6):1493-500. Epub 2010 Feb 22. Kawahara H, Noguchi K, Katayama K, Mitsuhashi J, Sugimoto Y
Pharmacological interaction with sunitinib is abolished by a germ-line mutation (1291T>C) of BCRP/ABCG2 gene.
Cancer Sci. 2010 Jun;101(6):1493-500. Epub 2010 Feb 22., [PMID:20345483]

Abstract [show]
Sunitinib malate (Sutent, SU11248) is a small-molecule multitargeted tyrosine kinase inhibitor (TKI) used for the treatment of renal cell carcinoma and imatinib-resistant gastrointestinal stromal tumors. Some TKIs can overcome multidrug resistance conferred by ATP-binding cassette transporter, P-glycoprotein (P-gp)/ABCB1, multidrug resistance-associated protein 1 (MRP1)/ABCC1, and breast cancer resistance protein (BCRP)/ABCG2. Here, we analyzed the effects of sunitinib on P-gp and on wild-type and germ-line mutant BCRPs. Sunitinib remarkably reversed BCRP-mediated and partially reversed P-gp-mediated drug resistance in the respective transfectants. The in vitro vesicle transport assay indicated that sunitinib competitively inhibited BCRP-mediated estrone 3-sulfate transport and P-gp-mediated vincristine transport. These inhibitory effects of sunitinib were further analyzed in Q141K-, R482G-, R482S-, and F431L-variant BCRPs. Intriguingly, the F431L-variant BCRP, which is expressed by a germ-line mutant allele 1291T>C, was almost insensitive to both sunitinib- and fumitremorgin C (FTC)-mediated inhibition in a cell proliferation assay. Sunitinib and FTC did not inhibit (125)I-iodoarylazidoprazosin-binding to F431L-BCRP. Thus, residue Phe-431 of BCRP is important for the pharmacological interaction with sunitinib and FTC. Collectively, this is the first report showing a differential effect of a germ-line variation of the BCRP/ABCG2 gene on the pharmacological interaction between small-molecule TKIs and BCRP. These findings would be useful for improving our understanding of the pharmaceutical effects of sunitinib in personalized chemotherapy.

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5 These inhibitory effects of sunitinib were further analyzed in Q141K-, R482G-, R482S-, and F431L-variant BCRPs. Login to comment
20 (14) We also reported variant BCRP SNP cDNAs harboring 421C>A (amino acid substitution Q141K),(15) and this SNP is physiologically important because the pharmacokinetics of diflomotecan, a new camptothecin-derivative anticancer agent, and the risk of adverse reactions, such as gefitinib-induced diarrhea, was affected in patients heterozygous for the A421 allele. Login to comment
109 (13,35-37) The Q141K variant, a widespread SNP in Japanese individuals, is associated with the low protein expression of BCRP,(15) and the F431L variant, also a germ-line mutation of BCRP, shows a low level of resistance to SN-38. Login to comment
173 Reversal ratios are shown for wild-type (gray circles), and Q141K (open squares), F431L (filled circles), R482S (open diamonds), and R482G (open triangles) BCRP variant-expressing cells. Login to comment

[hide] Placental P-glycoprotein and breast cancer resista... Placenta. 2010 May;31(5):351-7. Epub 2010 Mar 27. Hutson JR, Koren G, Matthews SG
Placental P-glycoprotein and breast cancer resistance protein: influence of polymorphisms on fetal drug exposure and physiology.
Placenta. 2010 May;31(5):351-7. Epub 2010 Mar 27., [PMID:20347140]

Abstract [show]
Recent studies have illustrated the importance of placental drug transport proteins, such as P-glycoprotein (Pgp) and breast cancer resistance protein (BCRP) in limiting fetal exposure to drugs and toxins. Moreover, increasing evidence supports a role for Pgp and BCRP in the normal development and physiological function of the placenta. Several single nucleotide polymorphisms (SNPs) in the genes encoding Pgp and BCRP have been described and are associated with altered protein expression, transporter activity, and clinical outcome in studies focusing on tissues other than the placenta. This review aims to summarize current research regarding the association between these polymorphisms and expression and function in the placenta. The influence of these genotypes on fetal drug exposure and altered placental physiology or development is also presented. To date, evidence suggests that SNPs in both ABCB1 and ABCG1 can alter expression of their respective protein; however, the functional significance of these polymorphisms is less clear. An understanding of this genotype-phenotype relationship will allow for prediction of susceptible or favorable genotypes in order to personalize medication choices to minimize fetal exposure to teratogens, or to maximize pharmacological therapy to the fetus.

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142 The G34A and C421 SNPs result in amino acid changes (V12M and Q141K respectively). Login to comment
357 [60] Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, Tsuruo T, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Polymorphisms within the folate pathway predict fo... Pharmacogenet Genomics. 2010 Jun;20(6):367-76. Stamp LK, Chapman PT, O'Donnell JL, Zhang M, James J, Frampton C, Barclay ML, Kennedy MA, Roberts RL
Polymorphisms within the folate pathway predict folate concentrations but are not associated with disease activity in rheumatoid arthritis patients on methotrexate.
Pharmacogenet Genomics. 2010 Jun;20(6):367-76., [PMID:20386493]

Abstract [show]
OBJECTIVES: To determine whether genetic polymorphisms within the folate pathway are associated with red blood cell (RBC) methotrexate (MTX) polyglutamate concentrations, RBC folate concentrations and MTX efficacy/toxicity. METHODS: Disease activity in 200 rheumatoid arthritis patients on MTX was assessed by swollen and tender joint counts and Disease Activity Score 28. Genetic polymorphisms shown to have an effect on gene expression or protein function were examined. RESULTS: RBC folate concentrations were significantly associated with MTHFR 677C>T (P=0.002), MTRR 66A>G (P<0.0001), MTHFD1 1958G>A (P=0.001) and SHMT 1420C>T (P=0.012), whereas no association of these polymorphisms with disease activity was observed. None of the polymorphisms tested predicted RBC MTX polyglutamate concentrations. There were weak associations between central nervous system adverse effects and AMPD1 34C>T (P=0.04) and between gastrointestinal adverse effects and MTHFD1 1958G>A (P=0.03) and ABCC2 IVS23+56T>C (P=0.045). There was a stronger association between any adverse effect and ABCG2 914C>A (P=0.004). CONCLUSION: Although RBC folate concentrations are associated with genetic polymorphisms within the folate pathway, these variants do not predict disease activity. To accurately evaluate whether any polymorphisms are reliable predictors of MTX efficacy or toxicity, large prospective clinical trials are required. Furthermore, application of a genome-wide association strategy is likely to uncover novel predictors of MTX response.

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57 With the exception of ABCG2 (rs2231142; Q141K) and ITPA 94C > A 368 Pharmacogenetics and Genomics 2010, Vol 20 No 6 (rs41320251), patients were genotyped for each SNP by two-tube allele-specific polymerase chain reaction (PCR) (see Supplementary data Table S1, Supplemental digital content 1, http://links.lww.com/FPC/A155 for primer sequences). Login to comment
67 Patients were genotyped for ABCG2 (rs2231142; Q141K) using a predesigned TaqMan SNP genotyping assay (C__15854163_70) (Applied Biosystems, Foster City, California, USA). Login to comment

[hide] The rs2231142 variant of the ABCG2 gene is associa... Rheumatology (Oxford). 2010 Aug;49(8):1461-5. Epub 2010 Apr 25. Yamagishi K, Tanigawa T, Kitamura A, Kottgen A, Folsom AR, Iso H
The rs2231142 variant of the ABCG2 gene is associated with uric acid levels and gout among Japanese people.
Rheumatology (Oxford). 2010 Aug;49(8):1461-5. Epub 2010 Apr 25., [PMID:20421215]

Abstract [show]
OBJECTIVES: Recent genome-wide association and functional studies have shown that the ABCG2 gene encodes for a urate transporter, and a common causal ABCG2 variant, rs2231142, leads to elevated uric acid levels and prevalent gout among Whites and Blacks. We examined whether this finding is observed in a Japanese population, since Asians have a high reported prevalence of the T-risk allele. METHODS: A total of 3923 Japanese people from the Circulatory Risk in Communities Study aged 40-90 years were genotyped for rs2231142. Associations of the rs2231142 variant with serum uric acid levels and prevalence of gout and hyperuricaemia were examined. RESULTS: The frequency of the T-risk allele was 31% in this Japanese sample. Multivariable adjusted mean uric acid levels were 7-9 micromol/l higher for TG and TT than GG carriers (P-additive = 0.0006). The multivariable-adjusted odds ratio (OR) of prevalent gout was 1.37 (95% CI 0.68, 2.76) for TG and 4.37 (95% CI 1.98, 9.62) for TT compared with the GG carriers (P-additive = 0.001). When evaluating the combined outcome of hyperuricaemia and gout, the respective ORs were 1.40 (95% CI 1.04, 1.87) for TG and 1.88 (95% CI 1.23, 2.89) for TT carriers. The population attributable risk was 29% for gout and 19% for gout and/or hyperuricaemia. CONCLUSIONS: The association of the causal ABCG2 rs2231142 variant with uric acid levels and gout was confirmed in a sample of Japanese ancestry. Our study emphasizes the importance of this common causal variant in a population with a high risk allele frequency, especially as more Japanese adopt a Western lifestyle with a concomitant increase in mean serum uric acid levels.

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31 The rs2231142 variant in ABCG2 (Q141K) was genotyped in participants in community surveys in 2001 in Kyowa and in 2003 in Ikawa, as a part of the CIRCS. Login to comment
86 A recent functional and association study of Japanese people [8] has identified another causal SNP of ABCG2, Q126X, with less frequency of minor allele (1.8% among control), but stronger effect on gout/hyperuricaemia than presently analysed ABCG2 (Q141K). Login to comment

[hide] The genetic basis of hyperuricaemia and gout. Joint Bone Spine. 2011 Jan;78(1):35-40. Epub 2010 May 15. Merriman TR, Dalbeth N
The genetic basis of hyperuricaemia and gout.
Joint Bone Spine. 2011 Jan;78(1):35-40. Epub 2010 May 15., [PMID:20472486]

Abstract [show]
Gout results from elevated urate concentrations in the blood (hyperuricaemia). When super-saturation of urate is reached, monosodium urate crystals form within the joint. In some individuals, these crystals elicit a painful self-limiting inflammatory response that is characteristic of acute gouty arthritis. The most important cause of hyperuricaemia is reduced excretion of uric acid in the urine. Uric acid excretion is coordinated by a suite of urate transport molecules expressed in the renal collecting tubules, and is a key physiological checkpoint in gout. Other checkpoints in gout are hepatic production of urate, monosodium urate crystal formation, and initiation of the acute inflammatory response. Genome-wide association scans for genes regulating serum urate concentrations have identified two major regulators of hyperuricaemia- the renal urate transporters SLC2A9 and ABCG2. The risk variants at each gene approximately double the risk for gout in people of Caucasian ancestry, with SLC2A9 also resulting in higher risk for gout in people of Polynesian ancestry, a diverse population characterized by a high prevalence of gout. Ongoing genetic association studies are identifying and confirming other genes controlling serum urate concentrations; although genome-wide association studies in gout per se await recruitment of suitable case sample sets.

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84 ABCG2 A non-synonymous single nucleotide polymorphism (rs2231142; Q141K) is the likely etiological variant explaining the association of ABCG2 with gout [11,24]. Login to comment
152 Strong association of the ABCG2 Q141K variant with gout has been reported in people of Western Pacific (Samoa, Tonga) ancestry (OR > 2.5) but not in people of Eastern Pacific ancestry (Cook Island and New Zealand M¯aori) (OR < 1.4) [28]. Login to comment

[hide] A 'complexity' of urate transporters. Kidney Int. 2010 Sep;78(5):446-52. Epub 2010 Jul 7. Wright AF, Rudan I, Hastie ND, Campbell H
A 'complexity' of urate transporters.
Kidney Int. 2010 Sep;78(5):446-52. Epub 2010 Jul 7., [PMID:20613716]

Abstract [show]
Genetic variation in the SLC2A9 gene is a new genetic risk factor for low fractional excretion of uric acid, hyperuricemia, and gout. Its gene product, GLUT9, was previously known as a type II glucose/fructose transporter but is now known to function as a high-capacity uric acid transporter that is expressed in kidney, liver, and several other tissues. Follow-up meta-analyses, including one with data from 28,141 individuals, implicated a total of nine additional loci influencing serum urate concentrations, including six other membrane transporters (SLC17A1, SLC17A3, SLC22A11, SLC22A12, SLC16A9, and ABCG2). Variants in these genes together account for about 5% of the variance in serum urate, two-thirds of which is due to SLC2A9. Using these variants in 'Mendelian randomization' analyses provides a powerful means of dissecting the role of urate in cardiovascular and metabolic diseases, where cause-and-effect influences are difficult to discern due to potential confounding. The results highlight the complex interplay of membrane transporters involved in urate metabolism. They also show how variants of weak effect identified by genome-wide association studies can still be important in identifying novel pathways, including a 'complexity' of new and potentially druggable targets for modifying urate transport.

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35 ABCG2 encodes an ATP binding cassette transporter, expressed in the apical membranes of several tissues, including liver and human proximal renal tubular cells, where it transports a wide variety of substrates, including urate.21 Recently, ABCG2 has been shown to encode a high-capacity urate secretion transporter.21,22 Several loss-of-function ABCG2 variants have also been identified that confer an increased risk of gout, including a common missense mutation (Q141K) within the nucleotide binding domain (next to the equivalent phenylalanine residue commonly mutated in cystic fibrosis), which was shown to reduce urate efflux in Xenopus oocytes by 53%.21 This variant appears to be the causal ABCG2 variant associated with raised serum urate and gout in GWAS meta-analyses.20,23 It confers an adjusted odds ratio of 1.68 per allele for gout and has a minor allele frequency of 3% in blacks, 11% in whites, and about 30% in Asians. Login to comment

[hide] Production of cells with targeted integration of g... Methods Mol Biol. 2010;648:139-59. Wakabayashi-Nakao K, Tamura A, Koshiba S, Toyoda Y, Nakagawa H, Ishikawa T
Production of cells with targeted integration of gene variants of human ABC transporter for stable and regulated expression using the Flp recombinase system.
Methods Mol Biol. 2010;648:139-59., [PMID:20700710]

Abstract [show]
The vector-mediated introduction of cDNA into mammalian cells by calcium phosphate co-precipitation or permeation with lipofectamine is widely used for the integration of cDNA into genomic DNA. Such integration, however, of cDNA occurs randomly at unpredictable sites in the host's chromosomal DNA, and the number of integrated recombinant DNAs is not controllable. To overcome this problem, we developed the Flp-In method to integrate one single copy of cDNA encoding the human ABC transporter ABCG2 into FRT-tagged genomic DNA. Examination of more than 20 metaphase spreads for both fluorescence in situ hybridization (FISH) mapping and multicolor-FISH analysis revealed that ABCG2 cDNA was incorporated into the telomeric region of the short arm on one of chromosomes 12 in Flp-In-293 cells. By using those cells, we investigated the effect of genetic polymorphisms and post-translational modifications of human ABC transporter ABCG2 on the protein expression and degradation. On the basis of our experience, it has been concluded that the Flp recombinase system provides a useful tool to quantitatively analyze the protein stability and endoplasmic reticulum (ER)-associated degradation of proteins like the ABC transporter. This system is also applicable for similar studies of the biogenesis of other proteins using the secretory pathway as well as proteins with other cellular localizations.

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34 Furthermore, certain non-synonymous single nucleotide polymorphisms (SNPs), such as Q141K, F208S, and S441N, were also found to greatly affect the stability of ABCG2 in the ER and to enhance the protein degradation rate via ubiquitination and proteasomal proteolysis in Flp-In-293 cells (22-27) (Fig. 1). Login to comment

[hide] Influence of CYP3A5 and drug transporter polymorph... J Hum Genet. 2010 Nov;55(11):731-7. Epub 2010 Aug 19. Takahashi N, Miura M, Scott SA, Kagaya H, Kameoka Y, Tagawa H, Saitoh H, Fujishima N, Yoshioka T, Hirokawa M, Sawada K
Influence of CYP3A5 and drug transporter polymorphisms on imatinib trough concentration and clinical response among patients with chronic phase chronic myeloid leukemia.
J Hum Genet. 2010 Nov;55(11):731-7. Epub 2010 Aug 19., [PMID:20720558]

Abstract [show]
Imatinib mesylate (IM) trough concentration varies among IM-treated chronic myeloid leukemia (CML) patients. Although IM pharmacokinetics is influenced by several enzymes and transporters, little is known about the role of pharmacogenetic variation in IM metabolism. In this study, associations between IM trough concentration, clinical response and 11 single-nucleotide polymorphisms in genes involved in IM pharmacokinetics (ABCB1, ABCC2, ABCG2 CYP3A5, SLC22A1 and SLCO1B3) were investigated among 67 Japanese chronic phase CML patients. IM trough concentration was significantly higher in patients with a major molecular response than in those without one (P=0.010). No significant correlations between IM trough concentration and age, weight, body mass index or biochemical data were observed. However, the dose-adjusted IM trough concentration was significantly higher in patients with ABCG2 421A than in those with 421C/C (P=0.015). By multivariate regression analysis, only ABCG2 421A was independently predictive of a higher dose-adjusted IM trough concentration (P=0.015). Moreover, previous studies have shown that the ABCG2 421C>A (p.Q141K) variant is prevalent among Japanese and Han Chinese individuals and less common among Africans and Caucasians. Together, these data indicate that plasma IM concentration monitoring and prospective ABCG2 421C>A genotyping may improve the efficacy of IM therapy, particularly among Asian CML patients.

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231 44 Imai, Y., Nakane, M., Kage, K., Tsukahara, S., Ishikawa, E., Tsuruo, T. et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment
6 Moreover, previous studies have shown that the ABCG2 421C4A (p.Q141K) variant is prevalent among Japanese and Han Chinese individuals and less common among Africans and Caucasians. Login to comment
79 In addition, our study identified a significant correlation between ABCG2 421C4A (p.Q141K) and IM trough concentration, suggesting an important role for the BCRP efflux transporter in IM metabolism. Login to comment

[hide] Structure and function of the human breast cancer ... Curr Drug Metab. 2010 Sep;11(7):603-17. Ni Z, Bikadi Z, Rosenberg MF, Mao Q
Structure and function of the human breast cancer resistance protein (BCRP/ABCG2).
Curr Drug Metab. 2010 Sep;11(7):603-17., [PMID:20812902]

Abstract [show]
The human breast cancer resistance protein (BCRP/ABCG2) is the second member of the G subfamily of the large ATP-binding cassette (ABC) transporter superfamily. BCRP was initially discovered in multidrug resistant breast cancer cell lines where it confers resistance to chemotherapeutic agents such as mitoxantrone, topotecan and methotrexate by extruding these compounds out of the cell. BCRP is capable of transporting non-chemotherapy drugs and xenobiotiocs as well, including nitrofurantoin, prazosin, glyburide, and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine. BCRP is frequently detected at high levels in stem cells, likely providing xenobiotic protection. BCRP is also highly expressed in normal human tissues including the small intestine, liver, brain endothelium, and placenta. Therefore, BCRP has been increasingly recognized for its important role in the absorption, elimination, and tissue distribution of drugs and xenobiotics. At present, little is known about the transport mechanism of BCRP, particularly how it recognizes and transports a large number of structurally and chemically unrelated drugs and xenobiotics. Here, we review current knowledge of structure and function of this medically important ABC efflux drug transporter.

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22 Of these, Q141K is most extensively studied and has been found to be associated with inter-individual variations in the pharmacokinetics, response or toxicity of drugs [20-22] as well as genetic diseases such as gout [23]. Login to comment
244 The most important variant is Q141K, which occurs in Japanese and Chinese populations at high allele frequencies (30 - 60%) and in Caucasians and African-American populations at relatively low allele frequencies (5 - 10%) [111-112]. Login to comment
245 Several studies consistently revealed that Q141K had a lower protein expression level than wild-type BCRP in both transfected cells and human tissues [113-115]. Login to comment
246 The transport activity of Q141K in vivo would be expected to be decreased compared with wild-type BCRP owing to its lower level of protein expression. Login to comment
248 A recent study has revealed that Q141K undergoes increased lysosomal and proteasomal degradations than wild-type BCRP, possibly explaining the lower level of protein expression of the variant [119]. Login to comment

[hide] Fetoprotective activity of breast cancer resistanc... Drug Metab Rev. 2011 Feb;43(1):53-68. Epub 2010 Sep 20. Hahnova-Cygalova L, Ceckova M, Staud F
Fetoprotective activity of breast cancer resistance protein (BCRP, ABCG2): expression and function throughout pregnancy.
Drug Metab Rev. 2011 Feb;43(1):53-68. Epub 2010 Sep 20., [PMID:20854129]

Abstract [show]
The medical treatment of pregnant women, as well as their fetuses, has become a common clinical practice in developed countries. Therefore, detailed knowledge of maternofetal pharmacokinetics, including the role of drug-efflux transporters in the fetoplacental unit, is crucial to optimize drug choice and dosage schemes and to avoid or exploit possible drug-drug interactions on placental transporters in order to assure appropriate drug levels in the mother and/or fetus. Breast cancer resistance protein (BCRP, ABCG2) is the most recent member of ATP-binding cassette drug-efflux transporters that has been associated with resistance in cancer chemotherapy. Importantly, ABCG2 has also been localized in various normal tissues, affecting the pharmacokinetics of several xenobiotics as well as a number of physiological substances. Extensive expression of ABCG2 in tissue barriers, such as the blood-brain barrier, intestine, testis, or placenta, suggests that ABCG2 plays an important role in the protection of sensitive tissues against toxins. In the placenta, ABCG2 has been experimentally evidenced to actively pump its substrates in the fetal-to-maternal direction and to play an important role in transplacental pharmacokinetics, fetal protection, and detoxication. Further, ABCG2 expression in embryonic and fetal membranes over the course of pregnancy helps ensure proper function of the fetoplacental unit. In this review, we summarize the current knowledge regarding expression and function of ABCG2 in the fetoplacental unit during the development of the fetus and overview the aspects of transplacental pharmacokinetics, ABCG2 regulation, and clinical significance of the transporter for pharmacotherapy in pregnancy.

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183 The nonsynonymous C421A SNP that results in a glycine to lysine (Q141K) amino-acid change has been linked to decreased plasma membrane expression of ABCG2 (Imai et al., 2002; Morisaki et al., 2005) and, consequently, altered drug pharmacokinetics (Sparreboom et al., 2004, 2005; Zamboni et al., 2006). Login to comment
191 In a recent work by Pollex et al. (2010), glyburide transport was studied in wild-type (Arg482) and SNP (Q141K) HEK-293 cells over a range of substrate concentrations (0.2-100 μM). Login to comment
192 The investigators conclude that the Q141K variant of ABCG2 may have the potential to alter the placental pharmacokinetics of glyburide and other clinically used ABCG2 substrate drugs in pregnancy (Pollex et al., 2010). Login to comment

[hide] A strong role for the ABCG2 gene in susceptibility... Hum Mol Genet. 2010 Dec 15;19(24):4813-9. Epub 2010 Sep 21. Phipps-Green AJ, Hollis-Moffatt JE, Dalbeth N, Merriman ME, Topless R, Gow PJ, Harrison AA, Highton J, Jones PB, Stamp LK, Merriman TR
A strong role for the ABCG2 gene in susceptibility to gout in New Zealand Pacific Island and Caucasian, but not Maori, case and control sample sets.
Hum Mol Genet. 2010 Dec 15;19(24):4813-9. Epub 2010 Sep 21., 2010-12-15 [PMID:20858603]

Abstract [show]
Genetic variation in ABCG2 (rs2231142, Q141K), encoding a uric acid transporter, is associated with gout in diverse populations. The aim of this study was to examine a role for ABCG2 in gout susceptibility in New Zealand Maori, Pacific Island and Caucasian samples. Patients (n = 185, 173 and 214, for Maori, Pacific Island and Caucasian, respectively) satisfied the American College of Rheumatology gout classification criteria. The comparison samples comprised 284, 129 and 562 individuals, respectively, without gout. rs2231142 was genotyped and stratification accounted for using genomic control markers. Association of the minor allele of rs2231142 with gout was observed in the Pacific Island samples (OR = 2.80, P(STRAT) < 0.001 after accounting for effects of population structure), but not in the Maori samples (OR = 1.08, P(STRAT)= 0.70), with heterogeneity in association evident between the Maori and Pacific Island datasets (P(HET) = 0.001). A similar dichotomy in association was observed when samples were stratified into Western (Tonga, Samoa, Niue, Tokelau) versus Eastern Polynesian (Maori, Cook Island) origin (OR = 2.59, P(STRAT) < 0.001; OR = 1.12, P(STRAT)= 0.48, respectively; P(HET) = 0.005). Association with gout was observed in the Caucasian samples (OR = 2.20, P = 3.2 x 10(-8)). Unlike SLC2A9, which is a strong risk factor for gout in both Maori and Pacific Island people, ABCG2 rs2231142 has a strong effect only in people of Western Polynesian ancestry. Our results emphasize the need to account for sub-population differences when undertaking biomedical genetic research in a group defined by a geographical region and shared ancestry but characterized by migratory events that create bottlenecks and altered genetic structure in the founder populations.

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0 A strong role for the ABCG2 gene in susceptibility to gout in New Zealand Pacific Island and Caucasian, but not Ma¯ori, case and control sample sets Amanda J. Phipps-Green1, Jade E. Hollis-Moffatt1, Nicola Dalbeth3, Marilyn E. Merriman1, Ruth Topless1, Peter J. Gow4, Andrew A. Harrison5, John Highton2, Peter B.B. Jones3, Lisa K. Stamp6 and Tony R. Merriman1,* 1 Department of Biochemistry and 2 Department of Medicine, University of Otago, Dunedin, New Zealand, 3 Department of Medicine, University of Auckland, Auckland, New Zealand, 4 Department of Rheumatology, Middlemore Hospital, Auckland, New Zealand, 5 Department of Medicine, University of Otago, Wellington, New Zealand and 6 Department of Medicine, University of Otago, Christchurch, New Zealand Received June 24, 2010; Revised September 8, 2010; Accepted September 16, 2010 Genetic variation in ABCG2 (rs2231142, Q141K), encoding a uric acid transporter, is associated with gout in diverse populations. Login to comment
23 In addition, a missense single nucleotide polymorphism (SNP) (rs2231142; Q141K) in ABCG2 has been associated with hyperuricaemia and gout in Caucasian, Han Chinese, Japanese and African-American samples (4,7-10). Login to comment
25 The lysine allele of Q141K, which explains the genetic association in Caucasian populations, encodes a transporter with 50% reduced activity (7,9). Login to comment
26 Given the very high rate of gout in NZ Ma¯ori and Pacific Island people, here we tested whether or not ABCG2 [rs2231142 (Q141K)] conferred a strong risk for gout in case-control samples drawn from these populations, and in an NZ Caucasian case-control sample set. Login to comment
60 DISCUSSION Association of rs2231142 (Q141K) with serum urate levels and gout in Caucasian and Japanese is established (4,7,9,10), and has been reported in Han Chinese and African-American sample sets (4,8). Login to comment
64 Analysis of association of rs2231142 (ABCG2: Q141K) in NZ Ma¯ori, Pacific Island and Caucasian case-control samples Genotype, no. Login to comment
74 Given the associations of ABCG2 Q141K reported in sample sets drawn from diverse populations (Caucasian, Japanese, Han Chinese, African-American, Pacific Island; Table 1) (4,7-10), the lack of association observed in NZ Ma¯ori is notable. Login to comment
96 Given that ABCG2 Q141K determines serum urate levels, where a gender effect is observed (4), and that gout is a condition of extreme serum urate levels in both men and women, a gender effect of ABCG2 in gout would not be expected since all cases, whether male or female, are already hyperuricaemic. Login to comment
22 In addition, a missense single nucleotide polymorphism (SNP) (rs2231142; Q141K) in ABCG2 has been associated with hyperuricaemia and gout in Caucasian, Han Chinese, Japanese and African-American samples (4,7-10). Login to comment
24 The lysine allele of Q141K, which explains the genetic association in Caucasian populations, encodes a transporter with 50% reduced activity (7,9). Login to comment
59 DISCUSSION Association of rs2231142 (Q141K) with serum urate levels and gout in Caucasian and Japanese is established (4,7,9,10), and has been reported in Han Chinese and African-American sample sets (4,8). Login to comment
63 Analysis of association of rs2231142 (ABCG2: Q141K) in NZ Ma¯ori, Pacific Island and Caucasian case-control samples Genotype, no. Login to comment

[hide] Association of multi-drug resistance gene polymorp... Cancer. 2011 Feb 15;117(4):744-51. doi: 10.1002/cncr.25510. Epub 2010 Oct 4. Tanaka M, Okazaki T, Suzuki H, Abbruzzese JL, Li D
Association of multi-drug resistance gene polymorphisms with pancreatic cancer outcome.
Cancer. 2011 Feb 15;117(4):744-51. doi: 10.1002/cncr.25510. Epub 2010 Oct 4., 2011-02-15 [PMID:20922799]

Abstract [show]
BACKGROUND: The purpose of this study was to identify single nucleotide polymorphisms (SNPs) of multidrug resistance genes that are associated with clinical outcome in patients with potentially resectable pancreatic adenocarcinoma who were treated with preoperative gemcitabine-based chemoradiotherapy at M. D. Anderson Cancer Center. METHODS: We selected 8 SNPs of 7 drug resistance genes, including MDR1 (ABCB1), MRP1-5 (ABCC1-5), and BCRP (ABCG2), reported to be important in mediating drug resistance. Genotype was determined by the Taqman method. The associations of genotype with tumor response to therapy and overall survival (OS) were evaluated using log-rank test, Cox regression, and logistic regression models. RESULTS: MRP5 A-2G AA genotype showed significant association with OS (log-rank P = .010). The hazard ratio (95% confidence interval) was 1.65 (1.11-2.45) after adjusting for clinical predictors. The MRP2 G40A GG genotype had a weak association with reduced OS (log-rank P = .097). A combined effect of the two genotypes on OS was observed. Patients with none of the adverse genotypes had a median survival time (MST) of 34.0 months, and those with 1-2 deleterious alleles had a significantly lower MST of 20.7 months (log-rank P = .006). MRP2 G40A GG genotype was also significantly associated with poor histological response to chemoradiotherapy (P = .028). CONCLUSIONS: These observations suggest a potential role of polymorphic variants of drug resistance genes in predicting therapeutic efficacy and survival of patients with potentially resectable pancreatic cancer.

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75 Minor Allele Frequency Observeda /Reportedb MDR1 7q21.12a Ex27 -55T>C I1145I 1045642 0.49/0.47 MRP1 16p13.11a Ex28 36G>A S1219S 2239330 0.28/0.24 MRP2 10q24.2c Ex10 40G>A V417I 2273697 0.25/0.23 Ex28 -16C>T I1324I 3740066 0.35/0.35 MRP3 17q21.33b Ex26 -13C>T H1314H 2277624 0.24/0.17 MRP4 13q32.1a Ex8 40A>G R317R 2274406 0.39/0.37 MRP5 3q27.1b Ex10 -2A>G Q382Q 7636910 0.35/0.39 BCRP 4q22.1b Ex5 43C>A Q141K 2231142 0.12/0.10 SNP indicates single-nucleotide polymorphism; RS No., reference SNP identification number. Login to comment

[hide] Effect of ABCG2 genotypes on the pharmacokinetics ... Eur J Clin Pharmacol. 2011 Feb;67(2):129-34. Epub 2010 Oct 23. Kim KA, Joo HJ, Park JY
Effect of ABCG2 genotypes on the pharmacokinetics of A771726, an active metabolite of prodrug leflunomide, and association of A771726 exposure with serum uric acid level.
Eur J Clin Pharmacol. 2011 Feb;67(2):129-34. Epub 2010 Oct 23., [PMID:20972558]

Abstract [show]
OBJECTIVE: It has been reported that leflunomide and its active metabolite, A771726, are substrates of the ABCG2 (BCRP) transporter in vitro. Recent genome-wide association studies have shown that ABCG2 transporter modulates serum uric acid (UA) levels. We explored the role of ABCG2 genotypes in the pharmacokinetics of A771726 and the relationship between serum UA levels and pharmacokinetics of A771726 in healthy participants. METHODS: Twenty-four healthy individuals were recruited and genotyped for ABCG2. After administration of a single dose of 20 mg leflunomide, plasma concentrations of A771726 were measured. Serum UA levels were measured just before medication, and ABCG2 c.421C>A and c.34G> A polymorphism were genotyped. RESULTS: ABCG2 c.421C>A but not c.34G>A substantially influenced the pharmacokinetics of A771726. A771726 C(max) was 30% higher, area under the concentration-time curve (AUC) 83% larger, and oral clearance (CL/F) 41% lower in c.421C>A carriers than in noncarriers. Serum UA levels were also higher in carriers than in noncarriers and exhibited a strong and positive correlation with A771726 AUC (Spearman r = 0.6746, P = 0.0003), but a negative correlation was observed with A771726 CL/F (Spearman r = -0.6616, P = 0.0004). CONCLUSION: ABCG2 c.421C>A but not c.34G>A polymorphism appears to be a major determinant of interindividual variability in A771726 disposition. Additionally, serum UA levels exhibited a strong correlation with exposure to A771726.

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17 Several ABCG2 genetic variants have been reported; the most frequent ABCG2 polymorphisms detected among different ethnic groups are c.34G>A (rs2231137), which codes for V12M, and c.421C>A (rs2231142), which codes for Q141K [4]. Login to comment

[hide] ABCG2 polymorphisms, 34G>A and 421C>A in a Korean ... J Clin Pharm Ther. 2010 Dec;35(6):705-12. doi: 10.1111/j.1365-2710.2009.01127.x. Kim KA, Joo HJ, Park JY
ABCG2 polymorphisms, 34G>A and 421C>A in a Korean population: analysis and a comprehensive comparison with other populations.
J Clin Pharm Ther. 2010 Dec;35(6):705-12. doi: 10.1111/j.1365-2710.2009.01127.x., [PMID:21054463]

Abstract [show]
BACKGROUND AND OBJECTIVE: ABCG2, also known as Breast Cancer Resistance Peptide (BCRP) or mitoxantrone-resistant protein, is the second member of the G-family of ABC transporters. The frequencies of ABCG2 34G>A and 421C>A polymorphisms in a Korean population were assessed using a newly developed multiplex pyrosequencing method, and compared with the corresponding frequencies seen in other ethnic groups. METHOD: We designed a multiplex pyrosequencing method to simultaneously detect ABCG2 421C>A and 34G>A polymorphisms and analysed the allele frequencies of these polymorphisms in 250 Korean subjects. RESULTS: The results showed 100% concordance between single and multiplex pyrosequencing methods. We also validated the polymorphisms identified by pyrosequencing with a direct sequencing method using randomly selected samples. The allele frequencies of ABCG2 421C>A and 34G>A in the population tested were 0.298 and 0.190 respectively. The allele frequency of the 421C>A polymorphism is comparable to other Asian populations, including Japanese and Chinese. However, both frequencies are different from those of Caucasians and Africans. CONCLUSIONS: The multiplex pyrosequencing method used to detect two ABCG2 polymorphisms concurrently is a rapid and reliable genotyping method for the detection of important ABCG2 genetic polymorphisms. The ABCG2 34G>A and 421C>A polymorphisms are frequently found in the Korean population. The frequencies are similar to those seen in other Asian populations including Japanese and Chinese, but very different to those of Caucasian and African-American populations.

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21 The most frequent ABCG2 polymorphisms detected among different ethnic groups are 34G>A, which codes for V12M, and 421C>A, which codes for Q141K (14). Login to comment
81 Comparisons of ABCG2 allele frequencies with other ethnic groups SNP Amino acid change Population (n) Frequency (%) Reference G34A V12M Korea (250) 19Æ8 Present study Southeast Asians (20) 45Æ0 14 Chinese (20) 20Æ0 14 Japanese (124) 19Æ0 15 Japanese (120) 17Æ5 27 Japanese (20) 15Æ0 14 Caucasian (150) 10Æ3 19 Ashkenazi Jewish (20) 10Æ0 14 Mexicans (20) 10Æ0 14 Dutch (100) 6Æ5 37 African-American (150) 6Æ3 27 Middle Eastern (40) 5Æ0 14 Caucasian (150) 3Æ7 27 Swedish (60) 1Æ7 36 C421A Q141K Korea (250) 27Æ8 Present study Japanese (20) 35Æ0 14 Chinese (20) 35Æ0 14 Chinese (95) 34Æ2 22 Japanese (120) 32Æ7 27 Japanese (124) 26Æ6 15 Southeast Asians (20) 15Æ0 14 Middle Eastern (40) 13Æ0 14 Dutch (100) 12Æ0 37 Caucasian (172) 11Æ3 22 Mexicans (20) 10Æ0 14 Swedish (60) 10Æ0 36 Caucasian (150) 8Æ7 19 Ashkenazi Jewish (20) 5Æ0 14 African-American (150) 2Æ3 27 was demonstrated that 421C>A carriers have a 1Æ3-fold decrease in ATPase activity compared to wild-type (19), and the bioavailability of sulfasalazine, diflomotecan and topotecan was significantly elevated by this polymorphism (29-31). Login to comment

[hide] Decreased hepatic breast cancer resistance protein... Drug Metab Dispos. 2011 Mar;39(3):441-7. Epub 2010 Nov 24. Yue W, Lee JK, Abe K, Sugiyama Y, Brouwer KL
Decreased hepatic breast cancer resistance protein expression and function in multidrug resistance-associated protein 2-deficient (TR) rats.
Drug Metab Dispos. 2011 Mar;39(3):441-7. Epub 2010 Nov 24., [PMID:21106720]

Abstract [show]
Multidrug resistance-associated protein (Mrp) 2-deficient (TR(-)) Wistar rats have been used to elucidate the role of Mrp2 in drug disposition. Decreased breast cancer resistance protein (Bcrp) levels were reported in sandwich-cultured hepatocytes (SCH) from TR(-) rats compared with those from wild-type (WT) rats. This study was designed to characterize hepatic Bcrp expression and function in TR(-) rats, using nitrofurantoin and pitavastatin as substrates. Bcrp was knocked down by RNA interference in rat SCH. Antibody BXP53, but not BXP21, specifically detected Bcrp knockdown in SCH. Bcrp protein levels were decreased markedly in TR(-) but not Mrp2-deficient Sprague-Dawley [Eisai hyperbilirubinemic rats (EHBR)] rats. Bcrp mRNA levels were decreased significantly in TR(-) livers as determined by TaqMan real-time reverse transcriptase-polymerase chain reaction. Biliary excretion of nitrofurantoin, a specific Bcrp substrate, was decreased significantly in SCH and isolated perfused livers from TR(-) rats compared with those from WT controls, indicating that hepatic Bcrp function is decreased in TR(-) rats. In Bcrp knockdown SCH, the biliary excretion index and in vitro biliary clearance of pitavastatin were decreased significantly to approximately 58 and approximately 52% of control, respectively, indicating that Bcrp plays a role in pitavastatin biliary excretion. Pitavastatin biliary excretion was decreased significantly in perfused livers from TR(-) compared with those from WT rats. In conclusion, expression and function of hepatic Bcrp are decreased significantly in TR(-) rats. The potential role of both Bcrp and Mrp2 should be considered when data generated in TR(-) rats are interpreted. TR(-) and EHBR rats in combination may be useful in differentiating the role of Mrp2 and Bcrp in drug/metabolite disposition.

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15 A single nucleotide polymorphism in ABCG2 C421A (Q141K) has been associated with altered drug disposition in clinical studies (elevated plasma concentrations of diflomotecan after intravenous administration and of topotecan and rosuvastatin after oral administration) (Sparreboom et al., 2004, 2005; Zhang et al., 2006a). Login to comment

[hide] The ABCG family of membrane-associated transporter... Br J Pharmacol. 2010 Dec 22. doi: 10.1111/j.1476-5381.2010.01177.x. Kerr ID, Haider AJ, Gelissen IC
The ABCG family of membrane-associated transporters: you don't have to be big to be mighty.
Br J Pharmacol. 2010 Dec 22. doi: 10.1111/j.1476-5381.2010.01177.x., 2010-12-22 [PMID:21175590]

Abstract [show]
Along with many other mammalian ATP binding cassette (ABC) transporters, members of the ABCG group are involved in the regulated transport of hydrophobic compounds across cellular membranes. In humans, 5 ABCG family members have been identified, encoding proteins ranging from 638 to 678 amino acids in length. All 5 have been the subject of intensive investigation to better understand their physiological roles, expression patterns, interactions with substrates and inhibitors, and regulation at both the transcript and protein level. The principal substrates for at least 4 of the ABCG proteins are endogenous and dietary lipids, with ABCG1 implicated in particular in the export of cholesterol, and ABCG5 and G8 forming a functional heterodimer responsible for plant sterol elimination from the body. ABCG2 has a much broader substrate specificity and its ability to transport numerous diverse pharmaceuticals has implications for the absorption, distribution, metabolism, excretion, and toxicity (ADMETox) profile of these compounds. ABCG2 is one of at least three so-called multidrug resistant (MDR) ABC-transporters expressed in humans, and its activity is associated with decreased efficacy of anticancer agents in several carcinomas. In addition to its role in cancer, ABCG2 also plays a role in the normal physiological transport of urate and haem, the implications of which are described below. We summarize here data on all 5 human ABCG transporters and provide a current perspective on their roles in human health and disease.

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143 The effects in vivo are rather less well understood, with the exception of the SNP rs2231142 (resulting in a Gln141Lys change). Login to comment
145 Interestingly, previous analysis had suggested that the Gln141Lys variant of ABCG2 was only subtly different in terms of export of chemotherapeutic drugs compared to wild type ABCG2 (Tamura et al., 2007), which emphasises that studies on the impact of SNPs on ABCG2 function have to be taken into the context of the extreme diversity of transport substrates. Login to comment
167 The effects in vivo are rather less well understood, with the exception of the SNP rs2231142 (resulting in a Gln141Lys change). Login to comment
169 Interestingly, previous analysis had suggested that the Gln141Lys variant of ABCG2 was only subtly different in terms of export of chemotherapeutic drugs compared with wild-type ABCG2 (Tamura et al., 2007), which emphasizes that studies on the impact of SNPs on ABCG2 function have to be taken Apo-A1 HDL sitosterol lipid droplets urate lumen ABCG2 ABCG5/G8 Apo-A1 HDL sitosterol lipidpp droplets urate lumen ABCG2 ABCG5/G8 lumen ABCG5/G8 sitosterol Figure 2 Principal functions of human ABCG transporters. Login to comment

[hide] Posttranslational negative regulation of glycosyla... Biochem Biophys Res Commun. 2011 Jan 21;404(3):853-8. Epub 2010 Dec 22. Sugiyama T, Shuto T, Suzuki S, Sato T, Koga T, Suico MA, Kusuhara H, Sugiyama Y, Cyr DM, Kai H
Posttranslational negative regulation of glycosylated and non-glycosylated BCRP expression by Derlin-1.
Biochem Biophys Res Commun. 2011 Jan 21;404(3):853-8. Epub 2010 Dec 22., 2011-01-21 [PMID:21184741]

Abstract [show]
Human breast cancer resistance protein (BCRP)/MXR/ABCG2 is a well-recognized ABC half-transporter that is highly expressed at the apical membrane of many normal tissues and cancer cells. BCRP facilitates disposition of endogenous and exogenous harmful xenobiotics to protect cells/tissues from xenobiotic-induced toxicity. Despite the enormous impact of BCRP in the physiological and pathophysiological regulation of the transport of a wide variety of substrates, little is known about the factors that regulate posttranslational expression of BCRP. Here, we identified Derlin-1, a member of a family of proteins that bears homology to yeast Der1p, as a posttranslational regulator of BCRP expression. Overexpression of Derlin-1 suppressed ER to Golgi transport of wild-type (WT) BCRP that is known to be efficiently trafficked to the plasma membrane. On the other hand, protein expression of N596Q variant of BCRP, N-linked glycosylation-deficient mutant that preferentially undergoes ubiquitin-mediated ER-associated degradation (ERAD), was strongly suppressed by the overexpression of Derlin-1, whereas knockdown of Derlin-1 stabilized N596Q protein, suggesting a negative regulatory role of Derlin-1 for N596Q protein expression. Notably, knockdown of Derlin-1 also stabilized the expression of tunicamycin-induced deglycosylated WT BCRP protein, implying the importance of glycosylation state for the recognition of BCRP by Derlin-1. Thus, our data demonstrate that Derlin-1 is a negative regulator for both glycosylated and non-glycosylated BCRP expression and provide a novel posttranslational regulatory mechanism of BCRP by Derlin-1.

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29 Moreover, the certain single nucleotide polymorphism (SNP) variants of BCRP (Q141K, F208S and S441N), which protein expression was markedly low despite the functional expression of mRNA, were also degraded by ERAD [10,11]. Login to comment
162 These include naturally occurring SNPs variants of BCRP such as Q141K, F208S and S441N [10,11]. Login to comment

[hide] Key Role of Human ABC Transporter ABCG2 in Photody... Adv Pharmacol Sci. 2010;2010:587306. Epub 2010 Jul 8. Ishikawa T, Nakagawa H, Hagiya Y, Nonoguchi N, Miyatake S, Kuroiwa T
Key Role of Human ABC Transporter ABCG2 in Photodynamic Therapy and Photodynamic Diagnosis.
Adv Pharmacol Sci. 2010;2010:587306. Epub 2010 Jul 8., [PMID:21188243]

Abstract [show]
Accumulating evidence indicates that ATP-binding cassette (ABC) transporter ABCG2 plays a key role in regulating the cellular accumulation of porphyrin derivatives in cancer cells and thereby affects the efficacy of photodynamic therapy and photodynamic diagnosis. The activity of porphyrin efflux can be affected by genetic polymorphisms in the ABCG2 gene. On the other hand, Nrf2, an NF-E2-related transcription factor, has been shown to be involved in oxidative stress-mediated induction of the ABCG2 gene. Since patients have demonstrated individual differences in their response to photodynamic therapy, transcriptional activation and/or genetic polymorphisms of the ABCG2 gene in cancer cells may affect patients' responses to photodynamic therapy. Protein kinase inhibitors, including imatinib mesylate and gefitinib, are suggested to potentially enhance the efficacy of photodynamic therapy by blocking ABCG2-mediated porphyrin efflux from cancer cells. This review article provides an overview on the role of human ABC transporter ABCG2 in photodynamic therapy and photodynamic diagnosis.

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158 The most extensively studied among those SNPs with potential clinical relevance is 421 C > A resulting in a glutamine to lysine substitution (Q141K) in the ABCG2 protein. Login to comment
159 The Q141K SNP has been identified with varying frequencies in different ethnic groups and was found to be the most relevant in Japanese and Chinese populations (approximately 30% in the allele frequency). Login to comment
161 Q141K has been associated with lower levels of protein expression and impaired transport in vitro [79-84]. Login to comment
163 Furthermore, the Q141K SNP was reportedly associated with a higher incidence of diarrhea in nonsmall cell lung cancer patients treated with gefitinib [88]. Login to comment
164 It has been demonstrated that the reduced expression levels of the Q141K variant may be due to its ubiquitin-mediated proteasomal degradation [89]. Login to comment
167 Based on the currently available data on SNPs and acquired mutations, we have created a total of 18 variant forms of ABCG2 (V12M, G51C, Q126stop, Q141K, T153M, Q166E, I206L, F208S, S248P, E334stop, F431L, S441N, R482G, R482T, F489L, F571I, N590Y, and D620N) by site-directed mutagenesis and expressed them in insect cells [41, 90]. Login to comment
177 Gefitinib and imatinib are new anticancer drugs Outside Plasma membrane Inside H2N COOH V12M G51C Q126stop Q141K T153M R160Q Q166E I206L F208S S248P E334stop F431L F489L S441N R482G R482T F571I R575stop N590Y D620N T542A A528T D296H P269S ATP-binding cassette (a) 0 0.1 0.3 0.4 0.2 0.5 Mock WT V12M G51C Q126stop Q141K T153M Q166E I206L F208S S248P E334stop F431L S441N F489L F571I N590Y D620N R482G R482T ATP-dependenthematoporphyrin transport(nmol/min/mgprotein) (b) Figure 4: (a) Schematic illustration of human ABCG2 and its nonsynonymous polymorphisms. Login to comment

[hide] Ixabepilone, a novel microtubule-targeting agent f... J Pharmacol Exp Ther. 2011 May;337(2):423-32. Epub 2011 Jan 24. Shen H, Lee FY, Gan J
Ixabepilone, a novel microtubule-targeting agent for breast cancer, is a substrate for P-glycoprotein (P-gp/MDR1/ABCB1) but not breast cancer resistance protein (BCRP/ABCG2).
J Pharmacol Exp Ther. 2011 May;337(2):423-32. Epub 2011 Jan 24., [PMID:21262849]

Abstract [show]
Ixabepilone is the first epothilone to be approved for clinical use. Current data suggest the epothilones have a role in treating taxane-resistant cancers and ixabepilone is unaffected by at least some of the mechanisms underlying chemoresistance. Here, we report a series of cytotoxicity and transport studies to assess the potential role of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in ixabepilone resistance. A significant decrease in ixabepilone-mediated cytotoxicity was observed in Madin-Darby canine kidney cells transfected with human multidrug resistance 1 (MDR1) comparative with the parental cells (IC(50) > 2000 nM versus 90 nM). Overexpression of P-gp also resulted in significantly decreased cell susceptibility to docetaxel, paclitaxel, and vinblastine. Bidirectional transport of ixabepilone across monolayers of porcine kidney-derived cells expressing human MDR1 showed a significantly increased efflux ratio relative to the parental cells. A BCRP-overexpressing cell line was developed by transfecting human embryonic kidney (HEK)-293 cells with BCRP cDNA and confirmed by immunoblotting and bodipy prazosin and mitoxantrone uptake. Neither P-gp nor multidrug resistance protein 2 was detected in the cells by corresponding polyclonal antibodies. This HEK-BCRP cell line demonstrated resistance to docetaxel, paclitaxel, vinblastine, and mitoxantrone, in comparison with the parental cell line (7.3, 4.3, 2.9, and 11.9 resistance factor, respectively). Transport inhibition by BCRP inhibitor fumitremorgin C and broad efflux inhibitor N-(4-[2-(1,2,3,4-tetrahydro-6,7-dimethoxy-2-isoquinolinyl)ethyl]-phenyl)-9 ,10-dihydro-5-methoxy-9-oxo-4-acridine carboxamide (GF120918) restored drug sensitivity. In contrast, ixabepilone was far less susceptible to BCRP-mediated resistance, resulting in a resistance factor of only 1.2-fold. In summary, these results suggest that P-gp could cause resistance to ixabepilone in tumors and affect the disposition of the drug, but it is unlikely that BCRP mediates any drug resistance to ixabepilone.

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37 A significant association between survival beyond 15 months and the ABCG2 421CϾA (Q141K) polymorphism was observed in a clinical trial (p ϭ 0.05) of 64 patients with hormone-refractory prostate cancer randomized to receive docetaxel plus vinorelbine or docetaxel plus estramustine phosphate. Login to comment

[hide] Association of genetic polymorphisms in the influx... Ther Drug Monit. 2011 Apr;33(2):244-50. Yamakawa Y, Hamada A, Nakashima R, Yuki M, Hirayama C, Kawaguchi T, Saito H
Association of genetic polymorphisms in the influx transporter SLCO1B3 and the efflux transporter ABCB1 with imatinib pharmacokinetics in patients with chronic myeloid leukemia.
Ther Drug Monit. 2011 Apr;33(2):244-50., [PMID:21311410]

Abstract [show]
This study explored the association of 14 single nucleotide polymorphisms in three genes coding for influx transporters (SLC22A1, SLCO1B1, and SLCO1B3), two genes coding for efflux transporters (ABCB1 and ABCG2), and four genes coding for enzymes (CYP2C9, CYP2C19, CYP2D6, and CYP3A5) with the pharmacokinetics of imatinib in Japanese patients with chronic myeloid leukemia. Pharmacokinetic parameters were estimated by a population pharmacokinetic analysis based on 622 plasma samples from 34 patients at steady state. Approximately 4.6-fold variability in individual clearance was observed (range, 3.4-15.5 L/hr). The individual estimated clearance was significantly increased in patients with the SLCO1B3 334GG genotype (median value +/- standard deviation, 9.5 +/- 3.1 L/hr; n = 19) compared with SLCO1B3 334TT and TG genotypes (7.0 +/- 3.1 L/hr; n = 15) (P = 0.019). Patients with the ABCB1 3435CC genotype had significantly higher imatinib clearance (12.7 +/- 3.0 L/hr; n = 7) compared with patients with ABCB1 3435CT and TT genotypes (7.9 +/- 2.7 L/hr; n = 27) (P = 0.035). In conclusion, the present study suggests that single nucleotide polymorphisms of the influx transporter SLCO1B3 and the efflux transporter ABCB1 were functionally associated with individual variability of imatinib pharmacokinetics in Japanese patients with chronic myeloid leukemia.

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77 Individual Clearance of Imatinib as a Function of Variant Genotypes Allele Region Effect* Number of Patients Allele Frequency† WT Het Var p q SLC22A11022C.T Exon6 P341L 24 8 2 0.82 0.18 SLCO1B1521T.C Exon6 V174A 26 7 1 0.87 0.13 SLCO1B3334T.G Exon3 S112A 5 10 19 0.29 0.71 ABCB11236C.T Exon12 G412G 5 19 10 0.43 0.57 ABCB12677G.T/A Exon21 A893S/T 10 14 10 0.50 0.50 ABCB13435C.T Exon26 I1145I 7 21 6 0.51 0.49 ABCG2421C.A Exon5 Q141K 21 13 0 0.81 0.19 CYP2C9430C.T Exon3 R144C 34 0 0 1.00 0.00 CYP2C91075A.C Exon7 I359L 33 1 0 0.99 0.01 CYP2C19636G.A Exon5 W212X 27 7 0 0.90 0.10 CYP2C19681G.A Exon4 Splice defect 16 13 5 0.66 0.34 CYP2D6100C.T Exon1 P34S 13 16 5 0.62 0.38 CYP2D61846G.A Exon4 Splice defect 34 0 0 1.00 0.00 CYP3A56986A.G Intron3 Splice defect 2 12 20 0.24 0.76 Allele Clearance 6 SD (L/hr) P WT Het Var WT vs Het vs Var WT + Het vs Var WT vs Het + Var SLC22A11022C.T 8.8 6 2.9 8.2 6 3.5 8.3 6 3.5 0.973 0.913 0.926 SLCO1B1521T.C 8.7 6 3.1 8.3 6 2.0 5.5 0.304 0.235 0.288 SLCO1B3334T.G 5.7 6 2.3 7.4 6 2.4 9.5 6 3.1 0.046 0.019 0.071 ABCB11236C.T 11.7 6 3.1 8.1 6 2.7 8.2 6 3.2 0.277 0.642 0.196 ABCB12677G.T/A 9.8 6 3.2 8.1 6 2.5 8.0 6 3.3 0.494 0.445 0.270 ABCB13435C.T 12.7 6 3.0 7.9 6 2.4 9.2 6 3.5 0.058 0.581 0.035 ABCG2421C.A 7.9 6 3.1 9.5 6 2.8 N/A 0.446 N/A 0.462 CYP2C9430C.T 8.3 6 3.0 N/A N/A N/A N/A N/A CYP2C91075A.C 8.6 6 3.0 10.0 N/A 0.508 N/A 0.647 CYP2C19636G.A 8.1 6 2.8 9.7 6 3.3 N/A 0.242 N/A 0.257 CYP2C19681G.A 8.6 6 3.6 9.5 6 2.6 7.0 6 1.1 0.473 0.252 0.852 CYP2D6100C.T 7.3 6 3.2 8.2 6 2.4 12.7 6 3.3 0.190 0.089 0.276 CYP2D61846G.A 8.3 6 3.0 N/A N/A N/A N/A N/A CYP3A56986A.G 6.3 6 1.0 7.9 6 3.4 9.4 6 2.8 0.285 0.323 0.175 *Number represents amino acid codon. Login to comment

[hide] Genetic polymorphisms of ATP-binding cassette (ABC... Int J Mol Epidemiol Genet. 2010;1(3):201-7. Epub 2010 May 20. Hampras SS, Sucheston L, Weiss J, Baer MR, Zirpoli G, Singh PK, Wetzler M, Chennamaneni R, Blanco JG, Ford L, Moysich KB
Genetic polymorphisms of ATP-binding cassette (ABC) proteins, overall survival and drug toxicity in patients with Acute Myeloid Leukemia.
Int J Mol Epidemiol Genet. 2010;1(3):201-7. Epub 2010 May 20., [PMID:21311724]

Abstract [show]
The overall survival of patients with acute myeloid leukemia (AML) remains poor due to both intrinsic and acquired chemotherapy resistance. Over expression of ATP binding cassette (ABC) proteins in AML cells has been suggested as a putative mechanism of drug resistance. Genetic variation among individuals affecting the expression or function of these proteins may contribute to inter-individual variation in treatment outcomes. DNA from pre-treatment bone marrow or blood samples from 261 patients age 20-85 years, who received cytarabine and anthracycline-based therapy at Roswell Park Cancer Institute between 1994 and 2006, was genotyped for eight non-synonymous single nucleotide polymorphisms in the ABCB1, ABCC1 and ABCG2 drug transporter genes. Heterozygous (AG) or homozygous (AA) variant genotypes for rs2231137 (G34A) in the ABCG2 (BRCP) gene, compared to the wild type (GG) genotype were associated with both significantly improved survival (HR=0.44, 95%CI=0.25-0.79), and increased odds for toxicity (OR=8.41, 95%CI= 1.10-64.28). Thus genetic polymorphisms in the ABCG2 (BRCP) gene may contribute to differential survival outcomes and toxicities in AML patients via a mechanism of decreased drug efflux in both, AML cells and normal progenitors.

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119 [15] Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, Tsuruo T, Miki Y and Sugimoto Y. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Impact of ABCG2 polymorphisms on the clinical outc... Pharmacogenomics. 2011 Feb;12(2):159-70. Lemos C, Giovannetti E, Zucali PA, Assaraf YG, Scheffer GL, van der Straaten T, D'Incecco A, Falcone A, Guchelaar HJ, Danesi R, Santoro A, Giaccone G, Tibaldi C, Peters GJ
Impact of ABCG2 polymorphisms on the clinical outcome and toxicity of gefitinib in non-small-cell lung cancer patients.
Pharmacogenomics. 2011 Feb;12(2):159-70., [PMID:21332310]

Abstract [show]
AIMS: The current study investigates whether or not functional polymorphisms in the ATP-binding cassette transporter gene ABCG2 might affect gefitinib activity and/or toxicity in non-small-cell lung cancer (NSCLC) patients. MATERIALS & METHODS: Towards this end, ABCG2 polymorphisms and expression were assessed in DNA and tumors from 94 NSCLC patients treated with gefitinib, whereas their associations with toxicity/response and time-to-progression/overall survival were evaluated using Pearson-chi(2) and log-rank-test, respectively. RESULTS: Patients carrying an ABCG2 -15622T/T genotype or harboring at least one TT copy in the ABCG2 (1143C/T, -15622C/T) haplotype developed significantly more grade 2/3 diarrhea (p < 0.01). No associations were found between polymorphisms and outcome. Consistently, ABCG2 protein levels in tumors were not significantly different between patients harboring different ABCG2 variants. CONCLUSION: The ABCG2 -15622C/T polymorphism and ABCG2 (1143C/T, -15622C/T) haplotype resulted in a gefitinib-dependent, moderate-to-severe diarrhea suggesting that these pharmacogenetic markers should be considered to optimize NSCLC treatment.

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7 In particular, the nonsynonymous SNP ABCG2 421C/A, which results in a glutamine to lysine amino acid substitution at position 141 (Q141K), has been associated with markedly decreased levels of ABCG2 protein expression [11-13] and/or activity [14-16]. Login to comment
236 11 Imai Y, Nakane M, Kage K et al.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Mol. Cancer Ther. 1, 611-616 (2002). Login to comment

[hide] Gene-wide tagging study of the association between... Pharmacogenomics. 2011 Mar;12(3):319-25. Kwan P, Wong V, Ng PW, Lui CH, Sin NC, Wong KS, Baum L
Gene-wide tagging study of the association between ABCC2, ABCC5 and ABCG2 genetic polymorphisms and multidrug resistance in epilepsy.
Pharmacogenomics. 2011 Mar;12(3):319-25., [PMID:21449672]

Abstract [show]
AIM: To determine the association between polymorphisms of the multidrug transporter genes ABCC2, ABCC5 and ABCG2, and drug resistance in epilepsy by genotyping comprehensive sets of tagging SNPs. MATERIALS & METHODS: A total of 25 tagging SNPs from ABCC2, ABCC5 and ABCG2 genes were genotyped in a total of 590 Han Chinese epilepsy patients (262 drug resistant and 328 drug responsive). Genotype and allele distributions in drug-responsive and drug-resistant patients were compared. RESULTS: Genotype distributions of all the selected SNPs were consistent with Hardy-Weinberg equilibrium. None of the polymorphisms, either genotype or allele distributions, were significantly associated with drug resistance. For each gene, no haplotypes of over 1% frequency, and that included all SNPs of the gene, were associated with drug resistance. CONCLUSION: This gene-wide tagging study revealed no association between ABCC2, ABCC5 and ABCG2 genetic polymorphisms and multidrug resistance in epilepsy.

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34 For ABCG2, 77 kbp from positions 89,365,500 to 89,442,500 on chromosome 4 were tagged by 11 poly-morphisms (with forced inclusion of two coding SNPs: rs2231137 or V12M and rs2231142 or Q141K), capturing 29 of 39 alleles at r2 ≥ 0.8. Login to comment

[hide] The MRP-related and BCRP/ABCG2 multidrug resistanc... Curr Drug Metab. 2004 Feb;5(1):21-53. Haimeur A, Conseil G, Deeley RG, Cole SP
The MRP-related and BCRP/ABCG2 multidrug resistance proteins: biology, substrate specificity and regulation.
Curr Drug Metab. 2004 Feb;5(1):21-53., [PMID:14965249]

Abstract [show]
Several members of different families of the ATP-binding cassette (ABC) superfamily of transport proteins are capable of transporting an extraordinarily structurally diverse array of endo- and xenobiotics and their metabolites across cell membranes. Together, these transporters play an important role in the absorption, disposition and elimination of these chemicals in the body. In tumor cells, increased expression of these drug transporters is associated with resistance to multiple chemotherapeutic agents. In this review, current knowledge of the biochemical, physiological and pharmacological properties of nine members of the multidrug resistance protein (MRP)-related ABCC family (MRP1, MRP2, MRP3, MRP4, MRP5, MRP6, MRP7, ABCC11 and ABCC12) as well as the G family member, ABCG2/BCRP, are summarized. A focus is placed on the structural similarities and differences of these drug transporters as well as the molecular determinants of their substrate specificities and transport activities. Factors that regulate expression of the MRP-related proteins and ABCG2/BCRP are also reviewed.

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721 One of these polymorphisms, C421A, results in the substitution of a Lys residue for a Gln residue at amino acid position 141 (Gln141Lys). Login to comment

[hide] Influence of ABCB1, ABCC1, ABCC2, and ABCG2 haplot... Pharmacogenet Genomics. 2005 Sep;15(9):599-608. Colombo S, Soranzo N, Rotger M, Sprenger R, Bleiber G, Furrer H, Buclin T, Goldstein D, Decosterd L, Telenti A
Influence of ABCB1, ABCC1, ABCC2, and ABCG2 haplotypes on the cellular exposure of nelfinavir in vivo.
Pharmacogenet Genomics. 2005 Sep;15(9):599-608., [PMID:16041239]

Abstract [show]
OBJECTIVES: The human immunodeficiency virus protease inhibitor nelfinavir is substrate of polyspecific drug transporters encoded by ABCB1 (P-glycoprotein), ABCC1 (MRP1) and ABCC2 (MRP2), and an inhibitor of BCRP, encoded by ABCG2. Genetic polymorphism in these genes may be associated with changes in transport function. METHODS: A comprehensive evaluation of single nucleotide polymorphisms (39 SNPs in ABCB1, 7 in ABCC1, 27 in ABCC2, and 16 in ABCG2), and inferred haplotypes was done to assess possible associations of genetic variants with cellular exposure of nelfinavir in vivo. Analysis used peripheral mononuclear cells from individuals receiving nelfinavir (n=28). Key results were re-examined in a larger sample size (n=129) contributing data on plasma drug levels. RESULTS AND CONCLUSIONS: There was no significant association between cellular nelfinavir area under the curve (AUC) and SNPs or haplotypes at ABCC1, ABCC2, ABCG2. There was an association with cellular exposure for two loci in strong linkage disequilibrium: ABCB1 3435C>T; AUCTT>AUCCT>AUCCC (ratio 2.1, 1.4, 1, Ptrend=0.01), and intron 26 +80T>C; AUCCC> AUCCT > AUCTT (ratio 2.4, 1.3, 1, Ptrend=0.006). Haplotypic analysis using tagging SNPs did not improve the single SNP association values.

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78 - 19346T > A 50 upstream Epidauros bc-v-081 c.19 delG exon 2 p.K7fsX28 Epidauros bc-v-007 c.34G > A exon 2 p.V12M Honjo et al., 2001 bc-v-008 rs2231137 c.71C > T exon 2 p.A24V Epidauros bc-v-009 c.114T > C exon 2 synonymous (p.S38S) Zamber et al., 2003 bc-v-071 rs12721644 IVS 2 + 35 A > G intron 2 Honjo et al., 2001 bc-v-058 rs4148152 IVS 2-12 A > G intron 4 Epidauros bc-v-014 rs2231141 c.421C > A exon 5 p.Q141K Imai et al., 2002 bc-v-015 rs2231142 c.496C > G exon 5 p.Q166E Epidauros bc-v-016 rs1061017 IVS 5 + 14 A > T intron 5 Epidauros bc-v-017 rs2231143 Statistics Association between AUC values and genotypes at single SNP loci was evaluated using a Kruskal-Wallis rank test complemented by a Spearman rank test for trend. Login to comment

[hide] Comprehensive pharmacogenetic analysis of irinotec... J Clin Oncol. 2009 Jun 1;27(16):2604-14. Epub 2009 Apr 6. Innocenti F, Kroetz DL, Schuetz E, Dolan ME, Ramirez J, Relling M, Chen P, Das S, Rosner GL, Ratain MJ
Comprehensive pharmacogenetic analysis of irinotecan neutropenia and pharmacokinetics.
J Clin Oncol. 2009 Jun 1;27(16):2604-14. Epub 2009 Apr 6., 2009-06-01 [PMID:19349540]

Abstract [show]
PURPOSE: We aim to identify genetic variation, in addition to the UGT1A1*28 polymorphism, that can explain the variability in irinotecan (CPT-11) pharmacokinetics and neutropenia in cancer patients. PATIENTS AND METHODS: Pharmacokinetic, genetic, and clinical data were obtained from 85 advanced cancer patients treated with single-agent CPT-11 every 3 weeks at doses of 300 mg/m(2) (n = 20) and 350 mg/m(2) (n = 65). Forty-two common variants were genotyped in 12 candidate genes of the CPT-11 pathway using several methodologies. Univariate and multivariate models of absolute neutrophil count (ANC) nadir and pharmacokinetic parameters were evaluated. RESULTS: Almost 50% of the variation in ANC nadir is explained by UGT1A1*93, ABCC1 IVS11 -48C>T, SLCO1B1*1b, ANC baseline levels, sex, and race (P < .0001). More than 40% of the variation in CPT-11 area under the curve (AUC) is explained by ABCC2 -24C>T, SLCO1B1*5, HNF1A 79A>C, age, and CPT-11 dose (P < .0001). Almost 30% of the variability in SN-38 (the active metabolite of CPT-11) AUC is explained by ABCC1 1684T>C, ABCB1 IVS9 -44A>G, and UGT1A1*93 (P = .004). Other models explained 17%, 23%, and 27% of the variation in APC (a metabolite of CPT-11), SN-38 glucuronide (SN-38G), and SN-38G/SN-38 AUCs, respectively. When tested in univariate models, pretreatment total bilirubin was able to modify the existing associations between genotypes and phenotypes. CONCLUSION: On the basis of this exploratory analysis, common polymorphisms in genes encoding for ABC and SLC transporters may have a significant impact on the pharmacokinetics and pharmacodynamics of CPT-11. Confirmatory studies are required.

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115 % of Patients HWE Exact P (white)African American (n ϭ 11) White (n ϭ 67) Other (n ϭ 7) ABCB1 IVS9 -44AϾG 10276036 .012 A/A 45.4 37.3 0 A/G 36.4 34.3 57.1 G/G 18.2 28.4 42.9 ABCB1 1236CϾT 1128503 .012 C/C 45.5 38.8 0 C/T 54.5 34.3 57.1 T/T 0 26.9 42.9 ABCB1 IVS13 ϩ24CϾT 2235033 .027 C/C 27.3 29.9 0 C/T 45.4 35.8 57.1 T/T 27.3 34.3 42.9 ABCB1 IVS14 ϩ38AϾG 2235013 .027 A/A 18.2 29.9 0 A/G 54.5 35.8 57.1 G/G 27.3 34.3 42.9 ABCB1 2677GϾA/T (A893T/S) 2032582 .046 G/G 40.0 36.9 40.0 G/T 60.0 36.9 60.0 T/T 0 26.2 0 ABCB1 3435CϾT 1045642 .212 C/C 36.4 28.4 57.1 C/T 54.5 41.8 42.9 T/T 9.1 29.8 0 ABCG2 34GϾA (V12M) 2231137 .113 G/G 100 92.5 28.6 G/A 0 6.0 57.1 A/A 0 1.5 14.3 ABCG2 421CϾA (Q141K) 2231142 1.000 C/A 9.1 14.9 42.9 C/C 90.9 85.1 57.1 A/A 0 0 0 SLCO1B1*1b 388AϾG (N130D) 2306283 .624 A/A 18.2 31.3 0 A/G 63.6 46.3 71.4 G/G 18.2 22.4 28.6 SLCO1B1*5 521TϾC (V174A) 4149056 1.000 T/T 81.8 65.7 57.1 T/C 18.2 31.3 42.9 C/C 0 3.0 0 NOTE. Alleles for which a * nomenclature has not yet been assigned have been reported according to their genomic position related to the ATG start site. The reference sequences used for genotyping are the following: ABCG2, NM_004827; SLCO1B1, NM_006446; ABCB1, NM_000927; ABCC1, NM_004987; and ABCC2, NM_00039. Login to comment

[hide] Impact of CYP2C8*3 on paclitaxel clearance: a popu... Pharmacogenomics J. 2011 Apr;11(2):113-20. Epub 2010 Apr 6. Bergmann TK, Brasch-Andersen C, Green H, Mirza M, Pedersen RS, Nielsen F, Skougaard K, Wihl J, Keldsen N, Damkier P, Friberg LE, Peterson C, Vach W, Karlsson MO, Brosen K
Impact of CYP2C8*3 on paclitaxel clearance: a population pharmacokinetic and pharmacogenomic study in 93 patients with ovarian cancer.
Pharmacogenomics J. 2011 Apr;11(2):113-20. Epub 2010 Apr 6., [PMID:20368717]

Abstract [show]
The primary purpose of this study was to evaluate the effect of CYP2C8*3 and three genetic ABCB1 variants on the elimination of paclitaxel. We studied 93 Caucasian women with ovarian cancer treated with paclitaxel and carboplatin. Using sparse sampling and nonlinear mixed effects modeling, the individual clearance of unbound paclitaxel was estimated from total plasma paclitaxel and Cremophor EL. The geometric mean of clearance was 385 l h(1) (range 176-726 l h(1)). Carriers of CYP2C8*3 had 11% lower clearance than non-carriers, P=0.03. This has not been shown before in similar studies; the explanation is probably the advantage of using both unbound paclitaxel clearance and a population of patients of same gender. No significant association was found for the ABCB1 variants C1236T, G2677T/A and C3435T. Secondarily, other candidate single-nucleotide polymorphisms were explored with possible associations found for CYP2C8*4 (P=0.04) and ABCC1 g.7356253C>G (P=0.04).

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135 This effect on clearance of a 'non-fixed` variable provides a competing and dynamic biological explanation for clearance that certainly should be Table 4 Clearance of unbound paclitaxel as function of observed genotypes Gene/allelea Effectb Reference homozygote Heterozygote Variant homozygote P-valuee SNP IDf Nc CLd (10th-90th) Nc CLd (10th-90th) Nc CLd (10th-90th) Candidate SNPs for confirmative analysis CYP2C8 1196A4G(*3) K399R 74 395 (297-490) 19 350 (238-458) 0.03* (0.04) rs10509681 ABCB1 1236C4T G412G 29 391 (270-569) 45 393 (299-490) 19 359 (291-437) 0.25 (0.25) rs1128503 2677G4T/Ag A893S/T 26 387 (270-490) 42(GT) 396 (299-490) 20(TT) 356 (294-437) 0.20 (0.26) rs2032582 3435C4T I1145I 11 403 (326-548) 44 387 (282-490) 38 378 (297-468) 0.83 (0.43) rs1045642 Candidate SNPs for exploratory analysis CYP2C8 792C4G(*4) I264M 86 391 (297-490) 7 321 (270-374) 0.04* (0.03) rs1058930 15577956G4T (*1B) - 49 395 (298-552) 43 373 (291-478) 1 461 0.75 (0.36) rs7909236 15578055A4C (*1C) - 69 382 (291-478) 24 393 (300-552) 0.48 (0.62) rs17110453 ABCB1 À1A4G - 1 458 29 396 (270-592) 63 379 (297-477) 0.56 (0.3) rs2214102 61A4G N21D 63 384 (282-490) 29 386 (298-478) 1 437 0.52 (0.77) rs9282564 1199G4A S400N 83 385 (291-490) 10 386 (322-461) 0.74 (0.99) rs2229109 CYP3A4 24616372T4C (*1B) - 85 383 (296-490) 7 397 (270-641) 0.67 (0.72) rs2740574 CYP3A5 219-237G4A Frameshift 84 388 (297-490) 9 360 (176-726) 0.30 (0.36) rs776746 SLCO1B3 699G4A M233I 1 326 19 377 (299-481) 73 388 (291-490) 0.99 (0.46) rs7311358 767G4C G256A 67 386 (298-481) 26 383 (291-490) 0.63 (0.89) rs60140950 CYP1B1 1294C4G (*3) V432L 30 389 (270-530) 36 401 (298-490) 27 361 (300-470) 0.77 (0.24) rs1056836 ABCC1 7356253C4G - 65 394 (297-548) 27 368 (291-470) 1 332 0.04* (0.15) rs504348 ABCC2 1249G4A V417I 67 381 (291-490) 24 396 (297-552) 2 415 (368-468) 0.21 (0.39) rs2273697 3563T4A V1188E 87 386 (296-490) 5 370 (176-569) 0.7 (0.7) rs17222723 4544G4A C1515Y 75 389 (296-490) 3 355 (176-569) 0.72 (0.52) rs8187710 ABCG2 421C4A Q141K 61 374 (291-478) 32 408 (315-548) 0.4 (0.09) rs2231142 34G4A V12M 87 385 (291-490) 4 395 (296-726) 0.68 (0.83) rs2231137 ABCC10 2759T4C I920T 46 386 (297-478) 43 386 (291-548) 4 373 (326-467) 0.88 (0.89) rs2125739 Abbreviations: CL, clearance of unbound paclitaxel; SNP, single-nucleotide polymorphism. Login to comment

[hide] Ubiquitin-mediated proteasomal degradation of ABC ... J Pharm Sci. 2011 Sep;100(9):3602-19. doi: 10.1002/jps.22615. Epub 2011 May 12. Nakagawa H, Toyoda Y, Wakabayashi-Nakao K, Tamaki H, Osumi M, Ishikawa T
Ubiquitin-mediated proteasomal degradation of ABC transporters: a new aspect of genetic polymorphisms and clinical impacts.
J Pharm Sci. 2011 Sep;100(9):3602-19. doi: 10.1002/jps.22615. Epub 2011 May 12., [PMID:21567408]

Abstract [show]
The interindividual variation in the rate of drug metabolism and disposition has been known for many years. Pharmacogenomics dealing with heredity and response to drugs is a part of science that attempts to explain variability of drug responses and to search for the genetic basis of such variations or differences. Genetic polymorphisms of drug metabolizing enzymes and drug transporters have been found to play a significant role in the patients' responses to medication. Accumulating evidence demonstrates that certain nonsynonymous polymorphisms have great impacts on the protein stability and degradation, as well as the function of drug metabolizing enzymes and transporters. The aim of this review article is to address a new aspect of protein quality control in the endoplasmic reticulum and to present examples regarding the impact of nonsynonymous single-nucleotide polymorphisms on the protein stability of thiopurine S-methyltransferase as well as ATP-binding cassette (ABC) transporters including ABCC4, cystic fibrosis transmembrane conductance regulator (CFTR, ABCC7), ABCC11, and ABCG2. Furthermore, we will discuss the molecular mechanisms underlying posttranslational modifications (intramolecular and intermolecular disulfide bond formation and N-linked glycosylation) and ubiquitin-mediated proteasomal degradation of ABCG2, one of the major drug transporter proteins in humans.

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155 Effect of Mutations and Nonsynonymous SNPs on Protein Trafficking, Maturation, or ERAD of ABC Transporters Protein AA Mutation/SNP Effect on Protein Reference ABCA1 W590S Mutation Functional defect 115 R587W Mutation Impaired glycol processing 115 Q597R Mutation Impaired glycol processing, ERAD 115,116 Y1532C Mutation Altered protein trafficking 117 R1925Q Mutation Altered protein trafficking 118 ABCA3 R43L Mutation Altered protein trafficking 119 L101P Mutation Altered protein trafficking 119 R280C Mutation Altered protein trafficking 119 ABCA4 L541P Mutation Mislocalization 120 R602W Mutation Mislocalization 120 A1038V Mutation Mislocalization 120 C1490Y Mutation Mislocalization 120 ABCB1a G268V Mutation ERAD 121 G341C Mutation ERAD 121 I1196S Mutation Reduced glycosylation 122 ABCB4 I541F Mutation Accumulation in ER 123 ABCB11a E135K Mutation Reduced level of mature protein 124 L198P Mutation Reduced level of mature protein 124 E297G Mutation Reduced level of mature protein 124 L413W Mutation Reduced level of mature protein 124 R432T Mutation Reduced level of mature protein 124 D482G Mutation Immature protein in ER 124,125 N490D Mutation Reduced level of mature protein 124 A570T Mutation Reduced level of mature protein 124 T655I Mutation Reduced level of mature protein 124 Y818F SNP Moderate reduction of protein 124 G982R Mutation Retention in ER 125 R1153C Mutation ERAD 125 R1286Q Mutation Retention in ER 125 ABCC2a R768W Mutation Impaired protein trafficking 126 I1173F Mutation Impaired protein maturation 127 R1392 Mutation Impaired protein maturation 128 M1393 Mutation Impaired protein maturation 129 ABCC4a E757K SNP Altered protein trafficking 23 ABCC7 F508 Mutation Misfolding, ERAD 36-39,130 G85E Mutation Impaired protein maturation 130-132 G91R Mutation Impaired protein maturation 130-132 N1303K Mutation Impaired protein maturation 130-132 ABCC8 WT Wild type Ubiquitin-proteasome degradation 133 A116P Mutation Ubiquitin-proteasome degradation 133 V187D Mutation Ubiquitin-proteasome degradation 133 F1388 Mutation Impaired protein trafficking 134 L1544P Mutation Impaired protein trafficking 135,136 ABCC11a G180R SNP Ubiquitin-proteasome degradation 50 27 Mutation Ubiquitin-proteasome degradation 50 ABCG2a V12M SNP Altered protein localization 96 Q141K SNP Ubiquitin-proteasome degradation 102 F208S SNP Ubiquitin-proteasome degradation 78,99 S441N SNP Ubiquitin-proteasome degradation 78,99 Mutations of ABCA1, ABCA3, ABCA4, ABCB4, ABCB11, ABCC2, ABCC7 (CFTR), and ABCC8 are associated with Tangier disease, fatal surfactant deficiency, Stargardt disease, progressive familial intrahepatic cholestasis type 3 (PFIC-3), progressive familial intrahepatic cholestasis type 2 (PFIC-2), Dubin-Johnson syndrome, cystic fibrosis, and familial hyperinsulinism, respectively. Login to comment
76 Furthermore, the nonsynonymous SNPs of Q141K, F208S, and S441N (Fig. 4) have been found to greatly affect APCG2 protein levels in the plasma membrane. Login to comment
78 Q141K (Gln141Lys), F208S (Phe208Ser), and S441N (Ser441Asn) are nonsynonymous SNPs that cause the reduced expression of ABCG2 protein. Login to comment
126 Pharmaceutical and Clinical Impacts of SNP 421C>A (Q141K) in ABCG2 Gene The most extensively studied among those SNPs with potential clinical relevance is 421 C>A resulting in a glutamine to lysine substitution (Gln141Lys or Q141K) in the ABCG2 protein. Login to comment
127 Q141K has been associated with lower levels of protein expression (Fig. 7a) and impaired transport activities in vitro.93,96-99,101 Figure 6. Login to comment
141 The SNP 421C>A is a nonsynonymous polymorphism that leads to amino acid substitution; Gln141Lys (Q141K). Login to comment
145 (b) The allele frequencies of 421C (WT) and 421A (Q141K) among different ethnic populations. Login to comment
148 The reduced protein levels of the Q141K variant are due to ubiquitin-mediated proteasomal degradation as well as endosomal degradation (Fig. 7a).102 The Q141K SNP has been identified with varying frequencies in different ethnic groups and was found to be the most relevant in Japanese and Chinese populations (approximately 30% in the allele frequency) (Fig. 7b). Login to comment
149 It has been reported that patients carrying the Q141K SNP have elevated plasma levels of gefitinib, diflomotecan, and increased bioavailability of oral topotecan.103-105 The Q141K SNP was reportedly associated with a higher incidence of diarrhea in nonsmall cell lung cancer patients treated with gefitinib.106 Figure 8. Login to comment
153 Three panels depict the time courses of the SmartAmp assay reactions with ABCG2 allele-specific primers carrying 421C (WT) or 421A (Q141K) alleles; namely, C/C homozygote, C/A heterozygote, and A/A homozygote. Login to comment
168 As the expression levels of the Q141K variant is reduced by ubiquitin-mediated proteasomal degradation,102 renal excretion of serum uric acid via ABCG2 is impaired in humans who are carrying the 421A allele (Q141K variant). Login to comment

[hide] Single nucleotide polymorphisms in human P-glycopr... Expert Opin Drug Deliv. 2006 Jan;3(1):23-35. Dey S
Single nucleotide polymorphisms in human P-glycoprotein: its impact on drug delivery and disposition.
Expert Opin Drug Deliv. 2006 Jan;3(1):23-35., [PMID:16370938]

Abstract [show]
Drug efflux pumps belong to a large family of ATP-binding cassette transporter proteins. These pumps bind their substrate and export it through the membrane using energy derived from ATP hydrolysis. P-glycoprotein, the main efflux pump in this family, is expressed not only in tumour cells but also in normal tissues with excretory function (liver, kidney and the intestine). It has a broad specificity of substrates and plays an important role in drug delivery and disposition. Recently, genetic screening of P-glycoprotein has yielded multiple single nucleotide polymorphisms, which seem to alter transporter function and expression. This review discusses the various polymorphisms of this gene and its impact on drug disposition and diseases.

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518 IMAI Y, NAKANE M, KAGE K et al.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] A T3587G germ-line mutation of the MDR1 gene encod... Mol Cancer Ther. 2006 Apr;5(4):877-84. Mutoh K, Mitsuhashi J, Kimura Y, Tsukahara S, Ishikawa E, Sai K, Ozawa S, Sawada J, Ueda K, Katayama K, Sugimoto Y
A T3587G germ-line mutation of the MDR1 gene encodes a nonfunctional P-glycoprotein.
Mol Cancer Ther. 2006 Apr;5(4):877-84., [PMID:16648557]

Abstract [show]
The human multidrug resistance gene 1 (MDR1) encodes a plasma membrane P-glycoprotein (P-gp) that functions as an efflux pump for various structurally unrelated anticancer agents. We have identified two nonsynonymous germ-line mutations of the MDR1 gene, C3583T MDR1 and T3587G MDR1, in peripheral blood cell samples from Japanese cancer patients. Two patients carried the C3583T MDR1 allele that encodes H1195Y P-gp, whereas a further two carried T3587G MDR1 that encodes I1196S P-gp. Murine NIH3T3 cells were transfected with pCAL-MDR-IRES-ZEO constructs carrying either wild-type (WT), C3583T, or T3587G MDR1 cDNA and selected with zeocin. The resulting zeocin-resistant mixed populations of transfected cells were designated as 3T3/WT, 3T3/H1195Y, and 3T3/I1196S, respectively. The cell surface expression of I1196S P-gp in 3T3/I1196S cells could not be detected by fluorescence-activated cell sorting, although low expression of I1196S P-gp was found by Western blotting. H1195Y P-gp expression levels in 3T3/H1195Y cells were slightly lower than the corresponding WT P-gp levels in 3T3/WT cells. By immunoblotting analysis, both WT P-gp and H1195Y P-gp were detectable as a 145-kDa protein, whereas I1196S P-gp was visualized as a 140-kDa protein. 3T3/I1196S cells did not show any drug resistance unlike 3T3/H1195Y cells. Moreover, a vanadate-trap assay showed that the I1196S P-gp species lacks ATP-binding activity. Taken together, we conclude from these data that T3587G MDR1 expresses a nonfunctional P-gp and this is therefore the first description of such a germ-line mutation. We contend that the T3587G MDR1 mutation may affect the pharmacokinetics of MDR1-related anticancer agents in patients carrying this allele.

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220 Moreover, Q141K BCRP-expressing cells show low levels of BCRP expression compared with WT BCRP-expressing cells (34, 35). Login to comment
299 Imai Y, Nakane M, Kage K, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Pharmacogenetics of drug transporters in the enter... Pharmacogenomics. 2011 May;12(5):611-31. Stieger B, Meier PJ
Pharmacogenetics of drug transporters in the enterohepatic circulation.
Pharmacogenomics. 2011 May;12(5):611-31., [PMID:21619426]

Abstract [show]
This article summarizes the impact of the pharmacogenetics of drug transporters expressed in the enterohepatic circulation on the pharmacokinetics and pharmacodynamics of drugs. The role of pharmacogenetics in the function of drug transporter proteins in vitro is now well established and evidence is rapidly accumulating from in vivo pharmacokinetic studies, which suggests that genetic variants of drug transporter proteins can translate into clinically relevant phenotypes. However, a large amount of conflicting information on the clinical relevance of drug transporter proteins has so far precluded the emergence of a clear picture regarding the role of drug transporter pharmacogenetics in medical practice. This is very well exemplified by the case of P-glycoprotein (MDR1, ABCB1). The challenge is now to develop pharmacogenetic models with sufficient predictive power to allow for translation into drug therapy. This will require a combination of pharmacogenetics of drug transporters, drug metabolism and pharmacodynamics of the respective drugs.

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94 Gene name Transporter SNP Protein Population size (n) In vitro function Ref. Intestinal uptake transporters SLC15A1 PEPT1 p.P586L 44 Reduced Vmax [81] p.F28Y 247 Increased Km [82] Intestinal efflux transporters ABCB1 MDR1 c.571G>A p.G191R N/A Reduced drug resistance [201] c.1199G>A p.S440N N/A Reduced activity (substrate dependent) [202] c.11199G>A c.1199G>t p.S440N p.S440I N/A N/A Increased drug resistance Reduced drug resistance [203] c.1292-3GT>TG p.C431L N/A Reduced drug resistance [204] c.2005C>T p.R669C N/A Reduced substrate affinity [202] c.2547A>G p.I849M N/A Increased transport activity [202] c.2677G>T p.A893S 60 Lower intracellular digoxin accumulation [205] c.2677G>T c.2677G>A p.A893S p.A893T N/A N/A Unchanged Unchanged [206] c.2677G>T p.A893S 46 No change in rhodamine 123 efflux from peripheral blood lymphocytes [207] c.2667G>T p.A893S N/A Reduced transport function [208] c.2667G>T c.2677G>A p.A893S p.A893T N/A N/A Increased transport function Increased transport function [209] c.2667G>T c.2677G>A p.A893S p.A893T N/A N/A Increased activity (substrate dependent) Increased substrate affinity and transport activity [202] c.2667G>T p.A893S 48 No change in rhodamine 123 efflux activity in peripheral blood mononuclear cells [210] c.2956A>G p.M986V N/A Increased transport activity [202] c.2995G>A p.A999T N/A Increased substrate affinity and transport activity [202] c.3151C>G p.P1051A N/A Increased transport activity (substrate dependent) [202] c.3188G>C p.G1063A N/A Increased transport activity [202] ABCG2 ABCG2 c.34G>A p.V12M N/A Low transport protein expression in vitro [211] c.34G>A p.V12M N/A Unchanged [212] c.34G>A p.V12M N/A No change in HEK-293, lowered transport activity in Sf9 cells in vitro [213] c.34G>A p.V12M N/A Unchanged [214] c.421C>A p.Q141K N/A Lower transport protein expression, normal transport activity [212] c.421C>A p.Q141K N/A Reduced drug resistance and lower ATPase activity [213] c.421C>A p.Q141K N/A Reduced drug extrusion [215] c.421C>A p.Q141K N/A Reduced drug resistance [216] c.421C>A p.Q141K N/A Unchanged [217] c.421C>A p.Q141K N/A No change of intracellular porphyrin accumulation [218] c.421C>A p.Q141K N/A Reduced transport activity [219] c.421C>A p.Q141K N/A Reduced transport activity [55] c.421C>A p.Q141K N/A Increased Km [220] For more information on members of the SLC superfamily of transporters please consult [301] and for more information of ABC transporters please consult [302]. Login to comment
739 219 Polgar O, Deeken JF, Ediriwickrema LS et al.: The 315-316 deletion determines the BXP-21 antibody epitope but has no effect on the function of wild type ABCG2 or the Q141K variant. Login to comment
742 220 Pollex EK, Anger G, Hutson J, Koren G, Piquette-Miller M: Breast cancer resistance protein (BCRP)-mediated glyburide transport: effect of the C421A/Q141K BCRP single-nucleotide polymorphism. Login to comment

[hide] Impact of functional ABCG2 polymorphisms on the ad... Cancer Chemother Pharmacol. 2010 Sep;66(4):691-8. Epub 2009 Dec 25. Akasaka K, Kaburagi T, Yasuda S, Ohmori K, Abe K, Sagara H, Ueda Y, Nagao K, Imura J, Imai Y
Impact of functional ABCG2 polymorphisms on the adverse effects of gefitinib in Japanese patients with non-small-cell lung cancer.
Cancer Chemother Pharmacol. 2010 Sep;66(4):691-8. Epub 2009 Dec 25., [PMID:20035425]

Abstract [show]
PURPOSE: ABCG2 is a half-size ATP-binding cassette transporter implicated in cellular gefitinib transport. Reportedly, the c.421C > A ABCG2 gene polymorphism was associated with gefitinib-induced diarrhea in Caucasian patients with non-small-cell lung cancer. Since c.421C > A ABCG2, resulting in p.Q141K substitution, is more prevalent in Asian populations, the putative relationship between gefitinib-induced adverse effects and this functional polymorphism was investigated in Japanese patients. c.376C > T, resulting in truncated, non-functional ABCG2, was also investigated. METHODS: ABCG2 gene polymorphisms were evaluated in 75 patients with non-small-cell lung cancer treated with gefitinib 250 mg/day orally, and results were correlated with treatment-related adverse effects. RESULTS: Forty (53.3%) patients harbored c.421A ABCG2 on at least one allele, while the remaining 35 (46.7%) were wild type for c.421C > A. No significant group difference was observed in frequency of gefitinib-related diarrhea or other adverse effects. In addition, the only one patient homozygous for the c.421A allele in this study was not affected with gefitinib-induced diarrhea or interstitial lung disease. Two patients (2.7%) were found to harbor the c.376T allele heterozygously. One of the two patients harbored both the c.376T and the c.421A genotypes on distinct alleles. Gefitinib-related interstitial lung disease and severe diarrhea were noted in neither of the two patients. CONCLUSIONS: In this Japanese population, we did not find an evident association between ABCG2 polymorphisms, c.376C > T and c.421C > A, and susceptibility to gefitinib-induced adverse effects.

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2 Since c.421C [ A ABCG2, resulting in p.Q141K substitution, is more prevalent in Asian populations, the putative relationship between gefitinib-induced adverse effects and this functional polymorphism was investigated in Japanese patients. Login to comment
97 In contrast, the c.421C [ A polymorphism results in a p.Q141K variant ABCG2, of which expression level is 3-to 5-fold lower than that of the wild type in spite of similar mRNA levels [22]. Login to comment

[hide] Xenobiotic, bile acid, and cholesterol transporter... Pharmacol Rev. 2010 Mar;62(1):1-96. Epub 2010 Jan 26. Klaassen CD, Aleksunes LM
Xenobiotic, bile acid, and cholesterol transporters: function and regulation.
Pharmacol Rev. 2010 Mar;62(1):1-96. Epub 2010 Jan 26., [PMID:20103563]

Abstract [show]
Transporters influence the disposition of chemicals within the body by participating in absorption, distribution, and elimination. Transporters of the solute carrier family (SLC) comprise a variety of proteins, including organic cation transporters (OCT) 1 to 3, organic cation/carnitine transporters (OCTN) 1 to 3, organic anion transporters (OAT) 1 to 7, various organic anion transporting polypeptide isoforms, sodium taurocholate cotransporting polypeptide, apical sodium-dependent bile acid transporter, peptide transporters (PEPT) 1 and 2, concentrative nucleoside transporters (CNT) 1 to 3, equilibrative nucleoside transporter (ENT) 1 to 3, and multidrug and toxin extrusion transporters (MATE) 1 and 2, which mediate the uptake (except MATEs) of organic anions and cations as well as peptides and nucleosides. Efflux transporters of the ATP-binding cassette superfamily, such as ATP-binding cassette transporter A1 (ABCA1), multidrug resistance proteins (MDR) 1 and 2, bile salt export pump, multidrug resistance-associated proteins (MRP) 1 to 9, breast cancer resistance protein, and ATP-binding cassette subfamily G members 5 and 8, are responsible for the unidirectional export of endogenous and exogenous substances. Other efflux transporters [ATPase copper-transporting beta polypeptide (ATP7B) and ATPase class I type 8B member 1 (ATP8B1) as well as organic solute transporters (OST) alpha and beta] also play major roles in the transport of some endogenous chemicals across biological membranes. This review article provides a comprehensive overview of these transporters (both rodent and human) with regard to tissue distribution, subcellular localization, and substrate preferences. Because uptake and efflux transporters are expressed in multiple cell types, the roles of transporters in a variety of tissues, including the liver, kidneys, intestine, brain, heart, placenta, mammary glands, immune cells, and testes are discussed. Attention is also placed upon a variety of regulatory factors that influence transporter expression and function, including transcriptional activation and post-translational modifications as well as subcellular trafficking. Sex differences, ontogeny, and pharmacological and toxicological regulation of transporters are also addressed. Transporters are important transmembrane proteins that mediate the cellular entry and exit of a wide range of substrates throughout the body and thereby play important roles in human physiology, pharmacology, pathology, and toxicology.

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6547 Chinese men with one mutant Q141K ABCG2 (BCRP) allele demonstrate higher plasma concentrations of rosuvastatin compared with those with both wild-type alleles (Table 15) (Zhang et al., 2006b). Login to comment
6548 Similar findings have been reported in Finnish volunteers with the Q141K ABCG2 (BCRP) SNP after atorvastatin or rosuvastatin administration (Keskitalo et al., 2009). Login to comment
6589 Absent C421A Q141K 2 Normal/reduced G445C A149P ↔ Normal G448A R163K ↔ Normal C496G Q166E ↔ Normal/reduced A616C I206L 2↔ Normal T623C F208S N.D. Reduced T742C S248P N.D. Normal C805T P269S 2↔ Normal T1291C F431L 2 Normal/reduced G1322A S441N 2 Reduced T1465C F489L 2↔ Normal/reduced A1768T N590Y 2↔ Increased G1858A D620N 2↔ Normal 2, reduced function; ↔, no change in function; N.D. not determined. Login to comment
6807 In support of these findings, humans with the Q141K ABCG2 (BCRP) polymorphism display increased oral bioavailability of sulfasalazine (Table 15) (Urquhart et al., 2008; Yamasaki et al., 2008). Login to comment
6918 A polymorphism in ABCG2 (BCRP) (Q141K) is clinically associated with diarrhea as well as elevated concentrations in heterozygous patients receiving another tyrosine kinase inhibitor, gefitinib (Table 15) (Cusatis et al., 2006; Li et al., 2007a). Login to comment
7031 It is noteworthy that a variant ABCG2 (BCRP) gene (Q141K) is associated with reduced BCRP protein accumulation in placenta, although the functional relevance of this polymorphism has not yet been determined (Table 15) (Kobayashi et al., 2005b). Login to comment

[hide] Single nucleotide polymorphism associations with r... Lancet Oncol. 2011 Nov;12(12):1143-50. Epub 2011 Oct 17. Garcia-Donas J, Esteban E, Leandro-Garcia LJ, Castellano DE, del Alba AG, Climent MA, Arranz JA, Gallardo E, Puente J, Bellmunt J, Mellado B, Martinez E, Moreno F, Font A, Robledo M, Rodriguez-Antona C
Single nucleotide polymorphism associations with response and toxic effects in patients with advanced renal-cell carcinoma treated with first-line sunitinib: a multicentre, observational, prospective study.
Lancet Oncol. 2011 Nov;12(12):1143-50. Epub 2011 Oct 17., [PMID:22015057]

Abstract [show]
BACKGROUND: Sunitinib is a tyrosine kinase inhibitor with proven efficacy in renal-cell carcinoma, but some patients do not respond or need dose reductions due to toxicity. Because there are no validated molecular predictors of response or toxicity to sunitinib, we aimed to identify genetic markers predictive of outcome and toxic effects. METHODS: In our observational, prospective study we enrolled previously untreated adults (>/= 18 years) with clear-cell renal-cell carcinoma at 15 institutions in the Spanish Oncology Genitourinary Group in Spain. Patients received sunitinib according to local practice guidelines. We assessed RECIST response, progression-free survival (PFS), overall survival, and toxicity of sunitinib with 16 key polymorphisms in nine genes: VEGFR2 (rs2305948 and rs1870377), VEGFR3 (rs307826, rs448012, and rs307821), PDGFR-alpha (rs35597368), VEGF-A (rs2010963, rs699947, and rs1570360), IL8 (rs1126647), CYP3A4 (rs2740574), CYP3A5 (rs776746), ABCB1 (rs1045642, rs1128503, and rs2032582), and ABCB2 (rs2231142). We assessed associations with efficacy and toxicity by use of univariable and multivariable analyses (with clinical factors associated with outcomes as covariates). We adjusted for multiplicity using the Bonferroni method; p values of less than 0.0031 before adjustment were deemed to still be significant after adjustment. FINDINGS: We enrolled 101 patients between Oct 10, 2007, and Dec 13, 2010. 95 of these patients were included in toxicity analyses and 89 in the efficacy analyses. Two VEGFR3 missense polymorphisms were associated with reduced PFS with sunitinib on multivariable analysis: rs307826 (hazard ratio [HR] per allele 3.57, 1.75-7.30; p(unadjusted)=0.00049, p(adjusted)=0.0079) and rs307821 (3.31, 1.64-6.68; p(unadjusted)=0.00085, p(adjusted)=0.014). The CYP3A5*1 (rs776746) high metabolising allele was associated in a multivariable analysis with an increased risk of dose reductions due to toxicity (HR per allele 3.75, 1.67-8.41; p(unadjusted)=0.0014, p(adjusted)=0.022). No other SNPs were associated with sunitinib response or toxicity. INTERPRETATION: Polymorphisms in VEGFR3 and CYP3A5*1 might be able to define a subset of patients with renal-cell carcinoma with decreased sunitinib response and tolerability. If confirmed, these results should promote interventional studies testing alternative therapeutic approaches for patients with such variants. FUNDING: Pfizer.

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57 See Online for webappendix Single nucleotide polymorphism Variation Patients* Homozygous wild-type Heterozygous Homozygous variant Minor allele frequency VEGFR2 rs2305948 C>T V297I 95 78 15 2 0·100 VEGFR2 rs1870377T>A Q472H 95 53 34 8 0·263 VEGFR3 rs307826 A>G T494A 95 80 15 0 0·079 VEGFR3 rs448012 C>G H890Q 94 32 48 14 0·404 VEGFR3 rs307821 G>T R1324L 95 78 16 1 0·095 PDGFR-α rs35597368T>C S478P 95 77 16 2 0·105 VEGF-A rs2010963 G>C 5´-UTR 95 47 37 11 0·311 VEGF-A rs699947 A>C Promoter 95 27 47 21 0·468 VEGF-A rs1570360 G>A Promoter 95 45 41 9 0·311 IL8 rs1126647 A>T 3´-UTR 94 35 48 11 0·372 CYP3A4 rs2740574 A>G Promoter 94 89 5 0 0·027 CYP3A5 rs776746 G>A Splicing 94 82 12 0 0·064 ABCB1 rs1045642 C>T I1145I 94 27 51 16 0·441 ABCB1 rs1128503 C>T G412G 95 36 45 14 0·384 ABCB1 rs2032582 G>T A893S 92 38 39 15 0·375 ABCG2 rs2231142 C>A Q141K 95 85 10 0 0·053 UTR=untranslated region. Login to comment

[hide] Germline genetic polymorphisms may influence chemo... Cancer. 2012 Apr 1;118(7):1856-67. doi: 10.1002/cncr.26472. Epub 2011 Sep 1. Windsor RE, Strauss SJ, Kallis C, Wood NE, Whelan JS
Germline genetic polymorphisms may influence chemotherapy response and disease outcome in osteosarcoma: a pilot study.
Cancer. 2012 Apr 1;118(7):1856-67. doi: 10.1002/cncr.26472. Epub 2011 Sep 1., [PMID:21887680]

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BACKGROUND: Osteosarcoma is the most common malignant bone tumor in children and young people. Efficacy of multiagent MAP (methotrexate, doxorubicin [Adriamycin], cisplatin) chemotherapy may be influenced by multiple cellular pathways. This pilot study aimed to investigate the association of 36 candidate genetic polymorphisms in MAP pathway genes with histological response, survival, and grade 3-4 chemotherapy toxicity in osteosarcoma. METHODS: Blood samples were obtained from 60 patients who had completed MAP chemotherapy. All patients were manually genotyped for 5 polymorphisms. The remaining 31 polymorphisms were genotyped in 50 patients using the Illumina 610-Quad microarray. Associations between candidate polymorphisms and histological response, progression-free survival, and toxicity were estimated using Pearson chi-square and Fisher exact tests, the Kaplan-Meier method, the log-rank test, and the Cox proportional hazards model. RESULTS: Poor histological response was increased in variants of ABCC2 c.24C>T (P = .011) and GSTP1 c.313A>G p.Ile(105)Val (P = .009), whereas MTHFD1 c.1958G>A p.Arg(653)Gln was protective (P = .03). Methotrexate toxicity was increased in variants of MTHFR c.1298A>C p.Glu(429)Ala (P = .038), ABCB1 c.3435T>C Ile(145)Ile (P = .027), and ABCC2 c.3563T>A p.Val(1188)Glu (P = .028). Variants of GSTP1 c.313A>G p.Ile(105)Val were at increased risk of myelosuppression (P = .024) and cardiac damage (P = .008). CONCLUSIONS: This pilot study represents the most comprehensive study to date examining the role of genetic polymorphisms in osteosarcoma. Although small and retrospective, it shows that several polymorphisms appear to significantly influence toxicity and clinical outcome. These deserve prospective validation in the hope of optimizing treatment for resistant disease and reducing the late effects burden.

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44 3R adverse prognosis in ALL.45 ABC efflux ABCB1 (MDR1) 7q21.12 MAP 1128503 1236C>T Gly412 Gly T allele : exposure to doxorubicin in breast cancer patients.4 No association in platinum-treated ovarian cancer.5 1045642 3435C>T Ile145 Ile CC : response to platinum chemotherapy in NSCLC.6 No association in platinum-treated ovarian cancer.5 ABCG2 (BCRP) 4q22.1 MAP 2231142 421C>A Gln141 Lys A ; OS in platinum-treated lung cancer.7 No association in platinum-treated ovarian cancer.5 ABCC1 (MRP1) 6p13.11 M 246240 A>G G ; methotrexate toxicity in psoriasis.8 3784862 A>G ABCC2 10q24.2 M 717620 24C>T T : response to platinum chemotherapy in NSCLC.9 No association in ovarian cancer.5 2273697 1249G>A Val417 Ile No association with response to platinum chemotherapy in NSCLC.9 17222723 3563T>A Val1188 Glu A : acute ACT, in 100% LD with Cis1515 Tyr.10 8087710 4544G>A Cis1515 Tyr DNA repair ERCC1 19q13.32 A, P 3212986 1510C>A (8092C>A)* Clinical effects conflicting. Login to comment

[hide] Recent advances in pharmacogenomics of ABC transpo... Pharmacogenomics. 2012 Apr;13(6):633-6. Ishikawa T
Recent advances in pharmacogenomics of ABC transporters involved in breast cancer therapy.
Pharmacogenomics. 2012 Apr;13(6):633-6., [PMID:22515603]

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43 The most extensively studied among those SNPs with potential clinical relevance is 421C>A resulting in a glutamic acid to lysine substitution (Q141K) in the ABCG2 protein. The expression level of the Q141K variant reduced compared with the wild-type, which is due to its ubiquitin-mediated proteasomal degradation [10]. Login to comment
45 Furthermore, the Q141K SNP was reportedly associated with a higher incidence of diarrhea in non-small-cell lung cancer patients treated with gefitinib [11]. Login to comment

[hide] Common variants in ABCB1, ABCC2 and ABCG2 genes an... Gynecol Oncol. 2012 Mar;124(3):575-81. Epub 2011 Nov 21. Tian C, Ambrosone CB, Darcy KM, Krivak TC, Armstrong DK, Bookman MA, Davis W, Zhao H, Moysich K, Gallion H, DeLoia JA
Common variants in ABCB1, ABCC2 and ABCG2 genes and clinical outcomes among women with advanced stage ovarian cancer treated with platinum and taxane-based chemotherapy: a Gynecologic Oncology Group study.
Gynecol Oncol. 2012 Mar;124(3):575-81. Epub 2011 Nov 21., [PMID:22112610]

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PURPOSE: Efflux transporters of the ATP-binding cassette (ABC) family are major determinants of chemoresistance in tumor cells. This study examined associations between functional variants in ABCB1, ABCC2 and ABCG2 genes and clinical outcomes in patients with epithelial ovarian/primary peritoneal cancer (EOC/PPC) following platinum and taxane-based chemotherapy. METHODS: Sequenom iPLEXTMGOLD Assay and MALDI-TOF platform were used to genotype the non-synonymous G2677T/A (rs2032582; encoding Ala893Ser/Thr) and synonymous C3435T (rs1045642; encoding Ile1145Ile) variants in ABCB1, the non-synonymous G1249A variant in ABCC2 (rs2273697; encoding Val417Ile), and the non-synonymous C421A variant in ABCG2 (rs2231142; encoding Q141K, Gln141Lys) in normal DNA from up to 511 women in Gynecologic Oncology Group (GOG) phase III trials, GOG-172 or GOG-182. Progression-free survival (PFS) and overall survival (OS) were analyzed in relation to genetic polymorphisms using Kaplan-Meier and Cox proportional hazards model. RESULTS: The C421A variant (CA+AA versus CC) in ABCG2 was associated with a 6-month longer median PFS (22.7 versus 16.8 months, p=0.041). In multivariate analysis, patients with variant genotypes were at a reduced risk of disease progression (hazard ratio [HR]=0.75, 95% confidence interval [CI]=0.59-0.96, p=0.022). The association between C421A and OS was not statistically significant (HR=0.88, 95% CI=0.67-1.15, p=0.356). None of the other variants measured in either ABCB1 or ABCC2 was associated with PFS or OS. CONCLUSION: The C421A variant in ABCG2, previously shown to be associated with enhanced protein degradation and drug sensitivity, was associated with longer PFS in advanced stage EOC/PPC patents treated with platinum+taxane-based chemotherapy. This finding requires further validation.

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4 Sequenom iPLEXTMGOLD Assay and MALDI-TOF platform were used to genotype the non-synonymous G2677T/A (rs2032582; encoding Ala893Ser/Thr) and synonymous C3435T (rs1045642; encoding Ile1145Ile) variants in ABCB1, the non-synonymous G1249A variant in ABCC2 (rs2273697; encoding Val417Ile), and the non-synonymous C421A variant in ABCG2 (rs2231142; encoding Q141K, Gln141Lys) in normal DNA from up to 511 women in Gynecologic Oncology Group (GOG) phase III trials, GOG-172 or GOG-182. Login to comment
109 The non-synonymous C421A polymorphism in ABCG2 results in an amino acid change, Gln141Lys. Login to comment

[hide] Irinotecan pharmacogenomics. Pharmacogenomics. 2010 Jul;11(7):1003-10. Marsh S, Hoskins JM
Irinotecan pharmacogenomics.
Pharmacogenomics. 2010 Jul;11(7):1003-10., [PMID:20602618]

Abstract [show]
Irinotecan is a camptothecin analog used as an anticancer drug. Severe, potentially life-threatening toxicities can occur from irinotecan treatment. Although multiple genes may play a role in irinotecan activity, the majority of evidence to date suggests that variation in expression of UGT1A1 caused by a common promoter polymorphism (UGT1A1*28) is strongly associated with toxicity; however, this link is dose dependent. Variations in other pharmacokinetic genes, particularly the transporter ABCC2, also contribute to irinotecan toxicity. In addition, recent studies have shown that pharmacodynamic genes such as TDP1 and XRCC1 can also play a role in both toxicity and response.

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36 ABCB1 1236C>T rs1128503 Decreased irinotecan clearance; risk of toxicity and reduced survival when in haplotype with 2677 and 3435 [56,58,59] ABCB1 2677G>A/T rs2032582 Risk of toxicity and reduced survival when in haplotype with 1236 and 3435 [58,59] ABCB1 3435C>T rs1045642 Increased toxicity; risk of toxicity and reduced survival when in haplotype with 1236 and 2677 [57-59] ABCC2 -24C>T rs717620 Increased response and survival with 3972; toxicity as part of ABCC2 haplotype in patients without GT1A1*28 [46,61-63] ABCC2 3972T>C rs3740066 Increased response and survival with -24; toxicity as part of ABCC2 haplotype in patients without UGT1A1*28 [43,46,60-63] ABCG2 34G>A rs2231137 Increased toxicity; no association with toxicity and outcome [60,61] ABCG2 421C>A; Q141K rs2231142 Reduced expression; irinotecan resistance; toxicity with IVS12 +49G>T [49,61,66,67] ABCG2 IVS12+49G>T rs3832043 Toxicity with 421C>A [49,61,66,67] CES1 -816A>C Unknown Altered CES1 promoter activity [22] CES2 830C>G; -171C>G rs11075646 No association with expression, catalytic activity, toxicity or outcome [20] CYP3A4 Activity Altered irinotecan dosing requirement [24] NR1I2 Expression Altered SN-38 glucuronidation [79] TDP1 IVS12+79T>G rs2401863 Response; no toxicity association at higher doses [74,75] UGT1A1 -3156G>A rs10929302 Increased risk of toxicity [32,33] UGT1A1 (TA)7 TAA, *28 rs8175347 Increased risk of toxicity; dose dependent [32,33,38-41] UGT1A1 G71R, *6 rs4148323 Increased risk of toxicity [44,47] UGT1A7 *2 (N129K and R131K) rs17868323, rs17868324 SN-38 glucuronidation; response [47,52] UGT1A7 *3 (N129K, R131K and W208R) rs17868323, rs17868324, rs11692021 SN-38 glucuronidation; increased risk of toxicity; altered response [47,52,54] UGT1A9 -118(dT) rs3832043 SN-30 glucuronidation; response [44,47,52,54] XRCC1 Haplotype (-1149delGGCC, R399Q) rs321329 rs25487 Response [74] XRCC1 R399Q rs25487 No association with toxicity; association with response [76,77] dbSNP: SNP database Asians to 45% in Africans. Login to comment
77 The ABCG2 variant 421C>A (Q141K) reduced ABCG2 gene expression and caused irinotecan resistance in cancer cell lines [66] and neutropenia in 55 patients receiving irinotecan monotherapy when assessed as a haplotype with ABCG2 IVS12 +49G>T [67]. Login to comment
268 66 Imai Y, Nakane M, Kage K et al.: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Molecular mechanism of ochratoxin a transport in t... Toxins (Basel). 2010 Jun;2(6):1381-98. Epub 2010 Jun 9. Anzai N, Jutabha P, Endou H
Molecular mechanism of ochratoxin a transport in the kidney.
Toxins (Basel). 2010 Jun;2(6):1381-98. Epub 2010 Jun 9., [PMID:22069643]

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The mycotoxin, ochratoxin A (OTA), is thought to be responsible for Balkan endemic nephropathy. OTA accumulates in several tissues, especially in the kidneys and liver. The excretion of OTA into urine is thought to be mainly by tubular secretion, presumably via the organic anion transport system. Recently, several families of multispecific organic anion transporters have been identified: organic anion transporters (OATs), organic anion-transporting polypeptides (OATPs), oligopeptide transporters (PEPTs), and ATP-binding cassette (ABC) transporters, such as MRP2 and BCRP. These renal transporters mediate the transmembrane transport of OTA and play a pivotal role in the development of OTA-induced nephrotoxicity.

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163 In addition, the finding that the Q141K polymorphism associated with gout leads to decreased urate excretion capacity suggests that ABCG2 contributes to urate excretion at the apical membrane of renal proximal tubules or at the luminal membrane of the intestine [56], or both. Login to comment

[hide] Efflux and uptake transporters as determinants of ... Expert Opin Drug Metab Toxicol. 2010 May;6(5):621-32. Rodrigues AC
Efflux and uptake transporters as determinants of statin response.
Expert Opin Drug Metab Toxicol. 2010 May;6(5):621-32., [PMID:20367534]

Abstract [show]
IMPORTANCE OF THE FIELD: The important role of drug transporters in drug absorption and disposition has been well documented. Statins are subjected to active transport of membrane proteins of the superfamilies ATP-binding cassette and solute carrier, and there is limited understanding of the mechanisms by which differences in transporter expression and activity contributes to variability of pharmacokinetics (PKs)/pharmacodynamics (PDs) of statins. AREAS COVERED IN THIS REVIEW: This review aims to discuss the roles of drug transporters in the PKs and PDs of statins, and in drug interactions with statins. WHAT THE READER WILL GAIN: A comprehensive summary of the literature on this subject including in vitro and in vivo observations. TAKE HOME MESSAGE: In vivo and in vitro studies have shown that efflux and uptake transporters modulate the PKs/PDs of statins. Until now organic anion transporting polypeptides (OATP)1B1 variants have been considered major factors in limiting the uptake of statins and increasing statin exposure, and, consequently, increasing risk of myopathy. Further studies in pharmacogenetics and in vitro models to assess statin disposition and toxicity are required to understand the contribution of others transporters, such as multidrug resistance-associated protein (MRP)1, MRP2, breast cancer resistance protein, OATP2B1, OAT1B3 and OATP1A2, in interindividual variability to statins efficacy and safety.

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126 Reduced BCRP transport activity has been associated with the presence of the ABCG2 c.421C>A SNP (rs2231142), resulting in an amino acid change p.Gln141Lys [53-55]. Login to comment

[hide] [Transporters involved in hepatobiliary transport ... Nihon Yakurigaku Zasshi. 2010 Feb;135(2):76-9. Maeda K
[Transporters involved in hepatobiliary transport of drugs].
Nihon Yakurigaku Zasshi. 2010 Feb;135(2):76-9., [PMID:20154415]

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1 Jpn.)135,76~79(2010) 化合物を医薬品にするために必要な薬物動態試験(その 4)排泄③ 前田和哉胆汁排泄とトランスポーター要約:近年,ヒト肝臓において非常に多くのトランスポーターが同定・機能解析されるにつれて,トランスポーターの遺伝子多型や薬物間相互作用による機能変動が薬物動態に与える影響を明らかにするための臨床研究も続々と報告されてきている．それに伴い,異物解毒システムの中でのトランスポーターの重要性が広く認知されてきた．代謝によって消失すると考えられてきた薬物の中にも肝取り込みトランスポーターの基質が含まれていることが明らかとなり,取り込みトランスポーターの機能変動が薬物動態の変化につながる事例が複数報告されてきている．本稿では,ヒト肝臓に発現する主な薬物トランスポーターを紹介すると共に,これらの遺伝子多型・薬物間相互作用が薬物動態や薬効・副作用に与える影響について概説することを目指した．はじめに肝臓は,腎臓とならび,種々の薬物の異物解毒器官として重要な役割を果たしている．肝臓における異物解毒システムの構成要素としては,種々の代謝酵素とトランスポーターが挙げられる(図 1)．代謝酵素は, 物質の構造変換を通じて薬物を不活性化へと導く役割を果たすのに対して,トランスポーターは,血液側から肝臓中への薬物の取り込み,ならびに肝臓内から胆汁中への薬物および代謝物の排出に寄与することにより,効率よく血液側から胆汁への方向性のある経細胞輸送(ベクトル輸送)を実現している．本稿では,肝臓に発現するトランスポーターの基本特性について概説すると共に,臨床薬物動態における重要性についても触れる． 1. 肝臓に発現する主な薬物トランスポーター 1)取り込みトランスポーター NTCP(Na+ -taurocholate cotransporting polypeptide)は,肝臓の血管側にのみ選択的に発現するトランスポーターで,様々な胆汁酸を Na+ イオンと共輸送する．胆汁酸以外にも,dehydrodpiandrosterone sulfate(DHEAS)や estrone-3-sulfate(E-sul)などステロイド抱合体や,肝機能検査薬ブロモスルホフタレイン,HMG-CoA 還元酵素阻害薬ロスバスタチンも基質とする(1, 2)． OATP(organic anion transporting polypeptide)ファミリートランスポーターは,Na+ イオン非依存的に基質を輸送するトランスポーターで,ヒト肝臓においては,特に OATP1B1 と OATP1B3 が有機アニオン化合物の肝取込みに重要な役割を果たすと考えられている．これらトランスポーターは共にヒト肝臓に選択的に発現しており,極めて多様な構造のアニオン系化合物を認識する．基質の中には,HMG-CoA 還元酵素阻害薬 (スタチン)やアンジオテンシン変換酵素阻害薬やアンジオテンシン II 受容体拮抗薬(サルタン),また種々の抗がん薬など臨床上汎用されている医薬品が多数見受けられる(3)．OATP1B1 と OATP1B3 のアミノ酸レベルでのホモロジーは 80%に達しており,基質認識性は極めて類似しているが,一部の基質は,片方のトラキーワード:薬物トランスポーター,遺伝子多型,薬物間相互作用東京大学大学院薬学系研究科分子薬物動態学教室(〒 113-0033 東京都文京区本郷 7-3-1) E-mail: kmaeda@mol.f.u-tokyo.ac.jp 原稿受領日:2009 年 11 月 11 日,依頼原稿 Title: Transporters involved in hepatobiliary transport of drugs. Author: Kazuya Maeda OCT1 NTCP Na+ OATP1B1 (OATP2) OATP1B3 (OATP8) OATP2B1 (OATP-B) OAT2 MRP3 MRP4 -OH -OX Phase I 代謝 Phase II ATP ATP ADPADP MRP2 BCRPMDR1 BSEP ATP ADP ATP ADP ADP ADP ATP ATP MRP6 ADP ATP Phase 0 取り込み代謝 Phase III 排泄図 1 肝臓における薬物の解毒システム肝臓における解毒といえば,かつては,Phase I 代謝(主に CYPs による酸化代謝),Phase II 代謝(主にグルクロン酸,グルタチオン,硫酸基などの抱合代謝)であったが,最近では,Phase 0 肝取り込み輸送や Phase III 胆汁排泄などトランスポーターを介した膜透過が一連の異物解毒の流れに組み込まれるようになった． 77胆汁排泄とトランスポーターンスポーターにのみ選択的に認識される．例えば,強心配糖体ジゴキシンやコレシストキニンオクタペプチド(CCK-8)は OATP1B3 に認識されるが OATP1B1 には認識されない(4, 5)．また,同じサルタン類であっても,バルサルタン,オルメサルタンは,OATP1B1, OATP1B3 両方の基質になるものの,テルミサルタンについては,OATP1B3 選択的に輸送されることが報告されている(6-9)．また,OATP1B1 は,ビリルビンを基質として輸送する(10, 11)．一部の薬物は,高ビリルビン血症を副作用として呈することがあるが, OATP1B1 を強力に阻害することにより血清中ビリルビン濃度の上昇を招いたと推測されるものが報告されている(12)．一方,有機カチオン系化合物の肝取込みにおいては, 主に OCT1(organic cation transporter 1)が重要であると考えられる．OCT1 は,分子量の小さな水溶性の高いカチオン性化合物(type I カチオン)を輸送することが知られている(13)．基質薬物としては,抗ウィルス薬(アシクロビル,ガンシクロビル),H2 ブロッカー(ファモチジン,ラニチジン)や経口抗糖尿病薬メトホルミンなどが挙げられる．Wang らは,Oct1 ノックアウトマウスを用いてメトホルミンの体内動態および副作用の変動について検討した結果,Oct1 ノックアウトマウスで,メトホルミンの肝臓中濃度が著しく減少しており,さらにメトホルミン投与後の血中乳酸濃度の上昇が著しく抑制されることを見出した(14, 15)．このことから,OCT1 はメトホルミンの重篤な副作用である乳酸アシドーシスの発現を決定する因子の 1 つであることが推察されている． 2)胆汁排泄トランスポーター MDR1(multidrug resistance 1; P-glycoprotein) は, 肝臓のみならず,腎臓・小腸・血液脳関門など生体内のあらゆる場所に発現し,細胞内からの化合物の排出を担うトランスポーターである．MDR1 は,比較的脂溶性の高い塩基性・中性化合物を主に幅広く基質として認識する．MDR1 の胆汁排泄への関与は,ノックアウトマウスにおいて,ジゴキシンやベクロニウムの胆汁排泄が有意に低下する報告から示されている(16, 17)．一方,MRP2(multidrug resistance-associated protein 2)は,種々の抱合代謝物を含む有機アニオンの胆汁排泄に重要な役割を果たす．MRP2 のin vivoにおける機能は,Mrp2 を遺伝的に欠損したラット EHBR (Eisai hyperbilirubinemic rat)や GY/TR- ラットを用いた胆汁排泄の検討により明らかにされてきた．基質薬物としては,スタチン類やサルタン類,抗悪性腫瘍薬メトトレキサートや環状ペプチドのエンドセリン受容体拮抗薬 BQ-123 など多岐にわたる．極めて興味深い知見として,肝取り込みに関与する OATP トランスポーターと MRP2 は,アミノ酸配列から見ると全く類似性は認められないものの,基質認識性は極めて類似している．その結果として,有機アニオン類はOATPs により肝取り込みされた後,MRP2 により速やかに胆汁中へと排泄され,両者の協調的な働きにより効率的な異物解毒が実現されている(18)．また,MRP2 は還元型グルタチオンやビリルビンなど種々の内因性基質の胆汁排泄にも関与している．特に MRP2 の機能欠損は,高ビリルビン血症を主徴とする Dubin-Johnson 症候群という遺伝病を引き起こすことが知られている． BCRP(breast cancer resistance protein)も,種々の硫酸抱合体やグルクロン酸抱合体のみならず,非抱合の化合物として種々の抗菌薬(ニトロフラントイン, スルファサラジン,フルオロキノロン類など)や抗がん薬(エトポシド,カンプトテシン類,インドールカルバゾール類,イマチニブなど),フラボノイド類(ゲニステイン,クエルセチンなど)等,多岐にわたる構造の化合物を基質として認識すると共に,MDR1 や MRP2 と基質特異性がかなりオーバーラップしている (19, 20)．これまでプラバスタチンは,Mrp2 欠損ラットEHBRにおいて著しく胆汁排泄が低下することから, Mrp2 がスタチン類の胆汁排泄に重要な役割を果たすと信じられてきた(21)が,一方,同じスタチンのピタバスタチンでは,EHBR で胆汁排泄の低下が見られず, Bcrp ノックアウトマウスにおいて胆汁排泄が著しく低下することが見いだされた(22)．また,ロスバスタチンでは,EHBR,Bcrp ノックアウトマウスの両方で胆汁排泄の低下が見られ,MRP2,BCRP 両方の関与が考えられた(23)．このことは,類似の構造を有する同系統の薬物で共に胆汁排泄により消失する薬物であっても,それに関与する分子機構は異なるということを示した好例であるといえる．一方,BSEP(bile salt export pump)は,専ら胆汁酸を排出するトランスポーターであり,NTCP による肝取り込みと協調的に機能して,胆汁酸を効率よく血管側から胆汁へと排出している．BSEP は,進行性家族性肝内胆汁うっ滞 2 型(PFIC2)という致死性の遺伝病の原因遺伝子であり,胆汁酸が肝臓から排出できなくなり,肝臓内に胆汁酸が蓄積することにより発症する．また,薬剤誘導性胆汁うっ滞という副作用が知られている薬物のうち,抗糖尿病薬グリベンクラミドや抗結核薬リファンピシン,肺動脈性高血圧症治療薬ボセンタン,また致死的な肝障害が原因で市場から撤退した糖尿病治療薬トログリタゾンの硫酸抱合体などは BSEP を低濃度で強力に阻害することが in vitro 実験から明らかとなっている(24, 25)． 2. トランスポーターの遺伝子多型が薬物動態に与えるインパクトトランスポーターの重要性が認知されるに従い,薬物トランスポーターに関する遺伝子多型解析が急ピッチに進められ,臨床研究を通じて薬物動態や薬効・副作用との関連が明らかにされつつある． SLCO1B1(OATP1B1)の遺伝子多型については,特に頻תが高い A388G(N130D)と T521C(V174A)に 78 前田和哉焦点をあてた臨床研究が複数展開されている(26)． Nishizato らは,日本人における SLCO1B1 変異解析を行い,T521C 変異が高頻度に A388G 変異とリンクしていることを見出した(*15 アレル)(27)．日本人では *15 は,約 15% 程度のアレル頻度で存在する．またプラバスタチンの血中濃度が *15 アレル保持者において有意に高いことを見出し,世界に先駆けて SLCO1B1 の変異が薬物動態を変動させることをヒトで直接示した(27)．その後,表 1 に示すような多くの基質薬物について,T521C 変異保持者では血中濃度が上昇するという統一した結果が得られている．中にはこれまで代謝により消失すると考えられていた薬物も含まれており,取り込みトランスポーターと代謝酵素の両方の基質の場合,解毒の第 1 ステップである肝取り込み過程の機能変化は,固有クリアランスに直接影響することに留意すべきである．薬効や副作用との関連についてもいくつか報告がある．例えば,レパグリニドの血糖降下作用が T521C 変異保持者で上昇することが報告されている(28)．また,シンバスタチンにより筋毒性を誘発した患者についてゲノムワイドな多型解析を実施したところ,唯一 SLCO1B1 T521C 変異が統計的に有意なリスクファクターとして抽出されており, T521C 変異をホモで持つ患者は,野生型の患者と比較して 16.9 倍筋毒性を発症するリスクが高いことが見出されている(29)．一方,A388G 変異は,OATP1B1 の機能が亢進し,肝クリアランスの上昇につながることが示唆される臨床研究のデータが複数出されている (30, 31)が事例はまだ少ないことから,さらなる検討が求められる． BCRP の遺伝子多型では,主にアジア人で頻度の高い(アジア人:35%, 白人:10%)変異である C421A(Q141K)が注目されている．これまでにジフロモテカン,ロスバスタチン,スルファサラジンなど複数の基質薬物について C421A 変異保持者で血中濃度の上昇が見られている(32-34)．一方で,当研究室では in vitro 実験より C421A変異が単位タンパク質量あたりの輸送活性には影響を与えないことを見出しており(35), また,ヒト胎盤における BCRP の発現量を遺伝子型ごとに比較したところ, C421A変異を有している検体で,発現量が有意に低いことが示された(36)．よって臨床試験の結果は, 主に小腸に発現する BCRP 発現量の低下が原因であろうと推察されているが,ジフロモテカンにおいては静脈内投与時でも血中濃度に差が見られている(34)ことから,肝臓の胆管側の BCRP の機能低下も同時に起こっている可能性が考えうる． 3. トランスポーターを介した薬物間相互作用が薬物動態に与えるインパクト肝臓において,薬物間相互作用の標的としてのトランスポーターの重要性を広く知らしめるきっかけとなったのは,セリバスタチンとシクロスポリン A(CsA) の相互作用である．セリバスタチンは,致死的な横紋筋融解症を引き起こしたため市場から撤退を余儀なくされたが,その事例の多くで,ゲムフィブロジルなど他剤との併用が報告されていた．一方,CsA もセリバスタチンの血中濃度を上昇させることが臨床報告されていた(37)ことから,Shitara らは,そのメカニズム解明のため検討を行った(38)(図 2)．セリバスタチンは,主に CYP2C8,一部 CYP3A4 により代謝され消失することから,当初は CsA による代謝酵素の阻害が疑われていたが,肝ミクロソームによるセリバスタチンの代謝は,30μM CsA 存在下においても半分以下の阻害しか見られず,一方,セリバスタチンのヒト凍結肝細胞や OATP1B1 発現細胞における取り込みを CsA は Ki＝0.3 ~ 0.7μM と低濃度で阻害することが示された．これらよりセリバスタチンと CsA の相互作用は,CsA によるセリバスタチンの OATP1B1 を介した肝取り込み過程の阻害が原因であることが示唆された(図 2)．一方,ゲムフィブロジルとの相互作用については,主にはゲムフィブロジルグルクロニドによる CYP2C8 の mechanism-based inhibition であることが示されたが,一部ゲムフィブロジルグルクロニドによる肝取り込み過程の阻害も寄与することが示唆されている(39, 40)(図 2)．これ以後,CsA との併用投与により,複数の OATP 基質薬物(スタチン類,レパグリニド,ボセンタンなど)の血中濃度が上昇するという臨床報告がなされている．一方,in vitro 試験により臨床投与条件での肝臓入り口の最大のタンパク質非結合型血中濃度の見積もり値と OATP1B1,OATP1B3 の阻害定数 (Ki)との比較を行ったところ,OATP1B1 については, CysA に加えて抗結核薬リファンピシン,リファマイシン SV,抗 HIV 治療薬のインジナビル,リトナビル, 抗生物質クラリスロマイシンが,OATP1B3 については,CysA とリファンピシンがそれぞれ阻害しうることが示唆された(41, 42)．最大濃度を仮定した時の相互作用リスク評価ではあるが,これら薬剤と OATP 基質との相互作用の報告も実際にあることから,今後注意する必要があると思われる．おわりに以上,肝臓の肝胆系輸送に関与するトランスポータ表 1 OATP1B1 の遺伝子多型が薬物動態に与える影響 ~ OATP1B1*15 により血中濃度が上昇した事例~ HMG-CoA 還元酵素阻害薬プラバスタチンシンバスタチン(acid 体) ピタバスタチンアトルバスタチンロスバスタチン高血糖治療薬レパグリニドナテグリニド抗アレルギー治療薬フェキソフェナジン肺動脈性高血圧治療薬アトラセンタン抗がん薬イリノテカン(SN-38) コレステロール吸収阻害薬エゼチミブループ利尿薬トラセミドアンジオテン̋ンII受容体拮抗薬オルメサルタン 79胆汁排泄とトランスポーターーと,それらが臨床薬物動態に与える影響について概説した．従来,代謝酵素により消失すると考えられてきた薬物のうち,肝取り込みトランスポーターの基質にもなる薬物が次第に増加傾向にあり,この場合,遺伝子多型や薬物間相互作用による取り込みトランスポーターの機能変動は,直接肝固有クリアランスに影響を与えることから,留意が必要である．現在ではヒト肝臓試料(凍結肝細胞・肝ブロック)が入手可能となり,ヒトにおける肝取り込み・胆汁排泄の予測が可能となりつつある．一方,ヒトでは容易に測定できない肝臓中濃度についても PET や SPECT などイメージング技術を利用することで非侵襲的に定量化することが出来ることから,予測の妥当性をより精度よく行える環境が整いつつある．今後,ヒト体内動態予測のための in vitro 実験・モデル構築と妥当性評価のための臨床研究が並行して行われ,予測事例がさらに集積する必要性があると考えている．文献 1)Ho RH, et al. Gastroenterology. 2006;130:1793-1806. 2)Meier PJ, et al. Hepatology. Login to comment

[hide] Genetic polymorphisms of uptake (OATP1B1, 1B3) and... Expert Opin Drug Metab Toxicol. 2009 Jul;5(7):703-29. Ieiri I, Higuchi S, Sugiyama Y
Genetic polymorphisms of uptake (OATP1B1, 1B3) and efflux (MRP2, BCRP) transporters: implications for inter-individual differences in the pharmacokinetics and pharmacodynamics of statins and other clinically relevant drugs.
Expert Opin Drug Metab Toxicol. 2009 Jul;5(7):703-29., [PMID:19442037]

Abstract [show]
Recent pharmacogenomic/pharmacogenetic studies have disclosed important roles of drug transporters in the pharmacokinetic/pharmacodynamic (PK/PD) profiles of some clinically relevant drugs. It has concurrently been explained that variations in the drug transporter genes are associated with not only inter-individual but also inter-ethnic differences in PK/PD profiles of these drugs. This review focuses on two uptake and two efflux transporters. Organic anion transporting polypeptide (OATP) 1B1 and OATP1B3 are uptake transporters, specifically expressed in the liver, and considered important for drugs, particularly as their pharmacological target organ is the liver. Two ATP-binding cassette transporters, multi-drug resistance-associated protein 2 and breast cancer resistance protein, are efflux transporters, expressed in various human tissues, and considered particularly important for intestinal drug absorption and hepatic drug elimination. All 3-hydroxyl-3-methylglutaryl-CoA reductase inhibitors (statins) except fluvastatin are substrates for OATP1B1, but hepatobiliary (canalicular) efflux transporters differ among statins. In this review, we update the pharmacogenomic/pharmacogenetic properties of these transporters and their effects on PK/PD profiles of statins and other clinically relevant drugs. In addition, we describe a physiologically-based pharmacokinetic model for predicting the effects of changes in transporter activities on systemic and hepatic exposure to pravastatin.

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546 Kim HS, Sunwoo YE, Ryu JY, et al. The effect of ABCG2 V12M, Q141K and Q126X, known functional variants in vitro, on the disposition of lamivudine. Login to comment

[hide] Pharmacogenetic pathway analysis of irinotecan. Clin Pharmacol Ther. 2008 Sep;84(3):393-402. Epub 2008 Apr 16. Rosner GL, Panetta JC, Innocenti F, Ratain MJ
Pharmacogenetic pathway analysis of irinotecan.
Clin Pharmacol Ther. 2008 Sep;84(3):393-402. Epub 2008 Apr 16., [PMID:18418374]

Abstract [show]
Irinotecan, a chemotherapeutic agent against various solid tumors, is a prodrug requiring activation to SN-38. Irinotecan's complex pharmacokinetics potentially allow for many genetic sources of variability. We explored relationships between pharmacokinetic pathways and polymorphisms in genes associated with irinotecan's metabolism and transport. We fitted a seven-compartment pharmacokinetic model with enterohepatic recirculation (EHR) to concentrations of irinotecan and metabolites SN-38, SN-38 glucuronide (SN-38G), and aminopentanoic acid (APC). Principal component analysis (PCA) of patient-specific parameter estimates produced measures interpretable along pathways. Nine principal components provided good characterization of the overall variation. Polymorphisms in genes UGT1A1, UGT1A7, and UGT1A9 had strong associations with a component corresponding to the irinotecan-to-SN-38 pathway and SN-38 recirculation and to a component relating to SN-38-to-SN-38G conversion and elimination of SN-38G. The component characterizing irinotecan's compartments was associated with HNF1alpha and ABCC2 polymorphisms. The exploratory analysis with PCA in this pharmacogenetic analysis was able to identify known associations and may have allowed identification of previously uncharacterized functional polymorphisms.

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97 Table 3 Associations between the principal components and polymorphisms Polymorphism PC1 PC2 PC3 PC4 PC5 PC6 PC7 PC8 PC9 UGT1A1, -53A(TA)6>7TAA, PROMOTER 0.086/0.5 0.301/0.2 0.007/4.9 0.019/1.8 0.216/0.2 0.009/3.4 0.314/0.2 0.204/0.2 0.123/0.4 UGT1A1, -3279G>T (UGT1A1*60), PBREM 0.021/1.7 0.056/0.7 0.043/0.9 0.027/1.3 0.588/0.1 0.001/24.8 0.482/0.1 0.527/0.1 0.046/0.8 UGT1A1, -3156G>A, PBREM 0.008/4.2 0.136/0.3 0.014/2.6 0.017/2.0 0.322/0.2 0.005/6.7 0.031/1.1 0.268/0.2 0.083/0.5 UGT1A7, 387G>T (N129K), EXON 1 0.007/4.6 0.139/0.3 0.001/26.7 0.050/0.8 0.050/0.8 0.103/0.4 0.116/0.4 0.186/0.3 0.114/0.4 UGT1A7, 622T>C (W208R), EXON 1 0.000/77.5 0.392/0.2 0.002/17.3 0.019/1.8 0.332/0.2 0.003/11.2 0.289/0.2 0.055/0.7 0.270/0.2 UGT1A9, -118(T)9>10, UGT1A9*1b, PROMOTER 0.003/12.3 0.258/0.2 0.001/36.4 0.017/2.0 0.054/0.7 0.025/1.4 0.067/0.6 0.279/0.2 0.066/0.6 UGT1A9, -2152C>T, PROMOTER 0.809/0.1 0.453/0.1 0.293/0.2 0.703/0.1 0.328/0.2 0.473/0.1 0.615/0.1 0.238/0.2 0.784/0.1 UGT1A9, -275T>A, PROMOTER 0.632/0.1 0.217/0.2 0.297/0.2 0.764/0.1 0.148/0.3 0.583/0.1 0.527/0.1 0.245/0.2 0.944/0.1 HNF1α, 79A>C (I27L), EXON 1 0.625/0.1 0.001/37.3 0.154/0.3 0.434/0.1 0.527/0.1 0.423/0.2 0.517/0.1 0.366/0.2 0.213/0.2 CYP3A4, -392A>G, CYP3A4*1B, 5ʹ-UTR 0.414/0.2 0.556/0.1 0.337/1.2 0.967/0.4 0.721/0.1 0.323/0.2 0.772/0.2 0.487/0.3 0.923/0.1 CYP3A5, 6986A>G, CYP3A5*3, INTRON 3 0.861/0.4 0.179/0.9 0.255/0.5 0.480/0.1 0.124/0.4 0.704/0.1 0.536/0.1 0.822/0.1 0.443/ 0.1 SLCO1B1, 388A>G (N130D), SLCO1B1*1b, EXON 4 0.079/0.5 0.106/0.4 0.023/1.6 0.097/0.4 0.580/0.1 0.379/0.2 0.317/0.2 0.038/1.0 0.269/0.2 SLCO1B1, 521T>C (V174A), SLCO1B1*15, EXON 5 0.878/0.1 0.614/0.6 0.600/0.1 0.433/0.2 0.751/0.2 0.159/0.5 0.942/0.1 0.145/0.3 0.066/0.6 ABCC2, -1549A>G, 5ʹ-Flanking region 0.383/0.2 0.001/47.4 0.301/0.2 0.308/0.2 0.171/0.3 0.749/0.1 0.705/0.1 0.253/0.2 0.643/0.1 ABCC2, -1019A>G, 5ʹ-Flanking region 0.583/0.1 0.002/15.4 0.254/0.2 0.249/0.2 0.398/0.2 0.732/0.1 0.681/0.1 0.226/0.2 0.809/0.1 ABCC2, -24C>T, 5ʹ-UTR 0.985/0.1 0.013/2.7 0.575/0.2 0.950/1.1 0.054/0.9 0.221/0.7 0.402/0.2 0.641/0.1 0.366/0.6 ABCC2, 1249G>A (V417I), EXON 10 0.443/0.1 0.045/0.9 0.934/0.1 0.358/0.2 0.521/0.1 0.329/0.2 0.495/0.1 0.002/14.9 0.706/0.1 ABCC2, -34T>C, INTRON 26 0.469/0.1 0.258/0.2 0.963/0.1 0.167/0.3 0.639/0.1 0.829/0.1 0.049/0.8 0.734/0.1 0.345/0.2 ABCC2, 3972C>T (I1324I), EXON 28 0.250/0.2 0.011/3.1 0.224/0.2 0.103/0.4 0.013/2.5 0.144/0.3 0.175/0.3 0.200/0.3 0.149/0.3 ABCC1, 1062T>C (N354N), EXON 9 0.136/0.3 0.179/0.3 0.221/0.2 0.120/0.4 0.139/0.3 0.684/0.1 0.013/2.3 0.228/0.2 0.082/0.5 ABCC1, -48C>T, INTRON 11 0.302/0.2 0.187/0.3 0.840/0.2 0.175/0.3 0.105/0.4 0.748/0.5 0.577/0.2 0.642/0.1 0.084/0.6 ABCC1, 1684T>C (L562L), EXON 13 0.405/0.2 0.018/2.0 0.414/0.2 0.098/0.4 0.579/0.1 0.436/0.1 0.805/0.1 0.037/1.0 0.233/0.2 ABCC1, -30C>G, INTRON 18 0.188/0.3 0.004/8.0 0.362/0.2 0.155/0.3 0.879/0.1 0.620/0.1 0.526/0.1 0.061/0.6 0.177/0.3 ABCC1, 4002G>A (S1334S), EXON 28 0.001/29.4 0.022/1.7 0.300/0.2 0.195/0.3 0.416/0.2 0.096/0.4 0.184/0.3 0.064/0.6 0.072/0.6 ABCC1, +18A>G, INTRON 30 0.023/1.6 0.198/0.3 0.424/0.2 0.825/0.1 0.365/0.2 0.296/0.2 0.217/0.2 0.403/0.2 0.236/0.2 ABCB1, -129T>C, 5ʹ-UTR 0.559/0.5 0.811/0.1 0.610/0.3 0.977/0.2 0.725/0.9 0.807/0.4 0.163/0.3 0.177/0.3 0.009/3.5 ABCB1, -25G>T, INTRON 4 0.229/0.3 0.774/0.1 0.832/0.5 0.826/1.1 0.635/0.1 0.877/0.2 0.368/0.2 0.661/0.1 0.832/0.1 ABCB1, -44A>G, INTRON 9 0.147/0.3 0.605/0.1 0.618/0.1 0.570/0.1 0.109/0.4 0.156/0.3 0.096/0.4 0.338/0.2 0.051/0.8 ABCB1, 1236C>T (G412G), EXON 12 0.182/0.3 0.437/0.1 0.382/0.2 0.482/0.1 0.090/0.5 0.280/0.2 0.106/0.4 0.376/0.2 0.153/0.3 ABCB1, +24C>T, INTRON 13 0.725/0.1 0.439/0.1 0.491/0.1 0.540/0.1 0.532/0.1 0.076/0.5 0.100/0.4 0.306/0.2 0.016/2.1 ABCB1, +38A>G, INTRON 14 0.627/0.1 0.538/0.1 0.669/0.1 0.540/0.1 0.532/0.1 0.054/0.7 0.100/0.4 0.306/0.2 0.033/1.1 ABCB1, 2677G>A/T (A893T/S), EXON 21 0.302/0.2 0.543/0.1 0.491/0.1 0.962/0.1 0.943/0.1 0.309/0.2 0.210/0.2 0.890/0.1 0.004/6.9 ABCB1, 3435C>T (I1145I), EXON 26 0.319/0.2 0.441/0.1 0.531/0.1 0.631/0.1 0.664/0.1 0.644/0.1 0.402/0.2 0.226/0.2 0.013/2.5 ABCG2, 34G>A (V12M), EXON 2 0.479/0.1 0.828/0.1 0.139/0.3 0.348/0.2 0.588/0.2 0.673/0.1 0.087/0.5 0.219/0.2 0.780/0.1 ABCG2, 421C>A (Q141K), EXON 5 0.565/0.1 0.397/0.2 0.421/0.2 0.435/0.1 0.628/0.1 0.256/0.2 0.708/0.1 0.533/0.1 0.787/0.1 CES2, -363C>G, 5ʹ-UTR 0.999/2.3 0.546/0.2 0.028/1.9 0.624/0.1 0.872/0.1 0.899/0.1 0.379/0.6 0.92/.1 0.586/0.1 CES2, +1361A>G, INTRON 1 0.381/1.0 0.549/0.2 0.616/0.8 0.118/0.4 0.546/0.1 0.629/0.1 0.275/0.3 0.26/0.2 0.352/0.2 Split to SN-38 and SN-38 to bile to gut to SN-38 IRN compartments SN-38 to SN-38G and SN-38G elimination Split to APC from IRN central compartment APC elimination EHR SN-38 recir-- culation without EHR SN-38 elimination IRN elimination The table shows the P values and Bayes factors, respectively, separated by a"/. Login to comment
193 UGT1A1, -53A(TA)6>7TAA, PROMOTER 0.709(6):0.286(7) 0.62(6):0.38(7) 15,32 UGT1A1, -3279G>T (UGT1A1*60), PBREM 0.47(G):0.53(T) 0.85(G):0.15(T) UGT1A1, -3156G>A, PBREM 0.69(G):0.31(A) 0.715(G):0.285(A) UGT1A1, 211G>A (G71R), UGT1A1*6, EXON 1 0(A):1(G) 0(A):1(G) UGT1A1, 686C>A (P229Q), UGT1A1*27, EXON 1 0(A):1(C) 0(A):1(C) UGT1A7, 387G>T (N129K), EXON 1 0.646(G):0.354(T) 0.522(G):0.478(T) Supplementary Data S1 onlinea UGT1A7, 391C>A (R131K), EXON 1 0.646(G):0.354(T) 0.522(G):0.478(T) UGT1A7, 622T>C (W208R), EXON 1 0.521(T):0.479(C) 0.729(T):0.271(C) UGT1A9, -118(T)9>10, UGT1A9*1b, PROMOTER 0.59(9):0.41(10) 0.56(9):0.44(10) 42 UGT1A9, -2152C>T, PROMOTER 0.91(C):0.09(T) Unknownb UGT1A9, -275T>A, PROMOTER 0.91(T):0.09(A) Unknownb HNF1α, 79A>C (I27L), EXON 1 0.75(A):0.25(C) Unknownb Supplementary Data S1 onlinea CYP3A4, -392A>G, CYP3A4*1B, 5ʹ-UTR 0.977(A):0.023(G) 0.321(A):0.679(G) 43 CYP3A5, 6986A>G, CYP3A5*3, INTRON 3 0.023(A):0.977(G) 0.633(A):0.367(G) SLCO1B1, 388A>G (N130D), SLCO1B1*1b, EXON 4 0.396(C):0.604(T) 0.717(C):0.283(T) Supplementary Data S1 onlinea SLCO1B1, 521T>C (V174A), SLCO1B1*15, EXON 5 0.083(C):0.917(T) 0.022(C):0.978(T) ABCC1, 1062T>C (N354N), EXON 9 0.458(C):0.542(T) 0.643(C):0.357(T) 44 ABCC1, +8A>G, INTRON 9 0.643(A):0.357(G) 0.433(A):0.567(G) ABCC1, -48C>T, INTRON 11 0.146(T):0.854(C) 0(T):1.0(C) ABCC1, 1684T>C (L562L), EXON 13 0.917(C):0.083(T) 0.848(C):0.152(T) ABCC1, -30C>G, INTRON 18 0.042(C):0.958(G) 0.217(C):0.783(G) ABCC1, 4002G>A (S1334S), EXON 28 0.688(C):0.312(T) 0.955(C):0.045(T) ABCC1, +18A>G, INTRON 30 0.213(T):0.787(C) 0.042(T):0.958(C) ABCC2, -1549A>G, 5ʹ-Flanking region 0.43(A):0.57(G) 0.485(A):0.515(G) 44 ABCC2, -1019A>G, 5ʹ-Flanking region 0.43(G):0.57(A) 0.365(G):0.635(A) ABCC2, -24C>T, 5ʹ-UTR 0.230(A):0.770(G) 0.06(A):0.940(G) ABCC2, 1249G>A (V417I), EXON 10 0.146(A):0.854(G) 0.239(A):0.761(G) ABCC2, -34T>C, INTRON 26 0.17(C):0.83(T) 0.25(C):0.75(T) ABCC2, 3972C>T (I1324I), EXON 28 0.380(A):0.620(G) 0.280(A):0.720(G) ABCB1, -129T>C, 5ʹ-UTR 0.620(C):0.938(T) 0.043(C):0.957(T) 44 ABCB1, -25G>T, INTRON 4 0.273(T):0.737(G) 0.385(T) :0.615(G) ABCB1, -44A>G, INTRON 9 0.409(C):0.591(T) 0.20(C):0.80(T) ABCB1, 1236C>T (G412G), EXON 12 0.523(C):0.477(T) 0.864(C):0.136(T) ABCB1, +24C>T, INTRON 13 0.50(T):0.50(C) 0.467(T):0.533(C) ABCB1, +38A>G, INTRON 14 0.429(A):0.571(G) 0.389(A):0.611(G) ABCB1, 2677G>A/T (A893T/S), EXON 21 0.614(G):0.386(T) 0.923(G):0.077(T) ABCB1, 3435C>T (I1145I), EXON 26 0.375(C):0.625(T) 0.848(C):0.152(T) ABCG2, 34G>A (V12M), EXON 2 0.017(A):0.983(G) 0.071(A):0.929(G) Supplementary Data S1 onlinea ABCG2, 421C>A (Q141K), EXON 5 0.045(A):0.955(C) 0.023(A):0.977(C) CES2, -363C>G, 5ʹ-UTR 0.810(C):0.190(G) 0.733(C):0.267(G) Supplementary Data S1 onlinea CES2, +1361A>G, INTRON 1 0.143(G):0.857(A) 0.438(G):0.562(A) CES2, 108C>G, 3ʹ-UTR 0.004(G):0.996(C) 0(G):1.0(C) PBREM, phenobarbital-responsive enhancer module; UTR, untranslated region. Login to comment

[hide] Apical/basolateral surface expression of drug tran... Pharm Res. 2005 Oct;22(10):1559-77. Epub 2005 Sep 22. Ito K, Suzuki H, Horie T, Sugiyama Y
Apical/basolateral surface expression of drug transporters and its role in vectorial drug transport.
Pharm Res. 2005 Oct;22(10):1559-77. Epub 2005 Sep 22., [PMID:16180115]

Abstract [show]
It is well known that transporter proteins play a key role in governing drug absorption, distribution, and elimination in the body, and, accordingly, they are now considered as causes of drug-drug interactions and interindividual differences in pharmacokinetic profiles. Polarized tissues directly involved in drug disposition (intestine, kidney, and liver) and restricted distribution to naive sanctuaries (blood-tissue barriers) asymmetrically express a variety of drug transporters on the apical and basolateral sides, resulting in vectorial drug transport. For example, the organic anion transporting polypeptide (OATP) family on the sinusoidal (basolateral) membrane and multidrug resistance-associated protein 2 (MRP2/ABCC2) on the apical bile canalicular membrane of hepatocytes take up and excrete organic anionic compounds from blood to bile. Such vectorial transcellular transport is fundamentally attributable to the asymmetrical distribution of transporter molecules in polarized cells. Besides the apical/basolateral sorting direction, distribution of the transporter protein between the membrane surface (active site) and the intracellular fraction (inactive site) is of practical importance for the quantitative evaluation of drug transport processes. The most characterized drug transporter associated with this issue is MRP2 on the hepatocyte canalicular (apical) membrane, and it is linked to a genetic disease. Dubin-Johnson syndrome is sometimes caused by impaired canalicular surface expression of MRP2 by a single amino acid substitution. Moreover, single nucleotide polymorphisms in OATP-C/SLC21A6 (SLCO1B1) also affect membrane surface expression, and actually lead to the altered pharmacokinetic profile of pravastatin in healthy subjects. In this review article, the asymmetrical transporter distribution and altered surface expression in polarized tissues are discussed.

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212 Kondo et al. (119) also examined the cellular localization of a total of seven SNP variants of BCRP (V12M, Q141K, A149P, R163K, Q166E, P269S, and S441N) in LLC-PK1. Login to comment
213 As a result, reduced protein expression levels of Q141K and S441N were observed compared with the wild-type BCRP. Login to comment

[hide] Functional hot spots in human ATP-binding cassette... Protein Sci. 2010 Nov;19(11):2110-21. Kelly L, Fukushima H, Karchin R, Gow JM, Chinn LW, Pieper U, Segal MR, Kroetz DL, Sali A
Functional hot spots in human ATP-binding cassette transporter nucleotide binding domains.
Protein Sci. 2010 Nov;19(11):2110-21., [PMID:20799350]

Abstract [show]
The human ATP-binding cassette (ABC) transporter superfamily consists of 48 integral membrane proteins that couple the action of ATP binding and hydrolysis to the transport of diverse substrates across cellular membranes. Defects in 18 transporters have been implicated in human disease. In hundreds of cases, disease phenotypes and defects in function can be traced to nonsynonymous single nucleotide polymorphisms (nsSNPs). The functional impact of the majority of ABC transporter nsSNPs has yet to be experimentally characterized. Here, we combine experimental mutational studies with sequence and structural analysis to describe the impact of nsSNPs in human ABC transporters. First, the disease associations of 39 nsSNPs in 10 transporters were rationalized by identifying two conserved loops and a small alpha-helical region that may be involved in interdomain communication necessary for transport of substrates. Second, an approach to discriminate between disease-associated and neutral nsSNPs was developed and tailored to this superfamily. Finally, the functional impact of 40 unannotated nsSNPs in seven ABC transporters identified in 247 ethnically diverse individuals studied by the Pharmacogenetics of Membrane Transporters consortium was predicted. Three predictions were experimentally tested using human embryonic kidney epithelial (HEK) 293 cells stably transfected with the reference multidrug resistance transporter 4 and its variants to examine functional differences in transport of the antiviral drug, tenofovir. The experimental results confirmed two predictions. Our analysis provides a structural and evolutionary framework for rationalizing and predicting the functional effects of nsSNPs in this clinically important membrane transporter superfamily.

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72 Predictions of the Functional Effects of 40 nsSNPs in ABC Transporters Comon name HUGO name Mutation NBD Prediction BSEP ABCB11 E592Q NBD1 Neutral BSEP ABCB11 N591S NBD1 Neutral BSEP ABCB11 Q558H NBD1 Neutral BSEP ABCB11 V444A NBD1 Neutral BSEP ABCB11 E1186K NBD2 Disease MDR1 ABCB1 P1051A NBD2 Neutral MDR1 ABCB1 S1141T NBD2 Neutral MDR1 ABCB1 T1256K NBD2 Disease MDR1 ABCB1 V1251I NBD2 Neutral MDR1 ABCB1 W1108R NBD2 Disease MRP2 ABCC2 I670T NBD1 Disease MRP2 ABCC2 L849R NBD1 Disease MRP2 ABCC2 C1515Y NBD2 Disease MRP3 ABCC3 D770N NBD1 Neutral MRP3 ABCC3 K718M NBD1 Neutral MRP3 ABCC3 T809M NBD1 Disease MRP3 ABCC3 V765L NBD1 Disease MRP3 ABCC3 Q1365R NBD2 Disease MRP3 ABCC3 R1297H NBD2 Disease MRP3 ABCC3 R1348C NBD2 Disease MRP3 ABCC3 R1381S NBD2 Disease MRP4 ABCC4 G487E NBD1 Disease MRP4 ABCC4 K498E NBD1 Neutral MRP4 ABCC4 R1220Q NBD2 Neutral MRP4 ABCC4 T1142M NBD2 Neutral MRP4 ABCC4 V1071I NBD2 Neutral MRP6 ABCC6 I1330L NBD1 Neutral MRP6 ABCC6 I742V NBD1 Neutral MRP6 ABCC6 P664S NBD1 Neutral MRP6 ABCC6 R724K NBD1 Neutral MRP6 ABCC6 R769K NBD1 Neutral MRP6 ABCC6 A1291T NBD2 Neutral MRP6 ABCC6 E1369K NBD2 Neutral MRP6 ABCC6 G1327E NBD2 Disease MRP6 ABCC6 L1416R NBD2 Disease MRP6 ABCC6 R1268Q NBD2 Disease MRP6 ABCC6 R1461H NBD2 Disease MXR ABCG2 I206L NBD1 Neutral MXR ABCG2 P269S NBD1 Disease MXR ABCG2 Q141K NBD1 Neutral nsSNPs. Login to comment

[hide] Signatures of recent positive selection at the ATP... Hum Mol Genet. 2007 Jun 1;16(11):1367-80. Epub 2007 Apr 5. Wang Z, Wang J, Tantoso E, Wang B, Tai AY, Ooi LL, Chong SS, Lee CG
Signatures of recent positive selection at the ATP-binding cassette drug transporter superfamily gene loci.
Hum Mol Genet. 2007 Jun 1;16(11):1367-80. Epub 2007 Apr 5., [PMID:17412754]

Abstract [show]
Members of the ATP-binding cassette (ABC) superfamily of transporters have been implicated as major players in drug response. Single nucleotide polymorphisms (SNPs) in the ABC transporter genes may account for variation in drug response between individuals. Given the abundance of SNPs within the human genome, identification of functionally important SNPs is difficult. Here, we utilized signatures of recent positive selection (RPS) to identify SNPs in ABC genes that have potential functional significance by using the long-range-haplotype test to search for signatures of RPS at 18 ABC genes involved in drug transport. From the genotype data of these 18 ABC genes in four populations extracted from the HapMap database, at least one SNP in each of these genes displayed genomic signatures of RPS in at least one population. However, only 13 SNPs in 10 ABC genes from three populations retained statistical significance after Type I error reduction. The functional significance of six of these RPS SNPs, including those that failed multiple testing correction (MTC), has been reported previously. We experimentally confirmed a functional effect for two SNPs, including one that failed to show evidence of RPS after MTC. These observations suggest that Type I error reduction may inadvertently increase Type II error. Although the remaining positively selected SNPs have yet to be functionally validated, our study illustrates the feasibility of using this strategy to identify SNPs within 'adaptive' genes that may confer functional effect, prior to testing their roles in individual/population drug response variation or in complex disease susceptibility.

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211 SNP e5/C421A, which we found to be under RPS even after multiple test corrected (Table 2), results in a non-synonymous amino acid change (Q141K) and resides in the functionally important ATP-binding region between the Walker A and B motifs. Login to comment

[hide] Intestinal transporters for endogenic and pharmace... J Pharm Pharmacol. 2012 Nov;64(11):1523-48. doi: 10.1111/j.2042-7158.2012.01505.x. Epub 2012 Mar 30. Grandvuinet AS, Vestergaard HT, Rapin N, Steffansen B
Intestinal transporters for endogenic and pharmaceutical organic anions: the challenges of deriving in-vitro kinetic parameters for the prediction of clinically relevant drug-drug interactions.
J Pharm Pharmacol. 2012 Nov;64(11):1523-48. doi: 10.1111/j.2042-7158.2012.01505.x. Epub 2012 Mar 30., [PMID:23058041]

Abstract [show]
Objectives This review provides an overview of intestinal human transporters for organic anions and stresses the need for standardization of the various in-vitro methods presently employed in drug-drug interaction (DDI) investigations. Key findings Current knowledge on the intestinal expression of the apical sodium-dependent bile acid transporter (ASBT), the breast cancer resistance protein (BCRP), the monocarboxylate transporters (MCT) 1, MCT3-5, the multidrug resistance associated proteins (MRP) 1-6, the organic anion transporting polypetides (OATP) 2B1, 1A2, 3A1 and 4A1, and the organic solute transporter alpha/beta (OSTalpha/beta) has been covered along with an overview of their substrates and inhibitors. Furthermore, the many challenges in predicting clinically relevant DDIs from in-vitro studies have been discussed with focus on intestinal transporters and the various methods for deducting in-vitro parameters for transporters (K(m) /K(i) /IC50, efflux ratio). The applicability of using a cut-off value (estimated based on the intestinal drug concentration divided by the K(i) or IC50) has also been considered. Summary A re-evaluation of the current approaches for the prediction of DDIs is necessary when considering the involvement of other transporters than P-glycoprotein. Moreover, the interplay between various processes that a drug is subject to in-vivo such as translocation by several transporters and dissolution should be considered.

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517 Kim HS et al. The effect of ABCG2 V12M, Q141K and Q126X, known functional variants in vitro, on the disposition of lamivudine. Login to comment

[hide] Deletion of abcg2 has differential effects on excr... J Pharmacol Exp Ther. 2012 Nov;343(2):316-24. doi: 10.1124/jpet.112.197046. Epub 2012 Aug 6. Huang L, Be X, Tchaparian EH, Colletti AE, Roberts J, Langley M, Ling Y, Wong BK, Jin L
Deletion of abcg2 has differential effects on excretion and pharmacokinetics of probe substrates in rats.
J Pharmacol Exp Ther. 2012 Nov;343(2):316-24. doi: 10.1124/jpet.112.197046. Epub 2012 Aug 6., [PMID:22869929]

Abstract [show]
This study was designed to characterize breast cancer resistance protein (Bcrp) knockout Abcg2(-/-) rats and assess the effect of ATP-binding cassette subfamily G member 2 (Abcg2) deletion on the excretion and pharmacokinetic properties of probe substrates. Deletion of the target gene in the Abcg2(-/-) rats was confirmed, whereas gene expression was unaffected for most of the other transporters and metabolizing enzymes. Biliary excretion of nitrofurantoin, sulfasalazine, and compound A [2-(5-methoxy-2-((2-methyl-1,3-benzothiazol-6-yl)amino)-4-pyridinyl)-1,5,6,7-tetr ahydro-4H-pyrrolo[3,2-c]pyridin-4-one] accounted for 1.5, 48, and 48% of the dose in the Abcg2(+/+) rats, respectively, whereas it was decreased by 70 to 90% in the Abcg2(-/-) rats. Urinary excretion of nitrofurantoin, a significant elimination pathway, was unaffected in the Abcg2(-/-) rats, whereas renal clearance of sulfasalazine, a minor elimination pathway, was reduced by >90%. Urinary excretion of compound A was minimal. Systemic clearance in the Abcg2(-/-) rats decreased 22, 43 (p < 0.05), and 57%, respectively, for nitrofurantoin, sulfasalazine, and compound A administered at 1 mg/kg and 27% for compound A administered at 5 mg/kg. Oral absorption of nitrofurantoin, a compound with high aqueous solubility and good permeability, was not limited by Bcrp. In contrast, the absence of Bcrp led to a 33- and 11-fold increase in oral exposure of sulfasalazine and compound A, respectively. These data show that Bcrp plays a crucial role in biliary excretion of these probe substrates and has differential effects on systemic clearance and oral absorption in rats depending on clearance mechanisms and compound properties. The Abcg2(-/-) rat is a useful model for understanding the role of Bcrp in elimination and oral absorption.

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15 ABCG2 421CϾA SNP results in a lysine to glutamine acid change at codon 141 (Q141K), which leads to decreased protein expression level or reduced drug resistance to anticancer agents in transfected cells (Imai et al., 2002; Mizuarai et al., 2004; Morisaki et al., 2005; Tamura Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. Login to comment
21 The Q141K variant was associated with increased risk for gefitinib-induced diarrhea (Cusatis et al., 2006). Login to comment

[hide] Q141K polymorphism of ABCG2 protein is associated ... Haematologica. 2012 Oct 12. Tiribelli M, Fabbro D, Franzoni A, Fanin R, Damante G, Damiani D
Q141K polymorphism of ABCG2 protein is associated with poor prognosis in adult acute myeloid leukemia treated with idarubicin-based chemotherapy.
Haematologica. 2012 Oct 12., [PMID:23065526]

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0 Q141K polymorphism of ABCG2 protein is associated with poor prognosis in adult acute myeloid leukemia treated with idarubicin-based chemotherapy by Mario Tiribelli, Dora Fabbro, Alessandra Franzoni, Renato Fanin, Giuseppe Damante, and Daniela Damiani Haematologica 2012 [Epub ahead of print] Citation: Tiribelli M, Fabbro D, Franzoni A, Fanin R, Damante G, and Damiani D. Q141K polymorphism of ABCG2 protein is associated with poor prognosis in adult acute myeloid leukemia treated with idarubicin-based chemotherapy. Login to comment
13 Q141K polymorphism of ABCG2 protein is associated with poor prognosis in adult acute myeloid leukemia treated with idarubicin‐based chemotherapy Running title: ABCG2 Q141K polymorphism in acute myeloid leukemia Mario Tiribelli1,2 , Dora Fabbro3 , Alessandra Franzoni3 , Renato Fanin1,2 , Giuseppe Damante3,4 and Daniela Damiani1,2 1 Division of Hematology and Bone Marrow Transplantation, Azienda Ospedaliero‐Universitaria Udine, Udine, Italy; 2 Department of Experimental and Clinical Medical Sciences, University of Udine, Udine, Italy; 3 Institute of Genetics, Azienda Ospedaliero‐Universitaria Udine, Udine, Italy, and 4 Department of Medical and Biological Sciences, University of Udine, Udine, Italy Correspondence Daniela Damiani, MD, Clinica Ematologica Azienda Ospedaliero‐Universitaria, Piazzale S. M. Misericordia, 15, 33100 Udine, Italy. Login to comment
19 (4) Among them, the 421C>A (Q141K), that is also the commonest in Caucasian ethnicity (around 10% (5), has been shown to alter protein function and to modify in vitro sensitivity to many anticancer drugs (6-7), including classical anthracyclines and mitoxantrone. Login to comment
20 A recent report on the prognostic value of six ABCG2 gene polymorphisms in 184 Chinese acute leukemia patients seems to suggest that the presence of Q141K is associated with worse outcome. Login to comment
21 (8) Aim of this study was to evaluate the frequency of the Q141K variant in a cohort of Caucasian patients with acute myeloid leukemia (AML) and to assess the impact of this polymorphism on the outcome of a treatment strategy that included idarubicin as the only anthracycline. Login to comment
27 (1) ABCG2 genotype was analyzed with the TaqMan SNP Q141K assay (Applied Biosystems, C_15854163_70) and the test was performed in duplicate. Login to comment
28 Q141K polymorphism was detected in 29/163 patients (18%), and two patients were homozygous for the mutation. Login to comment
29 Of the 125 patients receiving chemotherapy, 26 displayed Q141K variant while 99 were ABCG2 wt. Login to comment
32 No impact on CR was demonstrated for ABCG2 overexpression and Q141K polymorphism. Login to comment
35 Again, ABCG2 Q141K polymorphism had no impact on relapse: among Q141K+ patients, relapse occurred in 7/16 (44%), while 20 of the 57 patients (%) with wt ABCG2 who attained CR relapsed; 3-years DFS was 47% [CI: 30-64] in Q141+ cases, and 61% [CI: 47-75] in wt patients (p = 0.). Login to comment
36 However, stratifying patients in three groups according to ABCG2 expression (low or high) and presence of Q141K polymorphism, a negative impact of Q141K emerged. Login to comment
41 As for DFS, Q141K polymorphism per se did not affect survival, but stratifying patients in the three groups, patients with low ABCG2 and wt gene had a longer OS compared to patients with Q141K ABCG2 or with high ABCG2 expression (55%, CI: 38-72 vs 40%, CI: 20-60 vs 27%, CI: 13-41, respectively) (χ2 = 7.5, p = 0.02) (Figure 1B). Login to comment
43 Q141K-ABCG2 is the most frequent polymorphism in Caucasians, and we detected it in 18%, slightly higher than what is reported in the literature (5, 9). Login to comment
47 (6) In our AML patients receiving a therapeutic program that includes idarubicin as the only anthracycline, Q141K is associated with poor outcome, comparable to that of patients over-expressing wild type ABCG2 protein. Login to comment
1 A recent report on the prognostic value of six ABCG2 gene polymorphisms in 184 Chinese acute leukemia patients seems to suggest that the presence of Q141K is associated with worse outcome.8 The aim of this study was to evaluate the frequency of the Q141K variant in a cohort of Caucasian patients with acute myeloid leukemia (AML) and to assess the impact of this polymorphism on the outcome of a treatment strategy that included idarubicin as the only anthracycline. Login to comment
5 ABCG2 expression was studied on blast cells by flow cytometry, as previously described.1 ABCG2 genotype was analyzed with the TaqMan SNP Q141K assay (Applied Biosystems, C_15854163_70) and the test was performed in duplicate. Login to comment
6 Q141K polymorphism was detected in 29 of 163 patients (18%), and 2 patients were homozygous for the mutation. Login to comment
7 Of the 125 patients receiving chemotherapy, 26 displayed Q141K variant while 99 were ABCG2 wt. Login to comment
10 No impact on CR was demonstrated for ABCG2 overexpression and Q141K polymorphism. Login to comment
14 However, when patients were stratified in three groups according to ABCG2 expression (low or high) and presence of Q141K polymorphism, a negative impact of Q141K emerged. Login to comment
17 Clinical characteristics of the 125 patients treated for AML, divided according to ABCG2 Q141K polymorphism. Login to comment
18 Q141K + (n = 26) Q141K - (n = 99) P Age: median (range) 56 (20-75) 60 (25-84) 0.36 WBC x109 /L): median (range) 10.2 (1.4-258.0) 16.0 (0.5-265.0) 0.63 FAB subtype: n. (%) 0.70 M0-M1 7/26 (27) 32/99 (32) M2 9/26 (35) 24/99 (24) M4-M5 10/26 (38) 43/99 (44) Karyotype: n. (%) 0.13 Favorable / Intermediate 19/22 (86) 55/82 (67) Unfavorable 3/22 (14) 27/83 (33) CD34+ : n. (%) 15/26 (58) 59/99 (60) 1.00 CD56+ : n. (%) 1/26 (4) 18/99 (18) 0.13 ABCG2 0.95 overexpression: n. (%) 15/26 (58) 54/99 (54%) mRNA: mean &#b1; SD 0.96&#b1;2.00 1.17&#b1;2.76 protein expression: mean &#b1; SD 0.37&#b1;0.28 0.30&#b1;0.19 Figure 1. Login to comment
26 Regarding DFS, Q141K polymorphism per se did not affect survival, but stratifying patients in the three groups, patients with low ABCG2 and wt gene had a longer OS compared to patients with Q141K ABCG2 or with high ABCG2 expression (55%, CI: 38-72% vs. 40%, CI: 20-60% vs. 27%, CI: 13-41, respectively) (c7;2 = 7.5, P=0.02) (Figure 1B). Login to comment
31 In a group of 184 Chinese patients with acute leukemia, C421A polymorphism was associated with worse survival.6 In our AML patients receiving a therapeutic program that includes idarubicin as the only anthracycline, Q141K is associated with poor outcome, comparable to that of patients over-expressing wild-type ABCG2 protein. Login to comment

[hide] The role of OATP1B1 and BCRP in pharmacokinetics a... Cardiovasc Ther. 2012 Oct;30(5):e234-41. doi: 10.1111/j.1755-5922.2011.00290.x. Epub 2011 May 25. Hua WJ, Hua WX, Fang HJ
The role of OATP1B1 and BCRP in pharmacokinetics and DDI of novel statins.
Cardiovasc Ther. 2012 Oct;30(5):e234-41. doi: 10.1111/j.1755-5922.2011.00290.x. Epub 2011 May 25., [PMID:21884024]

Abstract [show]
The aim of this review is to provide useful information not only for studying the effect of OATP1B1 and/or BCRP gene mutation on pharmacokinetics of novle statins of pitavastatin and rosuvastatin but also for studying drug-drug interactions (DDI) between the novle statins and other substrates of OATP1B1 and/or BCRP. Intra- and inter-ethnic differences in pharmacokinetic profiles of clinically relevant drugs are important issues reported in many papers not only for scenes of appropriate drug used in clinical settings but also for those of the drug development. Pharmacogenomics is extremely useful for understanding these racial differences. Recent pharmacogenetics study have disclosed important roles of drug transporters in the pharmacokinetic (PK) profiles of some clinically relevant drugs. In this presentation, we introduce single nucleotide polymorphisms (SNPs) of OATP1B1 and BCRP and review the contribution of genetic polymorphisms of the transporters to the pharmacokinetics of dual substrates as pitavastatin and rosuvastatin from recent study. At the same time, the DDIs between pitavastatin or rosuvastatin and other drug have been extensively concerned because of inhibiting OATP1B1-mediated hepatic uptake or BCRP-mediated hepatic efflux of pitavastatin and rosuvastatin. This review summarized the current studies about the role of OATP1B1 and BCRP in DDIs between pitavastatin or rosuvastatin and other clinically relevant drugs. The role of OATP1B1 and BCRP gene mutation can affect the PK profiles of pitavastatin and rosuvastatin. The DDIs between the novle statins and other substrates of OATP1B1 or BCRP may occur and cause change in the pharmacokinetic of the novle statins.

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43 Of these, the nonsynonymous 421C>A SNP that results in a glycine to lysine (Q141K) amino acid change has been studied most extensively. The Q141K SNP has been linked to decreased plasma membrane expression of BCRP, decreased drug transport, or reduced ATPase activity [20]. Login to comment
44 The study suggested that Q141K SNP has functional consequences in the resulting BCRP protein [21]. Login to comment
46 Thus, the Q141K SNP may lead to higher drug toxicity in some patient populations [23]. Login to comment
73 Of these, the nonsynonymous 421C>A SNP that results in a glycine to lysine (Q141K) amino acid change has been studied most extensively. The Q141K SNP has been linked to decreased plasma membrane expression of ABCG2, decreased drug transport or reduced ATPase activity [20,38]. Login to comment

[hide] The genetics of hyperuricaemia and gout. Nat Rev Rheumatol. 2012 Oct;8(10):610-21. doi: 10.1038/nrrheum.2012.144. Epub 2012 Sep 4. Reginato AM, Mount DB, Yang I, Choi HK
The genetics of hyperuricaemia and gout.
Nat Rev Rheumatol. 2012 Oct;8(10):610-21. doi: 10.1038/nrrheum.2012.144. Epub 2012 Sep 4., [PMID:22945592]

Abstract [show]
Gout is a common and very painful inflammatory arthritis caused by hyperuricaemia. This Review provides an update on the genetics of hyperuricaemia and gout, including findings from genome-wide association studies. Most of the genes that associated with serum uric acid levels or gout are involved in the renal urate-transport system. For example, the urate transporter genes SLC2A9, ABCG2 and SLC22A12 modulate serum uric acid levels and gout risk. The net balance between renal urate absorption and secretion is a major determinant of serum uric acid concentration and loss-of-function mutations in SLC2A9 and SLC22A12 cause hereditary hypouricaemia due to reduced urate absorption and unopposed urate secretion. However, the variance in serum uric acid explained by genetic variants is small and their clinical utility for gout risk prediction seems limited because serum uric acid levels effectively predict gout risk. Urate-associated genes and genetically determined serum uric acid levels were largely unassociated with cardiovascular-metabolic outcomes, challenging the hypothesis of a causal role of serum uric acid in the development of cardiovascular disease. Strong pharmacogenetic associations between HLA-B(*)5801 alleles and severe allopurinol-hypersensitivity reactions were shown in Asian and European populations. Genetic testing for HLA-B(*)5801 alleles could be used to predict these potentially fatal adverse effects.

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36 An analysis of the Atherosclerosis Risk in Com-muni- ties study showed that at least 10% of all gout cases in white individuals were attributable to the Gln141Lys causal variant.62 However, the frequency of the Gln141Lys risk allele has been reported to be as high as 32% in the Asian population.32,60 Although both Maoris and Pacific Islanders are known to have an increased risk of gout, the Gln141Lys variant is strongly associated with gout only in the western Polynesian population.61 This distribu-tion might be the result of chance depletion of the allele dur-ing eastward migration, as the frequency of the Gln141Lys variant in white populations has been reported to be <12%.37,61 SLC22A12 The SLC22A12 gene encodes URAT1, an essential urate transporter in proximal tubules that has a role in the apical absorption of urate66 and is a target of both urico-suric and antiuricosuric agents (Figure 2). Login to comment
58 In the liver, GCKR regulates glucokinase75,85 by mediating the phosphorylation of glucose to glucose‑6-phosphate, a precursor of liver glycogen synthesis and of de novo purine synthesis.75,85 A deficiency in the activity of glucose‑6-phosphatase causes glycogen storage disease type 1 (also known as von Gierke disease), which is characterized by hypertriglyceridaemia and hyperuricaemia.75,85 The GCKR SNP rs780094 might affect both serum uric acid and triglyceride levels (which are associated with insulin resistance) via a common mediator.75 Interestingly, GCKR is associated with insulin resistance,32 glucose level, triglyceride level, and C‑reactive protein,86 which are components of metabolic syndrome.75 Table 2 | SLC2A9, ABCG2 and SLC22A12 variants associated with serum uric acid levels, FeUA and gout Variant Location Phenotype Populations SLC2A9 (chromosome 4) rs1014290 Intron 3 SUA, FeUA, gout European ancestry34 rs6449213 Intron 4 SUA, FeUA, gout White,34-37,43 African American51,72 rs16890979 Exon 6 SUA, gout White,37,42,44 African American,72 Amish55 rs734553 Intron 6 SUA, gout White,32,39,44 Icelandic,13 African American72 rs7442295 Intron 6 SUA, gout White 35,38,39,44 rs737267 Intron 7 SUA, FeUA, gout European ancestry34,44 rs6855911 Intron 7 SUA, gout White,35,38,39,44 African American72 rs13129697 Intron 7 SUA, gout White,33,44 African American72 rs2241480 Intron 8 SUA, gout European ancestry72 rs7663032 Intron 9 SUA, gout African American,72 Croatian44 rs3775948 Intron 9 SUA Croatian,44 African American51 rs16890979 Intergenic SUA, gout White,37 Amish,55 Croatian,44 Pacific Islander,42 New Zealander42 rs717615 Intergenic SUA Croatian44 rs6856396 Intergenic SUA African American51 rs10489070 Intergenic Gout Amish55 ABCG2 (chromosome 4) rs2231137 Exon 2 SUA Japanese63 rs72552713 (Q126X) Exon 4 SUA, gout Japanese63 rs2231142 (Q141K) Exon 5 FeUA, SUA, gout White,32,37,39,44,62 African,37,72 Chinese,60 Icelandic,13 Japanese,59,63 Pacific Islander,61 New Zealander31,61 rs2199936 Intergenic SUA White 32,33,44 SLC22A12 (chromosome 11) rs11231825 Exon 1 FeUA, SUA Chinese,70 White,32,68 African American72 rs3825016 Exon 2 FeUA German68 rs12800450 Exon 2 SUA African American72 rs161109885 Intron 3 SUA Chinese73 rs893006 Intron 4 SUA Japanese,67 Chinese71 rs1529909 Intron 4 FeUA, SUA Korean74 rs475688 Intron 4 Gout Chinese,70 Solomon Islander70 rs17300741 Intron 4 SUA European32,75 rs7932775 Exon 8 SUA, FeUA, gout German,68 Chinese,70,73 Solomon Islander70 rs505802 Intergenic SUA European,32,44 African American72 rs11602903 Intergenic FeUA, SUA German,68 Chinese73 Abbreviations: FeUA, fractional excretion of urate; SUA, serum uric acid. Login to comment

[hide] The contribution of the ABCG2 C421A polymorphism t... BMC Cancer. 2012 Sep 1;12:383. doi: 10.1186/1471-2407-12-383. Chen P, Zhao L, Zou P, Xu H, Lu A, Zhao P
The contribution of the ABCG2 C421A polymorphism to cancer susceptibility: a meta-analysis of the current literature.
BMC Cancer. 2012 Sep 1;12:383. doi: 10.1186/1471-2407-12-383., [PMID:22937733]

Abstract [show]
ABSTRACT: BACKGROUND: ABCG2, also known as BCRP, is a half ATP-binding cassette (ABC) transporter that localizes to plasma membranes. Recently, a number of studies have investigated the relationship between the C421A polymorphism in ABCG2 and cancer risk in multiple populations and various types of cancers; however, this relationship remains unclear. Therefore, we performed a meta-analysis to further explore this association. METHODS: The meta-analysis incorporated 10 studies involving a total of 3593 cases and 5875 controls. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated based on the date extracted from the studies to evaluate the strength of association. We also analyzed the heterogeneity and sensitivity of each report and the publication bias of the studies. RESULTS: Overall, our results showed that there appeared to be a significant association between the ABCG2 C421A polymorphism and decreased cancer susceptibility (heterozygote-AC versus CC: OR = 0.759, 95%CI = 0.620-0.930; dominant effects model-AA/AC versus CC: OR = 0.771, 95%CI = 0.634-0.938; additive effects model-A allele versus C allele: OR = 0.809, 95%CI = 0.687-0.952). Similarly, decreased cancer risk was also found after stratification of the SNP data by cancer type, ethnicity and source of controls in heterozygote model, dominant effects model and additive effects model. CONCLUSIONS: We found that the ABCG2 C421A polymorphism is a protective factor for developing cancer. The same relationship was found when the studies were stratified by cancer type, ethnicity and source of controls.

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31 Researches had shown that there were two frequently polymorphic SNPs in the BCRP gene: one in exon2 (G34A, resulting in a V12M change) and the other in exon5 (C421A, resulting in a Q141K substitution) respectively [15]. Login to comment
24 Researches had shown that there were two frequently polymorphic SNPs in the BCRP gene: one in exon2 (G34A, resulting in a V12M change) and the other in exon5 (C421A, resulting in a Q141K substitution) respectively [15]. Login to comment
207 Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, Tsuruo T, Miki Y, Sugimoto Y: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Lack of ABCG2 shortens latency of BRCA1-deficient ... Cancer Prev Res (Phila). 2012 Aug;5(8):1053-60. Epub 2012 Jul 5. Zander SA, Kersbergen A, Sol W, Gonggrijp M, van de Wetering K, Jonkers J, Borst P, Rottenberg S
Lack of ABCG2 shortens latency of BRCA1-deficient mammary tumors and this is not affected by genistein or resveratrol.
Cancer Prev Res (Phila). 2012 Aug;5(8):1053-60. Epub 2012 Jul 5., [PMID:22767648]

Abstract [show]
In addition to their role in drug resistance, the ATP-binding cassette (ABC) transporters ABCG2 and ABCB1 have been suggested to protect cells from a broad range of substances that may foster tumorigenesis. Phytoestrogens or their metabolites are substrates of these transporters and the influence of these compounds on breast cancer development is controversial. Estrogen-like properties might accelerate tumorigenesis on the one hand, whereas their proposed health-protective properties might antagonize tumorigenesis on the other. To address this issue, we used a newer generation mouse model of BRCA1-mutated breast cancer and examined tumor latency in K14cre;Brca1(F/F); p53(F/F), Abcb1a/b(-/-);K14cre;Brca1(F/F); p53(F/F), or Abcg2(-/-);K14cre;Brca1(F/F); p53(F/F) animals, fed with genistein- or resveratrol-supplemented diets. Ovariectomized K14cre;Brca1(F/F); p53(F/F) animals were included to evaluate whether any estrogen-mimicking effects can restore mammary tumor development in the absence of endogenous estrogens. Compared with the ABC transporter proficient model, ABCG2-deficient animals showed a reduced median tumor latency of 17.5 days (P < 0.001), whereas no significant difference was observed for ABCB1-deficient animals. Neither genistein nor resveratrol altered this latency reduction in Abcg2(-/-);K14cre;Brca1(F/F); p53(F/F) animals. Ovariectomy resulted in nearly complete loss of mammary tumor development, which was not restored by genistein or resveratrol. Our results show that ABCG2 contributes to the protection of genetically instable epithelial cells against carcinogenesis. Diets containing high levels of genistein or resveratrol had no effect on mammary tumorigenesis, whether mice were lacking ABCG2 or not. Because genistein and resveratrol only delayed skin tumor development of ovariectomized animals, we conclude that these phytoestrogens are no effective modulators of mammary tumor development in our mouse model.

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132 The common SNP rs2231142 (421C>A), which encodes a Q141K loss of function mutation, causes at least 10% of all gout cases in the Caucasian American population of the United States (34). Login to comment
130 The common SNP rs2231142 (421C>A), which encodes a Q141K loss of function mutation, causes at least 10% of all gout cases in the Caucasian American population of the United States (34). Login to comment

[hide] EGF receptor-targeted therapy in non-small-cell lu... Future Oncol. 2012 Aug;8(8):1015-29. doi: 10.2217/fon.12.89. Galvani E, Peters GJ, Giovannetti E
EGF receptor-targeted therapy in non-small-cell lung cancer: role of germline polymorphisms in outcome and toxicity.
Future Oncol. 2012 Aug;8(8):1015-29. doi: 10.2217/fon.12.89., [PMID:22894673]

Abstract [show]
Conventional chemotherapeutic regimens have limited impact against most solid tumors and deal with significant toxicity. Over the last 10 years, novel anticancer treatments targeting specific molecules or genes involved in cancer progression have been developed to improve outcome and reduce side effects. In particular, the tyrosine kinase inhibitors gefitinib and erlotinib have been approved for the treatment of non-small-cell lung cancer. Their clinical activity has been related to different clinical and biological parameters, such as EGFR-activating mutations. However, not all clinical outcomes, including tolerability, are explained, and the identification/validation of novel biomarkers is a viable area of research. Germline polymorphisms can be easily assessed in blood samples, and candidate polymorphisms in EGFR and ABCG2 have been correlated with outcome and toxicity in non-small-cell lung cancer patients treated with gefitinib or erlotinib. However, differences in study population and design resulted in several controversial findings, while the prognostic versus predictive role of the polymorphisms still needs to be validated within larger prospective studies. More studies on the relationship of the genotype with drug pharmacokinetics and mechanism of action are also warranted. These future studies, as well as further development and application of novel technologies to decipher genetic alterations, might contribute to the validation of selected polymorphisms as molecular markers predictive of drug activity and help in the selection of tyrosine kinase inhibitors best suited to the individual patient.

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35 Polymorphism Location Proposed functional effect EGFR CA repeat Chrom. 7, intron 1 EGFR gene transcription declines with increasing number of CA repeats EGFR R497KA/G Chrom. 7, exon 13 Substitution of arginine by lysine is associated with decreased EGFR activity EGFR -216G/T Chrom. 7, promoter T allele is associated with higher EGFR promoter activity EGFR -191C/A Chrom. 7, exon 1 A allele is associated with increased EGFR protein production AKT1-SNP4 Chrom. 14, exon 11 A allele is associated with reduced AKT1 mRNA ABCG2 421C/A (Q141K) Chrom. 4, exon 5 A allele associated with reduced transport of TKIs ABCG2 -15622C/T Chrom. 4, promoter T allele is associated with lower ABCG2 expression ABCG2 1143C/T Chrom. 4, exon 4 T allele is associated with lower ABCG2 expression CYP3A5*3 Chrom. 7, exon 7 CYP3A5*3 variant is associated with significant reduction in CYP3A5 protein expression and activity CYP3A4*1B Chrom. 7, exon 7 CYP3A4*1B variant is associated with twofold higher promoter activity Chrom.: Chromosome; TKI: Tyrosine kinase inhibitor. Login to comment
57 Study Patients (n) Studied polymorphisms Effects Ref. Cusatis et al. (2006) 124 ABCG2: 421C>A (Q141K) Heterozygosis is associated with diarrhea [67] Brugger et al. (2011) 889 EGFR: Intr1, mut, copy-num, expression; KRAS: mut EGFR mut are associated with longer PFS KRAS mut are associated with reduced PFS [70] Hamada et al. (2012) 50 ABCB1: 1236TT, 2677TT, 3435TT 1236TT, 2677TT and 3435TT are associated with higher plasma concentration and risk of developing toxicity [73] copy-num: Gene copy-number; DCR: Disease control rate; Intr1: Intron 1; mut: Mutations; OS: Overall survival; PFS: Progression-free survival; PR: Partial response; RR: Response rate; SD: Stable disease; SNP: Single nucleotide polymorphism; TTP: Time to progression. Login to comment
68 In agreement with the previous study, EGFR activating muta‑ tions were associated with sensitivity to gefit‑ inib and OS, and short CA-repeat status was also correlated with better response and longer EGFR EGFEGF EGF ATP ATP P P Cell growth, proliferation, invasion, angiogenesis, metastasis and inhibition of apoptosis ABCG2/ABCB1 RAS RAF MEK ERK PI3K Akt mTor AKT AKT1-SNP4: A allele associated with reduced AKT1 mRNA Cytoplasmic membrane Cytoplasm Drug target CA repeat: EGFR gene transcription declines with increasing number of CA repeats R497KA/G: the substitution of arginine by lysine is associated with decreased EGFR activity 216G/T: T allele associated with higher EGFR promoter activity 191C/A: A allele associated with increased EGFR protein production Drug transporters ABCG2 421C/A (Q141K): A allele is related to reduced TKI transport ABCG2 15622C/T and 1143C/T: T allele is related to lower ABCG2 expression ABCB1 2677G>T/A: associated with reduced TKI clearance ABCB1 1236TT, 2677TT and 3435TT: genotype related to higher plasma concentration and risk of developing higher toxicity CYP450 CYP3A5*3: variant associated with significant reduction in protein activity CYP3A4*1B: variant associated with twofold higher promoter activity CYP2D6*1, *2xn: variants associated with extensive drug metabolizers CYP2D6*3, *4, *5, *6, *9, *14 and *29: variants associated with poor drug metabolizers CYP1A1*2A: variant correlated with EGFR activating mutations and EGFR TKI response CYPs Erlotinib Gefitinib Nucleus Figure 1. Login to comment
116 In particular, the ABCG2 421C/A polymorphism, resulting in a glutamine-to-lysine amino acid change at position 141 (Q141K), has been correlated with the downregulation of ABCG2 and with increase in accumulation of both gefitinib and erlotinib [53]. Login to comment

[hide] Histone deacetylase inhibitors influence chemother... Cancer Res. 2012 Jul 15;72(14):3642-51. Epub 2012 Apr 3. Basseville A, Tamaki A, Ierano C, Trostel S, Ward Y, Robey RW, Hegde RS, Bates SE
Histone deacetylase inhibitors influence chemotherapy transport by modulating expression and trafficking of a common polymorphic variant of the ABCG2 efflux transporter.
Cancer Res. 2012 Jul 15;72(14):3642-51. Epub 2012 Apr 3., [PMID:22472121]

Abstract [show]
Histone deacetylase inhibitors (HDI) have exhibited some efficacy in clinical trials, but it is clear that their most effective applications have yet to be fully determined. In this study, we show that HDIs influence the expression of a common polymorphic variant of the chemotherapy drug efflux transporter ABCG2, which contributes to normal tissue protection. As one of the most frequent variants in human ABCG2, the polymorphism Q141K impairs expression, localization, and function, thereby reducing drug clearance and increasing chemotherapy toxicity. Mechanistic investigations revealed that the ABCG2 Q141K variant was fully processed but retained in the aggresome, a perinuclear structure, where misfolded proteins aggregate. In screening for compounds that could correct its expression, localization, and function, we found that the microtubule-disrupting agent colchicine could induce relocalization of the variant from the aggresome to the cell surface. More strikingly, we found that HDIs could produce a similar effect but also restore protein expression to wild-type levels, yielding a restoration of ABCG2-mediated specific drug efflux activity. Notably, HDIs did not modify aggresome structures but instead rescued newly synthesized protein and prevented aggresome targeting, suggesting that HDIs disturbed trafficking along microtubules by eliciting changes in motor protein expression. Together, these results showed how HDIs are able to restore wild-type functions of the common Q141K polymorphic isoform of ABCG2. More broadly, our findings expand the potential uses of HDIs in the clinic.

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3 As one of the most frequent variants in human ABCG2, the polymorphism Q141K impairs expression, localization, and function, thereby reducing drug clearance and increasing chemotherapy toxicity. Login to comment
4 Mechanistic investigations revealed that the ABCG2 Q141K variant was fully processed but retained in the aggresome, a perinuclear structure, where misfolded proteins aggregate. Login to comment
8 Together, these results showed how HDIs are able to restore wild-type functions of the common Q141K polymorphic isoform of ABCG2. Login to comment
17 The most studied ABCG2 polymorphism is the nonsynonymous single-nucleotide polymorphism (SNP) C421A, which results in a glutamic acid to lysine substitution at amino acid 141 (Q141K), localized in the ATP-binding domain. Login to comment
19 ABCG2 harboring Q141K has impaired protein expression, incomplete trafficking to the plasma membrane, and decreased function. Login to comment
20 Clinically, the Q141K polymorphism has been linked to reduced clearance of some ABCG2 substrate drugs including diflomotecan, irinotecan, topotecan, gefitinib, rosuvastatin, atorvastatin, fluvastatin, simvastatin, and sulfasalazine (6). Login to comment
21 In addition to its impact on pharmacokinetics, the Q141K SNP has recently been linked to at least 10% of all gout cases, as the decreased ABCG2 function reduces urate elimination (7). Login to comment
22 Because of the clinical significance of the Q141K polymorphism in anticancer drug pharmacokinetics and potential involvement in carcinogenesis, we studied processing and trafficking of the intracellular trapped variant protein and ways of rescuing Q141K ABCG2 to restore it to its wild-type (WT) phenotype. Login to comment
35 WT and Q141K ABCG2 sublines were created by transfection of the pcDNA5/FRT/V5-His-TOPO vector (Invitrogen) expressing WT or Q141K ABCG2 into Flp-In-293 cells, an HEK293 subline bearing a single integrated Flp Recombination Target site (Invitrogen), according to the manufacturer's instructions. Login to comment
59 Results Determination of Q141K ABCG2 trafficking Flp-In-293 cells were selected for this study, as one copy of the vector is known to insert at a unique specific site, resulting in comparable RNA levels. Login to comment
60 To evaluate ABCG2 WT and Q141K variant localization in cells, immunofluorescent experiments were carried out. Login to comment
61 As previously reported (15-17), we saw that Q141K ABCG2 proteins had incomplete trafficking and cytoplasmic retention, whereas WT proteins were mainly localized HDI Impact on the Expression and Trafficking of Q141K ABCG2 www.aacrjournals.org Cancer Res; 72(14) July 15, 2012 3643 American Association for Cancer ResearchCopyright (c) 2012 on October 25, 2012cancerres.aacrjournals.orgDownloaded from Published OnlineFirst April , in the cell membrane (Fig. 1A). Login to comment
62 Also noted was staining of large aggregates of proteins, suggesting that the Q141K ABCG2 variant was accumulated in a localized cellular area. Login to comment
63 Although Q141K and WT ABCG2 mRNA levels were equal, variant protein levels were 4-fold lower, suggesting that a posttranslational mechanism regulates expression (Fig. 1B). Login to comment
64 Accordingly, the Q141K variant showed 5-fold decreased surface expression compared with WT. Login to comment
65 We hypothesized that Q141K variant underexpression was due to protein degradation. Login to comment
66 To explore whether Q141K ABCG2 was degraded by the ubiquitin-proteasome, the endosome-lysosome, or the autophagic pathways, we treated thecells with the proteasome inhibitor MG132; bafilomycin A1, which inhibits lysosomal degradation; and 3-methyladenine, which inhibits autophagosome formation. Login to comment
67 Bafilomycin and MG132 treatment increased total protein expression (between 150% and 180%) for WT and Q141K protein (Fig. 1C). Login to comment
68 These data suggest that in normal processing, a small portion of WT and Q141K ABCG2 are degraded via the proteasomal pathway, whereas the fully processed proteins thatreach thesurface aredegraded by the lysosome, as observed for other transmembrane proteins (18). Login to comment
69 Interestingly, the inhibition of the autophagic pathway induced a 3-fold increase in Q141K ABCG2 levels but barely affected WT protein expression. Login to comment
70 This indicates that autophagy is the main posttranslational mechanism, leading to a loss of Q141K variant expression. Login to comment
72 Determination of Q141K ABCG2 localization, processing, and degradation pathways. Login to comment
80 We also observed that a higher proportion of Q141K variant is ubiquitinated than in the WT protein (Supplementary Fig. S2). Login to comment
82 To examine the processing of Q141K ABCG2, whole-cell lysates from Flp-In-293 cells were deglycosylated with PNGase F and Endo H. ABCG2 contains a polysaccharide chain on asparagine 596 (19). Login to comment
84 No change in molecular mass was observed after Endo H digestion (Fig. 1D), suggesting that both WT and Q141K ABCG2 were completely processed and not retained in the endoplasmic reticulum. Login to comment
87 Results showed that Q141K was not retained in Golgi (Fig. 1E). Login to comment
88 We then asked whether Q141K ABCG2 proteins were retained in aggresomes. Login to comment
91 Thus, the major fraction of Q141K variant accumulated in the aggresomes. Login to comment
94 The Q141K ABCG2 variant reached its mature form but was mainly sequestered in the aggresome and degraded via autophagy. Login to comment
98 The effects of mitoxantrone were thus assayed in Q141K ABCG2 variant trafficking. Login to comment
100 Treatment with 5 mmol/L mitoxantrone for 24 hours caused a small increase in WT and Q141K total ABCG2 expression (around 125% for mRNA and 120% for protein compared with untreated cells). Login to comment
101 Surface expression showed a 160% induction in WT cells and a 250% induction in Q141K cells (Fig. 2A). Login to comment
103 Q141K variant showed a marked homogeneous staining pattern, with loss of aggresome localization, whereas Flp-In-293 WT cells showed no change (Fig. 2B). Login to comment
104 Finally, ABCG2 net efflux was determined by fluorescence-activated cell sorting (FACS) in Flp-In-293 WT and Q141K cells after mitoxantrone treatment (Fig. 2C). Login to comment
105 Pretreatment with mitoxantrone for 24 hours caused a weak increase of substrate net efflux (113% and 128% for WT and Q141K ABCG2, respectively). Login to comment
112 Total and surface protein expressions were similarly increased (210%-310% for WT ABCG2 and 260%-375% for Q141K ABCG2). Login to comment
116 Again, basal plasma membrane localization of WT far exceeded that in Flp-In-293 Q141K cells, where ABCG2 was localized in aggresomes (Fig. 3B). Login to comment
117 After exposure to HDIs, WT ABCG2 showed a minimal change in localization, whereas a dramatic change was seen for Q141K ABCG2. Login to comment
119 In Fig. 3C, we observed in detail that in romidepsin-treated Flp-In-293 Q141K cells, the aggresome-localized ABCG2 had almost disappeared in favor of a strong surface localization. Login to comment
120 ABCG2- specific efflux was then determined by FACS after HDI treatment in Flp-In-293 WT and Q141K cells. Login to comment
122 However, the change in transport ability following exposure to the HDIs in Flp-In-293 Q141K cells was notably higher: 200% for romidepsin, 163% for panobinostat, and 210% for vorinostat, compared with untreated Flp-In-293 Q141K cells (Fig. 3D). Login to comment
123 Increased Q141K ABCG2 function was confirmed by a cell death assay using the ABCG2 substrate, pheophorbide A (Fig. 3E). Login to comment
124 Indeed, Flp-In-293 Q141K cells pretreated for 24 hours with HDI Impact on the Expression and Trafficking of Q141K ABCG2 www.aacrjournals.org Cancer Res; 72(14) July 15, 2012 3645 American Association for Cancer ResearchCopyright (c) 2012 on October 25, 2012cancerres.aacrjournals.orgDownloaded from Published OnlineFirst April 3, romidepsin showed a decrease in celldeath after pheophorbide A treatment compared with cells not pretreated. Login to comment
126 Altogether, these results indicate that romidepsin, vorinostat, and panobinostat induced an increase in WT and Q141K ABCG2 expression that was associated with an improved trafficking to the cell surface in the Q141K variant. Login to comment
127 The increase also came with a gain of Q141K ABCG2 function, indicating that the surface variant was functional. Login to comment
128 Transcriptional effects of HDIs on Q141K ABCG2 rescue Our goal was to determine the mechanism by which HDI treatment leads to improvement of ABCG2 mutant trafficking. Login to comment
132 We then observed, in both Flp-In-293 WT and Q141K cells, that the mitoxantrone- and HDI-induced increases in surface expression were abolished by inhibition of protein synthesis (Fig. 4A), and the same results were obtained with the transcription inhibitor actinomycin D (Supplementary Fig. S5). Login to comment
135 This suggests that mitoxantrone only modified the trafficking of the neosynthesized ABCG2 and would not act on Q141K ABCG2 already trapped in the aggresome. Login to comment
139 We compared the data from mitoxantrone and HDI effect on Q141K ABCG2 localization and function. Login to comment
141 HDIs A Surface B C ProteinANRm WT Q141K %Comparedwithuntreatedcells %Comparedwithuntreatedcells WT Q141K Flp-ln-293 WT Flp-ln-293 Q141K WT Q141K WT Q141K * 0 50 100 150 200 250 300 350 0 50 100 150 200 250 300 350 0 50 100 150 200 250 300 350 Compared with WT Compared with WT Compared with WT Ctl MX Ctl MX ABCG2 relative efflux ABCG2 expression * * 0 50 100 150 200 250 300 350 Ctl MX Figure 2. Login to comment
144 Q141K mRNA, total protein, and surface protein expression was 93%, 28%, and 21%, respectively, of WT ABCG2. Login to comment
145 B, ABCG2 was shown by confocal microscopy in Flp-In-293 WT and Q141K cells after 24-hour treatment with mitoxantrone. Login to comment
147 Q141K relative efflux was 39.5% of WT ABCG2. Login to comment
148 Ctl, control. Basseville et al. Cancer Res; 72(14) July 15, 2012 Cancer Research3646 induced mainly Q141K ABCG2 trafficking to the membrane, accompanied by a greater increase in protein function. Login to comment
152 Nontranscriptional mechanisms of HDIs inducing Q141K ABCG2 rescue Considering ABCG2 acetylation as a possible mechanism of rescue, we immunoprecipitated ABCG2 and immunoblotted for acetylated lysine but saw no effect of HDIs on the transporter acetylation state (data not shown). Login to comment
153 We observed that Q141K ABCG2 was retained in aggresomes. Login to comment
160 Impact of HDIs on WT and Q141K ABCG2 protein expression, localization, and function. Login to comment
161 A, ABCG2 mRNA, total and surface protein were, respectively, quantified by qPCR, immunoblotting, and FACS analysis in Flp-In-293 WT and Q141K cells following 24-hour exposure to 46 nmol/L romidepsin (RD), 100 nmol/L panobinostat (PN), 10 mmol/L vorinostat (VR), or 1 mmol/L valproic acid (VA). Login to comment
163 C, ABCG2 and g-tubulin were observed in Q141K cells after 24-hour RD treatment (magnified example). Login to comment
166 E, cells were pretreated or not with RD dilutions for 24 hours, then treated for 24 hours with pheophorbide A (pheo A; 1 mmol/L for WT and 0.5 mmol/L for Q141K cells), and cell death was measured by flow cytometry. Login to comment
168 Ctl RD PN VR VA Flp-In-293 WT Flp-In-293 Q141K ABCG2/γ-tub/nucleus in RD-treated Q141K cells %Comparedwithuntreatedcells %Comparedwithuntreatedcells Protein SurfaceAN A Rm C B WT Q141K Ctl RD PN VR VA D ABCG2 relative efflux 0 50 100 150 200 250 300 350 400 450 500 0 50 100 150 200 250 300 350 400 450 500 0 50 100 150 200 250 300 350 400 450 500 WT Q141K WT Q141K WT Q141K E Pheo A-induced cell death %Celldeathcomparedwith pheophorbideAuntreatedcells ** compared with WT compared with WT compared with WT * ** * ** * ** * ** * ** * ** * ** * ** * ** * 0 100 200 300 400 500 600 700 WT Q141K Ctl RD PN VR VA not pretreat. 24-h RD pretreat.: 9 nmol/L 18 nmol/L 46 nmol/L 0 50 100 150 200 250 300 350 400 450 HDI Impact on the Expression and Trafficking of Q141K ABCG2 www.aacrjournals.org Cancer Res; 72(14) July 15, 2012 3647 American Association for Cancer ResearchCopyright (c) 2012 on October 25, 2012cancerres.aacrjournals.orgDownloaded from Published OnlineFirst April 3, We next examined the role of microtubules, known to be involved in retrograde transport, in ABCG2 localization. Login to comment
175 To evaluate HDAC6 involvement in Q141K ABCG2 rescue, we treated Flp-In-293 Q141K cells with the HDAC6 inhibitor tubastatin (28). Login to comment
184 To confirm these data, we conducted an siRNA assay to inhibit the expression of dynamitin, and we assessed the effect of HDIs on ABCG2 Q141K rescue after dynamitin knockdown. Login to comment
185 As shown in Fig. 5F, dynamitin knockdown blunted the effect of HDIs on Q141K ABCG2 rescue with a significant effect for romidepsin. Login to comment
186 Discussion We saw that the Q141K variant was retained in aggresomes and then degraded by the autophagic pathway. Login to comment
189 Aggresome formation facilitates the capture of aggregated proteins by the autophagic pathway (30-33), which we observed to be the case for Q141K ABCG2. Login to comment
190 A previous study had shown in the same Flp-In-293 cell model that Q141K ABCG2 was subjected to ubiquitin-mediated proteasomal and lysosomal degradation, with a 2-fold increase in expression after treatment with both inhibitors (17). Login to comment
193 It is interesting to note that the expressed WT and Q141K ABCG2 variants are found in a fully mature form, contrary to what has been observed with other ABC transporters. Login to comment
196 We sought to rescue Q141K ABCG2 trafficking using HDIs. Login to comment
198 Transcriptional effects of HDIs on ABCG2 Q141K rescue. Login to comment
199 A, quantification of ABCG2 surface expression by FACS in Flp-In-293 WT and Q141K cells after 24-hour exposure to mitoxantrone (MX) and HDIs in presence or absence of cycloheximide (CHX). Login to comment
202 Basseville et al. Cancer Res; 72(14) July 15, 2012 Cancer Research3648 expression with a conserved ratio for WT as for Q141K variant. Login to comment
203 More importantly, HDIs caused a strong relocalization of the Q141K variant from aggresome to cell surface and increased Q141K-mediated efflux. Login to comment
204 We sought to understand why these HDI-induced Q141K proteins, instead of accumulating in the aggresomes, were able to traffic to the cell surface. Login to comment
215 We observed that colchicine was also able to rescue Q141K ABCG2 trafficking. Login to comment
217 We deduced that HDIs similarly inhibit Q141K retrograde transport toward the aggresome, likely via dynamitin overexpression. Login to comment
220 Nontranscriptional effects of HDIs on ABCG2 Q141K rescue. Login to comment
221 A, vimentin and g-tubulin localization were observed by confocal microscopy in Flp-in-293 Q141K cells after a 24-hour treatment with 46 nmol/L romidepsin (RD). Login to comment
222 B, the effect of colchicine (colch; 1 mmol/L, 24 hours) on ABCG2 localization was determined by immunofluorescence in Flp-In-293 Q141K ABCG2 cells. Login to comment
223 C, ABCG2 total expression was quantified by immunoblot in Flp-In-293 WT and Q141K cells after 24-hour exposure to 1 mmol/L colchicine. Login to comment
224 D, quantification of ABCG2 surface expression by FACS in Flp-In-293 Q141K cells after 24-hour exposure to HDIs in presence or absence of 2.5 mmol/L tubastatin (tuba). Login to comment
226 F, the knockdown of dynamitin was observed after 48-hour transfection in Q141K cells by immunoblotting (left). Login to comment
227 The effect of dynamitin knockdown was then assessed on ABCG2 Q141K protein expression (surface and total) after a subsequent 24-hour HDI treatment. Login to comment
229 HDI Impact on the Expression and Trafficking of Q141K ABCG2 www.aacrjournals.org Cancer Res; 72(14) July 15, 2012 3649 American Association for Cancer ResearchCopyright (c) 2012 on October 25, 2012cancerres.aacrjournals.orgDownloaded from Published OnlineFirst April 3, an inflammatory stimulus, HDAC1 competed with the adaptor proteins for binding to motor proteins that travel along the microtubules, impairing axonal transport in neurons. Login to comment
231 Nawrocki and colleagues (40) also showed that inhibition of HDAC6 led to disruption of the aggresomes, but our experiments suggested that this deacetylase was not involved in the Q141K ABCG2 rescue. Login to comment
232 On the basis of our results, we propose a multipathway mechanism for HDI-induced Q141K ABCG2 rescue to surface (Fig. 6). Login to comment
233 HDIs induced Q141K ABCG2 surface localization by an increase in ABCG2 transcription coupled to dynamitin overexpression, which reduced aggresome targeting by inhibition of dynein/microtubule retrograde transport. Login to comment
253 Proposed mechanism of multiple target HDI-mediated Q141K ABCG2 rescue. Login to comment
254 HDIs act on Q141K ABCG2 trafficking by inducing ABCG2 overexpression (1) and likely a trafficking/folding partner overexpression (3), as well as by inhibition of retrograde transport from cytoplasm to aggresome (2). Login to comment
294 Major SNP (Q141K) variant of human ABC transporter ABCG2 undergoes lysosomal and proteasomal degradations. Login to comment
349 HDI Impact on the Expression and Trafficking of Q141K ABCG2 www.aacrjournals.org Cancer Res; 72(14) July 15, 2012 3651 American Association for Cancer ResearchCopyright (c) 2012 on October 25, 2012cancerres.aacrjournals.orgDownloaded from Published OnlineFirst April 3, Login to comment
2 As one of the most frequent variants in human ABCG2, the polymorphism Q141K impairs expression, localization, and function, thereby reducing drug clearance and increasing chemotherapy toxicity. Login to comment
7 Together, these results showed how HDIs are able to restore wild-type functions of the common Q141K polymorphic isoform of ABCG2. Login to comment
16 The most studied ABCG2 polymorphism is the nonsynonymous single-nucleotide polymorphism (SNP) C421A, which results in a glutamic acid to lysine substitution at amino acid 141 (Q141K), localized in the ATP-binding domain. Login to comment
18 ABCG2 harboring Q141K has impaired protein expression, incomplete trafficking to the plasma membrane, and decreased function. Login to comment
34 WT and Q141K ABCG2 sublines were created by transfection of the pcDNA5/FRT/V5-His-TOPO vector (Invitrogen) expressing WT or Q141K ABCG2 into Flp-In-293 cells, an HEK293 subline bearing a single integrated Flp Recombination Target site (Invitrogen), according to the manufacturer's instructions. Login to comment
57 Results Determination of Q141K ABCG2 trafficking Flp-In-293 cells were selected for this study, as one copy of the vector is known to insert at a unique specific site, resulting in comparable RNA levels. Login to comment
58 To evaluate ABCG2 WT and Q141K variant localization in cells, immunofluorescent experiments were carried out. Login to comment
79 We also observed that a higher proportion of Q141K variant is ubiquitinated than in the WT protein (Supplementary Fig. S2). Login to comment
81 To examine the processing of Q141K ABCG2, whole-cell lysates from Flp-In-293 cells were deglycosylated with PNGase F and Endo H. Login to comment
149 (c) 2012 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from Published OnlineFirst April 3, induced mainly Q141K ABCG2 trafficking to the membrane, accompanied by a greater increase in protein function. Login to comment
154 We observed that Q141K ABCG2 was retained in aggresomes. Login to comment
162 A, ABCG2 mRNA, total and surface protein were, respectively, quantified by qPCR, immunoblotting, and FACS analysis in Flp-In-293 WT and Q141K cells following 24-hour exposure to 46 nmol/L romidepsin (RD), 100 nmol/L panobinostat (PN), 10 mmol/L vorinostat (VR), or 1 mmol/L valproic acid (VA). Login to comment
164 C, ABCG2 and g-tubulin were observed in Q141K cells after 24-hour RD treatment (magnified example). Login to comment
167 E, cells were pretreated or not with RD dilutions for 24 hours, then treated for 24 hours with pheophorbide A (pheo A; 1 mmol/L for WT and 0.5 mmol/L for Q141K cells), and cell death was measured by flow cytometry. Login to comment
169 Ctl RD PN VR VA Flp-In-293 WT Flp-In-293 Q141K ABCG2/b3;-tub/nucleus in RD-treated Q141K cells % Compared with untreated cells % Compared with untreated cells Protein Surface A N A R m C B WT Q141K Ctl RD PN VR VA D ABCG2 relative efflux 0 50 100 150 200 250 300 350 400 450 500 0 50 100 150 200 250 300 350 400 450 500 0 50 100 150 200 250 300 350 400 450 500 WT Q141K WT Q141K WT Q141K E Pheo A-induced cell death % Cell death compared with pheophorbide A untreated cells * * compared with WT compared with WT compared with WT * ** * ** * ** * ** * ** * ** * ** * ** * ** * 0 100 200 300 400 500 600 700 WT Q141K Ctl RD PN VR VA not pretreat. 24-h RD pretreat.: 9 nmol/L 18 nmol/L 46 nmol/L 0 50 100 150 200 250 300 350 400 450 HDI Impact on the Expression and Trafficking of Q141K ABCG2 www.aacrjournals.org Cancer Res; 72(14) July 15, 2012 3647 We next examined the role of microtubules, known to be involved in retrograde transport, in ABCG2 localization. Login to comment
176 To evaluate HDAC6 involvement in Q141K ABCG2 rescue, we treated Flp-In-293 Q141K cells with the HDAC6 inhibitor tubastatin (28). Login to comment
187 Discussion We saw that the Q141K variant was retained in aggresomes and then degraded by the autophagic pathway. Login to comment
191 A previous study had shown in the same Flp-In-293 cell model that Q141K ABCG2 was subjected to ubiquitin-mediated proteasomal and lysosomal degradation, with a 2-fold increase in expression after treatment with both inhibitors (17). Login to comment
194 It is interesting to note that the expressed WT and Q141K ABCG2 variants are found in a fully mature form, contrary to what has been observed with other ABC transporters. Login to comment
197 We sought to rescue Q141K ABCG2 trafficking using HDIs. Login to comment
200 A, quantification of ABCG2 surface expression by FACS in Flp-In-293 WT and Q141K cells after 24-hour exposure to mitoxantrone (MX) and HDIs in presence or absence of cycloheximide (CHX). Login to comment
205 More importantly, HDIs caused a strong relocalization of the Q141K variant from aggresome to cell surface and increased Q141K-mediated efflux. Login to comment
206 We sought to understand why these HDI-induced Q141K proteins, instead of accumulating in the aggresomes, were able to traffic to the cell surface. Login to comment
219 We deduced that HDIs similarly inhibit Q141K retrograde transport toward the aggresome, likely via dynamitin overexpression. Login to comment
225 C, ABCG2 total expression was quantified by immunoblot in Flp-In-293 WT and Q141K cells after 24-hour exposure to 1 mmol/L colchicine. Login to comment
228 F, the knockdown of dynamitin was observed after 48-hour transfection in Q141K cells by immunoblotting (left). Login to comment
234 On the basis of our results, we propose a multipathway mechanism for HDI-induced Q141K ABCG2 rescue to surface (Fig. 6). Login to comment
235 HDIs induced Q141K ABCG2 surface localization by an increase in ABCG2 transcription coupled to dynamitin overexpression, which reduced aggresome targeting by inhibition of dynein/microtubule retrograde transport. Login to comment
307 Furukawa T, Wakabayashi K, Tamura A, Nakagawa H, Morishima Y, Osawa Y, et al. Major SNP (Q141K) variant of human ABC transporter ABCG2 undergoes lysosomal and proteasomal degradations. Login to comment
384 HDI Impact on the Expression and Trafficking of Q141K ABCG2 www.aacrjournals.org Cancer Res; 72(14) July 15, 2012 3651 2012;72:3642-3651. Login to comment

[hide] Significant interaction between LRP2 rs2544390 in ... Gene. 2012 Jul 15;503(1):131-6. Epub 2012 Apr 28. Hamajima N, Naito M, Okada R, Kawai S, Yin G, Morita E, Higashibata T, Tamura T, Nakagawa H, Matsuo H, Mori A, Wakai K
Significant interaction between LRP2 rs2544390 in intron 1 and alcohol drinking for serum uric acid levels among a Japanese population.
Gene. 2012 Jul 15;503(1):131-6. Epub 2012 Apr 28., [PMID:22565184]

Abstract [show]
A genome-wide association study identified that LRP2 rs2544390 in intron 1 was associated with serum uric acid (SUA) levels among Japanese, as well as polymorphisms of SLC22A12, ABCG2, and SLC2A9. This study aimed to confirm the association of rs2544390 C/T with SUA, as well as another LRP2 polymorphism (rs3755166 G/A) in the promoter. Subjects were 5016 health checkup examinees (3409 males and 1607 females) aged 35 to 69years with creatinine<2.0mg/dL. The subjects with SLC22A12 258WW, SLC2A9 rs11722228C allele, ABCG2 126QQ and 141Q allele (2546 males and 1199 females) were selected for analysis. Mean SUA was 6.03mg/dL for CC, 6.18mg/dL for CT, and 6.19mg/dL for TT among males (p=0.012), and 4.49mg/dL, 4.45mg/dL, and 4.42mg/dL among females (not significant), respectively. No association was observed for rs3755166. The association with rs2544390 was stronger among male drinkers. The odds ratio of drinking >/=5/week relative to no drinking for hyperuricemia (SUA>/=7mg/dL and/or under medication for hyperuricemia) was 1.11 (95% confidence interval, 0.67-1.84) among CC males, 1.75 (1.22-2.51) among CT males, and 3.13 (1.80-5.43) among TT males. The interaction terms with drinking >/=5/week were 1.56 (p=0.156) for CT and 2.87 (p=0.005) for TT. This was the first report on the interaction between LRP2 genotype and alcohol drinking for SUA. Since the low density lipoprotein-related protein 2 (megalin) encoded by LRP2 is a multi-ligand endocytic receptor expressed in many tissues including the kidney proximal tubules, the association/interaction remained to be confirmed both epidemiologically and biologically.

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31 This study aimed to confirm the association of SUA with LRP2 rs2544390 at intron 1 and rs3755166 at the promoter region in a Japanese population, after eliminating the effects of SLC22A12 W258X SLC2A9 rs11722228 at intron 8, ABCG2 Q126X, and ABCG2 Q141K on SUA (Hamajima et al., 2011a, 2011b; Matsuo et al., 2009; Tabara et al., 2010). Login to comment
110 The difference in the mean SUA between CC and TT was 0.08 mg/dL among males with creatinineb2 mg/dL (G1 in Table 3), which was smaller than those among SLC22A12 W258X, SLC2A9 rs11722228, ABCG2 Q126X, and Q141K. Login to comment

[hide] Pharmacokinetic interaction study of sulphasalazin... Br J Pharmacol. 2012 Jul;166(6):1793-803. doi: 10.1111/j.1476-5381.2012.01887.x. Kusuhara H, Furuie H, Inano A, Sunagawa A, Yamada S, Wu C, Fukizawa S, Morimoto N, Ieiri I, Morishita M, Sumita K, Mayahara H, Fujita T, Maeda K, Sugiyama Y
Pharmacokinetic interaction study of sulphasalazine in healthy subjects and the impact of curcumin as an in vivo inhibitor of BCRP.
Br J Pharmacol. 2012 Jul;166(6):1793-803. doi: 10.1111/j.1476-5381.2012.01887.x., [PMID:22300367]

Abstract [show]
BACKGROUND AND PURPOSE An ATP-binding cassette (ABC) transporter, breast cancer resistance protein (BCRP)/ABCG2, limits oral bioavailability of sulphasalazine. Here we examined the effect of curcumin, the principal curcuminoid of turmeric, on oral bioavailability of microdoses and therapeutic doses of sulphasalazine in humans. EXPERIMENTAL APPROACH Effects of curcumin were measured on the ATP-dependent sulphasalazine uptake by hBCRP-expressing membrane vesicles and on oral bioavailability of sulphasalazine in wild-type and Bcrp(-/-) mice. Eight healthy Japanese subjects received an oral dose of sulphasalazine suspension (100 microg) or tablets (2 g) alone or after curcumin tablets (2 g). Uptake of sulphasalazine was studied in HEK293 cells transfected with the influx transporter (OATP)2B1. KEY RESULTS Curcumin was a potent hBCRP inhibitor in vitro (K(i) 0.70 +/- 0.41 microM). Curcumin increased the area under the curve (AUC)(0-8) of plasma sulphasalazine eightfold in wild-type mice at 300 and 400 mg.kg(-1), but not in Bcrp(-/-) mice. Curcumin increased AUC(0-24) of plasma sulphasalazine 2.0-fold at microdoses and 3.2-fold at therapeutic doses in humans. Non-linearity of the dose-exposure relationship was observed between microdoses and therapeutic doses of sulphasalazine. Sulphasalazine was a substrate for OATP2B1 (K(m) 1.7 +/- 0.3 microM). Its linear index (dose/K(m)) at the therapeutic dose was high and may saturate OATP2B1. CONCLUSIONS AND IMPLICATIONS Curcumin can be used to investigate effects of BCRP on oral bioavailability of drugs in humans. Besides the limited dissolution, OATP2B1 saturation is a possible mechanism underlying non-linearity in the dose-exposure relationship of sulphasalazine.

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20 Pharmacogenetic studies, which focused on the non-synonymous single nucleotide polymorphism (SNP) of BCRP (421C>A, Q141K) associated with lower protein expression of BCRP (Kondo et al., 2004; Kobayashi et al., 2005), have disclosed the importance of BCRP in limiting oral bioavailability of drugs in human small intestine. Login to comment

[hide] Role of breast cancer resistance protein (BCRP/ABC... Biochem Pharmacol. 2012 Apr 15;83(8):1084-103. Epub 2012 Jan 11. Natarajan K, Xie Y, Baer MR, Ross DD
Role of breast cancer resistance protein (BCRP/ABCG2) in cancer drug resistance.
Biochem Pharmacol. 2012 Apr 15;83(8):1084-103. Epub 2012 Jan 11., [PMID:22248732]

Abstract [show]
Since cloning of the ATP-binding cassette (ABC) family member breast cancer resistance protein (BCRP/ABCG2) and its characterization as a multidrug resistance efflux transporter in 1998, BCRP has been the subject of more than two thousand scholarly articles. In normal tissues, BCRP functions as a defense mechanism against toxins and xenobiotics, with expression in the gut, bile canaliculi, placenta, blood-testis and blood-brain barriers facilitating excretion and limiting absorption of potentially toxic substrate molecules, including many cancer chemotherapeutic drugs. BCRP also plays a key role in heme and folate homeostasis, which may help normal cells survive under conditions of hypoxia. BCRP expression appears to be a characteristic of certain normal tissue stem cells termed "side population cells," which are identified on flow cytometric analysis by their ability to exclude Hoechst 33342, a BCRP substrate fluorescent dye. Hence, BCRP expression may contribute to the natural resistance and longevity of these normal stem cells. Malignant tissues can exploit the properties of BCRP to survive hypoxia and to evade exposure to chemotherapeutic drugs. Evidence is mounting that many cancers display subpopulations of stem cells that are responsible for tumor self-renewal. Such stem cells frequently manifest the "side population" phenotype characterized by expression of BCRP and other ABC transporters. Along with other factors, these transporters may contribute to the inherent resistance of these neoplasms and their failure to be cured.

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3424 Notably, in genome-wide screening of genetic determinants of gout, the Q141K BCRP single nucleotide polymorphism (SNP) was found to be associated with high uric acid levels, suggesting that uric acid is a substrate of BCRP [57,58]. Login to comment
3476 The most frequently observed non-synonymous SNPs in the BCRP coding region occur in exon 2 (G34A, resulting in a V to M mutation in amino acid 12), and in exon 5 (C421A, resulting in a Q141K mutation). Login to comment
3640 With regard to BCRP SNPs, among 145 Korean patients with DLBCL treated with the R-CHOP regimen, there was no influence of BCRP SNPs on clinical characteristics or treatment outcomes, but patients with the Q141K polymorphism (QK or KK), but not the V12M polymorphism discussed above for AML and CML, had more chemotherapy-related diarrhea [201]. Login to comment
3748 The study included SNPs of 7 ABC transporters: MDR1/ABCB1, MRP1-5, and the BCRP exon 5 (C421A/Q141K) SNP. Login to comment

[hide] Decreased extra-renal urate excretion is a common ... Nat Commun. 2012 Apr 3;3:764. doi: 10.1038/ncomms1756. Ichida K, Matsuo H, Takada T, Nakayama A, Murakami K, Shimizu T, Yamanashi Y, Kasuga H, Nakashima H, Nakamura T, Takada Y, Kawamura Y, Inoue H, Okada C, Utsumi Y, Ikebuchi Y, Ito K, Nakamura M, Shinohara Y, Hosoyamada M, Sakurai Y, Shinomiya N, Hosoya T, Suzuki H
Decreased extra-renal urate excretion is a common cause of hyperuricemia.
Nat Commun. 2012 Apr 3;3:764. doi: 10.1038/ncomms1756., [PMID:22473008]

Abstract [show]
ABCG2, also known as BCRP, is a high-capacity urate exporter, the dysfunction of which raises gout/hyperuricemia risk. Generally, hyperuricemia has been classified into urate 'overproduction type' and/or 'underexcretion type' based solely on renal urate excretion, without considering an extra-renal pathway. Here we show that decreased extra-renal urate excretion caused by ABCG2 dysfunction is a common mechanism of hyperuricemia. Clinical parameters, including urinary urate excretion, are examined in 644 male outpatients with hyperuricemia. Paradoxically, ABCG2 export dysfunction significantly increases urinary urate excretion and risk ratio of urate overproduction. Abcg2-knockout mice show increased serum uric acid levels and renal urate excretion, and decreased intestinal urate excretion. Together with high ABCG2 expression in extra-renal tissues, our data suggest that the 'overproduction type' in the current concept of hyperuricemia be renamed 'renal overload type', which consists of two subtypes-'extra-renal urate underexcretion' and genuine 'urate overproduction'-providing a new concept valuable for the treatment of hyperuricemia and gout.

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19 Woodward et al. and our group independently found that ABCG2 transports urate and shows the reduced urate transport by a half-functional variant, Q141K (rs2231142)20,21. Login to comment
21 We also showed that common dysfunctional genotype combinations of ABCG2 gene (Q126X (rs72552713) and Q141K) are a major cause of gout21. Login to comment
28 The risk allele frequency of Q126X (risk allele, X) and Q141K (risk allele, K), among 644 male outpatients with hyperuricemia including 575 gout cases, was 4.1 and 45.9%, respectively. Login to comment
29 Those who had Q126X and Q141K variants were 8.1 and 71.9%, respectively, of all patients (Supplementary Table S2). Login to comment
30 Subsequent haplotype frequency analysis revealed that the minor alleles of Q126X and Q141K were in different haplotypes (Supplementary Table S3), which indicated that these variants were independent risks, as reported previously21. Login to comment
31 Therefore, we could estimate urate export function of ABCG2 by the combination of two common variants, non-functional Q126X and half-functional Q141K (Supplementary Fig. S1). Login to comment
82 Estimated transport activity Genotype N Frequency of OP hyperuricemia RR 95% CI P Adjusted RR† Adjusted 95% CI† Adjusted P† Q126X (rs72552713) Q141K (rs2231142) OP hyperuricemia* Non-OP hyperuricemia* ≤1/4 Function X/X Q/Q 26 3 0.897 2.35 1.86-2.97 3.32×10 - 7 2.30 1.31-3.90 2.65×10 - 3 Q/X Q/K 1/2 Function Q/X Q/Q 96 55 0.636 1.66 1.32-2.10 8.58×10 - 6 1.79 1.25-2.59 1.55×10 - 3 Q/Q K/K 3/4 Function Q/Q Q/K 160 147 0.521 1.36 1.09-1.71 4.55×10 - 3 1.42 1.03-2.00 0.035 Full function Q/Q Q/Q 60 97 0.382 1.00 Abbreviations: CI, confidence interval; OP, overproduction; RR, risk ratio. Login to comment
143 The export function of ABCG2 was then estimated from the combinations of ABCG2 variants, rs72552713 (Q126X) and rs2231142 (Q141K), and divided into four functional groups21; that is, full function, 3/4 function (mild dysfunction), 1/2 function (moderate dysfunction) and ≤1/4 function (severe dysfunction). Login to comment
146 The wild-type human ABCG2 cDNA (GenBank accession number NM_004827) or mutated (Q126X and Q141K) ABCG2 cDNA was inserted into the Nhe I and Apa I sites of pcDNA3.1( + ) vector plasmid, with a myc-tag sequence attached at the 5'-end21. Login to comment

[hide] ABCG2 transporter: therapeutic and physiologic imp... J Vet Pharmacol Ther. 2012 Apr;35(2):105-12. doi: 10.1111/j.1365-2885.2011.01313.x. Epub 2011 Jun 6. Mealey KL
ABCG2 transporter: therapeutic and physiologic implications in veterinary species.
J Vet Pharmacol Ther. 2012 Apr;35(2):105-12. doi: 10.1111/j.1365-2885.2011.01313.x. Epub 2011 Jun 6., [PMID:21645015]

Abstract [show]
Drug transporters significantly influence drug pharmacokinetics and pharmacodynamics. While P-glycoprotein, the product of the MDR1 (ABCB1) gene, is the most well-characterized ABC transporter, the pharmacological importance of a related transporter, ABCG2, is starting to be realized in veterinary medicine. Based primarily on human and rodent studies, a number of clinically relevant, structurally and functionally unrelated drugs are substrates for ABCG2. ABCG2 is expressed by a variety of normal tissues including the intestines, renal tubular cells, brain and retinal capillary endothelial cells, biliary canalicular cells, and others, where it functions to actively extrude substrate drugs. In this capacity, ABCG2 limits oral absorption of substrate drugs and restricts their distribution to privileged sites such as the brain and retina. ABCG2 is also expressed by tumor cells where it functions to limit the intracellular accumulation of cytotoxic agents, contributing to multidrug resistance. Several ABCG2 polymorphisms have been described in human patients, some of which result in altered drug disposition, increasing susceptibility to adverse drug reactions. Additionally, ABCG2 polymorphisms in humans have been associated with disease states such as gout. Feline ABCG2 has recently been demonstrated to have several amino acid differences at conserved sites compared with 10 other mammalian species. These amino acid differences adversely affect transport function of feline ABCG2 relative to that of human ABCG2. Furthermore, these differences appear to be responsible for fluoroquinolone-induced retinal toxicity in cats and may play a role in acetaminophen toxicity as well. Studies in rodents and sheep have determined that ABCG2 expressed in mammary tissue is responsible for the secretion of many compounds (both therapeutic and toxic) into milk. Finally, data in rodent models suggest that ABCG2 may play an important role in regulating a number of physiologic pathways involved in protecting erythrocytes from oxidative damage.

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134 c.421 C>A results in an amino acid change (Q141K) that substantially decreases transport efficacy of human ABCG2 (Morisaki et al., 2005). Login to comment
135 Interestingly, Q141K occurs within the ATP-binding domain as does one of the feline amino acid changes (E159M). Login to comment

[hide] Genome-wide association of serum uric acid concent... Ann Hum Genet. 2012 Mar;76(2):121-7. doi: 10.1111/j.1469-1809.2011.00698.x. Epub 2012 Jan 9. Karns R, Zhang G, Sun G, Rao Indugula S, Cheng H, Havas-Augustin D, Novokmet N, Rudan D, Durakovic Z, Missoni S, Chakraborty R, Rudan P, Deka R
Genome-wide association of serum uric acid concentration: replication of sequence variants in an island population of the Adriatic coast of Croatia.
Ann Hum Genet. 2012 Mar;76(2):121-7. doi: 10.1111/j.1469-1809.2011.00698.x. Epub 2012 Jan 9., [PMID:22229870]

Abstract [show]
A genome-wide association study of serum uric acid (SUA) laevels was performed in a relatively isolated population of European descent from an island of the Adriatic coast of Croatia. The study sample included 532 unrelated and 768 related individuals from 235 pedigrees. Inflation due to relatedness was controlled by using genomic control. Genetic association was assessed with 2,241,249 single nucleotide polymorphisms (SNPs) in 1300 samples after adjusting for age and gender. Our study replicated four previously reported SUA loci (SLC2A9, ABCG2, RREB1, and SLC22A12). The strongest association was found with a SNP in SLC2A9 (rs13129697, P=2.33x10(-19)), which exhibited significant gender-specific effects, 35.76 mumol/L (P=2.11x10(-19)) in females and 19.58 mumol/L (P=5.40x10(-5)) in males. Within this region of high linkage disequilibrium, we also detected a strong association with a nonsynonymous SNP, rs16890979 (P=2.24x10(-17)), a putative causal variant for SUA variation. In addition, we identified several novel loci suggestive of association with uric acid levels (SEMA5A, TMEM18, SLC28A2, and ODZ2), although the P-values (P<5x10(-6)) did not reach the threshold of genome-wide significance. Together, these findings provide further confirmation of previously reported uric-acid-related genetic variants and highlight suggestive new loci for additional investigation.

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67 ABCG2 is a confirmed uric-acid-associated gene (Dehghan et al., 2008; Kolz et al., 2009; Yang et al., 2010) and the variant, showing the highest signal (P = 5.14×10-6 ) is a nonsynonymous SNP (rs2231142, NP_004818.2, Gln141Lys) and the mutant allele associated with an average increase of 27.40 μmol/L (Supplementary Fig. S3). Login to comment

[hide] Recent advances in renal urate transport: characte... Clin Exp Nephrol. 2012 Feb;16(1):89-95. Epub 2011 Nov 1. Anzai N, Jutabha P, Amonpatumrat-Takahashi S, Sakurai H
Recent advances in renal urate transport: characterization of candidate transporters indicated by genome-wide association studies.
Clin Exp Nephrol. 2012 Feb;16(1):89-95. Epub 2011 Nov 1., [PMID:22038265]

Abstract [show]
Humans have higher serum uric acid levels than other mammalian species owing to the genetic silencing of the hepatic enzyme uricase that metabolizes uric acid into allantoin. Urate (the ionized form of uric acid) is generated from purine metabolism and it may provide antioxidant defense in the human body. Despite its potential advantage, sustained hyperuricemia has pathogenetic causes in gout and renal diseases, and putative roles in hypertension and cardiovascular diseases. Since the kidney plays a dominant role in maintaining plasma urate levels through the excretion process, it is important to understand the molecular mechanism of renal urate handling. Although the molecular identification of a kidney-specific urate/anion exchanger URAT1 in 2002 paved the way for successive identification of several urate transport-related proteins, the entire picture of effective renal urate handling in humans has not yet been clarified. Recently, several genome-wide association studies identified a substantial association between uric acid concentration and single nucleotide polymorphisms in at least ten genetic loci including eight transporter-coding genes. In 2008, we functionally characterized the facilitatory glucose transporter family member SLC2A9 (GLUT9), one of the candidate genes for urate handling, as a voltage-driven urate transporter URATv1 at the basolateral side of renal proximal tubules that comprises the main route of the urate reabsorption pathway, in tandem with URAT1 at the apical side. In this review, recent findings concerning these candidate molecules are presented.

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54 Moreover, ABCG2 SNPs have been indicated to be an important factor for adverse drug reactions in patients, e.g., Q141K, the most extensively studied ABCG2 Apical Basolateral MCs UA URAT1 UA GLUT9 (URATv1) MCs UA URAT1 UA A B UA UA Apical Basolateral Proximal tubular cells Proximal tubular cells GLUT9 (URATv1) Fig. 2 Novel concept for idiopathic renal hypouricemia. Login to comment
60 Above-mentioned hypoactive Q141K protein is associated with hyperuricemia. Login to comment

[hide] Rosuvastatin blocks hERG current and prolongs card... J Pharm Sci. 2012 Feb;101(2):868-78. doi: 10.1002/jps.22809. Epub 2011 Nov 11. Plante I, Vigneault P, Drolet B, Turgeon J
Rosuvastatin blocks hERG current and prolongs cardiac repolarization.
J Pharm Sci. 2012 Feb;101(2):868-78. doi: 10.1002/jps.22809. Epub 2011 Nov 11., [PMID:22081364]

Abstract [show]
Blocking of the potassium current I(Kr) [human ether-a-go-go related gene (hERG)] is generally associated with an increased risk of long QT syndrome (LQTS). The 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase inhibitor, rosuvastatin, is a methanesulfonamide derivative, which shows structural similarities with several I(Kr) blockers. Hence, we assessed the effects of rosuvastatin on cardiac repolarization by using in vitro, ex vivo, and in vivo models. Patch clamp experiments on hERG-transfected human embryonic kidney (HEK) 293 cells established the potency of rosuvastatin to block hERG [half maximal inhibitory concentration (IC(50) ) = 195 nM]. We showed in isolated guinea pig hearts that 195 nM rosuvastatin prolonged (basic cycle length of 250 ms; p < 0.05) the monophasic action potential duration at 90% repolarization (MAPD(90) ) by 11 +/- 1 ms. Finally, rosuvastatin (10 mg/kg, intraperitoneal) prolonged corrected QT interval (QTc) in conscious and unrestrained guinea pigs from 201 +/- 1 to 210 +/- 2 ms (p < 0.05). Thus, rosuvastatin blocks I(Kr) and prolongs cardiac repolarization. In additional experiments, we also show that hERG blockade in HEK 293 cells was modulated by coexpression of efflux [breast cancer resistance protein (BCRP), multidrug resistance gene (MDR1)] and influx [organic anion transporting polypeptide (OATP) 2B1] transporters involved in the disposition and cardiac distribution of the drug. Genetic polymorphisms observed for BCRP, MDR1, and OATP2B1, and IC(50) determined for hERG blocking lead us to propose that some patients may be at risk of rosuvastatin-induced LQTS.

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24 Transfection Procedure In order to evaluate the effects of different transporters on the block of hERG by rosuvastatin, the hERG-stably transfected HEK 293 cells were transiently transfected with 10 :g of a recombinant pIRES2 vector expressing the green fluorescent protein (GFP) and one or the other of the following transporters: efflux transporter BCRP (strong affinity for rosuvastatin), loss of function variants of BCRP (BCRP-F208S, BCRP-S441N, and BCRP-Q141K), the influx transporters OATP2B1 (strong affinity for rosuvastatin), and multidrug resistance gene (MDR1) (an efflux transporter showing weak affinity for rosuvastatin). Login to comment
28 Site-Directed Mutagenesis The three loss-of-function variants of BCRP (BCRP-F208S, BCRP-S441N, and BCRP-Q141K) were Figure 1. Login to comment
88 Then, we performed site-directed mutagenesis on BCRP and produced three variants associated with a loss of function of the protein: C421A (Q141K), T623C (F208S), and G1322A (S441N).34-36 The coexpression of hERG with one or the other of these variants blunted the effects observed with the native efflux transporter BCRP, that is, the block was in the order of 40%. Login to comment
89 Indeed, inhibition of hERG was 40 ± 4 (n = 5), 41 ± 7 (n = 7), and 41 ± 4 (n = 7) for Q141K, F208S, and S441N, respectively (all not different from control, but different from HEK 293-hERG cells with functional transporters; p < 0.05). Login to comment
111 Inhibition of current activity was also determined in cells expressing transporters with a loss of function in BCRP activity due to mutations (BCRP-F208S, BCRP-S441N, and BCRP-Q141K). Login to comment
138 We also tested the effects of rosuvastatin on hERG in the presence of three loss-of-function polymorphisms of BCRP (Q141K, F208S, and S441N) and observed a block comparable to the one found in the control cells (≈42%). Login to comment
139 These polymorphisms are found in humans with relatively high frequencies (0.3%-25%), especially for Q141K (25% of Caucasians).47 Patients sharing a loss of function in BCRP (ABCG2 421AA; 1.2%) and OATP1B1 (SLCO1B1 521CC; 2%), the major efflux transporters involved in the disposition of rosuvastatin, or a gain of function polymorphism in OATP2B1 (SLCO2B1 935GG; 10%) should use rosuvastatin with caution due to increased plasma and cardiac levels. Login to comment

[hide] Null alleles of ABCG2 encoding the breast cancer r... Nat Genet. 2012 Jan 15;44(2):174-7. doi: 10.1038/ng.1070. Saison C, Helias V, Ballif BA, Peyrard T, Puy H, Miyazaki T, Perrot S, Vayssier-Taussat M, Waldner M, Le Pennec PY, Cartron JP, Arnaud L
Null alleles of ABCG2 encoding the breast cancer resistance protein define the new blood group system Junior.
Nat Genet. 2012 Jan 15;44(2):174-7. doi: 10.1038/ng.1070., [PMID:22246505]

Abstract [show]
The breast cancer resistance protein, also known as ABCG2, is one of the most highly studied ATP-binding cassette (ABC) transporters because of its ability to confer multidrug resistance. The lack of information on the physiological role of ABCG2 in humans severely limits cancer chemotherapeutic approaches targeting this transporter. We report here that ABCG2 comprises the molecular basis of a new blood group system (Junior, Jr) and that individuals of the Jr(a-) blood type have inherited two null alleles of ABCG2. We identified five frameshift and three nonsense mutations in ABCG2. We also show that the prevalence of the Jr(a-) blood type in the Japanese and European Gypsy populations is related to the p.Gln126* and p.Arg236* protein alterations, respectively. The identification of ABCG2(-/-) (Jr(a-)) individuals who appear phenotypically normal is an essential step toward targeting ABCG2 in cancer and also in understanding the physiological and pharmacological roles of this promiscuous transporter in humans.

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73 To pursue this goal, we first evaluated the plasma levels of urate in Jr(a-) individuals, as genome-wide association studies (GWAS) of large cohorts have recently identified the minor allele at rs2231142 in ABCG2 (encoding the defective Q141K variant18,21) as a risk factor for hyperuricemia and gout22,23 and revealed the capacity of ABCG2 to transport urate24. Login to comment
174 Imai, Y. et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Population differences in major functional polymor... Drug Metab Pharmacokinet. 2012;27(1):9-54. Epub 2011 Nov 29. Kurose K, Sugiyama E, Saito Y
Population differences in major functional polymorphisms of pharmacokinetics/pharmacodynamics-related genes in Eastern Asians and Europeans: implications in the clinical trials for novel drug development.
Drug Metab Pharmacokinet. 2012;27(1):9-54. Epub 2011 Nov 29., [PMID:22123129]

Abstract [show]
Drug lag, recently discussed extensively in Japan, can be divided into two phases: clinical development time and application review time. The former factor is still an important problem that might be improved by promoting multi-regional clinical trials and considering the results from other similar populations with Japanese, such as Koreans and Chinese. In this review, we compare the allelic or genotype frequencies of 30 relatively common functional alleles mainly between Eastern Asians and Europeans as well as among 3 major populations in Eastern Asian countries, Japan, Korea, and China, in 12 pharmacokinetics (PK)/pharmacodynamics (PD)-related genes; CYP2C9 (*2 and *3), CYP2C19 (*2, *3 and *17), 13 CYP2D6 haplotypes including *4, *5 and *10, CYP3A5 (*3), UGT1A1 (*28 and *6), NAT2 (*5, *6 and *7), GSTM1 and GSTT1 null genotypes, SLCO1B1 521T>C, ABCG2 421C>A, and HLA-A*31:01 and HLA-B*58:01. In this review, differences in allele frequencies (AFs) or genotype frequencies (GFs) less than 0.1 (in the cases of highest AF (GF) >/=0.1) or less than 0.05 (in the cases of lowest AF (GF) <0.1) were regarded as similar. Between Eastern Asians and Europeans, AFs (or GFs) are regarded as being different for many alleles such as CYP2C9 (*2), CYP2C19 (*2, *3 and *17), CYP2D6 (*4 and *10), CYP3A5 (*3), UGT1A1 (*28 and *6), NAT2 (*5*7), GSTT1 null and ABCG2 421C>A. Among the 3 Eastern Asian populations, however, only AFs of CYP2C19*3, CYP2D6*10, HLA-A*31:01 and HLA-B*58:01 are regarded as dissimilar. For CYP2C19*3, the total functional impact on CYP2C19 could be small if the frequencies of the two null alleles CYP2C19*2 and *3 are combined. Regarding CYP2D6*10, frequency difference over 0.1 is observed only between Japanese and Chinese (0.147). Although environmental factors should be considered for PK/PD differences, we could propose that among Japan, Korea, and China, genetic differences are very small for the analyzed common PK-related gene polymorphisms. On the other hand, AFs of the two HLA alleles important for cutaneous adverse drug reactions are diverse even among Eastern Asians and thus should be taken into account.

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170 Frequencies of ABCG2 421ChA in different ethnic populations Country ¤Population¥ Allele frequencya Number of subjectsb Reference421ChA, Q141K Transporter activity Decreased Asia Eastern 0.301 1,602 Japan 0.313 411 0.350 10 322¥ 0.355 100 323¥ 0.266 124 324¥ 0.319 177 325¥ Korea 0.284 670 0.278 250 326¥ 0.280 275 327¥ 0.300 145 328¥ China 0.315 521 0.291 191 327¥ ¤Han¥ 0.342 95 329¥ ¤Han¥ 0.323 235 330¥ South-Eastern 0.295 237 Vietnam 0.311 140 327¥ Singapore ¤Chinese¥c 0.277 94 191¥ ¤Malay¥ 0.273 97 191¥ ¤Indian¥c 0.154 94 191¥ Europe 0.103 1,611 Northern Sweden 0.100 60 331¥ Western 0.107 1,314 Netherlandsd 0.120 100 332¥ Germany 0.100 349 333¥ 0.108 865 334¥ Southern Italy ¤Tuscany¥ 0.062 88 HapMape Eastern Hungary 0.094 149 335¥ Caucasians 0.105 479 Continued on next page: Continued: Country ¤Population¥ Allele frequencya Number of subjectsb Reference421ChA, Q141K Transporter activity Decreased European-Caucasians 0.107 84 329¥ Caucasians 0.087 150 336¥ 0.142 88 322¥ European-Americans 0.080 69 337¥ Caucasian-Americans 0.119 88 329¥ Africans 0.027 1,094 Africans ¤sub-Saharan¥ 0.009 938 329¥ African-Americans 0.053 94 329¥ 0.000 24 322¥ 0.039 38 337¥ The base A in the initiation codon ATG is denoted ¦1. a The total allele frequency in each population/region is calculated by dividing the sum of the each allele number by the total allele number in the population/region. b The subtotal number in each population/region is simply the sum of the subject numbers. c Excluded from total data in the South-Eastern Asia region. d The population included 1 African, 1 Asian and 5 mixed ethnic subjects ¤i.e., Caucasians were 93%¥. Login to comment

[hide] Functional significance of genetic polymorphisms i... Drug Metab Pharmacokinet. 2012;27(1):85-105. Epub 2011 Nov 29. Ieiri I
Functional significance of genetic polymorphisms in P-glycoprotein (MDR1, ABCB1) and breast cancer resistance protein (BCRP, ABCG2).
Drug Metab Pharmacokinet. 2012;27(1):85-105. Epub 2011 Nov 29., [PMID:22123128]

Abstract [show]
Recent pharmacogenomic/pharmacogenetic (PGx) studies have disclosed important roles for drug transporters in the human body. Changes in the functions of drug transporters due to drug/food interactions or genetic polymorphisms, for example, are associated with large changes in pharmacokinetic (PK) profiles of substrate drugs, leading to changes in drug response and side effects. This information is extremely useful not only for drug development but also for individualized treatment. Among drug transporters, the ATP-binding cassette (ABC) transporters are expressed in most tissues in humans, and play protective roles; reducing drug absorption from the gastrointestinal tract, enhancing drug elimination into bile and urine, and impeding the entry of drugs into the central nervous system and placenta. In addition to PK/pharmacodynamic (PD) issues, ABC transporters are reported as etiologic and prognostic factors (or biomarkers) for genetic disorders. Although a consensus has not yet been reached, clinical studies have demonstrated that the PGx of ABC transporters influences the overall outcome of pharmacotherapy and contributes to the pathogenesis and progression of certain disorders. This review explains the impact of PGx in ABC transporters in terms of PK/PD, focusing on P-glycoprotein and breast cancer resistance protein (BCRP).

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71 Impact of ABCG2 (BCRP) polymorphisms on PK/PD/disorders Gene marker ¤SNPs/haplotype¥ Functional effect of the study Drug Population Disease Ref.Pharmacokinetics Therapeutic efficacy Side effects SNPs assay Others ¤e.g., frequency and susceptibility¥ %15622CgT or ¤1143ChT, %15622ChT¥ haplotype * gefitinib patient NSCLC 68 421ChA * * A771726 HV 223 421ChA ¤141QhK, rs2231142¥ * patient gout 224 421ChA * imatinib patient CML 225 421ChA * patient uric acid 226 421ChA ¤141QhK, rs2231142¥ * patient gout 227 421ChA, 914ChA * methotrexate patient RA 228 421ChA * * sunitinib patient RCC 70 421ChA * rosuvastatin patient myocardial infarction 229 346GhA, 421ChA, 1143ChT, 15994GhA * danusertib patient 230 421ChA * rosuvastatin patient hypercholesterolemia 231 421ChA * patient gout 232 376ChT, 421ChA * gefitinib patient NSCLC 67 421ChA * telatinib patient 233 421ChA * 3 statines HV 234 rs2622604 * irinitecan patient myelosuppression 235 ¤%15622C/T, 1143C/T¥ haplotype * sunitinib patient 127 12VhM, 141QhK * mitoxantrone patient multiple sclerosis 236 421ChA * imatinib patient CML 237 421ChA * sulfasalazine HV 238 421ChA * erlotinib patient SCC 69 421ChA * patient gout 239 421ChA * atorvastatin, rosuvastatin HV 240 12VhM, 141QhK * R-CHOP patient DLBCL 241 12VhM * patient ischemic stroke 242 421ChA * imatinib patient solid malignancies 243 421ChA * patient gout 73 rs2622621, rs1481012 * patient colorectal cancer 244 421ChA * nitrofurantoin HV 245 421ChA * sulfasalazine HV 246 421ChA * doxorubicin patient breast cancer 168 421ChA * sulfasalazine HV 60 Q141K, V12M, Q126X * lamivudine HV 247 34ChA, 421ChA * patient DLBCL 248 421ChA * pitavastatin HV 64 Continued on next page: 141QhK¥ which has been associated with lower BCRP protein expression.52,55¥ Recently, 421ChA was found to greatly affect the stability of BCRP in the endoplasmic reticulum, leading to increased protein degradation via ubiquitination and proteasomal proteolysis.56,57¥ Therefore, 421ChA may lead to increased bioavailability after the oral administration of substrate drugs. Login to comment

[hide] Human ABCG2: structure, function, and its role in ... Int J Biochem Mol Biol. 2012;3(1):1-27. Epub 2011 Mar 30. Mo W, Zhang JT
Human ABCG2: structure, function, and its role in multidrug resistance.
Int J Biochem Mol Biol. 2012;3(1):1-27. Epub 2011 Mar 30., [PMID:22509477]

Abstract [show]
Human ABCG2 is a member of the ATP-binding cassette (ABC) transporter superfamily and is known to contribute to multidrug resistance (MDR) in cancer chemotherapy. Among ABC transporters that are known to cause MDR, ABCG2 is particularly interesting for its potential role in protecting cancer stem cells and its complex oligomeric structure. Recent studies have also revealed that the biogenesis of ABCG2 could be modulated by small molecule compounds. These modulators, upon binding to ABCG2, accelerate the endocytosis and trafficking to lysosome for degradation and effectively reduce the half-life of ABCG2. Hence, targeting ABCG2 stability could be a new venue for therapeutic discovery to sensitize drug resistant human cancers. In this report, we review recent progress on understanding the structure, function, biogenesis, as well as physiological and pathophysiological functions of ABCG2.

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889 The effect of ABCG2 V12M, Q141K and Q126X, known functional variants in vitro, on the disposition of lamivudine. Login to comment
895 The effect of ABCG2 V12M, Q141K and Q126X, known functional variants in vitro, on the disposition of lamivudine. Login to comment

[hide] ABCG2 is a direct transcriptional target of hedgeh... Oncogene. 2011 Dec 8;30(49):4874-86. doi: 10.1038/onc.2011.195. Epub 2011 May 30. Singh RR, Kunkalla K, Qu C, Schlette E, Neelapu SS, Samaniego F, Vega F
ABCG2 is a direct transcriptional target of hedgehog signaling and involved in stroma-induced drug tolerance in diffuse large B-cell lymphoma.
Oncogene. 2011 Dec 8;30(49):4874-86. doi: 10.1038/onc.2011.195. Epub 2011 May 30., [PMID:21625222]

Abstract [show]
Successful treatment of diffuse large B-cell lymphoma (DLBCL) is frequently hindered by the development of resistance to conventional chemotherapy resulting in disease relapse and high mortality. High expression of antiapoptotic and/or drug transporter proteins induced by oncogenic signaling pathways has been implicated in the development of chemoresistance in cancer. Previously, our studies showed that high expression of adenosine triphosphate-binding cassette drug transporter ABCG2 in DLBCL correlated inversely with disease- and failure-free survival. In this study, we have implicated activated hedgehog (Hh) signaling pathway as a key factor behind high ABCG2 expression in DLBCL through direct upregulation of ABCG2 gene transcription. We have identified a single binding site for GLI transcription factors in the ABCG2 promoter and established its functionality using luciferase reporter, site-directed mutagenesis and chromatin-immunoprecipitation assays. Furthermore, in DLBCL tumor samples, significantly high ABCG2 and GLI1 levels were found in DLBCL tumors with lymph node involvement in comparison with DLBCL tumor cells collected from pleural and/or peritoneal effusions. This suggests a role for the stromal microenvironment in maintaining high levels of ABCG2 and GLI1. Accordingly, in vitro co-culture of DLBCL cells with HS-5 stromal cells increased ABCG2 mRNA and protein levels by paracrine activation of Hh signaling. In addition to ABCG2, co-culture of DLBCL cells with HS-5 cells also resulted in increase expression of the antiapoptotic proteins BCL2, BCL-xL and BCL2A1 and in induced chemotolerance to doxorubicin and methotrexate, drugs routinely used for the treatment of DLBCL. Similarly, activation of Hh signaling in DLBCL cell lines with recombinant Shh N-terminal peptide resulted in increased expression of BCL2 and ABCG2 associated with increased chemotolerance. Finally, functional inhibition of ABCG2 drug efflux activity with fumitremorgin C or inhibition of Hh signaling with cyclopamine-KAAD abrogated the stroma-induced chemotolerance suggesting that targeting ABCG2 and Hh signaling may have therapeutic value in overcoming chemoresistance in DLBCL.

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25 One study identified two polymorphisms in ABCG2 gene, G34A (Val12Met) and C421A (Gln141Lys), and associated these polymorphisms with increased risk and poor overall survival of DLBCL (Hu et al., 2007). Login to comment

[hide] ABCG2/BCRP dysfunction as a major cause of gout. Nucleosides Nucleotides Nucleic Acids. 2011 Dec;30(12):1117-28. Matsuo H, Takada T, Ichida K, Nakamura T, Nakayama A, Suzuki H, Hosoya T, Shinomiya N
ABCG2/BCRP dysfunction as a major cause of gout.
Nucleosides Nucleotides Nucleic Acids. 2011 Dec;30(12):1117-28., [PMID:22132966]

Abstract [show]
Recent genome-wide association studies showed that serum uric acid (SUA) levels relate to ABCG2/BCRP gene, which locates in a gout-susceptibility locus revealed by a genome-wide linkage study. Together with the ABCG2 characteristics, we hypothesized that ABCG2 transports urate and its dysfunction causes hyperuricemia and gout. Transport assays showed ATP-dependent transport of urate via ABCG2. Kinetic analysis revealed that ABCG2 mediates high-capacity transport of urate (Km: 8.24 +/- 1.44 mM) even under high-urate conditions. Mutation analysis of ABCG2 in 90 Japanese hyperuricemia patients detected six nonsynonymous mutations, including five dysfunctional variants. Two relatively frequent dysfunctional variants, Q126X and Q141K, were then examined. Quantitative trait locus analysis of 739 Japanese individuals showed that Q141K increased SUA as the number of minor alleles of Q141K increased (p = 6.60 x 10(-5)). Haplotype frequency analysis revealed that there is no simultaneous presence of Q126X and Q141K in one haplotype. Becuase Q126X and Q141K are assigned to nonfunctional and half-functional haplotypes, respectively, their genotype combinations are divided into four functional groups. The association study with 161 male gout patients and 865 male controls showed that all of those with dysfunctional ABCG2 increased the gout risk, especially those with </=1/4 function (OR, 25.8; 95% CI, 10.3-64.6; p = 3.39 x 10(-21)). These genotypes were found in 10.1% of gout patients, but in only 0.9% of control. Our function-based clinicogenetic (FBCG) analysis showed that combinations of the two dysfunctional variants are major causes of gout, thereby providing a new approach for prevention and treatment of the gout high-risk population.

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11 Two relatively frequent dysfunctional variants, Q126X and Q141K, were Received 29 May 2011; accepted 17 October 2011. Login to comment
18 Quantitative trait locus analysis of 739 Japanese individuals showed that Q141K increased SUA as the number of minor alleles of Q141K increased (p = 6.60 × 10-5 ). Login to comment
19 Haplotype frequency analysis revealed that there is no simultaneous presence of Q126X and Q141K in one haplotype. Login to comment
20 Becuase Q126X and Q141K are assigned to nonfunctional and half-functional haplotypes, respectively, their genotype combinations are divided into four functional groups. Login to comment
38 1119 ABCG2 : ATP binding casse e G2 SNP : single nucleo de polymorphism QTL : quan ta ve trait locus OR : odds ra o ABCG2 as a urate secre on transporter in humans Gene c analysis Func onal analysis ABCG2 muta on analysis of 90 hyperuricemic cases (all coding regions) ABCG2 muta ons (with amino acid altera ons) 6 muta ons c d Func onal analysis of urate transport via wild type ABCG2 (vesicle studies) a Iden fica on of urate transport ac vi es via ABCG2 b Func onal analysis of urate transport via mutated ABCG2 6 mutants e No effect (V12M) g Dysfunc onal genotype combina ons of ABCG2 as major causes of gout q Dysfunc onal SNP with high frequency (>30%) (Q141K) QTL analysis in 739 Japanese individuals h i j n Gout / hyperuricemia with ABCG2 homozygous, n = 2 heterozygous, n = 24 Loss of func on (Q126X, G268R, S441N, F506Sfs) Reduced func on (~50%) (Q141K) f p Genotype combina on analysis 10.1% of gout with ≤1/4 ABCG2 func on OR = 25.8, p = 3.39×10-21 o Haplotype analysis 13.5% of gout with disease haplotype OR = 5.97, p = 4.10×10-12 Associa on analysis of hyperuricemia (Q126X) OR = 3.61, p = 2.91× 10-7 l m Associa on analysis of gout (Q126X) OR = 4.25, p =3.04 × 10-8 Genotyping of nonfunc onal SNP (Q126X) hyperuricemia, n=228 k FIGURE 1 Flowchart for molecular-function-based clinicogenetic (FBCG) analysis of gout patients with ABCG2 polymorphic variants. Login to comment
45 Quantitative trait locus (QTL) analysis of SUA concentrations was performed with genotyping of Q141K in 739 Japanese individuals. Login to comment
53 Using the site-directed mutagenesis technique, we constructed ABCG2 mutants (V12M, Q126X, Q141K, G268R, S441N, and F506SfsX4), which were used for urate transport analysis, on the expression vector for ABCG2. Login to comment
65 The following six nonsynonymous mutations, including three SNPs, were found: V12M, Q126X, Q141K, G268R, S441N, and F506SfsX4 (Figure 2A). Login to comment
70 Results are expressed as means ± S.D. (C) Quantitative trait locus (QTL) analysis of ABCG2 Q141K and serum uric acid levels. Login to comment
71 "C/C," "C/A," and "A/A" indicate wild-type subjects, heterozygous mutation carriers, and homozygous mutation carriers of Q141K, respectively. Login to comment
76 Maekawa et al.[17] reported that V12M, Q126X, and Q141K are quite common in the Japanese population, and allele frequencies for these SNPs were 31.9% for Q141K, 19.2% for V12M, and 2.8% for Q126X, respectively. Login to comment
77 Using Hardy-Weinberg equilibrium and these data on a Japanese population reported by Maekawa et al.,[17] we estimated that the frequencies of Japanese individuals with these minor alleles were 53.6% for Q141K, 34.7% for V12M, and 5.5% for Q126X. Login to comment
79 Among six mutants, ATP-dependent urate transport was reduced by approximately half (46.7%) in one mutant, Q141K, and was nearly eliminated in four mutants, Q126X, G268R, S441N, and F506SfsX4 (Figure 2B). Login to comment
81 ABCG2 Dysfunction Increases SUA In a random sample of 739 Japanese individuals, QTL analysis of SUA with the high-frequency dysfunctional variant Q141K revealed that SUA significantly increased as the number of minor alleles of Q141K increased (p = 6.60 × 10-5 ), and the corrected p value adjusted for sex is 2.02 × 10-6 (Figure 2C). Login to comment
83 Different from SUA, Q141K had no significant association with other clinical characteristics such as age, body mass index, or sex. Login to comment
90 The half-functional SNP Q141K also increased gout risk (OR, 2.23; 95% CI, 1.75-2.87; p = 5.54 × 10-11 ). Login to comment
91 Haplotype frequency analysis revealed no simultaneous presence of the minor alleles of Q126X and Q141K in one haplotype. Login to comment
93 Q141K is assigned to another independent risk haplotype. Login to comment
98 Because Q126X and Q141K are assigned to different risk haplotypes, nonfunctional and half-functional, respectively, their genotype combinations are divided into four functional groups on the basis of the estimated ABCG2 transport functions, i.e., full function, 3/4 function, 1/2 function, and ≤1/4 function. Login to comment
110 Together with P-glycoprotein 21.4% 45.3% 23.3% 10.1% 50.8% 35.6% 12.7% 0.9% other gout risks gout no function Q126X T/T Q141K C/C Full function C/C C/C 3/4 function 1/2 function C/C C/C T/C A/C A/A C/C Gout patients Normal control SUA ≤ 7.0 mg/dl 1/4 function T/C A/C 25.8-fold gout risk ≥3.02-fold gout risk (n = 865) (n = 161) FIGURE 3 ABCG2 transport dysfunction as a major gout risk. Login to comment
111 Genotype combinations of Q126X and Q141K are divided into several groups based on estimated ABCG2 transport functions. Login to comment
134 They also found that Q141K was associated with self-reported gout in Caucasians (OR, 1.74; 95% CI, 1.51-1.99), which is consistent with our finding from clinically diagnosed Japanese gout patients. Login to comment
135 A meta-analysis of GWAS of European descent also showed that the nine loci, including GLUT9 and ABCG2, influence SUA,[11] which confirms the findings of Dehghan et al.[10] In this study, we identified several nonfunctional ABCG2 mutations, including Q126X, which shows stronger effects on gout development than Q141K did in a previous study (OR < 2.0). Login to comment
138 Woodward et al.[19] and we[2] independently found a urate transport ability via ABCG2 to transport urate and characterized the effects of a half-functional variant Q141K using different methods. Login to comment

[hide] Identification of ABCG2 dysfunction as a major fac... Nucleosides Nucleotides Nucleic Acids. 2011 Dec;30(12):1098-104. Matsuo H, Takada T, Ichida K, Nakamura T, Nakayama A, Takada Y, Okada C, Sakurai Y, Hosoya T, Kanai Y, Suzuki H, Shinomiya N
Identification of ABCG2 dysfunction as a major factor contributing to gout.
Nucleosides Nucleotides Nucleic Acids. 2011 Dec;30(12):1098-104., [PMID:22132963]

Abstract [show]
The ATP-binding cassette, subfamily G, member 2 gene ABCG2/BCRP locates in a gout-susceptibility locus (MIM 138900) on chromosome 4q. Recent genome-wide association studies also showed that the ABCG2 gene relates to serum uric acid levels and gout. Since ABCG2 is also known as a transporter of nucleotide analogs that are structurally similar to urate, and is an exporter that has common polymorphic reduced functionality variants, ABCG2 could be a urate secretion transporter and a gene causing gout. To find candidate mutations in ABCG2, we performed a mutation analysis of the ABCG2 gene in 90 Japanese patients with hyperuricemia and found six non-synonymous mutations. Among the variants, ATP-dependent urate transport was reduced or eliminated in five variants, and two out of the five variants (Q126X and Q141K) were frequently detected in patients. Haplotype frequency analysis revealed that there is no simultaneous presence of Q126X and Q141K in one haplotype. As Q126X and Q141K are a nonfunctional and half-functional haplotype, respectively, their genotype combinations are divided into four estimated functional groups. The association study with 161 male gout patients and 865 male controls showed that all of those who had dysfunctional ABCG2 had an increased risk of gout, and that a remarkable risk was observed in those with </=1/4 function (OR, 25.8; 95% CI, 10.3-64.6; p = 3.39 x 10(-21)). In 2,150 Japanese individuals, the frequency of those with dysfunctional ABCG2 was more than 50%. Our function-based clinicogenetic analysis identified the combinations of dysfunctional variants of ABCG2 as a major contributing factor in Japanese patients with gout.

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16 Among the variants, ATP-dependent urate transport was reduced or eliminated in five variants, and two out of the five variants (Q126X and Q141K) were frequently detected in patients. Login to comment
17 Haplotype frequency analysis revealed that there is no simultaneous presence of Q126X and Q141K in one haplotype. Login to comment
18 As Q126X and Q141K are a nonfunctional and half-functional haplotype, respectively, their genotype combinations are divided into four estimated functional groups. Login to comment
36 Using the site-directed mutagenesis technique, we constructed mutants of ABCG2 (V12M, Q126X, Q141K, G268R, S441N, and F506SfsX4), which were used for urate transport analysis, on the expression vector for ABCG2. Login to comment
45 The following six non-synonymous mutations, V12M, Q126X, Q141K, G268R, S441N, and F506SfsX4, were found (Figure 1A), and the first three mutations were SNPs. Login to comment
46 Maekawa et al.[5] reported that allele frequencies for these SNPs, which are quite common in the Japanese population, were 31.9% for Q141K, 19.2% for FIGURE 1 Non-synonymous ABCG2 mutations in hyperuricemia patients. Login to comment
50 With the Hardy-Weinberg equilibrium and the data reported by Maekawa et al. of the Japanese population,[5] we calculated estimates of the minor allele frequencies of Japanese individuals to be 53.6% for Q141K, 34.7% for V12M, and 5.5% for Q126X. Login to comment
52 The ATP-dependent transport of urate was reduced by approximately half (46.7%) in Q141K and was nearly eliminated in Q126X, G268R, S441N, and F506SfsX4 mutants (Figure 1B). Login to comment
56 A total of 871 TABLE 1 Association analysis of ABCG2 genotype combination in gout patients Genotype Number Estimated transport Q126X Q141K Gout Control p value OR* 95% CI* ≤1/4 function T/T C/C 16 8 3.39 × 10-21 25.8 10.3-64.6 T/C A/C - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 1/2 function T/C C/C 37 110 2.23 × 10-9 4.34 2.61-7.24 C/C A/A - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 3/4 function C/C A/C 72 308 2.29 × 10-7 3.02 1.96-4.65 - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - Full function C/C C/C 34 439 1.00 *OR = odds ratio; 95% CI = 95% confidence interval. Login to comment
57 OR is obtained by comparing the non-risk genotype combination C/C (Q126X) and C/C (Q141K). Login to comment
58 Risk alleles for Q126X and Q141K are underlined. Login to comment
62 The half-functional SNP Q141K also increased gout (OR, 2.23; 95% CI, 1.75-2.87; p = 5.54 × 10-11 ). Login to comment
63 The haplotype frequency analysis revealed no simultaneous presence of the minor alleles of Q126X and Q141K in one haplotype. Login to comment
70 Because the nonfunctional variant Q126X and half-functional variant Q141K are assigned to different risk haplotypes, their genotype combinations are divided into four functional groups on the basis of the estimated ABCG2 transport functions, i.e., full function, 3/4 function, 1/2 function, and ≤1/4 function (Table 1). Login to comment
91 They also found that Q141K was associated with self-reported gout in Caucasians (OR, 1.74; 95% CI, 1.51-1.99), which is consistent with our findings from clinically diagnosed Japanese gout patients. Login to comment
92 A meta-analysis of GWAS in European descent also revealed that the nine loci, including GLUT9 and ABCG2, influence SUA, which confirms the findings of Dehghan et al.[3] The authors[6] and Woodward et al.[7] independently found a urate transport ability via ABCG2 to transport urate and characterized the effects of dysfunctional variant Q141K using different methods. Login to comment
93 In our study, we also identified several nonfunctional ABCG2 mutations including Q126X which shows stronger effects on gout development than Q141K did in a previous study (OR < 2.0). Login to comment

[hide] ABCG2 is a high-capacity urate transporter and its... Nucleosides Nucleotides Nucleic Acids. 2011 Dec;30(12):1091-7. Nakayama A, Matsuo H, Takada T, Ichida K, Nakamura T, Ikebuchi Y, Ito K, Hosoya T, Kanai Y, Suzuki H, Shinomiya N
ABCG2 is a high-capacity urate transporter and its genetic impairment increases serum uric acid levels in humans.
Nucleosides Nucleotides Nucleic Acids. 2011 Dec;30(12):1091-7., [PMID:22132962]

Abstract [show]
The ATP-binding cassette, subfamily G, member 2 (ABCG2/BCRP) gene encodes a well-known transporter, which exports various substrates including nucleotide analogs such as 3'-azido-3'-deoxythymidine (AZT). ABCG2 is also located in a gout-susceptibility locus (MIM 138900) on chromosome 4q, and has recently been identified by genome-wide association studies to relate to serum uric acid (SUA) and gout. Becuase urate is structurally similar to nucleotide analogs, we hypothesized that ABCG2 might be a urate exporter. To demonstrate our hypothesis, transport assays were performed with membrane vesicles prepared from ABCG2-overexpressing cells. Transport of estrone-3-sulfate (ES), a typical substrate of ABCG2, is inhibited by urate as well as AZT and ES. ATP-dependent transport of urate was then detected in ABCG2-expressing vesicles but not in control vesicles. Kinetic analysis revealed that ABCG2 is a high-capacity urate transporter that maintained its function even under high-urate concentration. The calculated parameters of ABCG2-mediated transport of urate were a Km of 8.24 +/- 1.44 mM and a Vmax of 6.96 +/- 0.89 nmol/min per mg of protein. Moreover, the quantitative trait locus (QTL) analysis performed in 739 Japanese individuals revealed that a dysfunctional variant of ABCG2 increased SUA as the number of minor alleles of the variant increased (p = 6.60 x 10(-5)). Because ABCG2 is expressed on the apical membrane in several tissues, including kidney, intestine, and liver, these findings indicate that ABCG2, a high-capacity urate exporter, has a physiological role of urate homeostasis in the human body through both renal and extrarenal urate excretion.

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28 For quantitative trait locus (QTL) analysis of SUA concentrations, genotyping of Q141K in 739 Japanese individuals was performed. Login to comment
34 With the site-directed mutagenesis technique, we constructed mutants of ABCG2 (V12M, Q126X, and Q141K), which were used for urate transport analysis, on the expression vector for ABCG2. Login to comment
47 We found the following six nonsynonymous mutations: V12M, Q126X, Q141K, G268R, S441N, and F506SfsX4, and the first three mutations are SNPs. Login to comment
48 Maekawa et al.[7] reported that these SNPs are quite common in the Japanese population, and allele frequencies for them are 31.9% for Q141K, 19.2% for V12M, and 2.8% for Q126X, respectively. Login to comment
49 Using Hardy-Weinberg equilibrium and these data on a Japanese population reported by Maekawa et al.,[7] the frequencies of Japanese individuals with these minor alleles were estimated to be 53.6% for Q141K, 34.7% for V12M, and 5.5% for Q126X, respectively. Login to comment
50 [8] To clarify how ABCG2 SNPs affect function, the urate transport capacity of these variants was examined in comparison with that of wild-type ABCG2. ATP-dependent transport of urate was reduced by approximately half (46.7%) in Q141K and was nearly eliminated in Q126X (Figure 2A). Login to comment
51 Common Variant of ABCG2 Increases SUA Levels in Humans With the high-frequency dysfunctional variant Q141K, QTL analysis of SUA was performed in a random sample of 739 Japanese individuals. Login to comment
52 The analysis revealed a significant increase in SUA as the number of minor alleles of Q141K increased (p = 6.60 × 10-5 ), and the corrected p value is 2.02 × 10-6 when adjusted for sex (Figure 2B). Login to comment
54 Unlike SUA, Q141K had no significant association with other clinical characteristics such as age, body mass index, or sex. Login to comment
58 Results are expressed as means ± S.D. (B) QTL analysis of ABCG2 Q141K and serum uric acid levels. Login to comment
59 "C/C," "C/A," and "A/A" indicate wild-type subjects, heterozygous mutation carriers, and homozygous mutation carriers of Q141K, respectively. Login to comment
62 We also demonstrated that Q141K, a dysfunctional variant of ABCG2, significantly affects human SUA levels. Login to comment

[hide] PDZK1 regulates breast cancer resistance protein i... Drug Metab Dispos. 2011 Nov;39(11):2148-54. Epub 2011 Aug 4. Shimizu T, Sugiura T, Wakayama T, Kijima A, Nakamichi N, Iseki S, Silver DL, Kato Y
PDZK1 regulates breast cancer resistance protein in small intestine.
Drug Metab Dispos. 2011 Nov;39(11):2148-54. Epub 2011 Aug 4., [PMID:21816982]

Abstract [show]
Transporter adaptor protein PDZK1 regulates several influx transporters for xenobiotics and nutrients in small intestine, and their expression on the apical membrane is diminished in pdzk1 gene knockout [pdzk1(-/-)] mice. In the present study, we initially attempted to use pdzk1(-/-) mice to functionally identify influx transporters responsible for intestinal absorption of cimetidine. Contrary to our expectation, the plasma concentration of cimetidine after oral administration to pdzk1(-/-) mice was higher than that in wild-type mice, and the double peaks of plasma concentration found in wild-type mice were not observed in pdzk1(-/-) mice. Western blot analysis of intestinal brush-border membranes revealed that expression of breast cancer resistance protein (BCRP) but not of P-glycoprotein is reduced in pdzk1(-/-) mice. This result was compatible with the reduction of apical localization of BCRP in pdzk1(-/-) mice assessed by immunohistochemical analysis. Transcellular transport of cimetidine in the basal-to-apical direction in Madin-Darby canine kidney II (MDCKII) cells stably expressing both BCRP and PDZK1 (MDCKII/BCRP/PDZK1) was higher than that in MDCKII cells stably expressing BCRP (MDCKII/BCRP) cells. Moreover, MDCKII/BCRP/PDZK1 cells are more resistant than MDCKII/BCRP cells to the cytotoxicity of the anticancer agent 7-ethyl-10-hydroxycamptothecin (SN-38), which is a substrate of BCRP. These results were consistent with the higher expression of BCRP on apical membranes in MDCKII/BCRP/PDZK1 cells. Pull-down and immunoprecipitation studies revealed a physical interaction between BCRP and PDZK1. Taken together, these findings demonstrate that PDZK1 plays a pivotal role in the apical localization of BCRP. This is the first identification of a regulatory protein that physically interacts with and regulates BCRP in small intestine in vivo.

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214 The genotype combination of two dysfunctional variants Q126X and Q141K results in increased serum uric acid concentration and increased risk of gout (Matsuo et al., 2009). Login to comment

[hide] Hyperuricemia cosegregating with osteogenesis impe... Hum Genet. 2011 Nov;130(5):671-83. Epub 2011 May 19. Kaneko H, Kitoh H, Matsuura T, Masuda A, Ito M, Mottes M, Rauch F, Ishiguro N, Ohno K
Hyperuricemia cosegregating with osteogenesis imperfecta is associated with a mutation in GPATCH8.
Hum Genet. 2011 Nov;130(5):671-83. Epub 2011 May 19., [PMID:21594610]

Abstract [show]
Autosomal dominant osteogenesis imperfecta (OI) is caused by mutations in COL1A1 or COL1A2. We identified a dominant missense mutation, c.3235G>A in COL1A1 exon 45 predicting p.G1079S, in a Japanese family with mild OI. As mutations in exon 45 exhibit mild to lethal phenotypes, we tested if disruption of an exonic splicing cis-element determines the clinical phenotype, but detected no such mutations. In the Japanese family, juvenile-onset hyperuricemia cosegregated with OI, but not in the previously reported Italian and Canadian families with c.3235G>A. After confirming lack of a founder haplotype in three families, we analyzed PRPSAP1 and PRPSAP2 as candidate genes for hyperuricemia on chr 17 where COL1A1 is located, but found no mutation. We next resequenced the whole exomes of two siblings in the Japanese family and identified variable numbers of previously reported hyperuricemia-associated SNPs in ABCG2 and SLC22A12. The same SNPs, however, were also detected in normouricemic individuals in three families. We then identified two missense SNVs in ZPBP2 and GPATCH8 on chromosome 17 that cosegregated with hyperuricemia in the Japanese family. ZPBP2 p.T69I was at the non-conserved region and was predicted to be benign by in silico analysis, whereas GPATCH8 p.A979P was at a highly conserved region and was predicted to be deleterious, which made p.A979P a conceivable candidate for juvenile-onset hyperuricemia. GPATCH8 is only 5.8 Mbp distant from COL1A1 and encodes a protein harboring an RNA-processing domain and a zinc finger domain, but the molecular functions have not been elucidated to date.

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182 Amino acid AF dbSNP II-2 II-3 VIII-2 1 AGL 100,336,361 C [ T Syn. 0.7 rs2230306 T/T -/T -/T 4 ABCG2b 89,034,551 G [ A Syn. 0.02 rs35622453 -/- -/- -/A 89,052,323 C [ A Q141K 0.31 rs2231142 -/A A/A -/- 89,061,114 G [ A V12M 0.19 rs2231137 -/A -/- -/A 4 SLC2A9b 9,909,923 C [ T P350L 0.33 rs2280205 -/T -/T -/- 9,922,130 G [ A R294H 0.72 rs3733591 -/A -/- -/A 9,998,440 G [ A Syn. 0.54 rs10939650 -/A A/A -/A 10,022,981 G [ A G25R 0.43 rs2276961 -/A -/A -/- 10,027,542 G [ A A17T 0.06 rs6820230 -/- -/A -/A 11 SLC22A12b 64,359,286 C [ T Syn. 0.21 rs3825016 -/T T/T -/T 64,360,274 C [ T Syn. 0.81 rs11231825 -/T -/- -/T 12 PFKM n.d. 16 UMODb n.d. 17 G6PC n.d. X HPRT1a n.d. X PRPS1a n.d. X MAOA 43,591,036 G [ T Syn. 0.3 rs6323 -/- T/T T/T a The gene is associated with purine metabolism b The gene is associated with renal excretion of urate - symbol represents in the patients` genotypes mean being identical to the reference nucleotides Syn synonymous nucleotide change, AF allelic frequency of the changed nucleotide, n.d. no SNPs are detected Discussion Phenotypic variabilities of OI mutations We identified a heteroallelic c.3235G[A mutation in COL1A1 exon 45 in a Japanese family with mild OI type I. Login to comment

[hide] Prognostic value of the multidrug resistance trans... Eur J Cancer. 2011 Sep;47(13):1990-9. Epub 2011 Apr 29. Wang F, Liang YJ, Wu XP, Chen LM, To KK, Dai CL, Yan YY, Wang YS, Tong XZ, Fu LW
Prognostic value of the multidrug resistance transporter ABCG2 gene polymorphisms in Chinese patients with de novo acute leukaemia.
Eur J Cancer. 2011 Sep;47(13):1990-9. Epub 2011 Apr 29., [PMID:21531129]

Abstract [show]
BACKGROUND: Functional polymorphisms of the ABCG2 gene may contribute to individual variability in drug response and the prognosis of patients. METHODS: In the present study, the genetic polymorphisms and expression of ABCG2 were analysed in blasts cells obtained from 184 Chinese patients with de novo acute leukaemia to investigate their possible association with clinical outcomes. RESULTS: A novel synonymous ABCG2-single nucleotide polymorphism (SNP) at exon 16 (13561218 C/T) and five known SNPs at exon 2 (13608835 G/A), exon 5 (13600044 C/A), intron 10 (13576005 C/T), intron 13 (13564503 C/T) and intron 14 (13563578 A/G) were identified with occurrence rates of 1.1%, 64.1%, 30.4%, 21.2%, 39.7% and 28.8%, respectively. We found that patients with the ABCG2 34GG genotype displayed longer disease free survival (DFS) (P<0.001) and overall survival (OS) (P<0.001) than those with the 34GA/AA genotypes. Furthermore, the DFS and OS were significantly diminished in bone marrow transplantation (BMT) patients with the 34GA/AA genotypes relative to those with the 34GG genotype. CONCLUSIONS: These results suggest that these highly prevalent ABCG2 34GA/AA genotypes are associated with poor prognosis of Chinese patients with acute leukaemia and BMT patients.

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17 In particular, two SNPs of ABCG2, G34A (V12M) in exon 2 and C421A (Q141K) in exon 5, were found to be the most prevalent in Asian population. Login to comment
80 Of the six polymorphisms, the allele frequencies of two published non-synonymous SNPs, G34A (Val12Met; exon 2) and C421A (Gln141Lys; exon 5), were 0.505 and 0.152 in our study. Login to comment
104 Position Location Change % Genotype frequency(n) Allele % Allele frequency NT-016354.18 dbSNP (NCBI) Nucleotide Amino acid Wildtype Heterozygote Homozygote 13608835 rs2231137 Exon 2 TCCCAG/ATGTCA Val12Met 35.9 (66) 27.2 (50) 36.9 (68) A 50.5 13600044 rs2231142 Exon 5 ACTTAC/AAGTTC Gln 141Lys 69.6 (128) 30.4 (56) 0 A 15.2 13561218a N.D. Exon 16 GGCATC/TGATCT Ile619Ile 98.9 (182) 1.1 (2) 0 T 0.5 13576005 rs2231149 Intron10 TCAAGC/TTTATT N/A 78.8 (145) 21.2 (39) 0 T 10.6 13564503 rs2231162 Intron13 TGACTC/TTTAGT N/A 60.3 (111) 39.7 (73) 0 T 19.8 13563578 rs2231164 Intron14 TTCTTA/GAAATT N/A 71.2 (131) 28.8 (53) 0 G 14.4 NCBI, National Cancer Center for Biotechnology Information; N.D., not determined; N/A, not applicable. Login to comment
256 Imai Y, Nakane M, Kage K, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] ABCG transporters and disease. FEBS J. 2011 Sep;278(18):3215-25. doi: 10.1111/j.1742-4658.2011.08171.x. Epub 2011 Jun 13. Woodward OM, Kottgen A, Kottgen M
ABCG transporters and disease.
FEBS J. 2011 Sep;278(18):3215-25. doi: 10.1111/j.1742-4658.2011.08171.x. Epub 2011 Jun 13., [PMID:21554546]

Abstract [show]
ATP-binding cassette (ABC) transporters form a large family of transmembrane proteins that facilitate the transport of specific substrates across membranes in an ATP-dependent manner. Transported substrates include lipids, lipopolysaccharides, amino acids, peptides, proteins, inorganic ions, sugars and xenobiotics. Despite this broad array of substrates, the physiological substrate of many ABC transporters has remained elusive. ABC transporters are divided into seven subfamilies, A-G, based on sequence similarity and domain organization. Here we review the role of members of the ABCG subfamily in human disease and how the identification of disease genes helped to determine physiological substrates for specific ABC transporters. We focus on the recent discovery of mutations in ABCG2 causing hyperuricemia and gout, which has led to the identification of urate as a physiological substrate for ABCG2.

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59 R L L A A M AT T T R V S G G G F I T Q R R V K K S G E A D RR V V K K L L G E E E I IN NN D H Q Q R V V V V V L L S G F E N M TT QD D S K R V K L L G F P C Y R K S G F P P C N N A V L L S G G G I N A D R K P P S GG G R V VK K KK L L L LL L S S S GG G PPE E IIII N NN M A A A T D D Y N E A I P E S I D L L F T LS G EI MT D I I P FC L R IH A N T T T T T G L D S S K K K L L L S G G G F F F F Q P P I M M A A A A D H G G LS S S V L L L L L R R RQ Q I I Y Y YS S HE E A T V V V V L Q I S F I I II A A L G G Y K F R S S E E I I L G Y YY Y V V K H S P C M M D R T I II L L L F F YV S S P F N T I A Q Q L L L G F Y Y H S S PR W C N M I I A A A L L G F V V K H W T L I F F C C C D D D A A A QQ Q Q Q G G G G G G G G G G FF FF F F FF Y Y Y Y Y V V V V V VVV K KKK KK K K E E E E P P P P R W W TT TT T TT T T NNNN N N N N N M MM M L L L L L L L L LL LL I I I I I I AA A A A A A S S S S S S S S SS L L L L LL L L LL V V F G GCC T Q Q Q Q Y Y Y KK K K K H H EE E E E E EEE E P P P P R R RW N N N II I I I I I I A AAA A A A S SS S S S S L L LL L L L V V V V F F F F F F F G GG G C TT T T T T K K K K KKKK N NN LL D DDD DS S 395 469 565 644 414 450 495 505 584 625 Signature Walker A WalkerBQ EP MI A V V VF FG GTN N NS S S S P F HE V FG CTT K NN LLD SS AAA I V12M N-terminus C-terminus M MM MM T A A A A L F F Y V V S S S F 524476 Y Q126X G268R S441N F506fs Q141K 44 288 PP AA DD Fig. 2. Login to comment
66 We therefore investigated whether urate is a physiological substrate of ABCG2, and whether the Q141K variant, encoded by rs2231142, leads to altered urate transport and as a consequence to elevated serum urate levels and increased risk of gout. Login to comment
81 Q141K is a functional variant in ABCG2 Several lines of evidence in the initial genome-wide association study by Dehghan et al. [21] suggested that the rs2231142 variant may be functional. Login to comment
83 Effect sizes of the ABCG2 rs2231142 (Q141K) variant on risk of gout and mean urate levels in study populations of different ancestry. Login to comment
85 the variant is located in exon 5 of ABCG2 and leads to a glutamine-to-lysine amino acid substitution (Q141K) in ABCG2. Login to comment
90 To test whether the rs2231142 is such a functional variant, the transport capacity of the encoded Q141K mutation was compared with that of wild-type ABCG2. Login to comment
91 Oocytes expressing ABCG2 Q141K showed 54% reduced urate transport rates compared with oocytes expressing wild-type ABCG2 (Fig. 3C). Login to comment
92 This is consistent with previous studies showing impaired transport of chemotherapeutic agents by ABCG2 Q141K [35,36] (and reviewed in [37]). Login to comment
93 While it is difficult to compare the results from different transport assays and substrates, the reduction of transport of the Q141K variant compared with wild-type ABCG2 appears to be of similar magnitude. Login to comment
94 The Q141 residue is located in the nucleotide binding domain of ABCG2 (Fig. 2), and Q141K ABCG2 expression is significantly lower than wild-type when overexpressed in mammalian cells [35,36,38,39]. Login to comment
96 And like the Q141K ABCG2 mutation, expression of the deleted F508 CFTR mutant is significantly lower than wild-type suggesting a common pathophysiology (Woodward, unpublished observations). Login to comment
97 0.0 0.5 1.0 1.5 Urateaccumulation pmolperoocyte·120min-1 Urateaccumulation pmolperoocyte·120min-1 H2O ABCG2 ** A 0 20 40 60 0.4 0.6 0.8 1.0 Relativeurateremaining Time (min) ** ** ** ** ** B Lumen Blood 0.0 0.3 0.6 0.9 WT Q141K ** C Others3 Others4 SLC2A9 URAT1 SLC2A9 Others1 Others2 UU- UU- UU-UU- UU- UU- UU- UU- U-UU- D ABCG2 Fig. 3. Login to comment
101 (C) Urate accumulation in oocytes expressing either the wild-type ABCG2 or the mutant Q141K ABCG2. Login to comment
108 In addition to Q141K, Q126X was identified as a novel loss-of-function variant. Login to comment
109 Q126X was assigned to a different haplotype than Q141K and shown to increase gout risk (odds ratio 5.97) to an even greater extent than the Q141K variant. Login to comment
110 In addition, 10% of the gout patients studied had genotype combinations of the Q141K and Q126X variants that resulted in more than a 75% reduction of ABCG2 function compared with patients that were homozygous for the non-risk allele at both variants (odds ratio 25.8, 95% confidence interval 10.3-64.6). Login to comment

[hide] Population heterogeneity in the genetic control of... Semin Nephrol. 2011 Sep;31(5):420-5. Merriman TR
Population heterogeneity in the genetic control of serum urate.
Semin Nephrol. 2011 Sep;31(5):420-5., [PMID:22000648]

Abstract [show]
Scanning of the genome in Caucasian cohorts for genes that control serum urate levels have revealed eight confirmed associations. Knowledge of genetic control of serum urate in other populations can be extrapolated from the study of gout, a condition of extreme hyperuricemia; three urate transport genes (SLC2A9, ABCG2, and SLC17A1/NPT1) have been studied in diverse populations (Caucasian, Asian, and Polynesian). Between-population heterogeneity is evident in allele frequency and association with gout, notably within Polynesian populations, which are defined by a geographic region and shared ancestry but also characterized by migratory events that create bottlenecks and alter genetic structure in the founder populations. Despite the large genome-wide studies in Caucasians, only 5% of the variance in serum urate levels has been explained. A more complete picture will be revealed by very large meta-analyses of genome-wide association scans in Caucasian and in other populations with less genetic heterogeneity than Caucasians.

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17 doi:10.1016/j.semnephrol.2011.08.005 Seminars in Nephrology, Vol 31, No 5, September 2011, pp 420-425420 Study using a SNP platform with an additional approximately 50 thousand SNPs identified association of the nonsynonymous Q141K (rs2231142) variant of the ABCG2 gene with serum urate levels in Caucasians.13 Similar to SLC2A9, ABCG2 was a previously undiscovered urate transporter.15 It is highly expressed in intestinal epithelial cells and also expressed in the apical membrane of the kidney proximal tubule. Login to comment
18 ABCG2 is an adenosine triphosphate-dependent uric acid secretory molecule with the risk allele (141K) encoding a molecule with approximately 50% reduced ability to transport uric acid.15-17 The intronic variants of SLC2A9 and ABCG2 Q141K also were associated with serum urate levels in African American subjects within the Atherosclerosis Risk in Communities study.13 The Dehghan et al13 study also identified a weaker association of a SNP within the SLC17A1/ NPT1 (sodium-phosphate transporter 1) locus with serum urate levels in Caucasians, but not African Americans. Login to comment
55 Association of Variants Controlling Serum Urate Levels in Caucasians With Gout and Other Ancestral Groups, in Samples Diagnosed Using ACR Criteria for Gout Population Prevalence of Gout, % SLC2A9 Intronic Variants SLC2A9 R263H (rs3735991) ABCG2 Q141K (rs2231142) NPT1 rs1183201 rs1165196* ReferencesOR† Frequency‡ OR Frequency OR Frequency OR Frequency Caucasians 1.4 2.1 (1.5-3.0) 0.69 1.2 (0.9-1.6) 0.81 2.2 (1.7-2.9) 0.13 1.5 (1.1-1.8) 0.54 24,32,33,39,40 Han Chinese 1.1 § Ͼ0.97 1.5 (1.0-2.2) 0.32 1.7 (1.3-2.2) 0.32 NR NR 30,34,41 Japanese 1-2 § 0.99 1.5 (1.2-2.0) 0.32 2.2 (1.8-2.9) 0.32 1.6 (1.1-2.4) 0.83 16,31,38 New Zealand Pacific Islanders¶ ʈ § 0.83 1.3 (0.9-2.0) 0.43 2.8 (1.9-4.0) 0.22 1.3 (0.9-2.0) 0.31 24,32,33,39 New Zealand Ma៮ori 6.4 5.1 (2.2-11.5) 0.70 1.0 (0.7-1.3) 0.71 1.1 (0.7-1.8) 0.10 1.7 (1.3-2.3) 0.38 24,32,33,39,42 Abbreviation: NR, not reported. Login to comment
63 ABCG2 With the notable exception of New Zealand Ma៮ori, there is consistent and strong association of the Q141K (rs2231142) variant of ABCG2 with gout in Caucasian, Asian, and Pacific Island people (Table 1),16,33-35 with ORs ranging from 1.7 in Han Chinese to 2.8 in Pacific Island people living in New Zealand. Login to comment
64 In contrast to the SLC2A9 variants discussed earlier, there is strong evidence that Q141K is the causative variant in Caucasians; rs2231142 is the most associated genetic variant in the region, and the 141K risk allele reduces the ability of ABCG2 to transport urate in the kidney and gut.15,16 In New Zealand Ma៮ori, however, despite an appreciable risk allele frequency (0.10, compared with 0.12 in Caucasian), there is no evidence for a genetic effect on the risk of gout (OR, 1.1).33 The heterogeneity between the New Zealand Ma៮ori and Pacific Island populations at ABCG2 is most notable. Login to comment
68 The difference between Eastern and Western Polynesia at ABCG2 is more likely to be owing to neutral processes such as migration and genetic drift than to adaptation.36 Why there is no apparent effect of ABCG2 Q141K on the risk of gout in New Zealand Ma៮ori is unclear, it is possible that ABCG2 does play a role in gout in New Zealand Ma៮ori, but other as yet unidentified variants within ABCG2 are important. Login to comment
69 Alternatively, expression of genetic association at Q141K may be dependent on a second risk factor (genetic or environmental) that is absent in New Zealand Ma៮ori. Login to comment

[hide] Urate transporters: an evolving field. Semin Nephrol. 2011 Sep;31(5):400-9. Anzai N, Endou H
Urate transporters: an evolving field.
Semin Nephrol. 2011 Sep;31(5):400-9., [PMID:22000646]

Abstract [show]
Urate (uric acid) is the end product of purine metabolism in human beings owing to the genetic loss of hepatic urate oxidase (uricase). Despite its potential advantage as an antioxidant, sustained hyperuricemia is associated with gout, renal diseases, hypertension, and cardiovascular diseases. Because the kidney plays a dominant role in maintaining serum urate levels through its excretion, it is important to understand the molecular mechanism of renal urate handling. Although molecular identification of the urate/anion exchanger URAT1 (SLC22A12) in 2002 paved the way for successive identification of several urate transport-related proteins, the entire picture of effective renal urate handling in human beings has not yet been clarified. Recently, several genome-wide association studies have revealed close associations between serum urate levels and single nucleotide polymorphisms in at least 10 genetic loci including eight transporter-related genes. These findings led us to consider the roles of urate transporters in extrarenal tissues such as the intestine. In this review, we discuss various aspects of transmembrane transport of urate in the human body.

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122 Breast Cancer Resistance Protein BCRP (ABCG2) Breast cancer resistance protein BCRP is an adenosine triphosphate-binding cassette (ABC) transporter (ABCG2) originally isolated from doxorubicin-resistant breast cancer cells and mediates the transport of various chemicals, natural compounds, and drugs including several anticancer drugs such as methotrexate, mitoxantrone, topotecan, imatinib, and gefitinib.52,53 Although overexpression of BCRP is related to anticancer drug resistance in cancer cells, its endogenous messenger RNA expression was highest in the placenta and was high in the liver and intestine. Although the human kidney is known to express many drug efflux pumps at the apical side of proximal tubules such as P-glycoprotein/MDR1 (ABCB1), MRP2 (ABCC2), and MRP4 (ABCC4),54 evidence for BCRP expression in the kidney is somewhat confusing: according to one report, a moderate level of BCRP protein expression was detected despite no messenger RNA expression in the human kidney.52 Because BCRP also has a high affinity to porphyrins, it is responsible for cellular homeostasis of porphyrins, photosensitivity, and photodynamic therapy.53 Moreover, ABCG2 SNPs have been indicated to be an important factor for patients` adverse drug responses: for example, the most extensively studied ABCG2 SNP Q141K, associated with lower protein expression level and impaired transport activity in vitro, is related to increased plasma levels of gefitinib, increased bioavailability of oral topotecan, and Figure 2. Login to comment
130 The earlier- mentioned hypoactive Q141K protein is associated with hyperuricemia. Login to comment
131 BCRP recently was reported by Woodward et al18 to transport urate and they suggested that BCRP (ABCG2) is involved in urate excretion in the luminal membrane of renal proximal tubules. However, given its expression pattern (ie, high in the liver and intestine) and pathophysiological role of Q141K polymorphism in pharmacokinetics, it is likely that the hypoactive variant of ABCG2 leads to decreased urate excretion into the intestine rather than decreased urate excretion from the kidney. Login to comment

[hide] Part 2: pharmacogenetic variability in drug transp... Oncologist. 2011;16(6):820-34. Epub 2011 May 31. Deenen MJ, Cats A, Beijnen JH, Schellens JH
Part 2: pharmacogenetic variability in drug transport and phase I anticancer drug metabolism.
Oncologist. 2011;16(6):820-34. Epub 2011 May 31., [PMID:21632461]

Abstract [show]
Equivalent drug doses in anticancer chemotherapy may lead to wide interpatient variability in drug response reflected by differences in treatment response or in severity of adverse drug reactions. Differences in the pharmacokinetic (PK) and pharmacodynamic (PD) behavior of a drug contribute to variation in treatment outcome among patients. An important factor responsible for this variability is genetic polymorphism in genes that are involved in PK/PD processes, including drug transporters, phase I and II metabolizing enzymes, and drug targets, and other genes that interfere with drug response. In order to achieve personalized pharmacotherapy, drug dosing and treatment selection based on genotype might help to increase treatment efficacy while reducing unnecessary toxicity. We present a series of four reviews about pharmacogenetic variability in anticancer drug treatment. This is the second review in the series and is focused on genetic variability in genes encoding drug transporters (ABCB1 and ABCG2) and phase I drug-metabolizing enzymes (CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP3A4, CYP3A5, DPYD, CDA and BLMH) and their associations with anticancer drug treatment outcome. Based on the literature reviewed, opportunities for patient-tailored anticancer therapy are presented.

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56 Particularly relevant SNPs in ABCG2 appear to be 421CϾA (Gln141Lys) and the nonsense SNP 376CϾT (Gln126stop). Login to comment
57 Particularly relevant SNPs in ABCG2 appear to be 421Cb0e;A (Gln141Lys) and the nonsense SNP 376Cb0e;T (Gln126stop). Login to comment

[hide] Drug transport by breast cancer resistance protein... Expert Opin Drug Metab Toxicol. 2010 Nov;6(11):1363-84. Epub 2010 Sep 27. Poguntke M, Hazai E, Fromm MF, Zolk O
Drug transport by breast cancer resistance protein.
Expert Opin Drug Metab Toxicol. 2010 Nov;6(11):1363-84. Epub 2010 Sep 27., [PMID:20873966]

Abstract [show]
IMPORTANCE OF THE FIELD: The ATP-binding cassette transporter ABCG2 is a well-known major mediator of multi-drug resistance in cancers. Beyond multi-drug resistance, experimental and recent clinical studies demonstrate a role for ABCG2 as a determinant of drug pharmacokinetic, safety and efficacy profiles. AREAS COVERED IN THIS REVIEW: The clinical evidence of the role of ABCG2 in pharmacokinetics and pharmacodynamics is reviewed. Key questions that arise from the perspective of preclinical drug evaluation are addressed, including the structure of ABCG2 and mechanisms of drug-transporter interactions, mechanisms responsible for the polyspecificity of ABCG2, methods suitable for studying drug-ABCG2 interactions in vitro and in silico prediction of ABCG2 substrates and inhibitors. WHAT THE READER WILL GAIN: An update on current knowledge of the importance of ABCG2 in drug disposition with special emphasis on drug development. TAKE HOME MESSAGE: The field of drug-ABCG2 interaction is rapidly advancing and beginning to expand into clinical practice. However, the structural understanding of drug binding and transport by ABCG2 is still incomplete. Incorporation of novel concepts of drug-transporter interactions such as electrostatic funneling might help explain the multispecificity of ABCG2 and enable in silico predictions.

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34 For example, the common ABCG2 421C>A (p.Gln141Lys; rs2231142) SNP causes reduced ABCG2 expression levels and altered substrate specificity, and has been associated with greater serum accumulation of orally administered ABCG2 substrates such as topotecan, diflomotecan and gefitinib [16-19]. Login to comment
75 SulfasalazineTheABCG2c.421C>ASNPisassociatedwithincreased AUC(C/C,C/Avs.A/A:3.5-fold)andCmaxofa2000mg singledosesulfasalazine Pharmacogeneticassociationstudy:37healthyvolunteers[72] Rituximabplushyper-CVAD regimen(cyclophosphamide, vincristine,adriamycin, dexamethasone) ExpressionofABCG2inbonemarrowsamplesappeared associatedwithaworsePFSandlevelsofthisgene paralleledthestatusofminimalresidualdisease 20patientswithMCLenrolled intoaPhaseIIstudy [12] GefitinibTumorexpressionofABCG2wasassociatedwithacquired resistancetogefitinib,independentofEGFRmutations Casereport:1patientwithadvancedlungcancerand acquiredresistancetogefitinib [11] Gefitinib7(44%)of16patientsheterozygousforABCG2 c.421C>A(Q141K)developeddiarrheavsonly13(12%) of108patientshomozygousforthewild-typesequence (p=0.0046) Pharmacogeneticassociationstudy:129caucasianpatients withNSCLC [84] TopotecanElacridar (interaction) Completeapparentoralbioavailabilityoftopotecanwas observedwithco-administrationofelacridarbeforeor simultaneouslywith2mgoraltopotecan PhaseI,randomized,open-label,parallel-cohort, dose-findingstudyofelacridarandoral topotecan:20cancerpatients [15] RosuvastatinTheABCG2c.421C>Apolymorphismmayplayan importantroleinthepharmacokineticsofrosuvastatin (CCvsCA/AAgenotype:increasedAUCandCmax) Pharmacogeneticassociationstudy:14healthyChinese malevolunteerswhoareSLCO1B1521TTand CYP2C9*1/*1wild-typehomozygotes [85] FluvastatinSimvastatinAUC0--¥offluvastatinwas97or72%largerin participantswiththeABCG2c.421AAgenotypethanin thosewiththeCAorCCgenotype.TheAUC0--¥of simvastatinlactonewas111%largerinparticipantswith theAAgenotypethaninparticipantswiththeCC genotype.TheABCG2genotypehadnosignificanteffect onsimvastatinacidpharmacokinetics Pharmacogeneticassociationstudy:Inacrossoverdesign, 5healthyvolunteerswiththeABCG2c.421AAgenotype, 4withthe421CAgenotypeand23withthec.421CC genotypeingestedasingle40mgdoseoffluvastatin, pravastatinandsimvastatin,withawashoutperiodof 1week [67] TopotecanHeterozygousABCG2c.421CAalleleobservedin 2patientswasassociatedwitha1.34-foldincreasedoral bioavailabilityoftopotecancomparedtothebioavailability in10patientswiththewild-typeallele Pharmacogeneticassociationstudy:12patientswith ovarianorsmall-celllungcancer [16] ALL:Acutelymphoblasticleukemia;AML:Acutemyelogenousleukemia;MCL:Mantlecelllymphoma;NSCLC:Nonsmallcelllungcancer;PFS:Progression-freesurvival;P-gp:P-glycoprotein;SNP:Singlenucleotide polymorphism. Poguntke, Hazai, Fromm & Zolk Expert Opin. Drug Metab. Toxicol. Login to comment
78 AMLinductionanthracycline ormitoxantroneincombination withcytarabine(insomecases additionalcytostaticdrugs) NodifferenceinABCG2mRNAexpressionbetween patientsrespondingtoinductiontreatmentand non-responders.Inthegroupofresponders,the 14patientswiththehighestexpressionhadsignificantly shorteroverallsurvival(mean38months)thanthe 14patientswiththelowestexpression Pharmacogeneticassociationstudy:40patientswithnewly diagnosedAML [86] TopotecanElacridar (interaction) Co-administrationoftheABCG2andP-gpinhibitor elacridarresultsinasignificantincreaseofthesystemic exposureoforaltopotecan:theapparentoral bioavailabilityincreasesfrom40to97.1%withoutand withelacridar,respectively Randomizedcontrolledtrial:CohortA:8patients randomizedtoreceiveoraltopotecanwithorwithout co-administrationofonesingleoraldoseof1000mg elacridar.CohortB:8otherpatientsrandomizedto receiveintravenoustopotecanwithorwithout1000mg oralelacridar [26] ClinicalevidenceagainstsignificantimpactofABCG2ondrugdispositionandeffect IrinotecanNosignificantchangesinirinotecanpharmacokinetics wereobservedinrelationtotheABCG2c.421C>A genotype Pharmacogeneticassociationstudy:88American Caucasians,94AfricanAmericans,938Africans,95Han Chinese,84Europeancaucasianpatientstreatedwith irinotecan [87] SulfasalazineNosignificantchangesinsulfasalazinepharmacokinetics wereobservedinrelationtotheABCG2c.421C>A genotype;pantoprazoleandfamotidinedidnotaffect sulfasalazinepharmacokineticsinanygenotypiccohort Pharmacogeneticassociationstudy,drug--druginteraction study:36malehealthyChinesesubjects [73] Taxaneandplatinum anticancerdrugs NoreproduciblesignificantassociationsbetweenABCG2 Q141Kgenotypeandoutcomeortoxicity Retrospectivepharmacogeneticanalysis:914ovarian cancerpatientsfromtheScottishRandomisedTrialin OvarianCancerPhaseIIItrial [88] PravastatinABCG2genotypeswerenotassociatedwithdifferencesin pravastatinpharmacokinetics Pharmacogeneticassociationstudy:107participants (69EuropeanAmericansand38AfricanAmericans) [63] IrinotecanCisplatinABCG2(c.34G>A,c.421C>A)SNPsdidnotcorrelate withirinotecan-PK,toxicity,tumorresponseandsurvival Pharmacogeneticassociationstudy:107patientswith advancedNSCLCtreatedwithirinotecanandcisplatin chemotherapies [89] LamivudineDispositionoflamivudinewasnotsignificantlyinfluenced byknowninvitrofunctionalvariantsofABCG2,Q141K, V12MandQ126X Pharmacogeneticassociationstudy:22healthymale Koreansubjects [90] ALL:Acutelymphoblasticleukemia;AML:Acutemyelogenousleukemia;MCL:Mantlecelllymphoma;NSCLC:Nonsmallcelllungcancer;PFS:Progression-freesurvival;P-gp:P-glycoprotein;SNP:Singlenucleotide polymorphism. Drug transport by breast cancer resistance protein 1368 Expert Opin. Drug Metab. Toxicol. Login to comment
529 The effect of ABCG2 V12M, Q141K and Poguntke, Hazai, Fromm & Zolk Expert Opin. Drug Metab. Toxicol. Login to comment

[hide] Common defects of ABCG2, a high-capacity urate exp... Sci Transl Med. 2009 Nov 4;1(5):5ra11. Matsuo H, Takada T, Ichida K, Nakamura T, Nakayama A, Ikebuchi Y, Ito K, Kusanagi Y, Chiba T, Tadokoro S, Takada Y, Oikawa Y, Inoue H, Suzuki K, Okada R, Nishiyama J, Domoto H, Watanabe S, Fujita M, Morimoto Y, Naito M, Nishio K, Hishida A, Wakai K, Asai Y, Niwa K, Kamakura K, Nonoyama S, Sakurai Y, Hosoya T, Kanai Y, Suzuki H, Hamajima N, Shinomiya N
Common defects of ABCG2, a high-capacity urate exporter, cause gout: a function-based genetic analysis in a Japanese population.
Sci Transl Med. 2009 Nov 4;1(5):5ra11., [PMID:20368174]

Abstract [show]
Gout based on hyperuricemia is a common disease with a genetic predisposition, which causes acute arthritis. The ABCG2/BCRP gene, located in a gout-susceptibility locus on chromosome 4q, has been identified by recent genome-wide association studies of serum uric acid concentrations and gout. Urate transport assays demonstrated that ABCG2 is a high-capacity urate secretion transporter. Sequencing of the ABCG2 gene in 90 hyperuricemia patients revealed several nonfunctional ABCG2 mutations, including Q126X. Quantitative trait locus analysis of 739 individuals showed that a common dysfunctional variant of ABCG2, Q141K, increases serum uric acid. Q126X is assigned to the different disease haplotype from Q141K and increases gout risk, conferring an odds ratio of 5.97. Furthermore, 10% of gout patients (16 out of 159 cases) had genotype combinations resulting in more than 75% reduction of ABCG2 function (odds ratio, 25.8). Our findings indicate that nonfunctional variants of ABCG2 essentially block gut and renal urate excretion and cause gout.

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3 This study identified Q141K as aABCG2gout in individuals carrying variants in a multifunctional transporter gene, based study supported an association between urate levels and-remained largely unclear. Login to comment
13 Quantitative trait locus analysis of 739 individuals showed that a common dysfunctional variant of ABCG2, Q141K, increases serum uric acid. Login to comment
14 Q126X is assigned to the different disease haplotype from Q141K and increases gout risk, conferring an odds ratio of 5.97. Login to comment
46 The following six nonsynonymous mutations were found: V12M, Q126X, Q141K, G268R, S441N, and F506SfsX4 (Table 1). Login to comment
48 Maekawa et al. reported that allele frequencies for these SNPs, which are quite common in the Japanese population, were 31.9% for Q141K, 19.2% for V12M, and 2.8% for Q126X, respectively (Table 1) (34). Login to comment
49 Using Hardy-Weinberg equilibrium and these data of a Japanese population reported by Maekawa et al. (34), we calculated estimates of the frequencies of Japanese individuals with these minor alleles to be 53.6% for Q141K, 34.7% for V12M, and 5.5% for Q126X. Login to comment
52 ATP-dependent transport of urate was reduced by approximately half (46.7%) in Q141K and was nearly eliminated in Q126X, G268R, S441N, and F506SfsX4 mutants (Fig. 2B). Login to comment
53 Western blot analysis showed that ABCG2 protein expression levels in the Q141K variant decreased by half (47.2%), whereas Q126X resulted in no protein on membrane vesicles (fig. S4). Login to comment
54 The decreased activity of Q141K is probably a result of decreased amounts of ABCG2 protein, consistent with our previous study on ES transport (29). Login to comment
57 Common variant of ABCG2 increases SUA concentrations Quantitative trait locus (QTL) analysis of SUA was performed with the high-frequency dysfunctional variant Q141K in ABCG2 in a random sample of 739 Japanese individuals. Login to comment
67 All results are expressed as means ± SD. SUA significantly increased as the number of minor alleles of Q141K increased (P = 6.60 × 10-5 ), and when adjusted for sex, the corrected P value is 2.02 × 10-6 (Fig. 2C). Login to comment
69 Different from SUA, Q141K had no significant association with other clinical parameters such as age, body mass index, or sex (table S2). Login to comment
76 The partially functional SNP Q141K also increased gout risk (OR, 2.23; 95% CI, 1.75-2.87; P = 5.54 × 10-11 ). Login to comment
77 The call rate, or the ability of the SNP to be reliably decoded, for V12M, Q126X, and LS N N SV FLC S P T AN FK G LM ETS S E V F I P Q G N T N G FV P A A AS LD V S N I C Y R V K K RKPVEKEILSNINGIKPGLNAILGPG GGKSSL LDVLA ARKDP S G T L S G D V L I G A P PR A N F K N S G Y Q D D V V M G T L T V R NE LV VC H Q F S A A A RL L T T TNEKNER HINRVIQELGLDKVADSKVGTQFIRGVG GERR KTSIGME L I T D P S I L F L D E P T T G L D S S T A N A V LL L L K R M S K Q G R I I F S T S I H Q P R Y M S I F K LFDSLTLLASGRLMFHGPAQEALGYFESAGYHCEAN YN T V A L N R E E D F K A T E II E P S K Q D K L I E L A EK I Y V N S S F Y K ETKAELHQLSGGEKKKKITVFKEISYTTSFCHQRWVK SRS AFFLDII N G D S A PD P L F K N LL G N P Q A S A I V G I I T L V A FI I Q V V L G Y AVEFLKNDST G I Q N R A G V L F F L T T Q C F S L V S S N G L S L M L I T P M S F I FV D L R P I C Y W L W Y I Y T Q S R F L NQ S L F P G A H E F Y S Y S E F R G Y I K V K S V Y I H L E V A S S V L M A A M F V A F M S Y M T M F K A T I M L H F I A V K G W I L V C A N L W V A T L M T C F VI F M M I F S G L L VNLTTIASAIAAGQS L S V V LKGL L F N Q L F P S L D Y G Q K V L C Y EEGTCTAYNCPNNGTAN G P G L K L L L K K SYF L Y D L G L M A P K Extracellular Intracellular 50 150 200 300 100 350 395 415 469 450 470 500 525 550 565 585 600 625 608 650 250 655 603 475 644 F506SfsX4 (F506fs) V12M Q126X Q141K S441N G268R V Q F S G Q # C signature Walker B motif Walker A motif C D E 4.0 4.5 5.0 5.5 6.0 C/C C/A A/A Male + female P= 2.02 x 10 -6 5.0 5.5 6.0 6.5 7.0 C/C C/A A/A Male P= 0.0144 Serumuric acid(mg/dl) 4.0 4.5 5.0 5.5 6.0 C/C C/A A/A Female P= 0.0137 (pmol/mgprotein) 0 20 40 60 80 100 120 140 160 180 200 + AMP + ATP B Serumuric acid(mg/dl) Serumuric acid(mg/dl) A [C]Uratetransport 14 G F M C-terminal N-terminal Fig. 2. Login to comment
84 Results are expressed as means ± SD. (C to E) QTL analysis of Q141K and SUA concentrations was performed in 739 Japanese individuals (C), including 245 male subjects (D) and 494 female subjects (E). Login to comment
85 "C/C," "C/A," and "A/A" indicate wild-type subjects, heterozygous mutation carriers, and homozygous mutation carriers of Q141K, respectively. Login to comment
89 Amino acid change SNP ID dbSNP (NCBI) Exon Type of mutation Number of hyperuricemia patients Allele frequency (%) (in hyperuricemia) Allele frequency* (%) (in Japanese population) Wild-type Heterozygote Homozygote Q141K rs2231142 5 Missense 29 47 14 41.67 31.9 V12M rs2231137 2 Missense 64 23 3 16.11 19.2 Q126X 4 Nonsense 80 10 0 5.56 2.8 G268R 7 Missense 89 1 0 0.56 N.D. S441N 11 Missense 89 1 0 0.56 0.3 F506SfsX4 13 Frameshift 89 1 0 0.56 0.3 * Data from Maekawa et al. (34). Login to comment
90 Q141K was 98.8%, 100%, and 99.2%, respectively. P values for Hardy-Weinberg equilibrium of V12M, Q126X, and Q141K were 0.08, 0.72, and 0.01, respectively. P values that suggested mistyping were not obtained. Login to comment
91 Haplotype frequency analysis revealed that there is no simultaneous presence of the minor alleles of Q126X and Q141K in one haplotype (Table 3). Login to comment
93 Q141K is assigned to another independent risk haplotype. Login to comment
99 Because Q126X and Q141K are assigned to different risk haplotypes, nonfunctional and half-functional haplotype, respectively, their genotype combinations are divided into four groups on the basis of the estimated ABCG2 transport functions, that is, full function, ¾ function, ½ function, and ≤¼ function (Table 4). Login to comment
115 Phenotype SNP Genotype* Allele frequency mode Case Control P value P value OR 95% CI 1/1 1/2 2/2 MAF 1/1 1/2 2/2 MAF Q126X 1 21 139 0.071 0 31 840 0.018 1.74 × 10-7 3.04 × 10-8 4.25 2.44-7.38 Gout Q141K 31 87 41 0.469 87 316 462 0.281 5.80 × 10-10 5.54 × 10-11 2.23 1.75-2.87 V12M 3 43 112 0.155 30 306 526 0.212 0.055 0.020 0.68 0.49-0.94 Q126X 2 24 202 0.061 0 31 840 0.018 1.91 × 10-6 2.91 × 10-7 3.61 2.14-6.08 Hyperuricemia Q141K 45 113 68 0.449 87 316 462 0.281 5.32 × 10-10 1.53 × 10-11 2.06 1.67-2.55 V12M 7 55 163 0.153 30 306 526 0.212 0.006 0.005 0.67 0.51-0.89 * Minor allele was referred to as allele 1 and major allele as 2. Login to comment
117 Allele 1 is A and allele 2 is C in Q141K. Login to comment
120 Haplotype frequency analysis of V12M, Q126X, and Q141K. Login to comment
122 Risk alleles for Q126X and Q141K are underlined. Login to comment
123 Allele Frequency P value OR 95% CI V12M Q126X Q141K Gout Control G C A 0.465 0.284 2.26 × 10-13 2.50 1.94-3.20 G T C 0.071 0.018 4.10 × 10-12 5.97 3.39-10.51 G C C 0.306 0.486 - - - A C C 0.155 0.212 - - - of ABCG2, such as rosuvastatin (42) and gefitinib (43), have been reported. Login to comment
144 OR is obtained by comparing with nonrisk genotype combination C/C(Q126X) and C/C(Q141K). Login to comment
145 Risk alleles for Q126X and Q141K are underlined. Login to comment
146 Estimated transport Genotype Number P value OR 95% CI Q126X Q141K Gout Control ≤¼ function T/T C/C 16 8 3.39 × 10-21 25.8 10.3-64.6 T/C A/C ½ function T/C C/C 37 110 2.23 × 10-9 4.34 2.61-7.24 C/C A/A ¾ function C/C A/C 72 308 2.29 × 10-7 3.02 1.96-4.65 Full function C/C C/C 34 439 Normal control UA ABCG2 transport Gout <UA 7.0 mg/dl 23.3% 10.1% 35.6% 12.7% 0.9% patients Gout no function Q126X T/T Q141K C/C 25% function 50% function =< T/C A/C 45.3% 50.8% 100% function 75% function 50% function C/C C/C T/C A/C A/A C/C 21.4%s 100% function C/C C/C Other gout risk G G G G G N G Fig. 3. Login to comment
148 The genotype combinations of Q126X and Q141K are divided into several groups based on estimated ABCG2 transport functions. Login to comment
154 They also found that a missense SNP (Q141K) was associated with self-reported gout in Caucasians (OR, 1.74; 95% CI, 1.51-1.99) (22), which is consistent with our finding from clinically diagnosed gout patients from a Japanese population. Login to comment
156 In this study, we identified several nonfunctional ABCG2 mutations, including Q126X, in Japanese gout patients, which shows stronger effects on gout development than Q141K did in a previous study (OR < 2.0) (22). Login to comment
160 We and Woodward et al. (55) independently found an ability of ABCG2 to transport urate and characterized the effects of a partially functional variant Q141K using different methods. Login to comment
172 For QTL analysis of SUA concentrations, genotyping of Q141K in 739 Japanese individualswas performed. Login to comment
181 Using the site-directed mutagenesis technique, we constructed mutants of ABCG2 (V12M, Q126X, Q141K, G268R, S441N, and F506SfsX4), which were used for urate transport analysis, on the expression vector for ABCG2. Login to comment
201 Association of Q141K in ABCG2 with clinical parameters. Login to comment

[hide] Pharmacological interplay between breast cancer re... Anticancer Res. 2009 Apr;29(4):1059-65. Katayama K, Shibata K, Mitsuhashi J, Noguchi K, Sugimoto Y
Pharmacological interplay between breast cancer resistance protein and gefitinib in epidermal growth factor receptor signaling.
Anticancer Res. 2009 Apr;29(4):1059-65., [PMID:19414346]

Abstract [show]
BACKGROUND: It has been previously shown that gefitinib reverses breast cancer resistance protein (BCRP)-mediated drug resistance. Here, the impact of BCRP on gefitinib-mediated inhibition in epidermal growth factor receptor (EGFR) signaling is evaluated. MATERIALS AND METHODS: Sensitivity to gefitinib was determined by growth inhibition assay, and intracellular gefitinib levels were measured with HPLC. Western blotting was performed to detect EGFR signaling molecules. RESULTS: BCRP reduced intracellular gefitinib levels and attenuated inhibitory activities of gefitinib to EGF-dependent EGFR signalings including downstream MAPK and Akt pathways in gefitinib-sensitive PC-9 cells. However, gefitinib did not inhibit MAPK and Akt signalings in KB-3-1 and HCT-116 cells, and BCRP-mediated gefitinib-resistance shown in PC-9 cells was not observed in gefitinib-insensitive KB-3-1 and HCT-116 cells. CONCLUSION: BCRP transports gefitinib and suppresses its inhibitory effects on EGFR phosphorylation. However, effects of BCRP on gefitinib activity in the EGFR signaling and on gefitinib-resistance were limited in the gefitinib-sensitive cells only.

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149 The expression levels of BCRP gene products harboring a C421A (Q141K) SNP are 5-fold lower than those of the wild-type gene, and the resistance of cells with a C421A BCRP SNP to SN-38 is also 5-fold lower than those with wild-type BCRP (25, 27). Login to comment
147 The expression levels of BCRP gene products harboring a C421A (Q141K) SNP are 5-fold lower than those of the wild-type gene, and the resistance of cells with a C421A BCRP SNP to SN-38 is also 5-fold lower than those with wild-type BCRP (25, 27). Login to comment

[hide] Structure, function, expression, genomic organizat... Int J Toxicol. 2006 Jul-Aug;25(4):231-59. Choudhuri S, Klaassen CD
Structure, function, expression, genomic organization, and single nucleotide polymorphisms of human ABCB1 (MDR1), ABCC (MRP), and ABCG2 (BCRP) efflux transporters.
Int J Toxicol. 2006 Jul-Aug;25(4):231-59., [PMID:16815813]

Abstract [show]
The ATP-binding cassette (ABC) transporters constitute a large family of membrane proteins, which transport a variety of compounds through the membrane against a concentration gradient at the cost of ATP hydrolysis. Substrates of the ABC transporters include lipids, bile acids, xenobiotics, and peptides for antigen presentation. As they transport exogenous and endogenous compounds, they reduce the body load of potentially harmful substances. One by-product of such protective function is that they also eliminate various useful drugs from the body, causing drug resistance. This review is a brief summary of the structure, function, and expression of the important drug resistance-conferring members belonging to three subfamilies of the human ABC family; these are ABCB1 (MDR1/P-glycoprotein of subfamily ABCB), subfamily ABCC (MRPs), and ABCG2 (BCRP of subfamily ABCG), which are expressed in various organs. In the text, the transporter symbol that carries the subfamily name (such as ABCB1, ABCC1, etc.) is used interchangeably with the corresponding original names, such as MDR1P-glycoprotein, MRP1, etc., respectively. Both nomenclatures are maintained in the text because both are still used in the transporter literature. This helps readers relate various names that they encounter in the literature. It now appears that P-glycoprotein, MRP1, MRP2, and BCRP can explain the phenomenon of multidrug resistance in all cell lines analyzed thus far. Also discussed are the gene structure, regulation of expression, and various polymorphisms in these genes. Because genetic polymorphism is thought to underlie interindividual differences, including their response to drugs and other xenobiotics, the importance of polymorphism in these genes is also discussed.

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573 Recently, Kondo et al. (2004) reported the effect of single nucleotide polymorphisms (SNPs) in ABCG2 gene on its localization, expression level, and transport activity of the BCRP protein. The cellular localization was identified using the wild-type and seven different SNP variants of BCRP protein (Val12Met, Gln141Lys, Ala149Pro, Arg163Lys, Gln166Glu, Pro269Ser, and Ser441Asn), following their expression in LLC-PK1 cells. Login to comment
575 The authors concluded that Gln141Lys variant of the BCRP protein may be associated with a lower expression level, and Ser441Asn variant may lower both the expression level and cellular localization. Login to comment
577 The effect of Gln141Lys variant of the BCRP protein function was also studied by other investigators, but the results are not consistent. Login to comment
579 The purpose of the study was to evaluate the ethnic distribution and potential functional consequence of the C421A SNP on ABCG2 gene function (that results in the Gln141Lys amino acid change in BCRP protein). Login to comment
581 No significant changes in irinotecan pharmacokinetics were observed in relation to the ABCG2 C421A genotype, i.e., the Gln141Lys variant of the BCRP protein. The authors concluded that the contribution of this genetic variant may have been obscured by a functional role of other polymorphic proteins. Login to comment
583 SNP analyses of the ABCG2 gene by Morisaki et al. (2005) revealed three nonsynonymous SNPs that resulted in amino acid substitution of the BCRP protein; these were Val12Met, Gln141Lys, and Asp620Asn. Login to comment
584 When human embryonic kidney cells (HEK-293) were stably transfected with wild-type or variants of ABCG2/BCRP, it was found that cells transfected with wild-type Arg482 ABCG2 showed IC(50) values up to 1.2-fold to 5-fold higher than cells expressing comparable levels of Gln141Lys variant of ABCG2, suggesting that the Gln141Lys SNP affects drug transport, such as that of mitoxantrone, topotecan, SN-38, or diflomotecan. Login to comment
585 This suggests that the Gln141Lys SNP affects the transport efficiency of ABCG2 that may result in altered pharmacokinetics or drug-resistance profiles in clinical oncology. Login to comment
586 The prevalence of C421A SNP in ABCG2 gene (resulting in Gln141Lys variant of the BCRP protein) has been recently confirmed by Kobayashi et al. (2005). Login to comment

[hide] ABCMdb: A database for the comparative analysis of... Hum Mutat. 2012 Nov;33(11):1547-56. doi: 10.1002/humu.22138. Epub 2012 Jul 11. Gyimesi G, Borsodi D, Saranko H, Tordai H, Sarkadi B, Hegedus T
ABCMdb: A database for the comparative analysis of protein mutations in ABC transporters, and a potential framework for a general application.
Hum Mutat. 2012 Nov;33(11):1547-56. doi: 10.1002/humu.22138. Epub 2012 Jul 11., [PMID:22693078]

Abstract [show]
To overcome the pathological phenomena caused by altered function of ABC (ATP Binding Cassette) proteins, their mechanisms of action are extensively investigated, often involving the design of mutant constructs for experiments. Designing mutagenetic constructs, interpreting the result of mutagenetic experiments, and finding individual genetic variants require an extensive knowledge of previously published mutations. To aid the recapitulation of mutations described in the literature, we set up a database of ABC protein mutations (ABCMdb) extracted from full-text papers using an automatic mining approach. We have also developed a Web application interface to compare mutations in different ABC proteins using sequence alignments and to interactively map the mutations to 3D structural models. Currently our database contains protein mutations published for ABCB1, ABCB11, ABCC1, ABCC6, ABCC7, and the proteins of the ABCG subfamily. The database will be extended to include other members and subfamilies, and to provide information on whether or not a mutation is disease causing, represents a high-incidence polymorphism, or was generated only in vitro. The ABCMdb database should already help to compare the effects of mutations at homologous positions in different ABC proteins, and its interactive tools aim to advance the design of experiments for a wider range of proteins. Hum Mutat 33:1547-1556, 2012. (c) 2012 Wiley Periodicals, Inc.

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134 The ABCG2 p.Gln141Lys mutation is shown. Login to comment

[hide] Genetic polymorphisms as predictive and prognostic... Gynecol Oncol. 2012 Feb;124(2):354-65. Epub 2011 Nov 6. Diaz-Padilla I, Amir E, Marsh S, Liu G, Mackay H
Genetic polymorphisms as predictive and prognostic biomarkers in gynecological cancers: a systematic review.
Gynecol Oncol. 2012 Feb;124(2):354-65. Epub 2011 Nov 6., [PMID:22063461]

Abstract [show]
PURPOSE: Numerous studies have explored the potential role of genetic polymorphisms as predictive or prognostic biomarkers in gynecologic malignancies. A systematic review for all eligible polymorphisms has not yet been reported. The aim of this study was to summarize the current status of the field and provide direction for future research. DESIGN: We searched literature databases (MEDLINE, EMBASE, Cochrane) from 2006 to April 2011 to identify studies evaluating the association between gene polymorphisms and clinical outcome in ovarian, endometrial, cervical, or vulvar cancer. The main outcome measures were overall survival (OS) and progression-free survival (PFS). Studies reporting relationships between polymorphisms and toxicity were also included. RESULTS: Sixty two studies met the inclusion criteria. The median sample size was 140. Most of the included studies (n=50, 81%) were conducted in ovarian cancer patients. Almost a third assessed potential predictive associations between gene polymorphism and outcome in ovarian cancer. The most commonly evaluated genes were ERCC1, VEGF, ABCB1 (MDR), and GSTP1. Most studies (n=44, 71%) were observational case-series. Only four studies (6%) included a validation arm and patient population ethnicity was explicitly stated only in 27% of included studies. CONCLUSION: No consistent association between any gene polymorphism and clinical outcome in gynecological cancers has been found across studies. There is incomplete adherence to the REMARK guidelines and inadequate methodology reporting in most studies. Moving forward, analysis of large trial-based clinical samples; adherence to the highest methodological standards, and focus on validation analyses are necessary to identify clinically useful pharmacogenomic biomarkers of outcome.

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1009 c Other polymorphisms evaluated in Marsh et al.: ABCC1 S1334S, ABCC1 IVS19-30C>G, ABCC2 24C>T, ABCC2 IVS12+148A>G, ABCC2 V417I, ABCG2 Q141K, CYP2C8 M264I, CYP2C8 R139K, CYP2C8 K399R, CYP3A4*1B, CYP3A4*3, CYP3A5*3C, ERCC1 17677G>T, MAPT P587P, MPO-463G>A, CDKN1A 10971C>T, CYP1B1*3. d The HR is outside of the 95% CI, authors were not able to resolve these contradictory numbers. Login to comment

[hide] Pharmacogenetics in ghana: reviewing the evidence. Ghana Med J. 2011 Jun;45(2):73-80. Kudzi W, Adjei GO, Ofori-Adjei D, Dodoo AN
Pharmacogenetics in ghana: reviewing the evidence.
Ghana Med J. 2011 Jun;45(2):73-80., [PMID:21857725]

Abstract [show]
Different clinical response of different patients to the same medicine has been recognised and documented since the 1950's. Variability in response of individuals to standard doses of drug therapy is important in clinical practice and can lead to therapeutic failures or adverse drug reactions. Pharmacogenetics seeks to identify individual genetic differences (polymorphisms) in drug absorption, metabolism, distribution and excretion that can affect the activity of a particular drug with the view of improving efficacy and reducing toxicity. Although knowledge of pharmacogenetics is being translated into clinical practice in the developed world, its applicability in the developing countries is low. Several factors account for this including the fact that there is very little pharmacogenetic information available in many indigenous African populations including Ghanaians. A number of genes including Cytochrome P450 (CYP) 2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, MDR1 and TPMT have been genotyped in the Ghanaian population since the completion of the Human genome project. There is however, an urgent need to increase pharmacogenetic research in Ghana to increase availability of data. Introducing Pharmacogenetics into the curriculum of Medical and Pharmacy training institutions will influence translating knowledge of pharmacogenetics into clinical practice. This will also equip health professionals with the skill to integrate genetic information into public health decision making.

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71 Table 2: Allele frequencies of Phase II genes in Ghanaian population Gene Allele No Variant Reference Phase II enzymes COMT 1947A 195 0.06 40 TPMT *3C *3C *8 719G 217 116 863 0.148 0.025 0.034 0.065 41 42 16 UGT1A1 -3156A 853 0.34 16 NAT2 *6 *14 850 800 0.27 0.10 16 16 GSTP1 I105V 837 0.50 16 Transporters ABCB1 3435T 3435T 3435T 3435T 194 206 172 861 0.11 0.17 0.17 0.12 36 43 44 16 ABCG2 Q141K 919 0.01 16 SDR5A2 129 0.19 38 Others TYMS 1494del 799 0.44 16 TNFα -238A 850 0.02 16 SGK1 I6C E8T 112 0.317 0.045 45 Two other phase 1 enzymes, CYP2A6 and CYP2B6, have been associated with efavirenz plasma concentration in HIV-infected patients. Login to comment

[hide] Influence of ethnicity on pharmacogenetic variatio... Pharmacogenomics J. 2009 Dec;9(6):373-9. Epub 2009 Jun 23. Yen-Revollo JL, Van Booven DJ, Peters EJ, Hoskins JM, Engen RM, Kannall HD, Ofori-Adjei D, McLeod HL, Marsh S
Influence of ethnicity on pharmacogenetic variation in the Ghanaian population.
Pharmacogenomics J. 2009 Dec;9(6):373-9. Epub 2009 Jun 23., [PMID:19546880]

Abstract [show]
It has been well established that the frequencies of genomic variants can vary greatly between the populations of different countries. We sought to quantify the intra-population variability in Ghana to determine the value of genotyping studies done at a nationwide level. Further, we investigated the differences between the Ghanaian and other African populations to determine the quality of genomic representation provided by a small subgroup within the continent with regard to the general population. We genotyped 934 unrelated Ghanaian individuals for 15 single nucleotide polymorphisms (SNPs) from genes defined as clinically relevant based on their reported roles in the transport of, metabolism of, or as targets of the medicines listed in the World Health Organization Essential Medicines list. Populations within Ghana and between nations in Western Africa were genetically cohesive. In contrast, populations in other areas of Africa were genetically divergent. Gene allele frequency also differed significantly between the populations in African nations and the United States for several of the SNPs. These results demonstrate that national populations in similar geographic regions, like Africa, may have widely varying genetic allele frequencies for clinically relevant SNPs. Further genotyping studies of specific populations are necessary to provide the best medical care to all individuals.

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27 Many existing US studies were performed using primarily Caucasian subjects (ABCG2 Q141K, CYP3A5*3C, DPYD*2A GSTP1 I105V, NAT2*6, TPMT 719A4G, TYMS 1494del).17-20 Some studies included allele frequencies from different ethnicities such as European-Americans, African-Americans, Asian- Americans, and Hispanic-Americans (ABCB1 3435C4T, CYP2C19*2, CYP2C9*2 and *3, CYP3A4*1B, and UGT1A1 - 3156G4A).18,21 Results All of the genotypes were found to be in Hardy-Weinberg equilibrium. Login to comment
42 Table2GenotypesandallelefrequenciesofpharmacogeneticallyrelevantpolymorphismsobservedintheGhanaianpopulation(%inparenthesis) ABCB1 3435C4T ABCG2 Q141K CYP2C19 *2 CYP2C9 *2 CYP2C9 *3 CYP3A4 *1B CYP3A5 *3C DPYD *2A GSTP1 I105V NAT2 *14 NAT2 *6 TNFa À238G4A TPMT 719A4G TYMS 1494del UGT1A1 À3156G4A Total# genotyped 861919828845835787864893837800850850863799853 #majorallele homozygous 671(78)905(98.5)578(70)843(99.8)834(99.9)34(4)644(75)893(100)224(27)643(80)467(55)820(96.5)751(87)287(36)382(45) #heterozygous176(20)13(1.4)224(27)2(0.2)1(0.1)271(34)201(23)0(0)392(47)150(19)314(37)27(3.2)106(12)325(41)360(42) #minorallele homozygous 14(2)1(0.1)26(3)0(0)0(0)482(61)19(2)0(0)221(26)7(1)69(8)3(0.3)6(1)187(23)111(13) Majorallele frequency 0.880.990.831.001.000.220.861.000.500.900.730.980.930.560.66 Minorallele frequency 0.120.010.170.000.000.780.140.000.500.100.270.020.070.440.34 Table3ComparisonofliteraturereportedallelefrequenciesofpharmacogeneticallyrelevantpolymorphismsinAfricannationscomparedwiththeGhanaian population MinoralleleABCB1 3435C4T ABCG2 Q141K CYP2C19 *2 CYP2C9 *2 CYP2C9 *3 CYP3A4 *1B CYP3A5 *3C DPYD *2A GSTP1 I105V NAT2 *14 NAT2 *6 TNFa -238G4A TPMT 719A4G TYMS 1494del UGT1A1 -3156G4A TAATCGGAGAAAGinsTTAAAGA Ghana(total)0.120.010.170.000.000.780.140.000.500.100.270.020.070.440.34 Yoruba (HapMap)28 0.110.167000.7460.1500.3750.0920.175N/A0.05 WestAfrica Ghana0.1718 0.8118 018 Benin0.1318 0a18 BurkinaFaso0.0222 Cameroon0.4618 Gabon0.10*23 Guinea-Bissau0.8118 Nigeria0.8718 0.0618 Senegal0.7818 0.3118 0.124 0.1724 TheGambia0.21*18 0.5318 0.06*30 EastAfrica Egypt0.4*18 0.06*18 0.18*a18 018 0.27*18 0.02*18 Ethiopia0.07*18 0.06*a18 Kenya0.1718 0.1518 0.0518 Sudan0.27*18 0.03*25 0.2925 0.08*31 Tanzania0.1818 0.16*18 0.1226 0.2126 SouthAfrica Namibia0.0718 SouthAfrica0.118 0.8418 0.1918 0.14*18 0.0432 Zimbabwe0.1318 0.78*18 0.24*18 0.1426 0.2126 NorthAfrica Algeria0.84*18 Tunisia018 0.34*18 0.2927 CentralAfrica CentralAfrican Republic 0.1118 Democratic Republicof theCongo 0.0718 UnitedStates0.54*18 0.14*17 0.1318 0.2*a18 0.04*18 0.91*18 018 0.28*18 0.2719 0.06*33 0.0418 0.32*20 0.3021 a CYP2C9*2/*3combinedfrequency. Login to comment
53 The results from this study strongly suggest that Ghanaian, Table 4 Self-reported ethnicity of 934 Ghanaian samples Tribe N Median Age (Min-Max) Male:Female Akwapim 90 25 (16-51) 51:39 Ashanti 103 23 (17-50) 54:49 Ewe 183 24 (17-51) 97:86 Fanti 160 24 (17-58) 85:75 Ga 183 25 (17-54) 113:70 Other 215 24 (17-57) 135:80 Total 934 24 (16-58) 535:399 Table5AllelefrequenciesofpharmacogeneticallyrelevantpolymorphismsinfiveGhanaiantribes MinoralleleABCB1 3435C4T ABCG2 Q141K CYP2C19 *2 CYP2C9 *2 CYP2C9 *3 CYP3A4 *1B CYP3A5 *3C DPYD *2A GSTP1 I105V NAT2 *14 NAT2 *6 TNFa -238G4A TPMT 719A4G TYMS 1494del UGT1A1 -3156G4A TAATCGGAGAAAGinsTTAAAGA Ghana(total)0.120.010.170.000.000.780.140.000.500.100.270.020.070.440.34 Akwapim0.150.010.160.000.000.730.140.000.490.080.210.020.060.400.33 Ashanti0.090.000.150.000.000.790.110.000.490.100.280.000.070.410.30 Ewe0.130.010.180.000.000.850.130.000.530.110.250.020.070.420.35 Fanti0.130.000.140.000.000.810.120.000.490.100.280.020.080.440.34 Ga0.090.000.180.000.000.750.150.000.490.120.270.030.060.480.35 and other West African populations, may have very different responses, particularly with regard to efficacy and toxicity, to drug treatment regimens that were optimized in US cohorts. Login to comment

[hide] Association of some rare haplotypes and genotype c... Leuk Res. 2008 Aug;32(8):1214-20. Epub 2008 Feb 19. Semsei AF, Erdelyi DJ, Ungvari I, Kamory E, Csokay B, Andrikovics H, Tordai A, Csagoly E, Falus A, Kovacs GT, Szalai C
Association of some rare haplotypes and genotype combinations in the MDR1 gene with childhood acute lymphoblastic leukaemia.
Leuk Res. 2008 Aug;32(8):1214-20. Epub 2008 Feb 19., [PMID:18243305]

Abstract [show]
To investigate their possible roles in disease susceptibility and some disease characteristics we genotyped C3435T and G2677T/A polymorphisms in multidrug resistance-1 (MDR1) gene with a single base extension method and the G34A and C421A polymorphisms of the breast cancer resistance protein gene with an allelic discrimination system in 396 children with acute lymphoblastic leukaemia (ALL) and 192 control patients. While the distribution of individual alleles and genotypes did not differ between patients and controls, there were significant differences in the frequencies of some rare haplotypes and genotype combinations in the MDR1 gene between the two groups.

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29 The two most frequently identified SNPs in the BCRP gene are in exon 2 (G34A, resulting in a V12M change) and exon 5 (C421A, resulting in a Q141K substitution). Login to comment

[hide] Role of P-glycoprotein in accumulation and cytotox... Biochem Pharmacol. 2008 Feb 15;75(4):973-80. Epub 2007 Oct 30. Hira A, Watanabe H, Maeda Y, Yokoo K, Sanematsu E, Fujii J, Sasaki J, Hamada A, Saito H
Role of P-glycoprotein in accumulation and cytotoxicity of amrubicin and amrubicinol in MDR1 gene-transfected LLC-PK1 cells and human A549 lung adenocarcinoma cells.
Biochem Pharmacol. 2008 Feb 15;75(4):973-80. Epub 2007 Oct 30., [PMID:18054347]

Abstract [show]
Amrubicin is a completely synthetic 9-aminoanthracycline agent for the treatment of lung cancer in Japan. The cytotoxicity of C-13 hydroxy metabolite, amrubicinol, is 10 to 100 times greater than that of amrubicin. The transporters responsible for the intracellular pharmacokinetics of amrubicin and amrubicinol remains unclear. This study was aimed to determine whether P-glycoprotein (P-gp) plays functional and preventive role in cellular accumulation and cytotoxicity of amrubicin and its active metabolite amrubicinol by in vitro transport and toxicity experiments. Cytotoxicity and intracellular accumulation of amrubicin and amrubicinol were evaluated by LLC-PK1 cells, MDR1 gene-transfected LLC-PK1 (L-MDR1) cells overexpressing P-gp, and human A549 lung adenocarcinoma cells. L-MDR1 cells showed 6- and 12-fold greater resistance to amrubicin and amrubicinol, respectively, than the parental LLC-PK1 cells. The intracellular accumulation of both drugs in L-MDR1 cells was significantly reduced compared to the LLC-PK1 cells. The basal-to-apical transepithelial transport of both drugs markedly exceeded, whereas the apical-to-basal transport of both drugs was significantly lower in L-MDR1 cells than LLC-PK1 cells. Cyclosporin A (CyA) restored the sensitivity, intracellular accumulation and transport activity for both drugs in L-MDR1 cells. In A549 cells, CyA significantly increased the intracellular accumulation and cytotoxicity of both drugs. These findings indicated that P-gp is responsible for cellular accumulation and cytotoxicity of both amrubicin and amrubicinol, therefore suggesting that the antitumor effect of amrubicin could be affected by the expression level of P-gp in lung cancer cells in chemotherapeutic treatments.

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239 [29] Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, Tsuruo T, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Development of lung cancer before the age of 50: t... Carcinogenesis. 2007 Jun;28(6):1287-93. Epub 2007 Jan 27. Gemignani F, Landi S, Szeszenia-Dabrowska N, Zaridze D, Lissowska J, Rudnai P, Fabianova E, Mates D, Foretova L, Janout V, Bencko V, Gaborieau V, Gioia-Patricola L, Bellini I, Barale R, Canzian F, Hall J, Boffetta P, Hung RJ, Brennan P
Development of lung cancer before the age of 50: the role of xenobiotic metabolizing genes.
Carcinogenesis. 2007 Jun;28(6):1287-93. Epub 2007 Jan 27., [PMID:17259654]

Abstract [show]
The role of genes coding for xenobiotic metabolizing enzymes (XMEs) and the risk of lung cancer is unclear. Under the assumption that these genes may be more important among people having a diagnosis of lung cancer at younger ages, we have investigated the role of single-nucleotide polymorphisms (SNPs) within phase I and phase II XME genes, and also genes involved in the metabolism of nucleic acids in a series of young onset patients and matched controls. We genotyped 299 lung cancer cases diagnosed before the age of 50 and 317 controls, from six countries of Central and Eastern Europe, by use of an oligonucleotide microarray and arrayed primer extension technique for 45 SNPs in 15 phase I XME genes, 46 SNPs in 17 phase II genes and 9 SNPs in 4 genes related to metabolism of nucleic acids. Heterozygote carriers of SNPs in CYP1A2 1545T>C, -164C>A and -740T>G; CYP2A6 -47A>C; MDR1 3435T>C; NAT1 1088T>A and 1095A>C; GSTA2 S112T; GSTM3 V224I and MTHFR A222V had altered risk of developing lung cancer. Phenotypes reconstructed after haplotype analyses showed that the carriers of the combined NAT1 fast+ NAT2 fast phenotypes were at lower risk when compared with those with the combined NAT1 slow + NAT2 slow acetylator phenotypes. Finally, extensive EPHX1 metabolizers showed an increased risk as compared with the poor metabolizers.

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154 Associations between lung cancer risk and SNPs belonging to genes involved in the phase II of xenobiotic metabolism SNP name rs no. Homozygotes common allele Heterozygotes Homozygotes rarer allele Ptrend Ca Co Ca Co ORa 95% CI Ca Co ORa 95% CI ABCG2: Q141K rs2231142 225 242 44 47 0.99 (0.61-1.63) 1 1 2.77 (0.15-52.00) 0.87 ABCG2: V12M rs2231137 171 190 15 10 1.91 (0.77-4.75) 0 1 - - 0.45 ALDH2: 348T.C-MspI rs440 187 202 94 96 0.94 (0.64-1.37) 9 7 1.18 (0.41-3.42) 0.91 ALDH2: 355G.A rs886205 184 204 99 93 1.04 (0.71-1.52) 10 9 0.99 (0.37-2.68) 0.88 ALDH2: 483T.C-HaeIII rs441 187 209 89 94 0.91 (0.62-1.34) 9 7 1.20 (0.41-3.46) 0.85 ALDH2: 69G.A rs4646777 191 213 89 89 0.98 (0.67-1.44) 10 8 1.07 (0.39-2.95) 0.99 COMT: IVS2-98A.G rs6269 123 125 136 134 1.17 (0.80-1.70) 36 57 0.74 (0.43-1.25) 0.52 COMT: 186C.G-L136L rs4818 123 124 138 135 1.17 (0.80-1.70) 34 50 0.76 (0.44-1.31) 0.64 COMT: H62H rs4633 60 77 151 149 1.14 (0.73-1.77) 80 78 1.04 (0.63-1.71) 0.92 COMT: V158M-472G.A rs4680 83 81 144 146 1.07 (0.70-1.62) 59 75 0.98 (0.60-1.62) 0.97 GSTA2: S112T rs1051739 70 90 74 84 1.19 (0.74-1.92) 63 49 1.88 (1.11-3.17) 0.02 GSTA4: 668C.T rs405729 86 87 129 148 1.04 (0.69-1.57) 74 72 1.12 (0.70-1.81) 0.64 GSTA4: Q117Q rs1802061 251 227 24 40 0.56 (0.31-1.01) 5 1 3.94 (0.43-36.38) 0.36 GSTM1: A/B and null rs1065411 82 97 18 15 1.63 (0.73-3.67) 50 42 1.39 (0.80-2.42) 0.24 GSTM3: 3 bp del-Mnl I rs1799735 113 132 41 39 1.29 (0.74-2.26) 2 5 0.30 (0.04-2.13) 0.88 GSTM3: IVS8À30 rs1537234 76 83 120 112 1.19 (0.76-1.85) 48 63 0.88 (0.52-1.50) 0.74 GSTM3: V224I rs7483 149 132 120 142 0.75 (0.52-1.09) 26 40 0.57 (0.32-1.04) 0.04 GSTP1: 313A.G-I105V rs947894 127 133 118 130 0.98 (0.67-1.43) 27 22 1.29 (0.67-2.49) 0.63 GSTP1: 341C.T-A114V rs1799811 244 265 47 47 1.08 (0.67-1.76) 6 1 8.47 (0.87-82.09) 0.20 GSTT2: 153 bp 3# of STP rs2719 60 72 107 132 1.14 (0.71-1.82) 57 54 1.55 (0.89-2.68) 0.13 GSTT2: M139I rs1622002 275 296 19 16 1.05 (0.50-2.21) 0 0 - - 0.89 MDR1: 2677G.T-S892A rs2032582 97 112 134 117 1.16 (0.78-1.73) 53 67 0.97 (0.60-1.58) 0.97 MDR1: 3435T.C rs1045642 62 86 164 141 1.56 (1.02-2.40) 65 81 1.18 (0.72-1.94) 0.50 MDR1: G411G rs1128503 96 106 134 125 1.03 (0.69-1.54) 46 64 0.82 (0.49-1.35) 0.50 MnSOD2: 1183C.T-V16A rs1799725 80 84 118 117 1.15 (0.75-1.77) 57 63 1.00 (0.60-1.66) 0.94 NAT1: 1088T.A rs1057126 190 180 77 101 0.67 (0.45-0.99) 7 12 0.87 (0.31-2.45) 0.08 NAT1: 1095A.C rs15561 155 140 74 104 0.63 (0.42-0.95) 18 22 0.83 (0.40-1.70) 0.10 NAT1: À344C.T rs4986988 219 219 14 17 0.85 (0.38-1.90) 2 1 1.23 (0.09-16.00) 0.80 NAT1: À40A.T rs4986989 248 256 17 23 0.81 (0.40-1.65) 2 1 1.64 (0.13-21.32) 0.75 NAT1: 445G.A-V149I rs4987076 243 259 18 26 0.75 (0.37-1.49) 3 1 1.85 (0.16-22.03) 0.64 NAT1: 459G.A-T153T rs4986990 245 264 17 25 0.78 (0.39-1.56) 2 1 1.62 (0.12-21.42) 0.65 NAT1: 559C.T-R187Stop rs5030839 227 238 1 2 0.90 (0.08-10.51) 0 0 - - 0.93 NAT1: 560G.A-R187Q rs4986782 270 300 5 3 2.17 (0.46-10.26) 0 0 - - 0.33 NAT2: 282C.T-Y94 rs1041983 135 144 114 120 1.13 (0.77-1.65) 34 33 1.28 (0.72-2.26) 0.36 NAT2: 341T.C-I114T rs1801280 69 73 101 96 1.16 (0.72-1.86) 51 47 1.02 (0.58-1.81) 0.86 NAT2: 481C.T-L161L rs1799929 113 105 127 161 0.73 (0.49-1.06) 43 36 1.07 (0.61-1.89) 0.65 NAT2: 590G.A-R197Q rs1799930 141 156 106 106 1.30 (0.88-1.91) 18 24 0.91 (0.46-1.83) 0.56 NAT2: 803A.G-L268R rs1208 96 88 117 139 0.73 (0.48-1.11) 52 64 0.69 (0.42-1.16) 0.13 NAT2: 857G.A-G286E rs1799931 244 258 15 12 1.34 (0.58-3.13) 0 0 - - 0.49 NQO1-DIA4: P187S rs1800566 202 207 75 97 0.74 (0.50-1.10) 17 11 1.89 (0.81-4.38) 0.96 NQO1-DIA4: R139W rs4986998 274 295 23 21 1.16 (0.59-2.27) 0 0 - - 0.67 SULT1A1: M223V rs1801030 282 302 0 1 - - 0 0 - - - SULT1A1: R213H-HaeII rs4149396 91 95 100 96 0.95 (0.61-1.47) 2 11 0.23 (0.05-1.18) 0.30 SULT1A2: 357 bp 3# of STP C.T rs11401 210 212 64 67 1.11 (0.72-1.71) 11 12 1.20 (0.48-2.98) 0.56 UGT1A7: N129K-R131K N/A 113 117 125 139 0.97 (0.65-1.43) 38 42 1.05 (0.61-1.81) 0.94 UGT1A7: W208R rs1126802 118 111 130 143 0.77 (0.52-1.12) 45 59 0.66 (0.39-1.10) 0.08 Ca, cases; Co, controls. ORa odd-ratio adjusted for age, sex, country and tobacco smoking. In bold, statistically significant results (P , 0.05). Login to comment

[hide] Predicting drug sensitivity and resistance: profil... Cancer Cell. 2004 Aug;6(2):129-37. Szakacs G, Annereau JP, Lababidi S, Shankavaram U, Arciello A, Bussey KJ, Reinhold W, Guo Y, Kruh GD, Reimers M, Weinstein JN, Gottesman MM
Predicting drug sensitivity and resistance: profiling ABC transporter genes in cancer cells.
Cancer Cell. 2004 Aug;6(2):129-37., [PMID:15324696]

Abstract [show]
For analysis of multidrug resistance, a major barrier to effective cancer chemotherapy, we profiled mRNA expression of the 48 known human ABC transporters in 60 diverse cancer cell lines (the NCI-60) used by the National Cancer Institute to screen for anticancer activity. The use of real-time RT-PCR avoided artifacts commonly encountered with microarray technologies. By correlating the results with the growth inhibitory profiles of 1,429 candidate anticancer drugs tested against the cells, we identified which transporters are more likely than others to confer resistance to which agents. Unexpectedly, we also found and validated compounds whose activity is potentiated, rather than antagonized, by the MDR1 multidrug transporter. Such compounds may serve as leads for development.

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233 C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K Received: March 12, 2004 protein and low-level drug resistance. Login to comment

[hide] A pharmacogenetic study of docetaxel and thalidomi... Pharmacogenomics J. 2010 Jun;10(3):191-9. Epub 2009 Dec 29. Deeken JF, Cormier T, Price DK, Sissung TM, Steinberg SM, Tran K, Liewehr DJ, Dahut WL, Miao X, Figg WD
A pharmacogenetic study of docetaxel and thalidomide in patients with castration-resistant prostate cancer using the DMET genotyping platform.
Pharmacogenomics J. 2010 Jun;10(3):191-9. Epub 2009 Dec 29., [PMID:20038957]

Abstract [show]
The anticancer agent docetaxel shows significant inter-individual variation in its pharmacokinetic and toxicity profile. Thalidomide is an active anticancer agent and also shows wide pharmacological variation. Past pharmacogenetic research has not explained this variation. Patients with prostate cancer enrolled in a randomized phase II trial using docetaxel and thalidomide versus docetaxel alone were genotyped using the Affymetrix DMET 1.0 platform, which tests for 1256 genetic variations in 170 drug disposition genes. Genetic polymorphisms were analyzed for associations with clinical response and toxicity. In all, 10 single-nucleotide polymorphisms (SNPs) in three genes were potentially associated with response to therapy: peroxisome proliferator-activated receptor-delta (PPAR-delta), sulfotransferase family, cytosolic, 1C, member 2 (SULT1C2) and carbohydrate (chondroitin 6) sulfotransferase 3 (CHST3). In addition, 11 SNPs in eight genes were associated with toxicities to treatment: spastic paraplegia 7 (pure and complicated autosomal recessive) (SPG7), CHST3, cytochrome P450, family 2, subfamily D, polypeptide 6 (CYP2D6), N-acetyltransferase 2 (arylamine N-acetyltransferase) (NAT2), ATP-binding cassette, sub-family C (CFTR/MRP), member 6 (ABCC6), ATPase, Cu++ transporting, alpha polypeptide (ATP7A), cytochrome P450, family 4, subfamily B, polypeptide 1 (CYP4B1) and solute carrier family 10 (sodium/bile acid cotransporter family), member 2 (SLC10A2). Genotyping results between drug metabolizing enzymes and transporters (DMET) and direct sequencing showed >96% of concordance. These findings highlight the role that non-CYP450 metabolizing enzymes and transporters may have in the pharmacology of docetaxel and thalidomide.

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125 Human microsome studies did not find a significant role for CYP2D6 and other CYP enzymes in thalidomide metabolism.54 It is also possible that certain CYP2D6 alleles may confer a multiple-chemical sensitivity phenotype that increases the likelihood of development of toxicities after drug treatment in general.55 Table 5 Concordance between DMET and direct sequencing genotyping Gene SNP Concordance CYP17 A1A1 100 CYP2C19 *2 97 CYP2C19 *3 100 CYP1B1 *3 96 CYP3A5 *3C 100 ABCB1 2677G4T 96 ABCG2 Q141K 100 PPAR-d Rs2076167 100 Abbreviations: DMET, drug-metabolizing enzymes and transporters; SNP, single-nucleotide polymorphism. Login to comment

[hide] ABC transporters in human lymphocytes: expression,... Expert Opin Drug Metab Toxicol. 2010 May;6(5):571-89. Giraud C, Manceau S, Treluyer JM
ABC transporters in human lymphocytes: expression, activity and role, modulating factors and consequences for antiretroviral therapies.
Expert Opin Drug Metab Toxicol. 2010 May;6(5):571-89., [PMID:20367109]

Abstract [show]
IMPORTANCE OF THE FIELD: ATP-binding cassette (ABC) transporters are a superfamily of efflux pumps that transport numerous compounds across cell membranes. These transporters are located in various human tissues including peripheral blood cells, in particular lymphocytes, and present a high variability of expression and activity. This variability may affect the intracellular concentrations and efficacy of drugs acting within lymphocytes, such as antiretroviral drugs. AREAS COVERED IN THIS REVIEW: This review focuses on the current knowledge about the expression, activity, roles and variability of ABC drug transporters in human lymphocytes. The identified modulating factors and their impact on the intracellular pharmacokinetics and efficacy of antiretroviral drugs are also detailed. WHAT THE READER WILL GAIN: Controversial data regarding the expression, activity and sources of variability of ABC transporters in lymphocytes are discussed. The modulating factors and their pharmacological consequences regarding antiretroviral therapies are also provided. TAKE HOME MESSAGE: Numerous studies have reported conflicting results regarding the expression and activity of ABC drug transporters in lymphocytes. Despite these discrepancies, which may partly result from heterogeneous analytical methods, ABCC1 appears to have the highest expression in lymphocytes and may thus play a predominant role in the resistance to antiretroviral drugs, particularly to protease inhibitors.

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204 Two common non-synonymous substitutions have been particularly investigated in various tissues, the 34 G > A (V12 M, 4% of Caucasians) and 421C > A (Q141K, 11% of Caucasians) polymorphisms. Login to comment
578 Imai Y, Nakane M, Kage K, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Genetic polymorphisms of drug-metabolising enzymes... Clin Pharmacokinet. 2006;45(3):253-85. Bosch TM, Meijerman I, Beijnen JH, Schellens JH
Genetic polymorphisms of drug-metabolising enzymes and drug transporters in the chemotherapeutic treatment of cancer.
Clin Pharmacokinet. 2006;45(3):253-85., [PMID:16509759]

Abstract [show]
There is wide variability in the response of individuals to standard doses of drug therapy. This is an important problem in clinical practice, where it can lead to therapeutic failures or adverse drug reactions. Polymorphisms in genes coding for metabolising enzymes and drug transporters can affect drug efficacy and toxicity. Pharmacogenetics aims to identify individuals predisposed to a high risk of toxicity and low response from standard doses of anti-cancer drugs. This review focuses on the clinical significance of polymorphisms in drug-metabolising enzymes (cytochrome P450 [CYP] 2C8, CYP2C9, CYP2C19, CYP2D6, CYP3A4, CYP3A5, dihydropyrimidine dehydrogenase, uridine diphosphate glucuronosyltransferase [UGT] 1A1, glutathione S-transferase, sulfotransferase [SULT] 1A1, N-acetyltransferase [NAT], thiopurine methyltransferase [TPMT]) and drug transporters (P-glycoprotein [multidrug resistance 1], multidrug resistance protein 2 [MRP2], breast cancer resistance protein [BCRP]) in influencing efficacy and toxicity of chemotherapy. The most important example to demonstrate the influence of pharmacogenetics on anti-cancer therapy is TPMT. A decreased activity of TPMT, caused by genetic polymorphisms in the TPMT gene, causes severe toxicity with mercaptopurine. Dosage reduction is necessary for patients with heterozygous or homozygous mutation in this gene. Other polymorphisms showing the influence of pharmacogenetics in the chemotherapeutic treatment of cancer are discussed, such as UGT1A1*28. This polymorphism is associated with an increase in toxicity with irinotecan. Also, polymorphisms in the DPYD gene show a relation with fluorouracil-related toxicity; however, in most cases no clear association has been found for polymorphisms in drug-metabolising enzymes and drug transporters, and pharmacokinetics or pharmacodynamics of anti-cancer drugs. The studies discussed evaluate different regimens and tumour types and show that polymorphisms can have different, sometimes even contradictory, pharmacokinetic and pharmacodynamic effects in different tumours in response to different drugs. The clinical application of pharmacogenetics in cancer treatment will therefore require more detailed information of the different polymorphisms in drug-metabolising enzymes and drug transporters. Larger studies, in different ethnic populations, and extended with haplotype and linkage disequilibrium analysis, will be necessary for each anti-cancer drug separately.

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2327 Characteristics of drug transporter genes (with the most important polymorphisms)[167-169] Gene Location Protein Exons Amino Polymorphisms Location Effect (chromosome) acids (exon) ABCB1 7q21 P-gp/MDR1 29 1280 *6 (C3435T) 26 Silent *7 (G2677T/A) 21 A893S *8 (C1236T) 12 Silent G1199A 11 S400N ABCC2 10q24 MRP2 32 1545 C-24T 5'-UTR Unknown C1249A 10 V417I C2302T 18 R768W C2366T 18 S789F T2439+2C 18 Splice site ND 26 W1254Y,A,C,F C3972T 28 Silent A4145G 29 Q1382R G4348A 31 G1440S ABCG2 4q22 BCRP 16 655 G34A 2 V12M C8825A 5 Q141K BCRP = breast cancer resistance protein; MDR1 = multidrug resistance 1; MRP2 = multidrug resistance protein 2; ND = no data; P-gp = P-glycoprotein. Login to comment
2381 [168] Overall, no clinical effects have been observed topoisomerase I inhibitor, of cells expressing V12M for the different MRP2 genotypes and pharmacoki- or Q141K was less than one-tenth compared with netics and pharmacodynamics of anti-cancer drugs. Login to comment
2385 In a study by Imai et al.[213] Japanese volunteers The ABCG2 gene is a recently described member with the mutant C8825A polymorphism (allele fre- of the G subfamily of ABC transporters that was quency of 46%) expressed a low amount of the previously called BCRP, mitoxantrone resistant Q141K ABCG2 protein. Login to comment
2400 polymorphisms G34A and C8825A, leading to an amino acid change of V12M and Q141K, respec- The levels of ABCG2 mRNA expression in cell tively (see table VII), on the transporter function of lines were found to be significantly correlated with the protein. Login to comment
2868 Kauffmann HM, Keppler D, Kartenbeck J, et al. Induction of low expression of Q141K protein and low-level drug resis- cMRP/cMOAT gene expression by cisplatin, 2-acetylami- tance. Login to comment

[hide] Genetic polymorphisms and gene-dosage effect in ov... J Exp Clin Cancer Res. 2015 Jan 16;34:2. doi: 10.1186/s13046-015-0124-y. Tecza K, Pamula-Pilat J, Kolosza Z, Radlak N, Grzybowska E
Genetic polymorphisms and gene-dosage effect in ovarian cancer risk and response to paclitaxel/cisplatin chemotherapy.
J Exp Clin Cancer Res. 2015 Jan 16;34:2. doi: 10.1186/s13046-015-0124-y., [PMID:25591549]

Abstract [show]
BACKGROUND: Ovarian malignancies are often diagnosed in advanced stage and at the same time resistance to treatment, both intrinsic and developed during treatment, is sometimes observed. These facts underscore the need for new markers of ovarian cancer risk, as well as markers of treatment effectiveness. METHODS: In this study we genotyped 225 ovarian cancer patients, 64 breast and ovarian cancer patients and 348 healthy controls. In total, 12 polymorphic variants and 2 deletions in PGR, ABCB1, ABCG2, GSTT1, GSTM1, GSTP1, ATM, TP53 and ATP7B genes were analyzed using ASA-PCR, RFLP-PCR, multiplex-PCR and sequencing. RESULTS: Ten genetic polymorphisms were significantly associated with the risk of developing ovarian carcinoma in at least one of the groups under study. Impact of PGR gene polymorphisms on ovarian cancer risk was specific only for the group of the BRCA1 mutation carriers (in presence of p.Val660Leu variant- OR 2,82; p = 0,010), which confirms the difference in modulation of ovarian cancer risk between sporadic and hereditary malignancies, including the breast-ovarian cancer group (as a cancer-prone group). The analyses showed also the importance of ATP7B gene in ovarian carcinogenesis, both studied variants of which significantly modulated the ovarian cancer risk in all groups excluding the group with BRCA1 mutation. Cumulative risk analysis revealed 3 unfavorable variants that increased significantly the risk of developing ovarian cancer (p.Ile1145 = ABCB1+ p.Asp1853Asn ATM+ p.Ser406Ala ATP7B- OR 7,47; p = 0,002) and significantly modified the progression free survival (PFS) of the patients, and also two favorable genotypes which protected against ovarian cancer (p.Arg952Lys ATP7B+ p.Arg72Pro TP53- OR 0,50; p = 0,008). PFS analysis for carriers of favorable versus unfavorable genotypes emphasized the impact of the regulation of cell cycle (p.Asp1853Asn ATM) and active transport of xenobiotics (p.Ser894Ala/Thr ABCB1) on the risk of disease progression (HR 3,81; p = 0,010) after paclitaxel/cisplatin chemotherapy. CONCLUSIONS: The unfavorable genetic variants could facilitate carcinogenic process and once their carriers developed malignancy, their chances of survival were smaller. Our analyses also showed a strong gene-dosage effect with the decrease of progression-free survival for the carriers of two unfavorable genetic factors.

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61 Genotyping of polymorphic variants in PGR (rs10895068 and p.Val660Leu), ABCB1 (p.Ile1145 = and p.Ser893Ala/ Trp), ABCG2 (p.Gln141Lys), ATM (p.Asp1853Asn), TP53 (p.Arg72Pro), GSTP1 (p.Ile105Val) genes, as well as detection of GSTT1/M1 gene deletions, were performed as described previously [9-17]. Login to comment
101 The polymorphisms, which in univariate analysis were modulating ovarian cancer risk, were included in the Table 2 Case-control analyses of ovarian and breast and ovarian cancer risk Ovarian cancer all patients Ovarian cancer BRCA1- Ovarian cancer BRCA1+ Breast and ovarian cancer Gene polymorphism Genotype Controls n(%) n(%) OR (&#b1;95% CI) p n(%) OR (&#b1;95% CI) p n(%) OR (&#b1;95% CI) p n(%) OR (&#b1;95% CI) p PGR p.Val660Leu rs1042838 GG 239 (69.3) 143 (63.9) 1(ref) 131 (66.5) 1(ref) 12 (44.4) 1(ref) 46 (75.4) 1 (ref) GT 95 (27.5) 74 (33.0) 1.03 (0.90-1.88) 0.160 60 (30.5) 1.15 (0.78-1.70) 0.473 14 (51.9) 2.94 (1.31-6.58) 0.009 14 (23.0) 0.77 (0.40-1.46) 0.416 TT 11 (3.2) 7 (3.1) 1.06 (0.40-2.81) 0.901 6 (3.0) 1.00 (0.36-2.75) 0.993 1 (3.70) 1.81 (0.22-15.20) 0.584 1 (1.6) 0.47 (0.06-3.75) 0.478 GT + TT 106 (30.7) 81 (36.1) 1.20 (0.88-1.63) 0.249 66 (33.5) 1.14 (0.78-1.65) 0.504 15 (55.6) 2.82 (1.28-6.23) 0.010 15 (24.6) 0.74 (0.39-1.38) 0.336 ABCB1 p.Ser893Ala p.Ser893Thr rs2032582 GG 117 (33.7) 65 (29.0) 1(ref) 56 (28.4) 1(ref) 9 (33.4) 1(ref) 16 (26.2) 1 (ref) GT 156 (45.0) 115 (51.4) 1.33 (0.90-1.95) 0.152 104 (52.8) 1.39 (0.93-2.09) 0.108 11 (40.7) 0.92 (0.37-2.28) 0.852 33 (54.1) 1.55 (0.71-2.94) 0.184 TT 60 (17.3) 37 (16.5) 1.11 (0.67-1.85) 0.688 32 (16.3) 1.11 (0.65-1.90) 0.691 5 (18.5) 1.08 (0.35-3.38) 0.890 9 (14.8) 1.10 (0.46-2.63) 0.836 GA 9 (2.6) 2 (0.9) 0.40 (0.08-1.91) 0.250 2 (1.0) 0.46 (0.10-2.24) 0.337 - - - 2 (3.3) 1.63 (0.32-8.31) 0.557 TA 5 (1.4) 5 (2.2) 1.80 (0.50-6.45) 0.367 3 (1.5) 1.25 (0.29-5.49) 0.763 2 (7.4) 5.20 (0.87-31.18) 0.069 1 (1.6) 1.46 (0.16-13.6) 0.736 AA - - - - - - - - - - - - - GG + GT + GA 282 (81.3) 182 (81.3) 1 (ref) 162 (82.2) 1 (ref) 20 (74.1) 1 (ref) 51 (83.6) 1 (ref) TT + TA 65 (18.7) 42 (18.7) 1.00 (0.92-1.09) 1.000 35 (17.8) 0.94 (0.60-1.48) 0.780 7 (25.9) 1.52 (0.61-3.76) 0.364 10 (16.4) 0.85 (0.41-1.77) 0.664 ABCB1 p.Ile1145= rs1045642 CC 83 (24.0) 44 (19.6) 1(ref) 35 (17.8) 1(ref) 9 (33.3) 1(ref) 16 (26.2) 1 (ref) CT 162 (47.0) 122 (54.5) 1.42 (0.92-2.19) 0.113 112 (56.8) 1.64 (1.03-2.60) 0.036 10 (37.1) 0.57 (0.22-1.46) 0.239 26 (42.6) 0.83 (0.42-1.64) 0.596 TT 100 (29.0) 58 (25.9) 1.09 (0.67-1.78) 0.718 50 (25.4) 1.18 (0.70-2.00) 0.522 8 (29.6) 0.74 (0.27-2.00) 0.549 19 (31.2) 0.98 (0.48-2.04) 0.969 CT + TT 262 (76.0) 180 (80.4) 1.02 (0.81-1.30) 0.827 162 (82.2) 1.47 (0.94-2.28) 0.089 18 (66.7) 0.63 (0.27-1.46) 0.285 45 (73.8) 0.89 (0.48-1.66) 0.716 ABCG2 p. Gln141Lys rs2231142 CC 276 (80.2) 191 (86.4) 1 (ref) 167 (85.6) 1 (ref) 24 (92.3) 1 (ref) 56 (87.5) 1 (ref) CA 68 (19.8) 30 (13.6) 0.64 (0.40-1.02) 0.059 28 (14.4) 0.68 (0.42-1.10) 0.116 2 (7.7) 0.34 (0.08-0.47) 0.147 8 (12.5) 0.58 (0.26-1.28) 0.175 ATM p. Asp1853Asn rs1801516 GG 254 (75.8) 153 (68.6) 1 (ref) 134 (68.4) 1 (ref) 19 (70.4) 1 (ref) 45 (70.3) 1 (ref) GA 76 (22.7) 64 (28.7) 1.40 (0.95-2.06) 0.091 57 (29.1) 1.42 (0.95-2.13) 0.083 7 (25.9) 1.23 (0.50-3.05) 0.651 15 (23.4) 1.11 (0.59-2.11) 0.740 AA 5 (1.5) 6 (2.7) 1.99 (0.60-6.66) 0.262 5 (2.5) 1.90 (0.54-6.69) 0.319 1 (3.7) 2.67 (0.39-24.29) 0.380 4 (6.3) 4.52 (1.16-17.56) 0.029 GA + AA 81 (24.2) 70 (31.4) 1.43 (0.98-2.09) 0.061 62 (31.6) 1.45 (0.98-2.15) 0.062 8 (29.6) 1.32 (0.73-2.40) 0.352 19 (29.7) 1.32 (0.56-3.14) 0.528 TP53 p.Arg72Pro rs1042522 GG 167 (49.0) 130 (57.8) 1 (ref) 115 (58.1) 1 (ref) 15 (55.6) 1 (ref) 29 (48.3) 1 (ref) GC 150 (44.0) 79 (35.1) 0.68 (0.47-0.97) 0.031 70 (35.3) 0.68 (0.47-0.98) 0.039 9 (33.3) 0.67 (0.28-1.57) 0.355 29 (48.3) 1.11 (0.64-1.95) 0.707 CC 24 (7.0) 16 (7.1) 0.86 (0.44-1.68) 0.652 13 (6.6) 0.79 (0.39-1.61) 0.511 3 (11.1) 1.39 (0.38-5.16) 0.621 2 (3.3) 0.48 (0.11-2.14) 0.336 GC + CC 174 (51.0) 95 (42.2) 0.80 (0.61-1.05) 0.104 83 (41.9) 0.69 (0.49-0.99) 0.042 12 (44.4) 0.77 (0.35-1.69) 0.511 31 (51.67) 1.03 (0.59-1.78) 0.927 ATP7B p.Ser406Ala rs1801243 TT 103 (30.8) 41 (19.0) 1 (ref) 35 (18.5) 1 (ref) 6 (22.2) 1 (ref) 13 (20.3) 1 (ref) TG 157 (47.0) 113 (52.3) 1.81 (1.17-2.80) 0.008 100 (52.9) 1.87 (1.18-2.97) 0.007 13 (48.2) 1.42 (0.52-3.88) 0.490 32 (50.0) 1.61 (0.81-3.23) 0.174 GG 74 (22.2) 62 (28.7) 2.10 (1.28-3.46) 0.003 54 (28.6) 2.15 (1.27-3.62) 0.004 8 (29.6) 1.86 (0.61-5.61) 0.270 19 (29.7) 2.03 (0.94-4.40) 0.069 TG + GG 231 (69.2) 175 (81.0) 1.90 (1.26-2.88) 0.002 154 (81.5) 1.96 (1.27-3.03) 0.002 21 (77.8) 1.56 (0.61-3.99) 0.352 51 (79.7) 1.75 (0.91-3.36) 0.093 Table 2 Case-control analyses of ovarian and breast and ovarian cancer risk (Continued) ATP7B p.Arg952Lys rs732774 AA 103 (30.7) 86 (38.9) 1 (ref) 76 (39.2) 1 (ref) 10 (37.0) 1 (ref) 25 (39.7) 1 (ref) AG 159 (47.5) 96 (43.4) 0.72 (0.49-1.06) 0.096 82 (42.3) 0.70 (0.47-1.04) 0.078 14 (51.9) 0.91 (0.39-2.11) 0.821 31 (49.2) 0.80 (0.45-1.44) 0.471 GG 73 (21.8) 39 (17.7) 0.64 (0.39-1.04) 0.070 36 (18.5) 0.67 (0.41-1.10) 0.112 3 (11.1) 0.42 (0.11-1.61) 0.203 7 (11.1) 0.40 (0.16-0.97) 0.041 AG + GG 232 (69.3) 135 (61.1) 0.70 (0.49-1.00) 0.047 118 (60.8) 0.69 (0.48-1.00) 0.049 17 (63.0) 0.75 (0.33-1.71) 0.499 38 (60.3) 0.67 (0.39-1.18) 0.165 GST T1/M1 Gene deletions wt/wt 118 (34.6) 82 (36.8) 1(ref) 72 (36.7) 1(ref) 10 (37.0) 1(ref) 20 (31.3) 1(ref) wt/null 49 (14.4) 18 (8.1) 0.53 (0.29-0.98) 0.040 17 (8.7) 0.57 (0.30-1.07) 0.076 1 (3.7) 0.24 (0.03-1.96) 0.180 6 (9.4) 0.72 (0.27-1.92) 0.512 null/wt 138 (40.5) 95 (42.6) 0.99 (0.64-1.54) 0.967 82 (41.8) 0.97 (0.65-0.47) 0.90 13 (48.2) 1.11 (0.47-2.64) 0.810 31 (48.4) 1.32 (0.72-2.45) 0.368 null/null 36 (10.5) 28 (12.5) 1.12 (0.63-1.98) 0.698 25 (12.8) 1.14 (0.63-2.06) 0.667 3 (11.1) 0.98 (0.24-4.04) 0.982 7 (10.9) 1.14 (0.45-2.95) 0.774 GSTP1 p.Ile105Val rs1695 AA 151 (45.2) 104 (46.4) 1 (ref) 93 (47.2) 1 (ref) 11 (40.7) 1 (ref) 32 (52.4) 1 (ref) AG 162 (48.5) 95 (42.4) 0.85 (0.60-1.22) 0.376 84 (42.6) 0.84 (0.58-1.22) 0.361 11 (40.7) 0.93 (0.39-2.22) 0.873 36 (36.1) 0.64 (0.36-1.15) 0.137 GG 21 (6.3) 25 (11.2) 1.73 (0.92-3.26) 0.090 20 (10.2) 1.54 (0.79-3.01) 0.199 5 (18.6) 3.27 (1.03-10.42) 0.044 7 (11.5) 1.57 (0.61-4.03) 0.342 AG + GG 183 (54.8) 120 (53.6) 0.95 (0.68-1.34) 0.776 104 (52.8) 0.92 (0.65-1.32) 0.656 16 (49.3) 1.20 (0.54-2.67) 0.653 43 (47.6) 0.75 (0.43-1.29) 0.296 Statistically significant analyses are in bold. groups of risk reductors (favorable genotypes) or risk enhancers (unfavorable genotypes). 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77 Polymorphism p.Gln141Lys in the ABCG2 gene, borderline significant (OR 0.64; 95% CI 0.40-1.02; p = 0.059), decreased the risk of ovarian cancer for heterozygotes. Login to comment
138 Also, in heterozygotes for ABCG2 polymorphism p.Gln141Lys we found a trend towards decreased risk of ovarian cancer. Login to comment

[hide] The epidermal growth factor tyrosine kinase inhibi... Oncol Rep. 2009 Feb;21(2):483-9. Shi Z, Parmar S, Peng XX, Shen T, Robey RW, Bates SE, Fu LW, Shao Y, Chen YM, Zang F, Chen ZS
The epidermal growth factor tyrosine kinase inhibitor AG1478 and erlotinib reverse ABCG2-mediated drug resistance.
Oncol Rep. 2009 Feb;21(2):483-9., [PMID:19148526]

Abstract [show]
ABCG2 is an important member of ATP-binding cassette (ABC) transporter shown to confer drug resistance in cancer cells. Recent studies show that an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), gefitinib, is able to modulate the function of ABCG2 and reverse ABCG2-mediated multidrug resistance (MDR) in cancer cells. Additionally, ABCG2 expression has been shown to impact treatment efficacy and development of side-effects in patients receiving gefitinib. However, it is unclear whether other EGFR TKIs interact with ABCG2 in a similar manner. In the present study, we investigated the interaction of two other EGFR TKIs, AG1478 and erlotinib, with ABCG2. Our data show that AG1478 and erlotinib potently sensitized drug-resistant cells overexpressing either wild-type or mutated ABCG2 to the ABCG2 substrate anti-cancer drugs flavopiridol and mitoxantrone. Neither AG1478 nor erlotinib sensitized ABCG2-overexpressing cells to drugs that are not substrates of ABCG2 nor did they impact drug sensitivity of parental cells. Furthermore, AG1478 and erlotinib were able to significantly enhance the intracellular accumulation of mitoxantrone in cells expressing either wild-type or mutated ABCG2. Additionally, they did not alter the protein expression of ABCG2 in the ABCG2-overexpressing cells. Taken together, we conclude that AG1478 and erlotinib potently reverse ABCG2-mediated MDR through directly inhibiting the drug efflux function of ABCG2 in the ABCG2-overexpressing cells. These results will be useful in the development of novel and more effective EGFR TKIs as well as the development of combinational chemotherapeutic strategies.

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149 One common single-nucleotide polymorphism of ABCG2 that has been shown to affect the function of ABCG2, Q141K (421Cėd;A), was reported to be associated with gefitinib-induced diarrhea, resulting in a high risk of diarrhea in patients treated with oral gefitinib (36), and was also found to be associated with greater accumulation at steady-state and toxicity of gefitinib (37). Login to comment

[hide] Pharmacokinetics of intravenous paracetamol in eld... Clin Pharmacokinet. 2011 Feb;50(2):121-9. doi: 10.2165/11537240-000000000-00000. Liukas A, Kuusniemi K, Aantaa R, Virolainen P, Niemi M, Neuvonen PJ, Olkkola KT
Pharmacokinetics of intravenous paracetamol in elderly patients.
Clin Pharmacokinet. 2011 Feb;50(2):121-9. doi: 10.2165/11537240-000000000-00000., [PMID:21241071]

Abstract [show]
BACKGROUND AND OBJECTIVES: Intravenous paracetamol (N-acetyl-paraminophenol, acetaminophen) is a widely used nonopioid analgesic which has become popular in the treatment of pain in many patient groups, including the elderly. Although intravenous paracetamol has been studied widely in clinical analgesia studies, there is little information on its pharmacokinetics in the elderly. We designed this study to determine the pharmacokinetics of intravenous paracetamol in very old patients and to compare them with that of younger patients. We also considered the effect of adenosine triphosphate-binding cassette G2 protein (ABCG2) genotype and renal function on paracetamol pharmacokinetics in these patients. METHODS: We compared the pharmacokinetics of intravenous paracetamol in four groups of ten patients, aged 20-40, 60-70, 70-80 and 80-90 years, undergoing orthopaedic surgery. Paracetamol 1000 mg was given by infusion over 15 minutes. Plasma concentrations of paracetamol and its glucuronide and sulphate conjugates were measured for 24 hours with a high-performance liquid chromatographic method and ABCG2 genotype was determined. Glomerular filtration rate (GFR) was estimated from age, sex and serum creatinine of the patient. RESULTS: In the group aged 80-90 years, the mean value of the area under the plasma concentration-time curve extrapolated to infinity (AUC(infinity)) of paracetamol was 54-68% higher than in the two youngest groups. Paracetamol clearance showed a statistically significant dependence on age group, whereas volume of distribution during elimination and elimination half-life were associated with age group and sex, respectively. Based on mean AUC(infinity) of paracetamol glucuronide and paracetamol sulphate, the oldest patients had 1.3- to 1.5-fold greater exposure to these metabolites than patients aged 20-40 years. ABCG2 genotype did not affect paracetamol pharmacokinetics. There was a linear correlation between the values of AUC(infinity) of paracetamol, its glucuronide and sulphate metabolites and GFR. CONCLUSION: Age and sex are important factors affecting the pharmacokinetics of paracetamol. The higher the age of the patient, the higher is the exposure to paracetamol. Female sex is associated with increased paracetamol concentrations but ABCG2 genotype does not seem to affect paracetamol pharmacokinetics. Trial registration number (EudraCT): 2006-001917-14.

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55 The subjects were genotyped for the ABCG2c.421C>A singlenucleotide polymorphism (p.Gln141Lys; rs2231142) by allelic discrimination with a TaqMan Drug Metabolism Genotyping Assay (assay ID: C__15854163_70; Applied Biosystems, Foster City, CA, USA). Login to comment

[hide] Analysis of the effect of the bovine adenosine tri... J Anim Sci. 2011 Dec;89(12):4325-38. doi: 10.2527/jas.2011-3841. Epub 2011 Aug 5. Real R, Gonzalez-Lobato L, Baro MF, Valbuena S, de la Fuente A, Prieto JG, Alvarez AI, Marques MM, Merino G
Analysis of the effect of the bovine adenosine triphosphate-binding cassette transporter G2 single nucleotide polymorphism Y581S on transcellular transport of veterinary drugs using new cell culture models.
J Anim Sci. 2011 Dec;89(12):4325-38. doi: 10.2527/jas.2011-3841. Epub 2011 Aug 5., [PMID:21821808]

Abstract [show]
In commercial dairy production, the risk of drug residues and environmental pollutants in milk from ruminants has become an outstanding problem. One of the main determinants of active drug secretion into milk is the ATP-binding cassette transporter G2/breast cancer resistance protein (ABCG2/BCRP). It is located in several organs associated with drug absorption, metabolism, and excretion, and its expression is highly induced during lactation in the mammary gland of ruminants, mice, and humans. As a consequence, potential contamination of milk could expose suckling infants to xenotoxins. In cows, a SNP for this protein affecting quality and quantity of milk production has been described previously (Y581S). In this study, our main purpose was to determine whether this polymorphism has an effect on transcellular transport of veterinary drugs because this could alter substrate pharmacokinetics and milk residues. We stably expressed the wild-type bovine ABCG2 and the Y581S variant in Madin-Darby canine kidney epithelial cells (MDCKII) and MEF3.8 cell lines generating cell models in which the functionality of the bovine transporter could be addressed. Functional studies confirmed the greater functional activity in mitoxantrone accumulation assays for the Y581S variant with a greater relative V(MAX) value (P = 0.040) and showed for the first time that the Y581S variant presents greater transcellular transport of the model ABCG2 substrate nitrofurantoin (P = 0.024) and of 3 veterinary antibiotics, the fluoroquinolone agents enrofloxacin (P = 0.035), danofloxacin (P = 0.001), and difloxacin (P = 0.008), identified as new substrates of the bovine ABCG2. In addition, the inhibitory effect of the macrocyclic lactone ivermectin on the activity of wild-type bovine ABCG2 and the Y581S variant was also confirmed, showing a greater inhibitory potency on the wild-type protein at all the concentrations tested (5 muM, P = 0.017; 10 muM, P = 0.001; 25 muM, P = 0.008; and 50 muM, P = 0.003). Differential transport activity depending on the genotype together with the differential inhibition pattern might have clinical consequences, including changes in substrate pharmacokinetics (and subsequently pharmacodynamics) and more specifically, changes in secretion of ABCG2 substrates into milk, potentially implying important consequences to veterinary therapeutics.

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217 As an example, the Q141K variant in humans has been related to altered pharmacokinetics of several substrates (Cusatis and Sparreboom, 2008; Keskitalo et al., 2009), and more recently, it has been reported as a loss-of-function mutation that causes hyperuricemia and gout in humans (Woodward et al., 2009). Login to comment

[hide] Intestinal drug transporters: an overview. Adv Drug Deliv Rev. 2013 Oct;65(10):1340-56. doi: 10.1016/j.addr.2012.09.042. Epub 2012 Oct 4. Estudante M, Morais JG, Soveral G, Benet LZ
Intestinal drug transporters: an overview.
Adv Drug Deliv Rev. 2013 Oct;65(10):1340-56. doi: 10.1016/j.addr.2012.09.042. Epub 2012 Oct 4., [PMID:23041352]

Abstract [show]
The importance of drug transporters as one of the determinants of pharmacokinetics has become increasingly evident. While much research has been conducted focusing the role of drug transporters in the liver and kidney less is known about the importance of uptake and efflux transporters identified in the intestine. Over the past years the effects of intestinal transporters have been studied using in vivo models, in situ organ perfusions, in vitro tissue preparations and cell lines. This review aims to describe up to date findings regarding the importance of intestinal transporters on drug absorption and bioavailability, highlighting areas in need of further research. Wu and Benet proposed a Biopharmaceutics Drug Disposition Classification System (BDDCS) that allows the prediction of transporter effects on the drug disposition of orally administered drugs. This review also discusses BDDCS predictions with respect to the role of intestinal transporters and intestinal transporter-metabolizing enzyme interplay on oral drug pharmacokinetics.

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1150 Recent studies have demonstrated that individuals with reduced BCRP expression levels (Q141K variant) are at increased risk for gefitinib-induced diarrhea and altered pharmacokinetics of 9-aminocamptothecin, diflomotecan, irinotecan, rosuvastatin, sulfasalazine and topotecan [1]. Login to comment

[hide] Influence of enzyme and transporter polymorphisms ... Ann Oncol. 2013 Mar;24(3):756-60. doi: 10.1093/annonc/mds532. Epub 2012 Oct 31. Seong SJ, Lim M, Sohn SK, Moon JH, Oh SJ, Kim BS, Ryoo HM, Chung JS, Joo YD, Bang SM, Jung CW, Kim DH, Park SY, Yoon SS, Kim I, Lee HG, Won JH, Min YH, Cheong JW, Park JS, Eom KS, Hyun MS, Kim MK, Kim H, Park MR, Park J, Kim CS, Kim HJ, Kim YK, Park EK, Zang DY, Jo DY, Lee HW, Yoon YR
Influence of enzyme and transporter polymorphisms on trough imatinib concentration and clinical response in chronic myeloid leukemia patients.
Ann Oncol. 2013 Mar;24(3):756-60. doi: 10.1093/annonc/mds532. Epub 2012 Oct 31., [PMID:23117072]

Abstract [show]
BACKGROUND: This study explored the impact of genetic polymorphisms in cytochrome P450 (CYP) enzymes and transporters on the plasma trough concentration of imatinib mesylate (IM) and clinical response in chronic myeloid leukemia (CML). PATIENTS AND METHODS: In total, 82 patients with CML who had been administered 400 mg IM daily for over 6 months were genotyped for 11 single-nucleotide polymorphisms in nine genes (CYP3A4, CYP3A5, CYP2C9, CYP2C19, CYP2D6, ABCB1, SLC22A1, SLC22A2 and ABCG2) using blood samples. The trough imatinib concentration and clinical responses were assessed 6 months after the initiation of IM therapy. RESULTS: The CC, CA and AA genotypes in ABCG2 421C>A gave significantly different frequencies for the major molecular response (MMR) (P = 0.02). However, no significant differences were found between the genotypes of the CYP enzymes and transporters identified in this study and the imatinib plasma trough concentrations and clinical response frequencies, except for the correlation of ABCG2 with MMR. CONCLUSIONS: The results of the present study may indicate that the ABCG 421C>A genetic polymorphism influences the MMR of imatinib in patients with CML.

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66 discussion ABCG2 421C>A in exon 5 is a non-synonymous SNP, where a glutamine is substituted with a lysine residue at codon 141 (Q141K). Login to comment
70 It has been reported that human embryonic kidney 293 cells transfected with ABCG2 Q141K showed a higher imatinib accumulation compared with wild-type ABCG2 in vitro [14]. Login to comment

[hide] Expression levels of the ABCG2 multidrug transport... PLoS One. 2012;7(11):e48423. doi: 10.1371/journal.pone.0048423. Epub 2012 Nov 15. Kasza I, Varady G, Andrikovics H, Koszarska M, Tordai A, Scheffer GL, Nemeth A, Szakacs G, Sarkadi B
Expression levels of the ABCG2 multidrug transporter in human erythrocytes correspond to pharmacologically relevant genetic variations.
PLoS One. 2012;7(11):e48423. doi: 10.1371/journal.pone.0048423. Epub 2012 Nov 15., [PMID:23166586]

Abstract [show]
We have developed a rapid, simple and reliable, antibody-based flow cytometry assay for the quantitative determination of membrane proteins in human erythrocytes. Our method reveals significant differences between the expression levels of the wild-type ABCG2 protein and the heterozygous Q141K polymorphic variant. Moreover, we find that nonsense mutations on one allele result in a 50% reduction in the erythrocyte expression of this protein. Since ABCG2 polymorphisms are known to modify essential pharmacokinetic parameters, uric acid metabolism and cancer drug resistance, a direct determination of the erythrocyte membrane ABCG2 protein expression may provide valuable information for assessing these conditions or for devising drug treatments. Our findings suggest that erythrocyte membrane protein levels may reflect genotype-dependent tissue expression patterns. Extension of this methodology to other disease-related or pharmacologically important membrane proteins may yield new protein biomarkers for personalized diagnostics.

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1 Our method reveals significant differences between the expression levels of the wild-type ABCG2 protein and the heterozygous Q141K polymorphic variant. Login to comment
27 Recently, a significant disease-association for a polymorphic ABCG2 variant (resulting in ABCG2-Q141K) has been observed in gout [24,25,26,27,28]. Login to comment
29 A common variant of ABCG2 (c.421C.A; Q141K), with a variable allele frequency between 5-30% in various ethnic groups (see ref. [29]), was shown to decrease membrane protein expression in model PLOS ONE | www.plosone.org 1 November 2012 | Volume 7 | Issue 11 | e48423 Hungarian Academy of Sciences ( ) cells, despite unchanged mRNA levels [30,31,32,33,34]. Login to comment
30 Still, a lower expression level of the ABCG2-Q141K variant has not been confirmed at physiologically relevant sites, given the difficulties in obtaining and processing human tissues. Login to comment
34 In this report we show that individuals heterozygous for the potentially miss-processed ABCG2 variant (Q141K) have significantly lower ABCG2 protein expression in their red cells than individuals carrying the wild-type ABCG2 gene. Login to comment
67 The most common SNPs in ABCG2 [V12M (c.34G.A, p.12Val.Met in exon 2, SNP database ID: rs2231137) and Q141K (c.421C.A, p.141Gln.Lys in exon 5, SNP database ID: rs2231142] were genotyped using the LightCycler480 (Roche Diagnostics, Basle, Switzerland) allelic discrimination system as described previously in detail [45]. Login to comment
89 For this purpose we quantified the expression of the erythrocyte ABCG2 in 47 unrelated, healthy individuals that were also screened for the presence of two most prevalent ABCG2 polymorphic variants found in the Caucasian population (V12M and Q141K) [29]. Login to comment
101 However, when the samples were grouped according to their genotypes, we found that the red blood cells of individuals carrying the heterozygous Q141K variant exhibited significantly lower expression of ABCG2 (5.2761.19), as compared to homozygous wild-type individuals (6.1360.61, p = 0.011) (Fig. 3). Login to comment
103 As a summary of the ABCG2 polymorphism analysis data, among the 47 donors we found 11 individuals with the heterozygous presence of the DNA sequence coding for the Q141K variant (carrier frequency: 23.4%, allele frequency: 11.766.6%), and 3 individuals with the heterozygous presence of the V12M variant (carrier frequency: 6.4%; allele frequency: 3.263.6%). Login to comment
136 Labels: individuals carrying wild-type ABCG2 (WT), polymorphic (Q141K, V12M) ABCG2 alleles, or a heterozygous stop mutation (STOP). Login to comment
185 Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, et al. (2002) C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment
209 Furukawa T, Wakabayashi K, Tamura A, Nakagawa H, Morishima Y, et al. (2009) Major SNP (Q141K) variant of human ABC transporter ABCG2 undergoes lysosomal and proteasomal degradations. Login to comment

[hide] The bovine ATP-binding cassette transporter ABCG2 ... Drug Metab Dispos. 2013 Mar;41(3):546-9. doi: 10.1124/dmd.112.049056. Epub 2012 Dec 10. Otero JA, Real R, de la Fuente A, Prieto JG, Marques M, Alvarez AI, Merino G
The bovine ATP-binding cassette transporter ABCG2 Tyr581Ser single-nucleotide polymorphism increases milk secretion of the fluoroquinolone danofloxacin.
Drug Metab Dispos. 2013 Mar;41(3):546-9. doi: 10.1124/dmd.112.049056. Epub 2012 Dec 10., [PMID:23230133]

Abstract [show]
The bovine adenosine triphosphate-binding cassette transporter G2 (ABCG2/breast cancer resistance protein) polymorphism Tyr581Ser (Y581S) has recently been shown to increase in vitro transepithelial transport of antibiotics. Since this transporter has been extensively related to the active secretion of drugs into milk, the potential in vivo effect of this polymorphism on secretion of xenobiotics in livestock could have striking consequences for milk production, the dairy industry, and public health. Our purpose was to study the in vivo effect of this polymorphism on the secretion of danofloxacin, a widely used veterinary antibiotic, into milk. Danofloxacin (1.25 mg/kg) was administered to six Y/Y 581 homozygous and six Y/S 581 heterozygous lactating cows, and plasma and milk samples were collected and analyzed by high-performance liquid chromatography. No differences were found in the pharmacokinetic parameters of danofloxacin in plasma between the two groups of animals. In contrast, Y/S heterozygous cows showed a 2-fold increase in danofloxacin levels in milk. In addition, the pharmacokinetic elimination parameters, mean residence time and elimination half-life, were significantly lower in the milk of the animals carrying the Y/S polymorphism. These in vivo results are in agreement with our previously published in vitro data, which showed a greater capacity of the S581 variant in accumulation assays, and demonstrate, for the first time, an important effect of the Y581S single-nucleotide polymorphism on antibiotic secretion into cow milk. These findings could be extended to other ABCG2 substrates, and may be relevant for the treatment of mastitis and for the design of accurate and novel strategies to handle milk residues.

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16 Several single-nucleotide polymorphisms (SNPs) have been studied in human ABCG2, and of these, Gln141Lys (Q141K) is one of the most important, causing impaired urate transport which may contribute to gout (Woodward et al., 2009). Login to comment
25 ABBREVIATIONS: ABC, ATP-binding cassette; ABCG2, adenosine triphosphate-binding cassette transporter G2; AUC, area under the curve; HPLC, high-performance liquid chromatography; Q141K, Gln141Lys; SNP, single-nucleotide polymorphism; Y581S, Tyr581Ser. Login to comment
73 Even human Q141K SNP does not affect the plasma disposition of all ABCG2 substrates (Kim et al., 2007; Adkison et al., 2008; Keskitalo et al., 2009). Login to comment

[hide] Genetics of hyperuricemia and gout: implications f... Curr Rheumatol Rep. 2013 Feb;15(2):309. doi: 10.1007/s11926-012-0309-8. George RL, Keenan RT
Genetics of hyperuricemia and gout: implications for the present and future.
Curr Rheumatol Rep. 2013 Feb;15(2):309. doi: 10.1007/s11926-012-0309-8., [PMID:23307580]

Abstract [show]
Gout is the most common inflammatory arthropathy and occurs in the setting of elevated serum urate levels. Gout is also known to be associated with multiple comorbidities including cardiovascular disease and the metabolic syndrome. Recent advances in research have increased our understanding and improved our knowledge of the pathophysiology of gout. Genome-wide association studies have permitted the identification of several new and common genetic factors that contribute to hyperuricemia and gout. Most of these are involved with the renal urate transport system (the uric acid transportasome), generally considered the most influential regulator of serum urate homeostasis. Thus far, SCL22A12, SCL2A9, and GLUT9 have been found to have the greatest variation and most influence on serum urate levels. However, genetics are only a part of the explanation in the development of hyperuricemia and gout. As results have been mixed, the role of known urate influential genes in gout's associated comorbidities remains unclear. Regardless, GWAS findings have expanded our understanding of the pathophysiology of hyperuricemia and gout, and will likely play a role in the development of future therapies and treatment of this ancient disease.

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121 Overall, the ABCG2 gene (SNP rs2231142, Q141K variant) had a stronger effect in men than women. Login to comment
129 The investigators found that common dysfunctional genotype combinations of ABCG2 gene (variants, Q126X and Q141K) are a major cause of gout [48]. Login to comment
135 The relationship between overproduction, hyperuricemia, and ABCG2 dysfunction led the authors to propose a new, testable model of hyperuricemia that takes into account population genotypes such as Q126X and Q141K affecting ABCG2 function. Login to comment
141 ABCG2 rs2231142 SNP (Q141K variant) decreased urate transport rates by approximately 50 % in vitro. Login to comment

[hide] Computational analysis and predictive modeling of ... Chem Cent J. 2013 Feb 4;7(1):23. doi: 10.1186/1752-153X-7-23. Lee Y, Jana S, Acharya G, Lee CH
Computational analysis and predictive modeling of polymorph descriptors.
Chem Cent J. 2013 Feb 4;7(1):23. doi: 10.1186/1752-153X-7-23., [PMID:23379683]

Abstract [show]
BACKGROUND: A computation approach based on integrating high throughput binding affinity comparison and binding descriptor classifications was utilized to establish the correlation among substrate properties and their affinity to Breast Cancer Resistant Protein (BCRP). The uptake rates of Mitoxantrone in the presence of various substrates were evaluated as an in vitro screening index for comparison of their binding affinity to BCRP.The effects of chemical properties of various chemotherapeutics, such as antiviral, antibiotic, calcium channel blockers, anticancer and antifungal agents, on their affinity to BCRP, were evaluated using HEK (human embryonic kidney) cells in which 3 polymorphs, namely 482R (wild type) and two mutants (482G and 482T) of BCRP, have been identified. The quantitative structure activity relationship (QSAR) model was developed using the sequential approaches of Austin Model 1 (AM1), CODESSA program, heuristic method (HM) and multiple linear regression (MLR) to establish the relationship between structural specificity of BCRP substrates and their uptake rates by BCRP polymorphs. RESULTS: The BCRP mutations may induce conformational changes as manifested by the altered uptake rates of Mitoxantrone by BCRP in the presence of other competitive binding substrates that have a varying degree of affinities toward BCRP efflux. This study also revealed that the binding affinity of test substrates to each polymorph was affected by varying descriptors, such as constitutional, topological, geometrical, electrostatic, thermodynamic, and quantum chemical descriptors. CONCLUSION: Descriptors involved with the net surface charge and energy level of substrates seem to be the common integral factors for defining binding specificity of selected substrates to BCRP polymorph. The reproducible outcomes and validation process further supported the accuracy of the computational model in assessing the correlation among descriptors involved with substrate affinity to BCRP polymorph. A quantitative computation approach will provide important structural insight into optimal designing of new chemotherapeutic agents with improved pharmacological efficacies.

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125 BCRP G34A (Val12Met) and C421A (Gln141Lys) polymorphisms occurred at high frequency in most ethnic populations and have been associated with the expression and activity of BCRP protein [48]. Login to comment
126 It has distinctive features including racial differences; for instance, BCRP V12M, Q141K, P269S and Q126Stop were detected in Korean at frequencies of 23, 28, 0.2 and 1.9%, respectively [49]. Login to comment
352 Pollex EK, Anger G, Hutson J, Koren G, Piquette-Miller M: Breast cancer resistance protein (BCRP)-mediated glyburide transport: effect of the C421A/Q141K BCRP single-nucleotide polymorphism. Login to comment

[hide] Gout-causing Q141K mutation in ABCG2 leads to inst... Proc Natl Acad Sci U S A. 2013 Mar 26;110(13):5223-8. doi: 10.1073/pnas.1214530110. Epub 2013 Mar 14. Woodward OM, Tukaye DN, Cui J, Greenwell P, Constantoulakis LM, Parker BS, Rao A, Kottgen M, Maloney PC, Guggino WB
Gout-causing Q141K mutation in ABCG2 leads to instability of the nucleotide-binding domain and can be corrected with small molecules.
Proc Natl Acad Sci U S A. 2013 Mar 26;110(13):5223-8. doi: 10.1073/pnas.1214530110. Epub 2013 Mar 14., [PMID:23493553]

Abstract [show]
The multidrug ATP-binding cassette, subfamily G, 2 (ABCG2) transporter was recently identified as an important human urate transporter, and a common mutation, a Gln to Lys substitution at position 141 (Q141K), was shown to cause hyperuricemia and gout. The nature of the Q141K defect, however, remains undefined. Here we explore the Q141K ABCG2 mutation using a comparative approach, contrasting it with another disease-causing mutation in an ABC transporter, the deletion of Phe-508 (DeltaF508) in the cystic fibrosis transmembrane conductance regulator (CFTR). We found, much like in DeltaF508 CFTR, that the Q141K mutation leads to instability in the nucleotide-binding domain (NBD), a defect that translates to significantly decreased protein expression. However, unlike the CFTR mutant, the Q141K mutation does not interfere with the nucleotide-binding domain/intracellular loop interactions. This investigation has also led to the identification of critical residues involved in the protein-protein interactions necessary for the dimerization of ABCG2: Lys-473 (K473) and Phe-142 (F142). Finally, we have demonstrated the utility of using small molecules to correct the Q141K defect in expression and function as a possible therapeutic approach for hyperuricemia and gout.

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0 Gout-causing Q141K mutation in ABCG2 leads to instability of the nucleotide-binding domain and can be corrected with small molecules Owen M. Woodwarda,1,2 , Deepali N. Tukayea,1 , Jinming Cuia , Patrick Greenwella , Leeza M. Constantoulakisa , Benjamin S. Parkera , Anjana Raoa , Michael K&#f6;ttgenb , Peter C. Maloneya , and William B. Gugginoa,2 a Department of Physiology, The Johns Hopkins University School of Medicine, Baltimore, MD 21205; and b Renal Division, University Medical Centre Freiburg, 79106 Freiburg, Germany Edited by Maurice B. Burg, National Heart, Lung, and Blood Institute, Bethesda, MD, and approved February 19, 2013 (received for review August 22, 2012) The multidrug ATP-binding cassette, subfamily G, 2 (ABCG2) transporter was recently identified as an important human urate transporter, and a common mutation, a Gln to Lys substitution at position 141 (Q141K), was shown to cause hyperuricemia and gout. Login to comment
1 The nature of the Q141K defect, however, remains undefined. Login to comment
2 Here we explore the Q141K ABCG2 mutation using a comparative approach, contrasting it with another disease-causing mutation in an ABC transporter, the deletion of Phe-508 (ƊF508) in the cystic fibrosis transmembrane conductance regulator (CFTR). Login to comment
3 We found, much like in ƊF508 CFTR, that the Q141K mutation leads to instability in the nucleotide-binding domain (NBD), a defect that translates to significantly decreased protein expression. Login to comment
4 However, unlike the CFTR mutant, the Q141K mutation does not interfere with the nucleotide-binding domain/intracellular loop interactions. Login to comment
6 Finally, we have demonstrated the utility of using small molecules to correct the Q141K defect in expression and function as a possible therapeutic approach for hyperuricemia and gout. Login to comment
14 Additionally, we characterized the single nucleotide polymorphism resulting in a Gln to Lys substitution at position 141 (Q141K), as causal for gout resulting from a loss of urate transport function in ABCG2 (10). Login to comment
15 The majority of work on ABCG2 [also know as the breast cancer resistance protein (BCRP)] has focused on its role as a multidrug transporter and specifically on its role as a xenotoxin and chemotherapeutic transport molecule, work that has included a description of the Q141K mutation`s effect on ABCG2 trafficking and function. Login to comment
16 Multiple groups reported that the Q141K mutation decreases protein expression and, coincidentally, xenotoxin transport function (11-16). Login to comment
17 This decrease of expression and function appears to be partially caused by the enhancement of the Q141K mutant`s susceptibility to proteasomal degradation (11-13). Login to comment
18 There is also recent evidence suggesting that a significant portion of the mutant Q141K protein accumulates in perinuclear aggresomes before being degraded (17). Login to comment
19 Conversely, the Q141K mutation has also been reported to reduce ABCG2 ATPase activity, suggesting that the mutation disrupts xenotoxin transport independently of expression or trafficking abnormalities (14, 16). Login to comment
20 However, how does the Q141K mutation affect urate transport? Login to comment
21 In our work, we found urate transport was significantly reduced by the Q141K mutation in Xenopus oocytes when transport was normalized to expression levels (10). Login to comment
22 In contrast, subsequent work by Matsuo et al. (18) confirmed ABCG2 as a urate transporter and that Q141K results in a loss of function. Login to comment
23 However, they postulated that the loss of urate transport function for the Q141K was specifically due to the reduction in protein expression (18) . Login to comment
24 Interestingly, the possibility that the Q141K mutation disrupts both function and trafficking/expression suggests intriguing parallels to other human disease causing ABC transporter mutations, namely the deletion of Phe-508 (ƊF508) in the cystic fibrosis transmembrane conductance regulator (CFTR). Login to comment
26 However, it remains unclear why the Q141K mutation causes increased ubiquitination and proteasomal degradation or accumulation in aggresomes. Login to comment
27 In this paper, we explore the nature of the Q141K defect and, specifically, its effect on expression in mammalian cells. Login to comment
30 Finally, we demonstrate that correcting the gout causing Q141K ABCG2 defect can be accomplished with small molecules in a manner similar to the successful correction of the CFTR molecule as a treatment for cystic fibrosis. Login to comment
41 13 | 5223-5228 Results Temperature Correction of Q141K. Login to comment
42 Previously, we showed that the Q141K mutation in ABCG2 was causal for gout and resulted in a 53% loss of urate transport function (10, 22). Login to comment
44 To investigate the opposing observations and to explore the possibility that both a loss of function and a expression defect could be caused by the Q141K mutation, we expressed our ABCG2 constructs in mammalian cell expression systems. Login to comment
45 We found, much like previous reports (12, 13, 15, 18), that, in multiple mammalian cell lines, transient expression of the Q141K mutation resulted in significantly less total protein (Fig. S1 A and B and Fig. 2D: 63.4 &#b1; 0.1% decrease), surface protein (Fig. 1D and Fig. S1 C and D), and function (Fig. S1E: 63.08 &#b1; 3.0% decrease) than wild-type ABCG2 [although with only slightly lower levels of unglycosylated protein (Fig. S2A): 27.3 &#b1; 0.1% reduction; see Fig. S2B for ABCG2 glycosylation states]. Login to comment
46 In both the oocytes and mammalian cell lines, we used the same human ABCG2 constructs; thus the differences in relative expression may give a clue as to the nature of the Q141K defect. Login to comment
48 To test the possibility that the Q141K mutant is being stabilized by the low incubation temperatures of the oocytes, we injected oocytes with mutant and wild-type ABCG2 mRNA and incubated the cells at either 16 &#b0;C or 32 &#b0;C (Fig. 1A). Login to comment
51 Fig. 1 B and C shows that the lowering of the incubation temperature of mammalian cells expressing wild-type ABCG2 and the Q141K mutant increases the expression of both; however, the percentage change in expression level is significantly greater in the mutant protein (Fig. 1C), a result consistent with low-temperature enhancement of folding and stabilizing mutant protein. Login to comment
53 Low temperature was also able to rescue the surface expression of the Q141K mutant (Fig. 1D), but again the majority of the rescued surface protein is the unglycosylated version of the protein (Fig. 1D, black arrows). Login to comment
56 We found that the low-temperature incubation again preferentially increased the dimer mutant Q141K protein expression compared with wild type (Fig. 1E: dimer Q141K expression increased 242.98 &#b1; 89.8% relative to wild type; n = 3). Login to comment
57 These results support the hypothesis that the Q141K mutant is unstable and that expression can be rescued with low-temperature incubation. Login to comment
58 Nature of the Q141K Defect. Login to comment
59 Human disease, loss of function, reduced expression of mature protein, and temperature correction are characteristics that the Q141K mutation shares with exactly one other mutant ABC protein: ƊF508 CFTR, the most common cystic fibrosis-causing mutation (25). Login to comment
61 However, the deletion of F508 in CFTR produces no mature glycosylated protein (25), whereas the Q141K defect in ABCG2 only reduces the amount of mature protein; thus the defects may be similar but not precisely homologous. Login to comment
65 These results support our model and the similar natures of the defects between Q141K ABCG2 and ƊF508 CFTR. Login to comment
70 To test whether or not Q141K decreases stability of the ABCG2 NBD, we introduced stabilizing mutations into the signature sequence to try to rescue protein expression. Login to comment
73 Temperature correction of Q141K ABCG2 expression defect. Login to comment
75 (B) Western blot of transient expression of wild-type (WT) and Q141K ABCG2 in CHO cells incubated for 48 h at 27 &#b0;C or 37 &#b0;C. Login to comment
76 (C) Summary data of temperature rescue of protein expression (WT: n = 6; Q141K: n = 8). Login to comment
78 (E) Dimer expression of WT and Q141K ABCG2 after 48 h of incubation at 27 &#b0;C or 37 &#b0;C (n = 4). Login to comment
81 In contrast, stabilizing the NBD domain of Q141K ABCG2 rescues protein expression and suggests that primarily NBD stability may underlie the Q141K defect. Login to comment
82 Even with considerable evidence suggesting that Q141K causes NBD instability, there remains the possibility that interdomain interactions may also be disrupted, as in the ƊF142 mutant and in the CFTR F508 deletion. Login to comment
89 Significantly, introduction of the K473W substitution on the Q141K background did not rescue total, mature, or dimer levels (all were actually significantly less) (Fig. 4 A-C and Fig. S5E). Login to comment
90 The failure of the K473W mutation to rescue Q141K expression supports the hypothesis that the Q141K mutation does not interfere with interdomain interactions or dimerization of the ABCG2 protein. Login to comment
91 Furthermore, we found that the Q141K dimer protein expression, as a fraction of wild-type expression, is greater than the total expression (Fig. 4D), not less, suggesting that the mutant dimerizes normally (or marginally more efficiently than wild type; P ࣘ 0.03; n = 7). Login to comment
92 Finally, modeling of ABCG2 with the Q141K substitution did not alter the F142 orientation or position in respect to the ICL residues, and K141 adopted the same orientation as Q141 (Fig. 4E). Login to comment
95 Taken together, our findings support the hypothesis that the Q141K mutation does not interfere with interdomain interactions or with dimerization of the ABCG2 protein. Login to comment
96 Small-Molecule Correction of Q141K. Login to comment
100 Q141K mutation occurs in mutational hotspot of ABC protein NBD. Login to comment
103 Western blots of transient total (B) and dimer expression (C) of WT, Q141K, and ƊF142 ABCG2 in HEK293 cells. Login to comment
108 Suppressor mutation correction of Q141K expression defect. Login to comment
112 Western blot of transient total (B) and dimer (C) expression in CHO cells of suppressor mutations in the Q141K background. Login to comment
113 Summary data of suppressor rescue of Q141K ABCG2 total (D; n = 10) and dimer protein expression (E; n = 4). Login to comment
114 (F) Surface expression of biotinylated ABCG2 and Q141K variants and Na+ /K+ ATPase, demonstrating strong rescue with the Q141K G188E mutation (n = 6). Login to comment
116 All means are &#b1; SEM; *P < 0.05; **P < 0.01. this principle and to learn more about the mechanism of dysfunction in the Q141K ABCG2 mutation, we attempted to correct the Q141K defect with small molecules. Login to comment
118 We found that application of 4-PBA increased significantly the expression of the Q141K protein (Fig. 5 A and B) but had no effect on the wild type, consistent with the hypothesis that the ERAD pathway targets the Q141K protein. Login to comment
119 The Q141K monomer expression levels, even after correction, were not at levels comparable to the wild type (Fig. 5A). Login to comment
120 Conversely, the dimer form of the Q141K protein when treated with 4-PBA was corrected to expression levels not different from wild type (P < 0.147, n = 4), suggesting that the effect of 4-PBA is critical for the creation of the dimer form of the protein (Fig. 5C). Login to comment
121 Interestingly, we found that 4-PBA was also able to increase the amount of Q141K/G188E protein (Fig. 5 A-C), demonstrating that the G188E suppressor mutation is not a full rescue and that some protein is still targeted for degradation. Login to comment
125 We found that incubation with the VRT-325 molecule significantly increased the amount of total (Fig. 5 D and E) and surface Q141K protein expression (Fig. S6A); VRT-325 had no effect on wild-type ABCG2 expression (P < 0.193, n = 3). Login to comment
126 Both low temperature and VRT-325 correction of Q141K produced increased levels of mature glycosylated monomer protein (Fig. 5D); however, unlike the low-temperature correction, VRT-325 did not increase the amount of immature unglycosylated protein (Fig. 5D and Fig. S6B). Login to comment
127 This suggests that VRT-325 may allow more of the Q141K protein to traffic normally, become glycosylated, and reach the cell membrane. Login to comment
128 In HEK293 cells expressing the Q141K mutant and treated with VRT-325, we also found that the accumulation of C-14-labeled uric acid was significantly less than in the untreated Q141K-expressing cells (Fig. 5F; P ࣘ 0.004; n = 11, 10), demonstrating partial rescue of function along with expression of the Q141K mutant. Login to comment
130 Surprisingly, corr-4a (Fig. 5 G and H) increased the total protein levels of the Q141K ABCG2 mutant, but not wild-type protein, although the corr-4a (20 bc;M) correction appears to be less than the VRT-325 (10 bc;M) correction (Fig. S6C). Login to comment
131 Taken together, our results show that the Q141K protein level can be rescued using small corrector molecules, which is consistent with the hypothesis that the Q141K mutation is a misfolding mutation that increases degradation and consistent with the principle that the gout-causing Q141K mutation can be corrected with small molecules. Login to comment
132 GAPDH monomer dimer 141K -- P E W 473: 141K -- P E W 473: * A B Normalized ABCG2 dimer protein 0.0 0.5 1.0 Q141K -- P E 473: W ** ** ** C ABCG2 Q141K Q141K protein expression as fraction of wild type 0 0.25 0.5 0.75 Dimer Total * D E Fig. 4. Login to comment
133 Q141K does not disrupt interdomain interactions or dimerization. Login to comment
134 Western blots of total (A) and dimer (B) expression of Q141K ABCG2 with secondary mutations at K473 demonstrating that K473W cannot rescue Q141K expression. Login to comment
135 Summary data of K473 mutation on total (C; n = 4-6) Q141K expression. Login to comment
136 (D) Expression of total or dimer Q141K as a fraction of wild type (total or dimer); a greater proportion of total Q141K exists as a dimer than does wild type (n = 7). Login to comment
137 (E) Superimposition of ABCG2 NBD model (green, with Q141 as orange sticks and F142 as red sticks) and the model with Q141K substitution (blue, with K141 and F142 as cyan sticks). Login to comment
140 76k ABCG2 GAPDH WT 141K 188E 193K -- -- +4-PBA WT 141K 188E 193K -- -- control A ** ** 0 100 200 300 WT 141K 188E -- % Change in expression after 4-PBA B 76k ABCG2 GAPDH 141K WT -- -- +VRT 325 +VRT 325 27&#b0;C 27&#b0;C D -50 0 100 200 % Change with VRT-325 141K WT * E WT 141K 188E 193K -- -- control +4-PBA WT 141K 188E 193K -- -- Ɗ142 Ɗ142 C 150k 76k ABCG2 GAPDH F Q141K Q141K + VRT-325 ** % Change in C-14 uric acid accumulation as compared to WT 0 20 40 WT + VRT-325 % Change with Corr-4a -50 0 50 100 150 141K WT ** H G WT -- +Corr 4a 76k ABCG2 GAPDH 141K -- +Corr 4a Fig. 5. Login to comment
141 Small-molecule correction of gout-causing Q141K ABCG2 expression defect. Login to comment
143 (B) Summary data of 4-PBA rescue of Q141K expression (n = 3-6); 4-PBA has no effect on WT ABCG2 expression (P < 0.28; n = 4) compared with zero. Login to comment
149 (G) Western blot and summary data (H) of total Q141K ABCG2 expression rescue by 24-h treatment with 20 bc;M corr-4a (n = 4). Login to comment
154 Seeing ABCG2 as an important urate transporter and the Q141K mutation as a loss-of-function mutation led naturally to a comparison with other disease-causing mutations in human ABC transporters, a comparison that lead inevitably to CFTR. Login to comment
155 The parallels between the Q141K mutation in ABCG2 and the deletion of the Phe-508 in CFTR, a mutation that occurs in ~90% of all individuals with cystic fibrosis (25), are striking. Login to comment
158 Here we tested homologous NBD mutations in Q141K ABCG2 and found the G188E mutation rescued Q141K expression, suggesting the Q141K mutation results in increased ABCG2 NBD instability. Login to comment
159 Interestingly, the R193K mutation, homologous to the suppressor mutation R555K in CFTR, didn`t increase Q141K ABCG2 expression; but did appear to decrease the insoluble fraction of Q141K protein (Fig. S6D). Login to comment
162 Our CFTR ƊF508 and Q141K ABCG2 comparison also revealed the importance of F142 in ABCG2. Login to comment
163 The F142 deletion causes a severe decrease in expression but, unlike the Q141K mutation, the ƊF142 cannot be rescued with temperature or with suppressor mutations. Login to comment
177 Interestingly, Basseville et al. (17) recently found that a different suite of HDAC inhibitors rescued Q141K ABCG2 function and expression levels but did not observe any changes in Hsp 70 or Hsp 90 levels. Login to comment
184 Thus, the VRT-325 molecule corrects in ƊF508 CFTR the exact defect that we describe here for Q141K ABCG2, and indeed, we found that VRT-325 significantly increased total, dimer, and surface expression of the Q141K protein. Login to comment
186 The success of the VRT-325 and corr-4a molecules in correcting mutant Q141K protein expression clearly demonstrates that a better understanding of the way in which ABCG2 folds and dimerizes can lead to drug discovery of new therapies for gout and hyperuricemia. Login to comment
187 Going forward, it will also be important to discern why the Q141K mutation leads to a functional defect, even after expression level is normalized (10, 16), and whether NBD instability reduces binding or hydrolysis of ATP. Login to comment
188 Finally, the similarities between ƊF508 CFTR and Q141K ABCG2 extend beyond the work presented here. Login to comment
191 For CFTR, the dysfunction of the ƊF508 mutation may provide some protection from cholera infection (54), whereas in ABCG2 the Q141K mutation may persist to increase blood urate levels of particular human populations beyond what even the loss of uricase function achieved. Login to comment
239 Furukawa T, et al. (2009) Major SNP (Q141K) variant of human ABC transporter ABCG2 undergoes lysosomal and proteasomal degradations. Login to comment
248 Imai Y, et al. (2002) C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] No association between MTHFR C677T and serum uric ... Nagoya J Med Sci. 2013 Feb;75(1-2):93-100. Hinohara Y, Naito M, Okada R, Yin G, Higashibata T, Tamura T, Kawai S, Morita E, Wakai K, Matsuo H, Mori A, Hamajima N
No association between MTHFR C677T and serum uric acid levels among Japanese with ABCG2 126QQ and SLC22A12 258WW.
Nagoya J Med Sci. 2013 Feb;75(1-2):93-100., [PMID:23544272]

Abstract [show]
Several genome-wide association studies (GWAS) have revealed that single nucleotide polymorphisms (SNPs) of ABCG2 and SLC22A12 were strongly associated with serum uric acid (SUA), but those of methylene tetrahydrofolate reductase (MTHFR) were not. However, there were several studies indicating the association with MTHFR C677T polymorphism. This study examined the association with the polymorphism, taking into account the genotypes of ABCG2 Q126X and SLC22A12 W258X. Subjects were 5,028 health checkup examinees of Seirei Preventive Health Care Center (3,416 males and 1,612 females) aged 35 to 69 years, who participated in the Japan Multi-Institutional Collaborative Cohort Study (J-MICC Study). Hyperuricemia was defined as SUA equal to 7 mg/dL or over. The genotype frequency was 35.9% for CC, 48.1% for CT, and 16.0% for TT, being in Hardy-Weinberg equilibrium (p=0.90). Among 4,425 participants with ABCG2 126QQ and SLC22A12 258WW who were not under medication for hyperuricemia, the mean SUA was 5.6 mg/dL, 5.6 mg/dL, and 5.7 mg/dL, respectively. When 114 participants with ABCG2 126QQ and SLC22A12 258WW under medication for hyperuricemia were included in hyperuricemia cases, the sex-age adjusted odds ratio (OR) of hyperuricemia was not significant; OR=1.00 (95% confidence interval, 0.89-1.24) for CT genotype and OR=0.98 (0.84-1.32) for TT genotype, relative to CC genotype. The present study indicated no association between SUA and MTHFR C677T genotype, after the influences of ABCG2 Q126X and SLC22A12 W258X were removed.

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12 Key Words: Serum uric acid, Urate transporter polymorphisms, MTHFR C677T INTRODUCTION It is well known that serum uric acid (SUA) levels are associated with various factors such as sex, age, body mass index (BMI), dietary habit and drinking habit.1-3) In addition, there is evidence that genetic traits influence SUA concentrations; the heritability was estimated to be up to 73%.4) A recent genome-wide association study performed in Japan showed strong associations of SUA with genetic polymorphisms of SLC22A12 coding uric acid transporter 1 (URAT1), Received: January 13, 2013; accepted: January 24, 2013 Corresponding author: Yukako Hinohara MD, MPH Department of Preventive Medicine, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan Phone: +81 52 744 2132, Fax: +81 52 744 2971, E-mail address: y.hinohara@soumu.go.jp SLC2A9 coding glucose transporter 9 (GLUT9), and ABCG2 coding ATP-binding cassette subfamily G member 2 (ABCG2),5) which were also reported to have associations in European ancestry.6) Among the polymorphisms, SLC22A12 W258X,7,8) SLC2A9 R380W and R198C,9) and ABCG2 Q126X and Q141K10,11) were confirmed to have associations with SUA, although the association of ABCG2 Q141K was relatively weak. Login to comment

[hide] [Transporter-mediated regulation of pharmacokineti... Yakugaku Zasshi. 2013;133(4):451-61. Takada T
[Transporter-mediated regulation of pharmacokinetics of lifestyle-related substances].
Yakugaku Zasshi. 2013;133(4):451-61., [PMID:23546589]

Abstract [show]
Recent studies revealed the importance of transporters in the behaviors of small molecules in the body. In mammals, the presence of a lot of transporters has been suggested, such as ATP-binding cassette (ABC) transporters and solute ligand carrier (SLC) transporters, some of which are clarified to be causative genes for various kinds of genetic disorders. In addition, a lot of transporters are known to mediate cellular import or export of drugs, to contribute to the pharmacokinetics of substrate drugs and to be involved in the interindividual differences of drug responses. In this review, I introduce our recent work on the transporter-mediated regulation of pharmacokinetics of lifestyle-related substances, such as cholesterol and urate.

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24 c3f; ⏚ ᣸ 3fa; c8; e9; f3; b9; dd; fc; bf; fc; ABCG2 Ĳe; SNPs ௼ ad8;c3f;⏚⊈Kc7;fb;Kdb;ba8;ea;b9;af; ˯f;d3b;fd2;᠓Kc5;௼ ௱௺Me5;఑Ĵc;Ĵb; ad8;c3f;⏚⊈Kc7;ఌKdb;ba8;Ĳf;,d05;ca2;Kc5;௼௤௦a00; ℁İc;௢Ĵb;ఐ௦Ĳb;,Pa5;e80;ఌ ad8;d7;ea;f3;bdf;Ĳe;ᤪbdf;Έe;ɏa;Ĳa;௽ Ĳe;Jb0;᛻⌕8e0;Ĳb;d77;8e0;௳Ĵb;௼ὃ௨఑Ĵc;௺௤ıf;İc;,fd1;e74;Ĳe; Ẇa76;Ĳb;ఐĴa;΋a;f1d;ḄĲa;⌕8e0;Ĳe;b58;ᙠఊ̙a;ᖂ௯Ĵc;௺௤ıf;&#ff0e; 2004 e74;,5f0;e7e;Ĳe;Ẇa76;b0;eb;fc;d7;ఐĴa;,d2;c8;b2c; 4 Cd3; ⁐f53;╩ΊĲb;ʠa;Me5;Ĳe;Kdb;ba8;Kc5;8e0;΋a;f1d;b50;İc;b58;ᙠ௳Ĵb;5ef;Pfd;ឋ İc;ᛇȠa;௯Ĵc;ıf;İc;, 26) ᎛Xdc;♚9df;Ĳb;Ĳf;ɏa;İf;Ĳe;΋a;f1d;b50;İc;Ȟb; ĳe;Ĵc;௺İa;Ĵa;,ᐹf53;ḄĲa;Kc5;8e0;΋a;f1d;b50;Ĳf;Ȝc;b9a;௯Ĵc;௺௤Ĳa; İb;௷ıf;&#ff0e;ıd;௭௻,d30;Pde;̳c;e0a;Ĳb;İa;௤௺f38;〈9fa;cea;Ĳe;᣸3fa; Ĳb;fc2;Ĵf;Ĵb; ABCG2/breast cancer resistance protein (BCRP) ΋a;f1d;b50;İc;௭Ĳe;♚9df;Ĳb;b58;ᙠ௳Ĵb;௭௼, ABCG2 Ĳb;Ĳf;a5f;Pfd;᜕4d5;ఔf34;௦ϗb;ea6;Ĳe; ad8;௤΋a;f1d;b50;ɏa;ɂb;İc;b58;ᙠ௳ Ĵb;௭௼,Ae2;Me5;Ĳe;f38;〈9fa;cea;௼Ĳe;bd4;f03;İb;఑c3f;⏚Ĳf; ABCG2 Ĳe;9fa;cea;Ĳb;Ĳa;Ĵa;f97;Ĵb;௼ὃ௨఑Ĵc;ıf;௭௼Ĳa;௽Ĳe;ᳮᵫĲb;ఐ Ĵa;,b46;ὅ఑Ĳf; ABCG2 Ĳb;_a2;௳Ĵb;ʳc;a0e;ఔ⍈ఉıf;(௵Ĳa; ĳf;Ĳb;,ıd;Ĳe;f8c;Ĳe; GWAS Ĳb;İa;௤௺ఊ⊈e05;c3f;⏚ᎠĲe;᜕ 4d5;Ĳb;_a2;⌿௳Ĵb;΋a;f1d;b50;௼௱௺ ABCG2 Ĳf;ᛇȠa;௯Ĵc;௺௤ Ĵb;) &#ff0e; 27&#e30f; 29) ABCG2/BCRP Ĳf;ıd;Ĳe;Ȝd;Ĳe;΅a;Ĵa;,ɏa;ᒐὊឋĲb;_a2;e0e; ௳Ĵb;8e0;b50;௼௱௺˿a;΂b;௯Ĵc;ıf;c8;e9;f3;b9;dd;fc;bf;fc;௻௢Ĵb; İc;,e83;bc4;Ĳa;d44;e54;ᑖe03;௼e83;௤9fa;cea;a8d;b58;ឋఔᨵ௳Ĵb;௭௼ İc;b21;b2c;Ĳb;ʔe;఑İb;௼Ĳa;Ĵa;,Ife;ᙠ௻Ĳf;ɏa;Ed8;Ĳa;⌤Fb9;İb;఑Ẇ a76;İc;⍈ఉ఑Ĵc;௺௤Ĵb;&#ff0e;Ae5;ʠc;eba;Ĳb;İa;௫Ĵb; ABCG2 ΋a;f1d; b50;ɏa;ɂb;Ĳe;a2;ec;eb;ϗb;ea6;Ĳf; ad8;İf;,˿a;Ife;[cf;5ca;ఁa5f;Pfd;᜕ᓄఔ f34;Ĵf;Ĳa;௤ 34G&#ff1e;A(V12M)Ĳf; 19.2%,bf;f3;d1;af;cea; ˿a;Ife;[cf;İc;d04;Ȗa;ᑖĲb;f4e;e0b;௳Ĵb; 421C&#ff1e;A(Q141K)Ĳf; 31.9%,d42;b62;b3;c9;f3;İc;˯f;௲a5f;Pfd;b20;ʀd;௼Ĳa;Ĵb; 376C&#ff1e;T (Q126X)Ĳf; 2.8%௻௢Ĵb;௼ᛇȠa;௯Ĵc;௺௤Ĵb;&#ff0e; 30) ௭Ĵc; ఑Ĳe;௦௵,ϗb;ea6;İc; ad8;İf;a5f;Pfd;f4e;e0b;ఔf34;௦ 421C&#ff1e;A (Q141K)Ĳb;௸௤௺Ĳf;Qe8;e8a;ḄĲb;ఊఐİf;Ẇa76;௯Ĵc;௺İa; Ĵa;,Uac;ᱥ4d5;ɦb;Ĳe;᜕4d5;௼௱௺Ĳf;,b9;eb;d5;a1;b5;e9;b8;f3;Ĳe; d88;ᓄba1;ᔾ5ce;Ĳe;e0a;᧏,ed;b9;d0;b9;bf;c1;f3;ఌd5;eb;d0;b9;bf;c1; f3;Ĳe;d4c;5e3;ᢗe0e;᧲Ĳe; AUC(Uac;ᱥ⊈e2d;fc3;ea6;ߟ᧲╹Bf2;dda; e0b;☢a4d;)ఌ Cmax(ᨬ ad8;⊈e2d;fc3;ea6;)Ĳe;e0a;᧏İc;Yb3;bdf;௯ Ĵc;௺௤Ĵb;&#ff0e;௭Ĳe;ఐ௦Ĳb;,Uac;ᱥc8;e9;f3;b9;dd;fc;bf;fc; ABCG2 Ĳe;Uac;ᱥ4d5;ɦb;ᑴfa1;8e0;b50;௼௱௺Ĳe;[cd;⌕ឋĲf;a8d;b58; ௯Ĵc;௺௤ıf;ఊĲe;Ĳe;,d2;c8;Ĳb;İa;௫Ĵb;˯f;ᳮḄ9fa;cea;ఌ˯f;ᳮ a5f;Pfd;Ĳb;௸௤௺Ĳf;e0d;ʔe;௻௢௷ıf;&#ff0e; ABCG2 Ĳb;ఐĴb;c3f;⏚f38;〈Ĳb;௸௤௺ʳc;a0e;௳Ĵb;ıf;ఉ, ABCG2 ˿a;Ife;d30;Pde;İb;఑abf;Xfd;௱ıf;d30;Pde;̳c;c0f;Pde;ఔᵨ௤ıf; f38;〈b9f; a13;ఔʹc;௷ıf;d50;ʧc;,ABCG2 Ĳf;˯f;ᳮḄfc3;ea6;௻Ĳf; bfd;Ȥc;௱Ĳa;௤ ad8;bb9;[cf;ឋfb;f4e;Yaa;Ȥc;ឋĲe;c3f;⏚f38;〈ఔ>c5;௦௭ ௼İc;ʔe;఑İb;௼Ĳa;௷ıf;(c3f;⏚Ĳe;eb6;Ye3;ea6;Ĳe;bd4;f03;Ḅ ad8;௤ ad8; pH e0b;௻c42;ఉ఑Ĵc;ıf; Km ᎠĲf; 8.24&#b1;1.44 mM ௻௢ Ĵa;,⊈e05;c3f;⏚Ꭰ(f8b;௨௾ 7.0 mg/dL&#ff1d;d04; 420 mM) ௼bd4;ఇ௺Ĳf;Ĵb;İb;Ĳb; ad8;௤) &#ff0e; 31) ĳe;ıf;,᜕ᶒf53;Ye3;᪆Ĳe;d50; ʧc;,ed6;Ĳe;f38;〈9fa;cea;௼Ȝc;Ed8;,c3f;⏚f38;〈Ĳb;İa;௤௺ఊ Q141K ௻Ĳf;a5f;Pfd;İc;Ȗa;e1b;,Q126X ௻Ĳf;a5f;Pfd;İc;d88;ᜫ௳ Ĵb;௭௼İc;̙a;௯Ĵc;ıf;&#ff0e; ௸௹௤௺,Ae5;ʠc;eba;Ĳe;Ꮙeb7;a3a;Aad;5d7;a3a;ὅĲe;b5;f3;d7;eb;ఔ ᵨ௤௺,⊈e05;c3f;⏚Ꭰ௼ ABCG2 ΋a;f1d;b50;ɏa;ɂb;Ĳe;_a2;fc2;Ĳb; ௸௤௺ʳc;a0e;௱ıf;d50;ʧc;,Q141K ᜕ᶒĲe;fdd;ᢝᦪİc;ɏa;௤ ĳb;௽,⊈e05;c3f;⏚Ꭰİc;e0a;᧏௱௺௤ıf;&#ff0e; 31) ĳe;ıf;,cf;d7;ed; bf;a4;d7;ϗb;ea6;Ye3;᪆Ĳb;ఐĴa;,Q126X ௼ Q141K Ĳe;e21;᜕ᶒ 4. Proposed Model of ABCG2-mediated Urate Secretion31) 458 Vol. 133 (2013) Ĳf;Ȝc;௲Cd3;⁐f53;e0a;Ĳb;Ĳf;b58;ᙠ௱Ĳa;௤௭௼İc;̙a;௯Ĵc;ıf;&#ff0e;ıd; ௭௻,e21;᜕ᶒĲe;ϗb;ea6;Ĳb;௸௤௺Ae5;ʠc;eba;ᵱឋĲe;Kdb;ba8;Kc7;f8b; ௼Ꮙe38;ὅ௻bd4;f03;௱ıf;d50;ʧc;,Q126X ௼ Q141K Ĳe;d44;ĳf; ᔠĴf;ıb;İb;఑?a8;b9a;௯Ĵc;Ĵb;c3f;⏚f38;〈d3b;ឋĲe;f4e;e0b;Ĳb;f34;௤, aa;c3;ba;bd4;௻̙a;௯Ĵc;Ĵb;Kdb;ba8;˿a;Kc7;ea;b9;af;İc;♿℉Ĳb; ad8;ĳe;Ĵb; ௭ ௼ İc; ʔe; ఑ İb; ௼ Ĳa; ௷ ıf; &#ff0e; 31) ௭ Ĵc; ఑ Ĳe; d50; ʧc; Ĳf; , ABCG2 İc;˯f;f53;ᑁĲb;İa;௫Ĵb;c3f;⏚Ĳe;f53;᜜ఆĲe;᣸cc4;Ĳb;_a2; e0e;௱௺İa;Ĵa;,ıd;Ĳe;a5f;Pfd;f4e;e0b;Ĳf;⊈e05;c3f;⏚Ꭰ31,32)5ca;ఁKdb; ba8;˿a;Kc7;ea;b9;af;31)Ĳe;e0a;᧏ఔఊıf;఑௳௭௼ఔ̙a;௳ఊĲe;௻ ௢௷ıf;&#ff0e;ABCG2 Ĳf;̩d;Qd3;,̮e;Qd3;,c0f;ῲĲa;௽Ĳe;♊aef;̳c; Ĳb;˿a;Ife;௱,f38;〈9fa;cea;Ĳe;f53;᜜ఆĲe;᣸3fa;Ĳb;fc2;Ĵf;Ĵb;௭௼İc; Me5;఑Ĵc;௺௤Ĵb;௭௼,c3f;⏚Ĳf;c3f;e2d;Ĳe;ĳf;Ĳa;఑ıa;cde;e2d;Ĳb;ఊ ᣸ cc4; ௯ Ĵc; Ĵb; ௭ ௼ İc; ᛇ Ƞa; ௯ Ĵc; ௺ ௤ ıf; ௭ ௼ İb; ఑ , ABCG2 Ĳf;௭Ĵc;఑Ĳe;d44;e54;İb;఑Ĳe;c3f;⏚ᑖccc;Ĳb;_a2;e0e;௱௺ ௤Ĵb;5ef;Pfd;ឋİc;ὃ௨఑Ĵc;ıf;(Fig. 4) &#ff0e; 31) 5-2. Login to comment

[hide] Association of functional polymorphism rs2231142 (... Am J Epidemiol. 2013 May 1;177(9):923-32. doi: 10.1093/aje/kws330. Epub 2013 Apr 3. Zhang L, Spencer KL, Voruganti VS, Jorgensen NW, Fornage M, Best LG, Brown-Gentry KD, Cole SA, Crawford DC, Deelman E, Franceschini N, Gaffo AL, Glenn KR, Heiss G, Jenny NS, Kottgen A, Li Q, Liu K, Matise TC, North KE, Umans JG, Kao WH
Association of functional polymorphism rs2231142 (Q141K) in the ABCG2 gene with serum uric acid and gout in 4 US populations: the PAGE Study.
Am J Epidemiol. 2013 May 1;177(9):923-32. doi: 10.1093/aje/kws330. Epub 2013 Apr 3., [PMID:23552988]

Abstract [show]
A loss-of-function mutation (Q141K, rs2231142) in the ATP-binding cassette, subfamily G, member 2 gene (ABCG2) has been shown to be associated with serum uric acid levels and gout in Asians, Europeans, and European and African Americans; however, less is known about these associations in other populations. Rs2231142 was genotyped in 22,734 European Americans, 9,720 African Americans, 3,849 Mexican Americans, and 3,550 American Indians in the Population Architecture using Genomics and Epidemiology (PAGE) Study (2008-2012). Rs2231142 was significantly associated with serum uric acid levels (P = 2.37 x 10(-67), P = 3.98 x 10(-5), P = 6.97 x 10(-9), and P = 5.33 x 10(-4) in European Americans, African Americans, Mexican Americans, and American Indians, respectively) and gout (P = 2.83 x 10(-10), P = 0.01, and P = 0.01 in European Americans, African Americans, and Mexican Americans, respectively). Overall, the T allele was associated with a 0.24-mg/dL increase in serum uric acid level (P = 1.37 x 10(-80)) and a 1.75-fold increase in the odds of gout (P = 1.09 x 10(-12)). The association between rs2231142 and serum uric acid was significantly stronger in men, postmenopausal women, and hormone therapy users compared with their counterparts. The association with gout was also significantly stronger in men than in women. These results highlight a possible role of sex hormones in the regulation of ABCG2 urate transporter and its potential implications for the prevention, diagnosis, and treatment of hyperuricemia and gout.

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2 A loss-of-function mutation (Q141K, rs2231142) in the ATP-binding cassette, subfamily G, member 2 gene (ABCG2) has been shown to be associated with serum uric acid levels and gout in Asians, Europeans, and European and African Americans; however, less is known about these associations in other populations. Login to comment
27 More importantly, the resultant missense polymorphism at codon 141 (glutamine to lysine, Q141K) has been shown to be a loss-of-function mutation in 2 independent functional studies (22, 23). Login to comment
134 DISCUSSION In a meta-analysis of 39,853 persons from 4 different population-based samples (European Americans, African Americans, Mexican Americans, and American Indians), we demonstrated that the functional variant rs2231142 (Q141K) in the ABCG2 gene was significantly associated with both serum uric acid level (P = 2.37 &#d7; 10-67 , P = 3.98 &#d7; 10-5 , P = 6.97 &#d7; 10-9 , and P = 5.33 &#d7; 10-4 in European Americans, African Americans, Mexican Americans, and American Indians, respectively) and the odds of Table 3. Login to comment
148 The Q141K (rs2231142) polymorphism occurs in a highly conserved region of the gene and has been shown to be a loss-of-function mutation in 2 independent functional studies (22, 23). Login to comment
156 systems are reflective of the human kidney, one might expect that the consequence of the Q141K loss-of-function polymorphism would be relatively less in persons with high estrogen levels (they had decreased ABCG2 mRNA expression to begin with) compared with those with low estrogen levels. Login to comment
176 Lastly, the Q141K variant is a well-studied functional polymorphism, thus providing a strong a priori hypothesis and biological plausibility. Login to comment
178 In conclusion, the common polymorphism rs2231142, which leads to a change from glutamine to lysine in codon 141 (Q141K) of the ABCG2 gene, is significantly associated with elevated serum uric acid levels and increased prevalence of gout in European Americans, African Americans, Mexican Americans, and Americans Indians. Login to comment

[hide] Levofloxacin-induced seizures in a patient without... Eur J Clin Pharmacol. 2013 Aug;69(8):1611-3. doi: 10.1007/s00228-013-1515-7. Epub 2013 Apr 25. Gervasoni C, Cattaneo D, Falvella FS, Vitiello P, Cheli S, Milazzo L, Clementi E, Riva A
Levofloxacin-induced seizures in a patient without predisposing risk factors: the impact of pharmacogenetics.
Eur J Clin Pharmacol. 2013 Aug;69(8):1611-3. doi: 10.1007/s00228-013-1515-7. Epub 2013 Apr 25., [PMID:23616064]

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43 This drug can readily cross the BBB [3], Table 1 Genotyping analyses performed in our case Protein Gene Variant Database SNP ID AA substitution MAFa Case genotypeb Functional effect of the variant P-glycoprotein 1 ABCB1 c.1236C>T rs1128503 T=0.451 CT Controversial results P-glycoprotein 1 ABCB1 c.2677G>T rs2032582 Ala893Ser T=0.469 GT Altered P-gp activity and expression P-glycoprotein 1 ABCB1 c.3435T>C rs1045642 C=0.429 TC Decreased mRNA and protein levels BCRP ABCG2 c.421C>A rs2231142 Gln141Lys A=0.111 CA Decreased protein levels OATP1A2 SLCO1A2 c.38T>C rs10841795 Ile13Thr C=0.159 TT Increased protein function OATP1A2 SLCO1A2 c.516A>C rs11568563 Glu172Asp C=0.083 AA Decreased protein function SNP, Single nucleotide polymorphism a Minor allele frequency in the CEU population of the International HapMap Project b All genotypes of our case were determined by Real Time PCR, using the TaqMan&#ae; SNP Genotyping Assays (Applied Biosystems, Foster City, CA) or SimpleProbe Assays (Roche, Italy) Eur J Clin Pharmacol (2013) 69:1611-1613 and thus its effect in the CNS can be rapidly observed. Login to comment

[hide] Overlapping functions of ABC transporters in topot... Mol Cancer Ther. 2013 Jul;12(7):1343-55. doi: 10.1158/1535-7163.MCT-13-0100. Epub 2013 May 1. Tiwari AK, Zhang R, Gallo JM
Overlapping functions of ABC transporters in topotecan disposition as determined in gene knockout mouse models.
Mol Cancer Ther. 2013 Jul;12(7):1343-55. doi: 10.1158/1535-7163.MCT-13-0100. Epub 2013 May 1., [PMID:23635651]

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It is established that efflux transporters of the ATP-binding cassette (ABC) superfamily can affect the pharmacokinetics of drugs through mechanisms pertaining to drug absorption, elimination, and distribution. To characterize the role of multiple transporters in topotecan's pharmacokinetics, total (lactone+carboxylate) and lactone forms were measured by liquid chromatography/tandem mass spectrometry (LC/MS-MS) in plasma, bile, urine, and feces following intravenous administration at doses of 1 and 4 mg/kg to eight mouse strains: C57BL/6 [wild-type (WT)], Abcb1(-/-), Abcc2(-/-), Abcc4(-/-), Abcg2(-/-), Abcc2;Abcb1(-/-), Abcc2;Abcg2(-/-), and Abcc4;Abcg2(-/-). Compared with WT mice and at both dose levels, the plasma areas under the curve for topotecan lactone were not significantly different in the Abcc2(-/-), Abcc4(-/-), and Abcb1(-/-) strains, whereas significant differences were found in Abcg2(-/-), Abcc2;Abcb1(-/-) (only at the high dose), Abcc4;Abcg2(-/-), and Abcc2;Abcg2(-/-) mice and ranged from 2.1- to 3.3-fold higher. Consistent with these changes, the fecal and biliary excretion of topotecan was reduced, whereas renal elimination was elevated in Abcg2(-/-)-based strains. Similarly, the Abcc2;Abcb1(-/-) strain also had elevated renal elimination and reduced fecal excretion of topotecan lactone. This was more pronounced at the 4 mg/kg dose level, suggesting possible saturation of Abcg2. The Abcc4 transporter was found not to be a major determinant of topotecan pharmacokinetics. It is concluded that Abcg2 has the most significant effect on topotecan elimination, whereas both Abcb1 and Abcc2 have overlapping functions with Abcg2. As such it is relevant to examine how polymorphisms in these transporters influence topotecan activity in patients and whether coadministration of transport modulators could positively affect efficacy without increasing toxicity.

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232 With respect to the latter, a pilot study indicated that the presence of single-nucleotide polymorphisms (SNP) in ABCG2 Q141K (ABCG2 421C>A) in a cohort of patients with cancer significantly altered topotecan bioavailability and increased plasma AUC by 1.35-fold (34). Login to comment

[hide] Hyperuricemia influences tryptophan metabolism via... Biochim Biophys Acta. 2013 Oct;1832(10):1715-22. doi: 10.1016/j.bbadis.2013.05.002. Epub 2013 May 9. Dankers AC, Mutsaers HA, Dijkman HB, van den Heuvel LP, Hoenderop JG, Sweep FC, Russel FG, Masereeuw R
Hyperuricemia influences tryptophan metabolism via inhibition of multidrug resistance protein 4 (MRP4) and breast cancer resistance protein (BCRP).
Biochim Biophys Acta. 2013 Oct;1832(10):1715-22. doi: 10.1016/j.bbadis.2013.05.002. Epub 2013 May 9., [PMID:23665398]

Abstract [show]
Hyperuricemia is related to a variety of pathologies, including chronic kidney disease (CKD). However, the pathophysiological mechanisms underlying disease development are not yet fully elucidated. Here, we studied the effect of hyperuricemia on tryptophan metabolism and the potential role herein of two important uric acid efflux transporters, multidrug resistance protein 4 (MRP4) and breast cancer resistance protein (BCRP). Hyperuricemia was induced in mice by treatment with the uricase inhibitor oxonic acid, confirmed by the presence of urate crystals in the urine of treated animals. A transport assay, using membrane vesicles of cells overexpressing the transporters, revealed that uric acid inhibited substrate-specific transport by BCRP at clinically relevant concentrations (calculated IC50 value: 365+/-13muM), as was previously reported for MRP4. Moreover, we identified kynurenic acid as a novel substrate for MRP4 and BCRP. This finding was corroborated by increased plasma levels of kynurenic acid observed in Mrp4(-/-) (107+/-19nM; P=0.145) and Bcrp(-/-) mice (133+/-10nM; P=0.0007) compared to wild type animals (71+/-11nM). Hyperuricemia was associated with >1.5 fold increase in plasma kynurenine levels in all strains. Moreover, hyperuricemia led to elevated plasma kynurenic acid levels (128+/-13nM, P=0.005) in wild type mice but did not further increase kynurenic acid levels in knockout mice. Based on our results, we postulate that elevated uric acid levels hamper MRP4 and BCRP functioning, thereby promoting the retention of other potentially toxic substrates, including kynurenic acid, which could contribute to the development of CKD.

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37 The vital role of transporters in uric acid homeostasis can clearly be observed in patients suffering from hyperuricemia due to single nucleotide polymorphisms (SNPs) that render the transporters inactive, such as the common SNP C421A encoding the Q141K mutation of BCRP [21,22,24] and several genetic variants for SLC2A9 [19]. Login to comment
224 For instance, Woodward et al. [22], described that the common single nucleotide polymorphism (SNP) rs2231142 encoding the Q141K mutation of BCRP caused hyperuricemia-based gout. Login to comment
225 Also in a Japanese population, Q141K was shown to be a common dysfunctional form of BCRP causing gout [21]. Login to comment

[hide] A multicenter clinical study evaluating the confir... Haematologica. 2013 Sep;98(9):1407-13. doi: 10.3324/haematol.2013.085167. Epub 2013 May 28. Shinohara Y, Takahashi N, Nishiwaki K, Hino M, Kashimura M, Wakita H, Hatano Y, Hirasawa A, Nakagawa Y, Itoh K, Masuoka H, Aotsuka N, Matsuura Y, Takahara S, Sano K, Kuroki J, Hata T, Nakamae H, Mugitani A, Nakane T, Miyazaki Y, Niioka T, Miura M, Sawada K
A multicenter clinical study evaluating the confirmed complete molecular response rate in imatinib-treated patients with chronic phase chronic myeloid leukemia by using the international scale of real-time quantitative polymerase chain reaction.
Haematologica. 2013 Sep;98(9):1407-13. doi: 10.3324/haematol.2013.085167. Epub 2013 May 28., [PMID:23716542]

Abstract [show]
Achievement of complete molecular response in patients with chronic phase chronic myeloid leukemia has been recognized as an important milestone in therapy cessation and treatment-free remission; the identification of predictors of complete molecular response in these patients is, therefore, important. This study evaluated complete molecular response rates in imatinib-treated chronic phase chronic myeloid leukemia patients with major molecular response by using the international standardization for quantitative polymerase chain reaction analysis of the breakpoint cluster region-Abelson1 gene. The correlation of complete molecular response with various clinical, pharmacokinetic, and immunological parameters was determined. Complete molecular response was observed in 75/152 patients (49.3%). In the univariate analysis, Sokal score, median time to major molecular response, ABCG2 421C>A, and regulatory T cells were significantly lower in chronic phase chronic myeloid leukemia patients with complete molecular response than in those without complete molecular response. In the multivariate analysis, duration of imatinib treatment (odds ratio: 1.0287, P=0.0003), time to major molecular response from imatinib therapy (odds ratio: 0.9652, P=0.0020), and ABCG2 421C/C genotype (odds ratio: 0.3953, P=0.0284) were independent predictors of complete molecular response. In contrast, number of natural killer cells, BIM deletion polymorphisms, and plasma trough imatinib concentration were not significantly associated with achieving a complete molecular response. Several predictive markers for achieving complete molecular response were identified in this study. According to our findings, some chronic myeloid leukemia patients treated with imatinib may benefit from a switch to second-generation tyrosine kinase inhibitors (ClinicalTrials.gov, UMIN000004935).

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223 Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, Tsuruo T, et al. C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Common dysfunctional variants in ABCG2 are a major... Sci Rep. 2013;3:2014. doi: 10.1038/srep02014. Matsuo H, Ichida K, Takada T, Nakayama A, Nakashima H, Nakamura T, Kawamura Y, Takada Y, Yamamoto K, Inoue H, Oikawa Y, Naito M, Hishida A, Wakai K, Okada C, Shimizu S, Sakiyama M, Chiba T, Ogata H, Niwa K, Hosoyamada M, Mori A, Hamajima N, Suzuki H, Kanai Y, Sakurai Y, Hosoya T, Shimizu T, Shinomiya N
Common dysfunctional variants in ABCG2 are a major cause of early-onset gout.
Sci Rep. 2013;3:2014. doi: 10.1038/srep02014., [PMID:23774753]

Abstract [show]
Gout is a common disease which mostly occurs after middle age, but more people nowadays develop it before the age of thirty. We investigated whether common dysfunction of ABCG2, a high-capacity urate transporter which regulates serum uric acid levels, causes early-onset gout. 705 Japanese male gout cases with onset age data and 1,887 male controls were genotyped, and the ABCG2 functions which are estimated by its genotype combination were determined. The onset age was 6.5 years earlier with severe ABCG2 dysfunction than with normal ABCG2 function (P = 6.14 x 10(-3)). Patients with mild to severe ABCG2 dysfunction accounted for 88.2% of early-onset cases (twenties or younger). Severe ABCG2 dysfunction particularly increased the risk of early-onset gout (odds ratio 22.2, P = 4.66 x 10(-6)). Our finding that common dysfunction of ABCG2 is a major cause of early-onset gout will serve to improve earlier prevention and therapy for high-risk individuals.

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17 We also showed that genotyping of only two dysfunctional variants, Q126X (rs72552713) and Q141K (rs2231142), is sufficient to estimate the severity of ABCG2 dysfunction; i.e. full function, 3/4 function (mild dysfunction), 1/2 function (moderate dysfunction), and # 1/4 function (severe dysfunction). Login to comment
39 Table 1 | ABCG2 functions of participants Estimated Function Genotype Combination Number (%) Q126X* (rs72552713) Q141K* (rs2231142) Gout Control #1/4 function T/T C/C 37 (5.2) 22 (1.2) T/C C/A 1/2 function T/C C/C 169 (24.0) 219 (11.6) C/C A/A 3/4 function C/C C/A 331 (47.0) 699 (37.0) Full function C/C C/C 168 (23.8) 947 (50.2) Total 705 (100.0) 1,887 (100.0) * Risk alleles (T for Q126X, A for Q141K) are indicated in bold type at four locations, respectively. Login to comment
73 Genotyping of Q126X (rs72552713) and Q141K (rs2231142) in ABCG2 gene by high-resolution melting (HRM) analysis was performed with a LightCycler 480 (Roche Diagnostics)41 . Login to comment

[hide] Effects of the gout-causing Q141K polymorphism and... Biochem Biophys Res Commun. 2013 Jul 19;437(1):140-5. doi: 10.1016/j.bbrc.2013.06.054. Epub 2013 Jun 22. Saranko H, Tordai H, Telbisz A, Ozvegy-Laczka C, Erdos G, Sarkadi B, Hegedus T
Effects of the gout-causing Q141K polymorphism and a CFTR DeltaF508 mimicking mutation on the processing and stability of the ABCG2 protein.
Biochem Biophys Res Commun. 2013 Jul 19;437(1):140-5. doi: 10.1016/j.bbrc.2013.06.054. Epub 2013 Jun 22., [PMID:23800412]

Abstract [show]
ABCG2 is an important multidrug transporter involved also in urate transport, thus its mutations can lead to the development of gout and may also alter general drug absorption, distribution and excretion. The frequent ABCG2 polymorphism, Q141K, is associated with an elevated risk of gout and has been controversially reported to reduce the plasma membrane expression and/or the transport function of the protein. In the present work we examined the stability and cellular processing of the Q141K ABCG2 variant, as well as that of the DeltaF142 ABCG2, corresponding to the DeltaF508 mutation in the CFTR (ABCC7) protein, causing cystic fibrosis. The processing and localization of full length ABCG2 variants were investigated in mammalian cells, followed by Western blotting and confocal microscopy, respectively. Folding and stability were examined by limited proteolysis of Sf9 insect cell membranes expressing these ABCG2 constructs. Stability of isolated nucleotide binding domains, expressed in and purified from bacteria, was studied by CD spectroscopy. We find that the Q141K variant has a mild processing defect which can be rescued by low temperature, a slightly reduced activity, and a mild folding defect, especially affecting the NBD. In contrast, the DeltaF142 mutant has major processing and folding defects, and no ATPase function. We suggest that although these mutations are both localized within the NBD, based on molecular modeling their contribution to the ABCG2 structure and function is different, thus rescue strategies may be devised accordingly.

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0 Effects of the gout-causing Q141K polymorphism and a CFTR DF508 mimicking mutation on the processing and stability of the ABCG2 protein Hajnalka Sarank&#f3; a,b,1 , Hedvig Tordai b,1 , &#c1;gnes Telbisz c , Csilla &#d6;zvegy-Laczka d , G&#e1;bor Erd} os a,b , Bal&#e1;zs Sarkadi b,c,d , Tam&#e1;s Heged} us a,b,d1; a MTA-SE Molecular Biophysics Research Group, Budapest, Hungary b Department of Biophysics and Radiation Biology, Semmelweis University, Budapest, Hungary c Institute of Molecular Pharmacology, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary d National Blood Center, Budapest, Hungary a r t i c l e i n f o Article history: Received 12 June 2013 Available online 22 June 2013 Keywords: ABCG2 Polymorphism Gout Cystic fibrosis Stability Structure a b s t r a c t ABCG2 is an important multidrug transporter involved also in urate transport, thus its mutations can lead to the development of gout and may also alter general drug absorption, distribution and excretion. Login to comment
1 The frequent ABCG2 polymorphism, Q141K, is associated with an elevated risk of gout and has been controversially reported to reduce the plasma membrane expression and/or the transport function of the protein. Login to comment
2 In the present work we examined the stability and cellular processing of the Q141K ABCG2 variant, as well as that of the DF142 ABCG2, corresponding to the DF508 mutation in the CFTR (ABCC7) protein, causing cystic fibrosis. Login to comment
6 We find that the Q141K variant has a mild processing defect which can be rescued by low temperature, a slightly reduced activity, and a mild folding defect, especially affecting the NBD. Login to comment
26 A frequent SNP in the ABCG2 gene produces a Q141K variant [5] of the protein. Login to comment
28 The Q141K variation has been reported to result in altered processing and a less efficient substrate transport by ABCG2 [6-8]. Login to comment
29 Expression of ABCG2 variants in single gene copy systems suggested that the Q141K variant has a reduced plasma membrane expression level and increased intracellular concentration [9,10]. Login to comment
30 In order to rescue the Q141K variant phenotype, which may be relevant in preventing gout and increasing protection against xenobiotics, the molecular details associated with this amino acid change have to be explored. Login to comment
31 The Q141K change is located in the NBD of ABCG2, adjacent to F142, which has an analogous position to F508 of the CFTR (ABCC7) protein (Figs. Login to comment
34 Therefore it has been suggested that the ABCG2 Q141K variant may cause similar changes to the DF508 mutation, disrupting the interface between the NBD and the TMD [2,11]. Login to comment
40 In the present study we compared the biochemical properties of the wild-type ABCG2 to those of the Q141K variant and the DF142 mutation. Login to comment
43 Our results indicate that the Q141K polymorphism reduces the efficient processing and membrane targeting, while has only a mild effect on the structure and function of ABCG2. Login to comment
80 The Q141K variant localizes to the plasma membrane, in contrast to the DF142 mutant In order to examine the effects of Q141K and DF142 alterations on the ABCG2 protein expression and localization, we transiently expressed these constructs in HEK cells. Login to comment
82 As shown in Fig. 1A, the Q141K variant exhibited significant level of maturation, although the amount of the fully glycosylated form was significantly lower, compared to that of the wild type ABCG2. Login to comment
87 Both the wild type and Q141K ABCG2 were found to be expressed mostly in the plasma membrane, although the Q141K variant exhibited a higher level of intracellular staining (Fig. 1B). Login to comment
90 These experiments confirmed lower cell surface expression of the Q141K variant compared to the wild type, whereas cells expressing the DF142 mutant did not bind the 5D3 antibody. Login to comment
91 Our results demonstrate that the effect of the Q141K variation causes a milder phenotype in ABCG2 than the DF142 mutation, while this latter mutation results in an expression deficiency similar to that of DF508 CFTR. Login to comment
93 Phenylbutyrate treatment increases the level of mature WT and Q141K ABCG2, while do not rescue DF142 ABCG2 To evaluate the potential folding and trafficking deficiency of the ABCG2 Q141K and DF142 proteins, their expression was characterized under more permissive conditions. Login to comment
97 As shown in Fig. 2, in transiently transfected HEK cells the level of the wild type and the Q141K proteins slightly decreased at low temperature (Fig. 2), indicating the adverse effect of low temperature on protein synthesis. Login to comment
100 Expression of ABCG2 WT, Q141K, and DF142 constructs in HEK cells. Login to comment
120 Interestingly, one of the rescue mutations, G188E has been reported to promote Q141K maturation [11]. Login to comment
121 Most likely, similarly to the homologous CFTR G550E [13,21], this mutation increases the thermostability of NBD in an extent that is sufficient to counteract the effect of the mild Q141K mutation, but not that of the DF142. Login to comment
128 The Q141K NBD is more stable than the DF142 NBD, indicated by limited proteolysis experiments Since the quality control system in insect cells is less stringent than in mammalian cells, Sf9 cells were used to express the Q141K, DF142, and WT ABCG2 for functional and biochemical studies. Login to comment
129 In Sf9 cells the expression levels of the wild type and the Q141K ABCG2 variant were similar, while the expression of the deletion mutant was significantly lower (Fig. 3A). Login to comment
130 The vanadate sensitive ATPase activity of the Q141K variant in isolated Sf9 membrane was approximately 70% of that of the wild type protein ([8] and Fig. S6A). Login to comment
134 Membranes expressing Q141K, DF142, and WT ABCG2 were Table 1 Cell surface expression of the ABCG2 constructs quantitated by flow cytometry. Login to comment
135 Geometric mean values HEK parent EGFP only ABCG2 WT ABCG2 Q141K ABCG2 DF142 ABCG2 3R ABCG2DF142 3R Isotype control 7.5 7.6 7.3 7.2 7 8.4 7 Isotype control + Ko143 6.99 7.3 7.3 7.1 7 8.3 7.44 5D3 9.45 12 110 78 10 146 8.4 5D3 + Ko143 30 29.5 210 150 27 152 27.4 Transfection efficiency EGFP FL1 0% 85% 93% 89% 89% 83% 89% Fig. 2. Login to comment
150 Although the rate of Q141K degradation was somewhat greater than that of the WT, it was clearly smaller than the rate of the DF142 fragment. Login to comment
153 (A) WT and Q141K ABCG2 are expressed at similar levels in membranes isolated from Sf9 insect cells, as detected by Western blotting using the BXP-21 antibody at 60 kDa. Login to comment
162 (A) Far UV CD spectra of WT, Q141K, and DF142 NBD fused to MBP, expressed in and isolated from E. coli. Login to comment
164 (B) Thermal unfolding of isolated MBP-fused NBD constructs was followed by measuring CD at 222 nm between 20 &#b0;C and 90 &#b0;C. Q141K variant may also be caused by a potential extra trypsin cleavage site introduced by the Q141K variation [11]. Login to comment
166 The thermal stability of the isolated Q141K NBD is similar to the WT, in contrast to the DF142 To identify the effects of the amino acid changes on the NBD conformation and dynamics, isolated NBD constructs fused to MBP, were purified from bacteria (the MBP could not be removed because of a rapid precipitation of the isolated NBDs). Login to comment
172 We found that the melting curve of the Q141K variant was similar to that of the wild type, while the melting profile of the DF142 differed significantly. Login to comment
173 As a summary, we found that the ABCG2 Q141K variant is expressed in the plasma membrane at a lower level and with a somewhat decreased function, as compared to the wild type protein. Login to comment
175 While the Q141K variation is located in a position adjacent to the F508 in CFTR, their structural roles and functions are significantly different based on our experiments and in silico analysis (Fig. S2). Login to comment
177 Moreover, studies focusing on positions similar to the ABCG2 Q141K variant in other ABC transporters may help to understand alterations at the protein level in various diseases, and promote the development of successful treatments. Login to comment

[hide] Association between the ABCG2 C421A polymorphism a... Neurosci Lett. 2013 Aug 29;550:51-4. doi: 10.1016/j.neulet.2013.06.044. Epub 2013 Jul 1. Feher A, Juhasz A, Laszlo A, Pakaski M, Kalman J, Janka Z
Association between the ABCG2 C421A polymorphism and Alzheimer's disease.
Neurosci Lett. 2013 Aug 29;550:51-4. doi: 10.1016/j.neulet.2013.06.044. Epub 2013 Jul 1., [PMID:23827224]

Abstract [show]
ATP binding cassette subfamily G member 2 (ABCG2) is involved in amyloid-beta transport and was found to be significantly up-regulated in Alzheimer's disease (AD) brain. A functional polymorphism of the ABCG2 gene (C421A; rs2231142) was genotyped in a sample of 299 Hungarian late-onset AD patients and 259 elderly, non-demented controls to investigate for the first time its association with AD, either alone or in combination with apolipoprotein E (APOE) varepsilon2/varepsilon3/varepsilon4 polymorphism. A significantly increased susceptibility to AD (OR=1.741, 95% CI: 1.075-2.819, p=0.024) associated with ABCG2 C/C genotype was found when compared with the variant allele containing genotypes (CA and AA) as the reference category. Logistic regression analysis revealed a significant interaction effect between the ABCG2 C/C genotype and APOE varepsilon4 allele on AD risk (p=0.003). It seems that the potential modest risk effect of the ABCG2 C/C genotype on AD risk is more pronounced in combination with the APOE varepsilon4 allele. Further independent replications of our findings are required.

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39 The C421A non-synonymous single nucleotide polymorphism (/SNP/Q141K, rs2231142) of the ABCG2 gene is reportedly associated with lower levels of protein expression, impaired transport, and enhanced susceptibility to ubiquitin-mediated proteosomal degradation [12,15,16,23,24]. Login to comment

[hide] Serum uric acid and non-alcoholic fatty liver dise... PLoS One. 2013 Jul 23;8(7):e67152. doi: 10.1371/journal.pone.0067152. Print 2013. Xie Y, Wang M, Zhang Y, Zhang S, Tan A, Gao Y, Liang Z, Shi D, Huang Z, Zhang H, Yang X, Lu Z, Wu C, Liao M, Sun Y, Qin X, Hu Y, Li L, Peng T, Li Z, Yang X, Mo Z
Serum uric acid and non-alcoholic fatty liver disease in non-diabetic Chinese men.
PLoS One. 2013 Jul 23;8(7):e67152. doi: 10.1371/journal.pone.0067152. Print 2013., [PMID:23935829]

Abstract [show]
Increased serum uric acid (SUA) levels may be involved in the development of non-alcoholic fatty liver disease (NAFLD) in men presenting with metabolic syndrome (MetS) and/or insulin resistance. We aimed to determine the independent relationship between SUA and NAFLD in non-diabetic Chinese male population, and to explore the determinants of SUA levels among indexes of adiposity, lipid, and genotypes pertaining to triglycerides metabolism, inflammation, oxidative stress, and SUA concentrations. A total of 1440 men, classified depending on the presence of ultrasonographically detected NAFLD, underwent a complete healthy checkup program. Genotypes were extracted from our previously established genome-wide association study database. After adjusting for age, smoking, drinking, body mass index, homeostasis model assessment of insulin resistance, C-reactive protein, creatinine, alanine aminotransferase (ALT) and components of metabolic syndrome, the odds ratio for NAFLD, comparing the highest with the lowest SUA quartile, was 2.81 (95% confidence interval 1.66-4.76). A stepwise multivariate linear regression analysis (R(2) = 0.238, P<0.001) retained age, waist circumference, serum creatinine, triglycerides, the Q141K variant in ABCG2 (rs2231142) and NAFLD as significant predictors of SUA levels (all P<0.001). Besides, ALT and Met196Arg variant in TNFRSF1B (rs1061622) additionally associated with SUA among individuls with NAFLD. Our data suggest that in Chinese men, elevated SUA is significantly associated with NAFLD, independent of insulin resistance and other metabolic disorders, such as central obesity or hypertriglyceridemia. Meanwhile, among subjects with NAFLD, index of liver damage, such as elevated ALT combined with genetic susceptibility to inflammation associated with increased SUA levels.

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5 A stepwise multivariate linear regression analysis (R2 = 0.238, P,0.001) retained age, waist circumference, serum creatinine, triglycerides, the Q141K variant in ABCG2 (rs2231142) and NAFLD as significant predictors of SUA levels (all P,0.001). Login to comment
88 Among all subjects, a final constructed model using a stepwise method (probability to enter #0.05; to remove $0.10), found age (b = 20.11, 95% CI 20.16 to 20.06), WC (b = 0.17, 95% CI 0.11-0.23), NAFLD (b = 0.15, 95% CI 0.09-0.21), log-transformed serum creatinine (b = 0.29, 95% CI, 0.24-0.34) and triglycerides (b = 0.11, 95% CI 0.05-0.16), as well as the Q141K variant in ABCG2 gene (b = 0.12, 95% CI 0.07-0.17) as significant predictors (all P,0.001) of the logarithm of SUA levels (R2 = 0.238, P,0.001). Login to comment
112 Our present result also suggested that the Q141K variant in ABCG2, leads to variable degree of SUA concentration in men, in conceptual agreement with the ABCG2`s function of altering uric acid transport in kidney proximal tubule cell and excretion in liver via the biliary system [37]. Login to comment
137 SUA are interrelated with age, waist circumference, NAFLD, creatinine, triglycerides, and the Q141K variant in ABCG2 in non-diabetic Chinese men. Login to comment

[hide] The FLT3 inhibitor quizartinib inhibits ABCG2 at p... PLoS One. 2013 Aug 14;8(8):e71266. doi: 10.1371/journal.pone.0071266. eCollection 2013. Bhullar J, Natarajan K, Shukla S, Mathias TJ, Sadowska M, Ambudkar SV, Baer MR
The FLT3 inhibitor quizartinib inhibits ABCG2 at pharmacologically relevant concentrations, with implications for both chemosensitization and adverse drug interactions.
PLoS One. 2013 Aug 14;8(8):e71266. doi: 10.1371/journal.pone.0071266. eCollection 2013., [PMID:23967177]

Abstract [show]
The oral second-generation bis-aryl urea fms-like tyrosine kinase 3 (FLT3) inhibitor quizartinib (AC220) has favorable kinase selectivity and pharmacokinetics. It inhibits mutant and wild-type FLT3 in vivo at 0.1 and 0.5 microM, respectively, and has shown favorable activity and tolerability in phase I and II trials in acute myeloid leukemia, with QT prolongation as the dose-limiting toxicity. Co-administration with chemotherapy is planned. We characterized interactions of quizartinib with the ATP-binding cassette (ABC) proteins ABCB1 (P-glycoprotein) and ABCG2 (breast cancer resistance protein). Its effects on uptake of fluorescent substrates and apoptosis were measured by flow cytometry, binding to ABCB1 and ABCG2 drug-binding sites by effects on [(1)(2)(5)I]iodoarylazidoprazosin ([(1)(2)(5)I]-IAAP) photolabeling and ATPase activity, and cell viability by the WST-1 colorimetric assay. Quizartinib inhibited transport of fluorescent ABCG2 and ABCB1 substrates in ABCG2- and ABCB1-overexpressing cells in a concentration-dependent manner, from 0.1 to 5 microM and from 0.5 to 10 microM, respectively, and inhibited [(1)(2)(5)I]-IAAP photolabeling of ABCG2 and ABCB1 with IC(5)(0) values of 0.07 and 3.3 microM, respectively. Quizartinib at higher concentrations decreased ABCG2, but not ABCB1, ATPase activity. Co-incubation with quizartinib at 0.1 to 1 microM sensitized ABCG2-overexpressing K562/ABCG2 and 8226/MR20 cells to ABCG2 substrate chemotherapy drugs in a concentration-dependent manner in cell viability and apoptosis assays. Additionally, quizartinib increased cellular uptake of the ABCG2 substrate fluoroquinolone antibiotic ciprofloxacin, which also prolongs the QT interval, in a concentration-dependent manner, predicting altered ciprofloxacin pharmacokinetics and pharmacodynamics when co-administered with quizartinib. Thus quizartinib inhibits ABCG2 at pharmacologically relevant concentrations, with implications for both chemosensitization and adverse drug interactions. These interactions should be considered in the design of treatment regimens combining quizartinib and chemotherapy drugs and in choice of concomitant medications to be administered with quizartinib.

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159 ABCG2 has been found to be a urate efflux transporter, with increased incidence of both hyperuricemia and gout in association with the Q141K ABCG2 single nucleoside polymorphism, which results in decreased urate transport [69]. Login to comment

[hide] Pharmacogenetic determinants associated with sunit... Cancer Chemother Pharmacol. 2013 Oct;72(4):825-35. doi: 10.1007/s00280-013-2258-y. Epub 2013 Sep 8. Kim HR, Park HS, Kwon WS, Lee JH, Tanigawara Y, Lim SM, Kim HS, Shin SJ, Ahn JB, Rha SY
Pharmacogenetic determinants associated with sunitinib-induced toxicity and ethnic difference in Korean metastatic renal cell carcinoma patients.
Cancer Chemother Pharmacol. 2013 Oct;72(4):825-35. doi: 10.1007/s00280-013-2258-y. Epub 2013 Sep 8., [PMID:24013576]

Abstract [show]
PURPOSE: The aim of this study was to investigate the pharmacogenetic determinants of sunitinib-related toxicity and ethnic difference in metastatic renal cell carcinoma (mRCC) among Korean patients. METHODS: A pharmacogenetic study was performed in 65 patients with mRCC treated with the standard schedule of sunitinib (50 mg orally once daily for 4 weeks-on/2 weeks-off). Detailed data regarding the toxicity of sunitinib, including thrombocytopenia, neutropenia, anemia, and hand-foot syndrome (HFS), were prospectively collected in a clinical trial program (n = 38) or standard oncology practice (n = 27). Total of 12 genetic polymorphisms in 8 candidate genes (CYP1A1, CYP3A5, ABCB1, ABCG2, PDGFRalpha, VEGFR2, RET, and FLT3) were analyzed for an association with treatment-related toxicity from sunitinib using Pearson chi (2) test. RESULTS: Common grade 3 or grade 4 treatment-related toxicities were thrombocytopenia (36.9 %, 24/65), neutropenia (18.4 %, 12/65), anemia (7.7 %, 5/65), and HFS (12.3 %, 8/65). Patients carrying an ABCG2 421 AA genotype developed significantly more grade 3 or grade 4 thrombocytopenia, neutropenia, and HFS adjusted for age, sex, and Eastern Cooperative Oncology Group performance status, and body surface area (odds ratio compared with AC/CC genotypes [OR] 9.90, P = 0.04, thrombocytopenia; OR 18.20, P = 0.02, neutropenia; and OR 28.46, P = 0.01, HFS). In addition, total and surface protein ABCG2 protein expression was decreased in ABCG2 421 AA mutant cells compared to wild type. CONCLUSION: Among 12 genetic polymorphisms, polymorphism in the ABCG2 421C>A gene may be mostly associated with the risk of sunitinib-related toxicity in mRCC patients. Considering the high frequency of 421C>A SNP in Asian, this may be related to differential toxicities among ethnic groups.

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97 With the median 23.8 (range 1.1-57.9) months of TableÊf;1ߒߙPolymorphism genotyped in the pharmacokinetic and pharmacodynamic pathways of sunitinib Gene rs number Polymorphism Region CYP1A1 Cytochrome P450 1A1 rs1048943 2,455 A>G Non-synonymous I462V CYP3A5 Cytochrome P450 3A5 rs776746 219-237 A>G Intron ABCB1 ATP binding cassette member B1 rs1128503 1,236 C>T Synonymous G412G rs2032582 2,677 G>T/A Non-synonymous A893S/T rs1045642 3,435 C>T Synonymous I1145I ABCG2 (BCRP) ATP binding cassette member G2 rs2231142 421 C>A Non-synonymous Q141K PDGFRb1; Platelet-derived growth factor receptor rs1800812 -537 G>T Promoter rs35597368 1,580 C>T Non-synonymous P478S VEGFR2 (KDR) Vascular endothelial growth factor receptors rs2305948 889 G>A Non-synonymous V297I rs1870377 1,416 T>A Non-synonymous H472Q RET Ret proto-oncogene rs1799939 2,071 G>A Non-synonymous G691S FLT3 Fms-like tyrosine kinase 3 receptor rs1933437 680 C>T Non-synonymous T227M follow-up, the median number of treatment cycle was 7 (range 1-38). Login to comment
113 Among the 12 candidate SNPs, the ABCG2 421C>A (Q141K) polymorphism showed a strong association with sunitinib-related toxicity, especially HFS, at the allele frequency level. Login to comment
125 TableÊf;3ߒߙDistribution of toxicity during first cycle a ߒ Toxicities were evaluated by NCI-CTC version 3.0 b ߒ "No" represents grade 0,1, or 2 toxicities c ߒ "Yes" represents grade 3 or 4 toxicities Toxicity gradea No % Thrombocytopenia 30 46.2 1 1 1.5 2 5 7.7 3 23 35.4 4 1 1.5 Neutropenia 30 46.2 1 8 12.3 2 10 15.4 3 11 16.9 4 1 1.5 Anemia 21 32.3 1 11 16.9 2 5 7.7 3 5 7.7 4 0 0 Any hematologic toxicity >grade 2 Nob 38 58.4 Yesc 27 41.6 Hand-foot syndrome 33 50.8 1 15 23.1 2 10 15.4 3 8 12.3 4 0 0 TableÊf;4ߒߙAssociation of each genetic variation with sunitinib-related toxicity in mRCC patients Gene name; polymorphic site, allele; amino acid; rs number Polymorphism Minor Thrombocytopenia Neutropenia Anemia Hand-foot syndrome Allele Odds ratio 95 % CI P value Odds ratio 95 % CI P value Odds ratio 95 % CI P value Odds ratio 95 % CI P value CYP1A1 c.2,455 A>G I462V G 0.6 0.24-1.49 0.27 0.45 0.12-1.64 0.22 0.61 0.14-2.54 0.44 1.22 0.36-4.13 0.75 CYP3A5 c.219-237 A>G Intron 3 A 0.42 0.17-1.08 0.07 0.60 0.19-1.92 0.39 1.46 0.35-6.03 0.69 0.43 0.09-2.00 0.36 ABCB1 c.1,236 C>T G412G C 1.12 0.54-2.31 0.77 0.56 0.21-1.46 0.23 1.0 0.26-3.73 0.63 0.65 0.21-1.99 0.45 ABCB1 c.2,677 G>T/A A893S/T T 1.00 0.48-2.11 0.99 1.12 0.45-2.80 0.81 0.29 0.06-1.45 0.18 1.11 0.37-3.28 0.85 ABCB1 c.3,435 C>T I1145I T 0.87 0.41-1.83 0.71 0.89 0.35-2.28 0.82 0.33 0.08-1.25 0.16 1.11 0.38-3.28 0.85 ABCG2 c.421 C>A Q141K A 1.74 0.81-3.75 0.15 1.89 0.76-4.75 0.17 1.85 0.38-9.14 0.72 6.76 2.16-21.13 0.00003 PDGFRb1; -573G>T Promoter T 0.508 0.19-1.38 0.18 0.347 0.076-1.590 0.244 2.04 0.24-16.95 0.69 0.264 0.03-2.10 0.302 PDGFRb1; c.1,580 C>T P478S C 0.638 0.23-1.77 0.386 0.416 0.090-1.923 0.362 1.80 0.22-15.01 0.49 0.313 0.04-2.51 0.467 VEGFR2 c.889 G>A V297I A 0.48 0.15-1.58 0.22 0.25 0.03-1.94 0.19 1.09 0.92-1.16 0.35 2.59 0.73-9.23 0.23 VEGFR2 c.1,416 T>A H472Q T 0.87 0.42-1.79 0.70 0.89 0.37-2.20 0.81 1.84 0.45-7.48 0.514 1.33 0.47-3.78 0.59 RET c.2,071 G>A G691S A 0.68 0.22-2.06 0.49 2.88 0.94-8.78 0.09 0.57 0.11-2.94 0.61 2.59 0.73-9.22 0.23 FLT3 c.680 C>T T227M C 1.24 0.57-2.71 0.59 1.671 0.658-4.246 0.277 0.56 0.15-2.14 0.46 0.818 0.25-2.72 1 Pharmacogenetic determinants for sunitinib treatment outcomes Among the 12 single-nucleotide polymorphisms in 8 candidate genes, only a few polymorphisms from pharmacodynamic genes were associated with OS. Login to comment
132 This finding implicates that patients with Q141K variation of ABCG2 show reduced ABCG2 transporter activity. Login to comment
142 Total and surface protein in the Q141K mutant transfected cells was decreased compared with those in the wild-type ABCG2 cells. Login to comment
161 Our in vitro data as well as previous data demonstrated that the non-synonymous SNP of ABCG2 421 C>A, which causes a glutamine-to-lysine amino acid substitution at position141 (Q141K), has been associated with markedly decreased levels of ABCG2 protein expression and/or activity [23, 31, 32]. Login to comment

[hide] Molecular pharmacology of ABCG2 and its role in ch... Mol Pharmacol. 2013 Nov;84(5):655-69. doi: 10.1124/mol.113.088609. Epub 2013 Sep 10. Stacy AE, Jansson PJ, Richardson DR
Molecular pharmacology of ABCG2 and its role in chemoresistance.
Mol Pharmacol. 2013 Nov;84(5):655-69. doi: 10.1124/mol.113.088609. Epub 2013 Sep 10., [PMID:24021215]

Abstract [show]
The ATP-binding cassette, subfamily G, isoform 2 protein (ABCG2) is an important member of the ABC transporter superfamily, which has been suggested to be involved in multidrug resistance (MDR) in cancer. Its diverse range of substrates includes many common chemotherapeutics such as imatinib, doxorubicin, and mitoxantrone. Physiologically, ABCG2 is highly expressed in areas such as the blood-brain barrier and gastrointestinal tract, where it is thought to play a role in protection against xenobiotic exposure. High ABCG2 expression has also been found in a variety of solid tumors and in hematologic malignancies and has been correlated with poorer clinical outcomes. Furthermore, ABCG2 expression is a characteristic feature of cancer stem cells, which are able to self-renew and differentiate. These cancer stem cells have been postulated to play an important role in MDR, where their inherent ABCG2 expression may allow them to survive chemotherapy and repopulate the tumor after exposure to chemotherapeutics. This observation raises the exciting possibility that by inhibiting ABCG2, cancer stem cells and other cancers may be targeted and eradicated, at which point conventional chemotherapeutics would be sufficient to eliminate the remaining tumor cells. Inhibitors of ABCG2, such as tyrosine kinase inhibitors, phosphodiesterase-5 inhibitors, and the fumitremorgin-type indolyl diketopiperazine, Ko143 [(3S,6S,12aS)-1,2,3,4,6,7,12,12a-octahydro-9-methoxy-6-(2-methylpropyl)-1,4-dioxo pyrazino[1',2':1,6]pyrido[3,4-b]indole-3-propanoic acid 1,1-dimethylethyl ester], could potentially be used for this purpose. However, these agents are still awaiting comprehensive clinical assessment.

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40 The two most frequent polymorphisms identified were the G34A (resulting in V12M) and C421A (resulting in a Q141K substitution) transitions (Fig. 3), found in 18 and 35.5% of the studied population, respectively (Kobayashi et al., 2005). Login to comment
46 This was not corroborated by Morisaki et al. (2005), who found that cells with the Q141K variant of ABCG2 had IC50 values 1.2-to 5-fold lower than cells transfected with wild-type (Arg482) ABCG2, suggesting that the Q141K SNP affects drug transport (Morisaki et al., 2005). Login to comment
47 Recently, studies aimed at evaluating the clinical relevance of the Q141K SNP in patients undergoing chemotherapy have also been reported. Login to comment
50 Additionally, the Q141K mutation has been demonstrated to decrease ATPase activity by 1.3-fold compared with wild-type ABCG2 (Mizuarai et al., 2004). Login to comment
51 Furthermore, kinetic analysis of ATPase activity showed that the Km value in Q141K cells was 1.4-fold higher than that of wild-type ABCG2 (Mizuarai et al., 2004). Login to comment
79 Val12M and Q141K are SNPs thought to affect the pharmacokinetics of ABCG2 substrates. Login to comment

[hide] Metabolic Interactions of Purine Derivatives with ... Pharmaceuticals (Basel). 2013 Nov 4;6(11):1347-60. doi: 10.3390/ph6111347. Ishikawa T, Aw W, Kaneko K
Metabolic Interactions of Purine Derivatives with Human ABC Transporter ABCG2: Genetic Testing to Assess Gout Risk.
Pharmaceuticals (Basel). 2013 Nov 4;6(11):1347-60. doi: 10.3390/ph6111347., [PMID:24287461]

Abstract [show]
In mammals, excess purine nucleosides are removed from the body by breakdown in the liver and excretion from the kidneys. Uric acid is the end product of purine metabolism in humans. Two-thirds of uric acid in the human body is normally excreted through the kidney, whereas one-third undergoes uricolysis (decomposition of uric acid) in the gut. Elevated serum uric acid levels result in gout and could be a risk factor for cardiovascular disease and diabetes. Recent studies have shown that human ATP-binding cassette transporter ABCG2 plays a role of renal excretion of uric acid. Two non-synonymous single nucleotide polymorphisms (SNPs), i.e., 421C>A (major) and 376C>T (minor), in the ABCG2 gene result in impaired transport activity, owing to ubiquitination-mediated proteosomal degradation and truncation of ABCG2, respectively. These genetic polymorphisms are associated with hyperuricemia and gout. Allele frequencies of those SNPs are significantly higher in Asian populations than they are in African and Caucasian populations. A rapid and isothermal genotyping method has been developed to detect the SNP 421C>A, where one drop of peripheral blood is sufficient for the detection. Development of simple genotyping methods would serve to improve prevention and early therapeutic intervention for high-risk individuals in personalized healthcare.

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81 The SNP 421C>A is a non-synonymous polymorphism that leads to amino acid substitution; Gln to Lys (Q141K) in the intracellular loop containing an ATP-binding cassette (ABC). Login to comment
82 608 592 603 592 Extracellular loop Outside Inside Plasma membrane 596 N-linked glycan Asn Cys ATP-binding cassette (ABC) Q141K Q141K 608 592 603 592 Extracellular loop Outside Inside Plasma membrane 596 N-linked glycan Asn Cys ATP-binding cassette (ABC) 608 592 603 592 Extracellular loop Outside Inside Plasma membrane 596 N-linked glycan Asn Cys 608 592 603 592 Extracellular loop Outside Inside Plasma membrane 596 N-linked glycan Asn Cys N-linked glycan Asn Cys ATP-binding cassette (ABC) Q141K Q141K ABCG2 expressed on the apical side of the proximal tubular cells in human kidney plays a pivotal role in renal excretion of serum uric acid. Login to comment
83 Introduction of the mutation Q141K encoded by the common SNP (rs2231142) by site-directed mutagenesis resulted in 53% reduced urate transport rates compared to wild-type ABCG2 (p < 0.001). Login to comment
84 The expression levels of the Q141K variant are reduced by ubiquitin-mediated proteasomal degradation [31-34] (Figure 5). Login to comment
85 Thus, renal excretion of serum uric acid via ABCG2 is impaired in persons who are carrying the 421A allele (Q141K variant). Login to comment
88 Effect of the SNP variant (Q141K) on the protein expression level and degradation of ABCG2. Login to comment
90 To assess the effect of Q141K variant on the protein expression level, Flp-In-293 cells expressing WT and the Q141K variant were incubated in the absence or presence of MG132 (2 bc;M) for 24 h. ABCG2 WT and Q141 variant proteins were analyzed by immunoblotting with the ABCG2-specific monoclonal antibody (BXP-21) after PNGase F treatment. Login to comment
94 (B) Correctly processed ABCG2 WT protein is destined to reach the plasma membrane and is then degraded by the endosome-lysosome pathway after remaining in the plasma membrane domain for a certain period. In contrast, the ABCG2 Q141K variant protein is recognized as a misfolded form and then undergoes ubiquitination-mediated proteasomal degradation. Bafilomycin A1 (BMA) and MG132 inhibit lysosomal and proteasomal degradation, respectively. Login to comment
95 Golgi Endosome ER Plasma membrane Folded ABCG2 Mature ABCG2 ATP ATP N N N C C Misfolded ABCG2 E3 E3 Ubiquitin Ubiquitin ligase Proteasome MG132 Cis Medial Trans CNX PDI Dol-P-P- Dol-P-P- BiP Chaperone proteins Ribosomes ERGIC Lysosome BMA H+ H+ Control MG132 Relative expression level 0 0.5 1.0 Lys141 (Q141K) Gln141 (WT) * Control MG132 Relative expression level 0 0.5 1.0 Lys141 (Q141K) Gln141 (WT) * A B 6. Login to comment
108 The allele frequencies of 421C (WT) and 421A (Q141K) among different ethnic populations. Login to comment
110 0 0.2 0.4 0.6 0.8 1 African African-American Mexican-American Caucasian Japanese Chinese ABCG2 allele frequency 421C (WT) 421A (Q141K) 0 0.2 0.4 0.6 0.8 1 0 0.2 0.4 0.6 0.8 1 - - 0 0.2 0.4 0.6 0.8 1 0 0.2 0.4 0.6 0.8 1 African African-American Mexican-American Caucasian Japanese Chinese ABCG2 allele frequency 421C (WT) 421A (Q141K) 0 0.2 0.4 0.6 0.8 1 0 0.2 0.4 0.6 0.8 1 - - An investigation of the expression level of ABCG2 in 99 Japanese placenta samples revealed that individuals homozygous for the Q141K variant showed significantly lower expression levels of this transporter protein, while the heterozygous samples displayed intermediate expression levels [44]. Login to comment
111 Based on those observations, it is assumed that the protein stability of the Q141K variant is significantly reduced without showing significant changes in its mRNA levels. Login to comment
112 Evidence has been provided to show that the Q141K variant of ABCG2 is degraded via ubiquitin-mediated proteasomal degradation [31-35]. Login to comment
113 The level of the Q141K variant protein could be recovered when proteosomal degradation was inhibited by MG132 in vitro (Figure 5A). Login to comment
114 The Q141K mutation does not interfere with the nucleotide-binding domain/intracellular loop interactions but greatly affect the protein-protein interactions necessary for the dimerization of ABCG2. Login to comment
116 In addition to the Q141K variant, a minor SNP in exon 4, 376C>T, substituting a stop codon for Gln126, was found in the Japanese population with an allele frequency of 2.4% [37]. Login to comment
119 Matsuo et al. [4] reported that the genotype combinations of Q126stop and Q141K are clinically important biomarkers to predict the possible risk of gout in the Japanese population. Login to comment
123 Their data suggest that at least 10% of all gout cases in Caucasians are attributable to SNP 421C>A (Q141K). Login to comment

[hide] Association of single nucleotide polymorphisms in ... Med Oncol. 2014 Jan;31(1):802. doi: 10.1007/s12032-013-0802-6. Epub 2013 Dec 13. Zhao J, Li W, Zhu D, Yu Q, Zhang Z, Sun M, Cai S, Zhang W
Association of single nucleotide polymorphisms in MTHFR and ABCG2 with the different efficacy of first-line chemotherapy in metastatic colorectal cancer.
Med Oncol. 2014 Jan;31(1):802. doi: 10.1007/s12032-013-0802-6. Epub 2013 Dec 13., [PMID:24338217]

Abstract [show]
Either oxaliplatin- or irinotecan-containing regimen could receive a good effectiveness in patients with metastatic colorectal cancer as the first-line chemotherapy, but not all patients would benefit from the treatment they have received. This study was to investigate the role of single nucleotide polymorphisms (SNPs) of methylenetetrahydrofolate reductase (MTHFR) and ATP-binding cassette sub-family G member 2 (ABCG2) in selecting the most appropriate treatment for individual patients. Ninety-two metastatic colorectal cancer patients treated with first-line 5-fluoropyrimidine (5-FU), leucovorin, and oxaliplatin (FOLFOX), capecitabine, and oxaliplatin (XELOX) and sixty-two patients receiving 5-FU, leucovorin, and irinotecan (FOLFIRI) were reviewed. The SNPs of MTHFR and ABCG2 were detected using gene sequencing method after DNA PCR amplification, which was extracted from peripheral blood karyocytes. Clinical characteristics and gene polymorphisms were evaluated in univariate and multivariate analysis as predictive factors for response rate (RR) and progression-free survival (PFS). In patients bearing 2-4 genotypes of MTHFR 677C/C, MTHFR 1298 A/C or C/C, ABCG2 34G/G, and ABCG2 421C/A or A/A, those who received oxaliplatin-based chemotherapy achieved a higher RR (41.7 vs. 18.8 %, P = 0.027) and longer median PFS (mPFS) than irinotecan-based therapy [8.9 vs. 7.1 m, FOLFIRI: hazard ratio (HR) = 1.722, 95 % confidence interval (CI) 1.026-2.892, P = 0.040, compared with FOLFOX/XELOX]; on the contrary, patients carrying 0 or 1 above genotype exhibited better outcomes after receiving FOLFIRI chemotherapy (mPFS: 9.3 vs. 6.4 m, FOLFIRI: HR = 0.422, 95 % CI 0.205-0.870, P = 0.019, compared with FOLFOX/XELOX). Combination of SNPs with MTHFR and ABCG2 may play a role in helping clinicians to select first-line chemotherapy for patients with metastatic colorectal cancer.

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32 Genotyping Genomic DNA was isolated from whole blood using the FUJIFILM DNA extraction kit. Based on previous published studies, the single nucleotide polymorphisms selected for testing were MTHFR 677C[T (rs1801133, Ala 222 Val) and 1298A[C (rs1801131, Glu 428 Ala), and ABCG2 34G[A (rs2231137, Val 12 Met) and 421C[A (rs2231142, Gln 141 Lys). Login to comment

[hide] Epigenetic modulation of the drug resistance genes... BMC Cancer. 2013 Dec 31;13:617. doi: 10.1186/1471-2407-13-617. Oberstadt MC, Bien-Moller S, Weitmann K, Herzog S, Hentschel K, Rimmbach C, Vogelgesang S, Balz E, Fink M, Michael H, Zeden JP, Bruckmuller H, Werk AN, Cascorbi I, Hoffmann W, Rosskopf D, Schroeder HW, Kroemer HK
Epigenetic modulation of the drug resistance genes MGMT, ABCB1 and ABCG2 in glioblastoma multiforme.
BMC Cancer. 2013 Dec 31;13:617. doi: 10.1186/1471-2407-13-617., [PMID:24380367]

Abstract [show]
BACKGROUND: Resistance of the highly aggressive glioblastoma multiforme (GBM) to drug therapy is a major clinical problem resulting in a poor patient's prognosis. Beside promoter methylation of the O6-methylguanine-DNA-methyltransferase (MGMT) gene the efflux transporters ABCB1 and ABCG2 have been suggested as pivotal factors contributing to drug resistance, but the methylation of ABCB1 and ABCG2 has not been assessed before in GBM. METHODS: Therefore, we evaluated the proportion and prognostic significance of promoter methylation of MGMT, ABCB1 and ABCG2 in 64 GBM patient samples using pyrosequencing technology. Further, the single nucleotide polymorphisms MGMT C-56 T (rs16906252), ABCB1 C3435T (rs1045642) and ABCG2 C421A (rs2231142) were determined using the restriction fragment length polymorphism method (RFLP). To study a correlation between promoter methylation and gene expression, we analyzed MGMT, ABCB1 and ABCG2 expression in 20 glioblastoma and 7 non-neoplastic brain samples. RESULTS: Despite a significantly increased MGMT and ABCB1 promoter methylation in GBM tissue, multivariate regression analysis revealed no significant association between overall survival of glioblastoma patients and MGMT or ABCB1 promoter methylation. However, a significant negative correlation between promoter methylation and expression could be identified for MGMT but not for ABCB1 and ABCG2. Furthermore, MGMT promoter methylation was significantly associated with the genotypes of the MGMT C-56 T polymorphism showing a higher methylation level in the T allele bearing GBM. CONCLUSIONS: In summary, the data of this study confirm the previous published relation of MGMT promoter methylation and gene expression, but argue for no pivotal role of MGMT, ABCB1 and ABCG2 promoter methylation in GBM patients' survival.

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277 Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, Tsuruo T, Miki Y, Sugimoto Y: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Impact of genetic variability in the ABCG2 gene on... Biochem Biophys Res Commun. 2014 Jan 24;443(4):1211-7. doi: 10.1016/j.bbrc.2013.12.119. Epub 2014 Jan 3. Deppe S, Ripperger A, Weiss J, Ergun S, Benndorf RA
Impact of genetic variability in the ABCG2 gene on ABCG2 expression, function, and interaction with AT1 receptor antagonist telmisartan.
Biochem Biophys Res Commun. 2014 Jan 24;443(4):1211-7. doi: 10.1016/j.bbrc.2013.12.119. Epub 2014 Jan 3., [PMID:24388985]

Abstract [show]
The ATP-binding cassette transporter ABCG2 plays a prominent role in cardiovascular and cancer pathophysiology, is involved in the pathogenesis of gout, and affects pharmacokinetics of numerous drugs. Telmisartan, a widely used AT1 receptor antagonist, inhibits the transport capacity of ABCG2 and may cause drug-drug interactions, especially in individuals carrying polymorphism that facilitate the telmisartan-ABCG2 interaction. Thus, the aim of this study was to identify ABCG2 polymorphisms and somatic mutations with relevance for the telmisartan-ABCG2 interaction. For this purpose, a cellular system for the conditional expression of ABCG2 was established. ABCG2 variants were generated via site-directed mutagenesis. Interaction of telmisartan with these ABCG2 variants was investigated in HEK293-Tet-On cells using the pheophorbide A efflux assay. Moreover, expression of ABCG2 variants was studied in these cells. Importantly, protein levels of the Q141K and F489L variant were significantly reduced, a phenomenon that was partly reversed by pharmacological proteasome inhibition. Moreover, basal pheophorbide A efflux capacity of S248P, F431L, and F489L variants was significantly impaired. Interestingly, inhibition of ABCG2-mediated pheophorbide A transport by telmisartan was almost abolished in cells expressing the R482G variant, whereas it was largely increased in cells expressing the F489L variant. We conclude that the arginine residue at position 482 of the ABCG2 molecule is of major importance for the interaction of telmisartan with this ABC transporter. Furthermore, individuals carrying the F489L polymorphism may be at increased risk of developing adverse drug reactions in multi-drug regimens involving ABCG2 substrates and telmisartan.

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6 Importantly, protein levels of the Q141K and F489L variant were significantly reduced, a phenomenon that was partly reversed by pharmacological proteasome inhibition. Login to comment
17 Interestingly, recently a role of ABCG2 in uric acid transport and in the pathogenesis of gout has been demonstrated and has been linked to single nucleotide polymorphisms (SNPs) within the ABCG2 gene, e.g. the Q141K (C421A) SNP [7]. Login to comment
37 Site-directed mutagenesis Non-synonymous ABCG2 single nucleotide polymorphisms (SNPs) G34A (V12M), C421A (Q141K), T742C (S248P), T1291C (F431L), T1465C (F489L) as well as somatic mutation A1444G (R482G) were inserted into the ABCG2 cDNA sequence in the pTRE-Tight-BI-AcGFP1-ABCG2 plasmid using the QuickChange&#d2; Lightning Site-Directed Mutagenesis Kit (Agilent Technologies, Waldbronn, Germany) with specific primers according to the manufacturer`s instructions (Supplemental Fig. 1). Login to comment
83 In contrast, ABCG2 protein levels of cells transfected with the Q141K and the F489L mutants, respectively, were lower than those transfected with ABCG2 wild-type (Fig. 2B and C). Login to comment
84 Concomitant treatment of transfected HEK293-Tet-On cells with increasing concentrations of the proteasome inhibitor PS-341 (bortezomib) for 24 h slightly increased protein levels of variants Q141K and F489L but not that of ABCG2 wild-type (Fig. 2D). Login to comment
91 The average PhA-associated fluorescence in non-induced HEK293-Tet-On cells transiently transfected with the various ABCG2 variants was not significantly different as compared with that observed in HEK293-Tet-On cells transfected with ABCG2 wild-type (wild-type (100 &#b1; 12.1%), V12M (106.7 &#b1; 2.0%), Q141K (97.1 &#b1; 9.3%), S248P (99.1 &#b1; 9.8%), F431L (104.7% &#b1; 10.9%), R482G A B C D Fig. 2. Login to comment
95 (D) Impact of increasing concentrations of the proteasome inhibitor PS-341 (bortezomib) on ABCG2 protein levels in doxycycline-treated (1 lg/mL; 24 h) HEK293-Tet-On cells transiently transfected with ABCG2 wild-type or the ABCG2 variants Q141K and F489L, respectively. Login to comment
99 PhA-associated fluorescence was similar in doxycycline-induced AcGFP1-positive HEK293-Tet-On cells transfected with the ABCG2 variants V12M (11.8 &#b1; 0.9%), Q141K (17.9 &#b1; 5.8%), and R482G (17.0 &#b1; 2.8%). Login to comment
104 Inhibitory efficacy of telmisartan was not altered by the ABCG2 polymorphisms V12M and Q141K but tended to be lower in the ABCG2 variants S248P and F431L (Fig. 4A and C). Login to comment
117 Although the mRNA expression levels of these ABCG2 variants were similar to ABCG2 wild-type, protein levels of the Q141K and the F489L variants were significantly reduced in our experimental system. These findings are in accordance with experimental data from other groups [19], although the reduction of the F489L variant was more pronounced in our cellular expression model. Login to comment
118 Moreover, a reduced protein level of the Q141K variant has been described in vitro and interestingly, the Q141K variant has been linked to an impaired uric acid efflux in vitro as well as gout disease in vivo [7,20,21]. Login to comment
121 The reason for the reduced protein levels of the Q141K and F489L variant are currently unclear, but enhanced proteasomal degradation has been proposed to participate in reduction of cellular Q141K protein content [22]. Login to comment
122 Indeed, inhibition of proteasomal activity using the proteasome inhibitor bortezomib slightly increased the protein levels of both the Q141K and the F489L variant in HEK293-Tet-On cells, thereby suggesting a role of the proteasome in decreasing the protein content of both ABCG2 variants. Login to comment
123 Surprisingly, the Q141K variant did not exhibit a reduced pheophorbide A (PhA) efflux in HEK293-Tet-On cells as evidenced by flow cytometric analyses. Login to comment
124 In addition, PhA efflux of the F489L variant was only marginally impaired in our experimental system. These data suggest that (1) the observed reduction in ABCG2 protein content caused by the Q141K or F489L polymorphisms may be compensated by a more efficient PhA transport Fig. 3. Login to comment
127 Nevertheless, data from clinical investigations indicate that the Q141K variant may be associated with a reduced ABCG2-mediated substrate clearance in affected individuals [8,9,21] and that thus, clinical investigations are needed to further explore this issue with respect to the F489L variant. Login to comment

[hide] ABCG2 dysfunction causes hyperuricemia due to both... Sci Rep. 2014 Jan 20;4:3755. doi: 10.1038/srep03755. Matsuo H, Nakayama A, Sakiyama M, Chiba T, Shimizu S, Kawamura Y, Nakashima H, Nakamura T, Takada Y, Oikawa Y, Takada T, Nakaoka H, Abe J, Inoue H, Wakai K, Kawai S, Guang Y, Nakagawa H, Ito T, Niwa K, Yamamoto K, Sakurai Y, Suzuki H, Hosoya T, Ichida K, Shimizu T, Shinomiya N
ABCG2 dysfunction causes hyperuricemia due to both renal urate underexcretion and renal urate overload.
Sci Rep. 2014 Jan 20;4:3755. doi: 10.1038/srep03755., [PMID:24441388]

Abstract [show]
Gout is a common disease which results from hyperuricemia. We have reported that the dysfunction of urate exporter ABCG2 is the major cause of renal overload (ROL) hyperuricemia, but its involvement in renal underexcretion (RUE) hyperuricemia, the most prevalent subtype, is not clearly explained so far. In this study, the association analysis with 644 hyperuricemia patients and 1,623 controls in male Japanese revealed that ABCG2 dysfunction significantly increased the risk of RUE hyperuricemia as well as overall and ROL hyperuricemia, according to the severity of impairment. ABCG2 dysfunction caused renal urate underexcretion and induced hyperuricemia even if the renal urate overload was not remarkable. These results show that ABCG2 plays physiologically important roles in both renal and extra-renal urate excretion mechanisms. Our findings indicate the importance of ABCG2 as a promising therapeutic and screening target of hyperuricemia and gout.

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17 Their functional ABCG2 activities were estimated from their genotype combinations of its two dysfunctional missense variants, Q126X (rs72552713) and Q141K (rs2231142). Login to comment
18 Because there is no simultaneous presence of the minor alleles of non-functional variant Q126X and half-functional variant Q141K in one hap- lotype5,7 , we defined three haplotype IDs as *1, *2, and *3, as shown in Figure 1a. Login to comment
26 When hyperuricemia was divided into three distinct types (i.e., RUE type, combined type, and ROL type as shown in Supplementary Fig. S1), severe ABCG2 dysfunction (#1/4 function) significantly raised the risk of combined and ROL types but not that of RUE type Figure 1 | Estimation of ABCG2 function from diplotype of Q126X and Q141K alleles. Login to comment
27 (a) ABCG2*2 or *3 represents a haplotype with Q141K or Q126X variant, respectively. Login to comment
28 ABCG2*1 indicates a haplotype with neither Q141K nor Q126X variant. Login to comment
29 Since Q141K is a half-functional variant and Q126X is a nonfunctional variant, relative function of ABCG2*1, *2, and *3 is 1, 1/2, and 0, respectively, which is visualized by black-indicated areas. Login to comment
32 Table 1 | ABCG2 functions of participants Estimated transport activity Diplotype of Q126X (rs72552713) and Q141K (rs2231142) alleles** Case{ Control N % N % #1/4 function *3/*3 or *2/*3 29 (26) 4.5 (4.7) 22 1.3 1/2 function *1/*3 or *2/*2 151 (135) 23.4 (23.5) 190 11.7 3/4 function *1/*2 307 (277) 47.7 (48.2) 600 37.0 Full function *1/*1 157 (136) 24.4 (23.7) 811 50.0 Total 644 (575) 100.0 (100.0) 1,887 100.0 **Haplotypes ''Q-Q``, ''Q-K``, and ''X-Q`` of two SNPs (Q126X and Q141K) are referred to as *1, *2, and *3, respectively. Login to comment
33 Risk alleles are X for Q126X, and K for Q141K. Login to comment
67 Genotyping of ABCG2 Q126X (rs72552713) and Q141K (rs2231142) was performed by high-resolution melting analysis with a LightCycler 480 (Roche Diagnostics)19 . Login to comment
68 From the haplotype analyses reported in the previous studies5,7 , there is no simultaneous presence of the minor alleles (risk alleles) of non-functional variant Q126X and half-functional variant Q141K in one haplotype. Login to comment
69 In this study, their haplotype IDs, *1, *2, and *3, were defined as Figure 1a; the combination of wild-type Q126X and Q141K alleles (''Q-Q``) was designated as ABCG2*1, which corresponds to the cDNA sequence of GenBank (accession number NM_004827). Login to comment
71 Based on the diplotype of Q126X and Q141K alleles (Fig. 1b)5,7 , ABCG2 function was estimated and divided into four groups5-7 ; i.e., full function, 3/4 function, 1/2 function, and #1/4 function (Table 1). Login to comment

[hide] Reduced ABCG2 and increased SLC22A1 mRNA expressio... Med Oncol. 2014 Mar;31(3):851. doi: 10.1007/s12032-014-0851-5. Epub 2014 Jan 29. de Lima LT, Vivona D, Bueno CT, Hirata RD, Hirata MH, Luchessi AD, de Castro FA, de Lourdes F Chauffaille M, Zanichelli MA, Chiattone CS, Hungria VT, Guerra-Shinohara EM
Reduced ABCG2 and increased SLC22A1 mRNA expression are associated with imatinib response in chronic myeloid leukemia.
Med Oncol. 2014 Mar;31(3):851. doi: 10.1007/s12032-014-0851-5. Epub 2014 Jan 29., [PMID:24469953]

Abstract [show]
Imatinib mesylate (IM) has become a standard of care in chronic myeloid leukemia (CML) therapy. Single nucleotide polymorphisms (SNPs) and altered expression in drug transporter genes may influence IM response. In order to investigate whether mRNA expression and SNPs in drug transporters are associated with IM resistance, we studied 118 chronic-phase CML patients receiving the standard dose of IM (400 mg/day). They were assigned as responders and non-responders according to European LeukemiaNet criteria (2009). mRNA expression in samples at diagnosis (without IM therapy) and outcomes after IM failure were also evaluated in subgroups of patients. Major molecular response (MMR), complete molecular response and primary and secondary resistance were all assessed. BCR-ABL1, ABCB1, ABCG2, SLC22A1 and SLCO1A2 mRNA expression and SNPs in ABCG2 and SLC22A1 genes were analyzed. ABCG2 mRNA expression in the non-responders was higher before and during IM therapy. Furthermore, ABCG2 was overexpressed in those who did not achieve MMR (P=0.027). In a subgroup of patients who switched to second-generation tyrosine kinase inhibitors, high mRNA expression of ABCG2 was associated with a risk of 24 times that of not achieving complete cytogenetic response (OR 24.00, 95% CI 1.74-330.80; P=0.018). In the responder group, patients who achieved MMR (P=0.009) presented higher mRNA levels of SLC22A1. The SNPs were not associated with mRNA expression of ABCG2 and SLC22A1. Our data suggest that elevated ABCG2 expression (an efflux transporter) could be associated with IM resistance and could impact on second-generation TKI response, whereas high SLC22A1 expression (an influx transporter) may be associated with a successful IM therapy in CML patients.

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174 Non-synonymous ABCG2c.421C[A SNP (Q141K) was described as responsible for lower levels of protein expression [41], impaired transport [42, 43], and enhanced susceptibility to ubiquitin-mediated proteasomal degradation [44]. Login to comment

[hide] Influence of the ABCG2 gout risk 141 K allele on u... Arthritis Res Ther. 2014 Jan 30;16(1):R34. doi: 10.1186/ar4463. Dalbeth N, House ME, Gamble GD, Pool B, Horne A, Purvis L, Stewart A, Merriman M, Cadzow M, Phipps-Green A, Merriman TR
Influence of the ABCG2 gout risk 141 K allele on urate metabolism during a fructose challenge.
Arthritis Res Ther. 2014 Jan 30;16(1):R34. doi: 10.1186/ar4463., [PMID:24476385]

Abstract [show]
INTRODUCTION: Both genetic variation in ATP-binding cassette sub-family G member 2 (ABCG2) and intake of fructose-containing beverages are major risk factors for hyperuricemia and gout. This study aimed to test the hypothesis that the ABCG2 gout risk allele 141 K promotes the hyperuricaemic response to fructose loading. METHODS: Healthy volunteers (n = 74) provided serum and urine samples immediately before and 30, 60, 120 and 180 minutes after ingesting a 64 g fructose solution. Data were analyzed based on the presence or absence of the ABCG2 141 K gout risk allele. RESULTS: The 141 K risk allele was present in 23 participants (31%). Overall, serum urate (SU) concentrations during the fructose load were similar in those with and without the 141 K allele (PSNP = 0.15). However, the 141 K allele was associated with a smaller increase in SU following fructose intake (PSNP <0.0001). Those with the 141 K allele also had a smaller increase in serum glucose following the fructose load (PSNP = 0.002). Higher fractional excretion of uric acid (FEUA) at baseline and throughout the fructose load was observed in those with the 141 K risk allele (PSNP <0.0001). However, the change in FEUA in response to fructose was not different in those with and without the 141 K risk allele (PSNP = 0.39). The 141 K allele effects on serum urate and glucose were more pronounced in Polynesian participants and in those with a body mass index >/=25 kg/m(2). CONCLUSIONS: In contrast to the predicted responses for a hyperuricemia/gout risk allele, the 141 K allele is associated with smaller increases in SU and higher FEUA following a fructose load. The results suggest that ABCG2 interacts with extra-renal metabolic pathways in a complex manner to regulate SU and gout risk. CLINICAL TRIALS REGISTRATION: The study was registered by the Australian Clinical Trials Registry (ACTRN12610001036000).

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139 This could be related to an influence of Q141K on hepatic conversion of fructose to glucose, estimated to occur at a rate up to 60% depending on sex and health status [18]. Login to comment
197 5. Woodward OM, Tukaye DN, Cui J, Greenwell P, Constantoulakis LM, Parker BS, Rao A, Kottgen M, Maloney PC, Guggino WB: Gout-causing Q141K mutation in ABCG2 leads to instability of the nucleotide-binding domain and can be corrected with small molecules. Login to comment

[hide] Genetic association analysis of ATP binding casset... PLoS One. 2014 Feb 21;9(2):e89253. doi: 10.1371/journal.pone.0089253. eCollection 2014. Balan S, Bharathan SP, Vellichiramal NN, Sathyan S, Joseph V, Radhakrishnan K, Banerjee M
Genetic association analysis of ATP binding cassette protein family reveals a novel association of ABCB1 genetic variants with epilepsy risk, but not with drug-resistance.
PLoS One. 2014 Feb 21;9(2):e89253. doi: 10.1371/journal.pone.0089253. eCollection 2014., [PMID:24586633]

Abstract [show]
Epilepsy constitutes a heterogeneous group of disorders that is characterized by recurrent unprovoked seizures due to widely different etiologies. Multidrug resistance remains a major issue in clinical epileptology, where one third of patients with epilepsy continue to have seizures. Role of efflux transporters in multidrug resistant epilepsy has been attributed to drug-resistant epilepsy although, with discrepant observation in genetic studies. These discrepancies could be attributed to variety of factors such as variable definition of the anti-epileptic drug (AED)-resistance, variable epilepsy phenotypes and ethnicities among the studies. In the present study we inquired the role of multidrug transporters ABCB1 and ABCG2 variants in determining AED-resistance and susceptibility to epilepsy in three well-characterized cohorts comprising of mesial temporal lobe epilepsy with hippocampal sclerosis (MTLE-HS) (prototype for AED-resistant epilepsy); juvenile myoclonic epilepsy (JME) (prototype for AED-responsive epilepsy); and healthy non-epileptic controls, in 738 subjects of Malayalam speaking south Indian ancestry. ABCB1 and ABCG2 variants were not found to be associated with drug resistance when AED-resistant and AED-responsive cohorts were compared. However, a significant association was observed between ABCB1 (C3435T) rs1045642 and risk of having epilepsy (MTLE-HS and JME pooled cohort; genotypic p-value = 0.0002; allelic p-value = 0.004). This association was seen persistent with MTLE-HS (genotypic p-value = 0.0008; allelic p-value = 0.004) and also with JME (genotypic p-value = 0.01; allelic p-value = 0.05) cohort individually. In-silico functional prediction indicated that ABCB1 rs1045642 has a deleterious impact on protein coding function and in splicing regulation. We conclude that the ABCB1 and ABCG2 variants do not confer to AED-resistance in the study population. However, ABCB1 rs1045642 increases vulnerability to epilepsy with greater tendency for MTLE-HS in south Indian ancestry from Kerala.

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62 For ABCG2 three functional variants viz: rs2231142 (Gln141Lys; missense), rs72552713 (Gln126Ter; stop gain) and rs2231137 (Val12Met; missense) were screened. Login to comment

[hide] Association between statin-induced creatine kinase... Eur J Clin Pharmacol. 2014 May;70(5):539-47. doi: 10.1007/s00228-014-1661-6. Epub 2014 Mar 6. Ferrari M, Guasti L, Maresca A, Mirabile M, Contini S, Grandi AM, Marino F, Cosentino M
Association between statin-induced creatine kinase elevation and genetic polymorphisms in SLCO1B1, ABCB1 and ABCG2.
Eur J Clin Pharmacol. 2014 May;70(5):539-47. doi: 10.1007/s00228-014-1661-6. Epub 2014 Mar 6., [PMID:24595600]

Abstract [show]
PURPOSE: Treatment with statins requires close monitoring of serum creatine kinase (CK) levels to prevent myopathy, a common and potentially serious dose-dependent adverse effect of these drugs. We have investigated the correlation between elevated CK levels and polymorphisms in the genes encoding transporters involved in statin disposition. METHODS: Patients with and without statin-induced elevated serum CK levels were genotyped for polymorphisms in SLCO1B1 (SLCO1B1 A388G and SLCO1B1 T521C), ABCB1 (ABCB1 C1236T and ABCB1 C3435T) and ABCG2 (ABCG2 C421A). RESULTS: Patients carrying SLCO1B1 T521C or ABCB1 C1236T single nucleotide polymorphisms (SNPs) had an odds ratio (OR) for statin-induced elevated serum CK levels of 8.86 (p<0.01) and 4.67 (p<0.05), respectively, while patients carrying the SLCO1B1 A388G SNP had an OR of 0.24 (p<0.05). An arbitrary score based on genotype combination discriminated patients with and without CK elevation at a specificity of 97 % and a sensitivity of 39 %. CONCLUSION: Genotyping of the SLCO1B1, ABCB1 and ABCG2 genes deserves consideration as a clinical approach to improve statin safety while concomitantly reducing the burden of blood tests for CK measurements.

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36 Finally, in the ABCG2 gene, a relatively common SNP is C421A (p.Gln141Lys; rs2231142); this SNP occurs in 10-15 % of Caucasians and has been associated with reduced ABCG2 activity [36] and increased statin bioavailability [23]. Login to comment

[hide] High-dose methotrexate in Egyptian pediatric acute... J Cancer Res Clin Oncol. 2014 Aug;140(8):1359-65. doi: 10.1007/s00432-014-1670-y. Epub 2014 Apr 10. El Mesallamy HO, Rashed WM, Hamdy NM, Hamdy N
High-dose methotrexate in Egyptian pediatric acute lymphoblastic leukemia: the impact of ABCG2 C421A genetic polymorphism on plasma levels, what is next?
J Cancer Res Clin Oncol. 2014 Aug;140(8):1359-65. doi: 10.1007/s00432-014-1670-y. Epub 2014 Apr 10., [PMID:24718721]

Abstract [show]
PURPOSE: High-dose methotrexate (HD-MTX) is a cornerstone antineoplastic drug in most treatment protocols of pediatric acute lymphoblastic leukemia (ALL). Among the membrane efflux transporters of MTX, the human breast cancer resistant protein is the second member of the G subfamily of ATP-binding cassette (ABC) efflux pump (ABCG2). A single-nucleotide polymorphism (SNP) in ABCG2, the exchange of C to A at position 421, represents 13 % in the Middle Eastern population. We studied the effect of this SNP on the plasma levels of HD-MTX in Egyptian pediatric ALL. METHODS: Two hundred ALL patients were recruited from Children's Cancer Hospital Egypt-57357, and all were treated according to the St Jude Total XV protocol. Determination of plasma MTX levels was done at 23, 42 and 68 h. Genotyping of C421A of ABCG2 was done by polymerase chain reaction-restriction fragment length polymorphism. RESULTS: We found 14.5 % of the variant allele of the ABCG2 C421A SNP. The statistical association between ABCG2 421C>A SNP and the cutoff toxic plasma level of 24 h HD-MTX infusion at different time points tested was not statistically significant. There was no statistical significance between steady-state plasma concentration in patients with and without with this SNP. CONCLUSION: To date, this is the largest study on Egyptian ALL patients for this SNP. This study shows that there is no effect of ABCG2 421C>A on plasma concentrations of HD-MTX. Replacing candidate gene association studies with genome-wide studies of HD-MTX is now mandatory and is part of our research blueprint.

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32 Out of more than 80 single-nucleotide polymorphisms (SNPs) in the ABCG2 gene, the C421A (Q141K) nonsynonymous polymorphism located on exon 5 is the most extensively studied SNP. Login to comment
34 The allelic frequency of C421A (Q141K) varies between different populations; it is present in approximately 13 % of Middle Eastern people (Zamber et al. 2003). Login to comment
57 Briefly, 200 ng of gDNA was amplified using primers flanking the regions of BCRB C421A polymorphism (rs2231142, Gln 141 Lys). Login to comment

[hide] The association between the polymorphism rs2231142... Clin Rheumatol. 2014 Dec;33(12):1801-5. doi: 10.1007/s10067-014-2635-x. Epub 2014 Apr 29. Lv X, Zhang Y, Zeng F, Yin A, Ye N, Ouyang H, Feng D, Li D, Ling W, Zhang X
The association between the polymorphism rs2231142 in the ABCG2 gene and gout risk: a meta-analysis.
Clin Rheumatol. 2014 Dec;33(12):1801-5. doi: 10.1007/s10067-014-2635-x. Epub 2014 Apr 29., [PMID:24777469]

Abstract [show]
Gout is a common metabolic disorder with high heritability. We tried to explore the association between rs2231142 and gout. We searched "rs2231142 or Q141K and gout" in four databases and scholar searching website until 1 June, 2013 and included data from 52,010 participants in meta-analysis and subgroup analysis. The T allele of rs2231142 was associated with increased gout susceptibility (odds ratio [OR] [95 % confidence interval (95 % CI)] = 1.73 [1.55-1.91], P < 0.001). It increased gout risk in Caucasians with OR (95 % CI) = 1.68 (1.50-1.87), P < 0.001; Asians with OR (95 % CI) = 1.93 (1.54-2.31), P < 0.001; Africans with OR (95 % CI) = 1.76 (1.15-2.36), P < 0.001; and New Zealand Pacific Islanders with OR (95 % CI) = 2.94 (1.72-4.15), P < 0.001, but not in New Zealand Maoris, with OR (95 % CI) = 1.12 (0.57-1.67), P = 0.061. No publish or other biases were observed. The T allele of rs2231142 was associated with increased risk of gout.

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2 We searched "rs2231142 or Q141K and gout" in four databases and scholar searching website until 1 June, 2013 and included data from 52,010 participants in meta-analysis and subgroup analysis. Login to comment
8 Q141K . Login to comment
10 Single-nucleotide polymorphism (SNP) rs2231142, known as Q141K in ABCG2, leading to a change from glutamine to lysine, is reported to be associated with gout in several populations, such as Atherosclerosis Risk in Communities (ARIC) Study [2], European Americans [3], African Americans [3], Mexican Americans [3], Americans Indians [3], German [4], Chinese [5, 6], Japanese [7], and New Zealand Pacific Islanders [8], but not in New Zealand Maoris [8]. Login to comment
17 Ling Guangdong Provincial Key Laboratory of Food, Nutrition and Health, Department of Nutrition, School of Public Health, Sun Yat-Sen University (Northern Campus), 74 Zhongshan Road 2, Guangzhou 510080, China Clin Rheumatol (2014) 33:1801-1805 DOI 10.1007/s10067-014-2635-x Methods Literature search Two researchers (Xiaofei Lv and Yuan Zhang) retrieved studies in PubMed (January 1966-June 2013), ISI Web of Knowledge (1950-2013), and OVID (Sun Yat-sen University Journals@Ovid and Journals@Ovid Full Text, 1860-2013) by using the terms "rs2231142 and gout" and "Q141K and gout." Login to comment
47 SNP rs2231142, the Q141K mutation in ABCG2 gene, was reported to be associated with gout by reducing renal urate secretion. Login to comment
62 It is already discovered that the Q141K (rs2231142) polymorphism is a loss-of-function mutation in two independent functional studies [1, 13]. Login to comment
64 The results showed that Q141K decreased ATPase activity by 1.3 below that of wild-type ABCG2. Login to comment
65 In addition, kinetic analysis of ATPase activity showed that the K(m) value in Q141K was 1.4-fold higher than that of wild-type ABCG2 [14]. Login to comment

[hide] Structure and function of BCRP, a broad specificit... Arch Toxicol. 2014 Jun;88(6):1205-48. doi: 10.1007/s00204-014-1224-8. Epub 2014 Apr 29. Jani M, Ambrus C, Magnan R, Jakab KT, Beery E, Zolnerciks JK, Krajcsi P
Structure and function of BCRP, a broad specificity transporter of xenobiotics and endobiotics.
Arch Toxicol. 2014 Jun;88(6):1205-48. doi: 10.1007/s00204-014-1224-8. Epub 2014 Apr 29., [PMID:24777822]

Abstract [show]
The discovery and characterization of breast cancer resistance protein (BCRP) as an efflux transporter conferring multidrug resistance has set off a remarkable trajectory in the understanding of its role in physiology and disease. While the relevance in drug resistance and general pharmacokinetic properties quickly became apparent, the lack of a characteristic phenotype in genetically impaired animals and humans cast doubt on the physiological importance of this ATP-binding cassette family member, similarly to fellow multidrug transporters, despite well-known endogenous substrates. Later, high-performance genetic analyses and fine resolution tissue expression data forayed into unexpected territories concerning BCRP relevance, and ultimately, the rise of quantitative proteomics allows putting observed interactions into absolute frameworks for modeling and insight into interindividual and species differences. This overview summarizes existing knowledge on the BCRP transporter on molecular, tissue and system level, both in physiology and disease, and describes a selection of experimental procedures that are the most widely applied for the identification and characterization of substrate and inhibitor-type interactions.

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87 Four variants are found with allele frequencies above 3 % in at least one of the studied population (African American, Asian, and Caucasian): Val12Met (V12M), Gln141Lys (Q141K), Phe208Ser (F208S), and Asp590Tyr (N590Y). Login to comment
91 Q141K and V12M are two variants that are much more highly represented across different ethnicities, although the V12M mutation is not known to alter significantly BCRP function or expression (Tamura et al. 2006; Kondo et al. 2004; Honjo et al. 2002). Login to comment
93 Variant Q141K is by far the most studied and characterized BCRP variant, and is found at frequencies around 30 % in Asians and around 10 % in Caucasians. Login to comment
94 A mechanistic study showed that the Q141K variant undergoes increased lysosomal and proteosomal degradation that affects the half-life of the BCRP protein (Furukawa et al. 2009), as well as targeting to the aggresome, a perinuclear structure where misfolded proteins accumulate (Basseville et al. 2012). Login to comment
95 Histone deacetylase inhibitors rescue newly synthesized transporter proteins and prevent aggresome targeting by disturbing TableÊf;1ߒߙMajor non-synonymous single-nucleotide polymorphisms found in the ABCG2 coding region Allele frequencies presented in this table do not reflect interethnic differences Mutation Position in BCRP Cellular effects of SNP Allele frequency % References 34G>A, V12M (rs2231137) N-terminus Lower expression, no impact on function 0-29.8 Tamura et al. (2006), Bosch et al. (2005), Mizuarai et al. (2004), Imai et al. (2002), Kobayashi et al. (2005), Backstrom et al. (2003), Honjo et al. (2002), Kondo et al. (2004) 151G>T, G51C N-terminus Slightly overexpressed, decreased transport activity 0.1 Tamura et al. (2006), Yoshioka et al. (2007) 376C>T, Q126X (rs7255271) NBD No expression, no activity 0-1.7 Tamura et al. (2006), Mizuarai et al. (2004), Itoda et al. (2003), Imai et al. (2002), Kobayashi et al. (2005), Kondo et al. (2004) 421C>A, Q141K (rs2231142) NBD Lower expression, decreased transport activity, substrate specificity altered 0-35.7 Tamura et al. (2006), Bosch et al. (2005), Mizuarai et al. (2004), Imai et al. (2002), Kobayashi et al. (2005), Backstrom et al. (2003), Honjo et al. (2002), Kondo et al. (2004) 458C>T, T153 M NBD Slightly lower expression, no impact on function 3.3 Tamura et al. (2006), Mizuarai et al. (2004) 479G>A, R160Q NBD Not determined 0.5 Bosch et al. (2005), Tamura et al. (2006) 496C>G, Q166E (rs1061017) NBD Slightly lower expression, no impact on function 0-1.1 Tamura et al. (2006), Kondo et al. (2004), Yoshioka et al. (2007) 616A>C, I206L (rs12721643) NBD Well expressed, decreased transport activity 0-10.0 Tamura et al. (2006), Zamber et al. (2003), Vethanayagam et al. (2005), Ieiri (2012a) 623T>C, F208 (rs1061018) NBD No expression, no transport activity 0.9-3.9 Tamura et al. (2006) 742T>C, S248P (rs3116448) NBD Well expressed, no transport activity 0.5 Tamura et al. (2006), Yoshioka et al. (2007) 1000G>T, E334X (rs3201997) NBD No expression, no transport activity Not determined Tamura et al. (2006), Ishikawa et al. (2005) 1291T>C F431L ECL1 Lower expression, substrate specificity altered 0.6-0.8 Tamura et al. (2006), Itoda et al. (2003), Yoshioka et al. (2007) 1322G>A, S441 N ECL1 Slightly lower expression, no transport activity 0.5 Tamura et al. (2006), Kobayashi et al. (2005), Kondo et al. (2004) 1465T>C, F489L TM3 Slightly lower expression, no transport activity 0.5-0.8 Tamura et al. (2006), Itoda et al. (2003), Kobayashi et al. (2005) 1515delC, F506S TM4 Not determined 0.5 Itoda et al. (2003), Kobayashi et al. (2005) 1515delC, F507L 1515delC, V508L 1515delC, M509X 1711T>A, F571I (rs9282571) TM5 Well expressed, substrate specificity altered 0.5 Tamura et al. (2006) 1723C>T, R575X TM5 Not determined 0.5 Tamura et al. (2006) 1768A>T, N590Y (rs34264773) ECL3 Slightly overexpressed, substrate specificity altered 0-9.7 Tamura et al. (2006), Mizuarai et al. (2004), Zamber et al. (2003), Vethanayagam et al. (2005) 1858G>A, D620 N (rs34783571) ECL3 Slightly overexpressed, substrate specificity altered 0-11.1 Tamura et al. (2006), Bosch et al. (2005), Honjo et al. (2002), Vethanayagam et al. (2005) the trafficking along microtubules (Basseville et al. 2012). Login to comment
96 A recent study revealed that the Q141K mutation leads to instability of the transporter NBD, and suggests that certain small molecules could rescue this phenotype (Woodward et al. 2013). Login to comment
97 BCRP substrates with a narrow therapeutic index and low bioavailability are the most likely to be affected by the Q141K polymorphism. Login to comment
99 However, the pharmacogenomic effect of the Q141K polymorphism seems to be substrate dependent, as irinotecan and pitavastatin pharmacokinetic (PK) profiles were not significantly different compared with wild-type BCRP (Ieiri et al. 2007; de Jong et al. 2004; Han et al. 2007). Login to comment
100 More recently, the Q141K variant, in combination with a specific CYP2C9 haplotype, was associated with fluvastatin-induced adverse drug reaction in renal transplant patients (Mirosevic Skvrce et al. 2013). Login to comment
101 Another study showed that Q141K is associated with longer survival in epithelial ovarian/primary peritoneal cancer patients treated with platinum-/taxane-based chemotherapy (Tian et al. 2012). Login to comment
102 More recently, the impact of BCRP polymorphism, particularly of mutation Q141K, on the PK profile of protein kinases inhibitors has been investigated. Login to comment
103 The frequency of diarrhea after treatment with gefitinib was found to be significantly higher in Caucasian non-small-cell lung cancer (NSCLC) patients with at least one allele carrying the Q141K mutation, than in homozygous wild-type patients (Cusatis et al. 2006). Login to comment
104 Conversely, in Japanese NSCLC patients, no association between Q141K and susceptibility to gefitinib-induced adverse effects could be established (Akasaka et al. 2010). Login to comment
107 Interestingly, genomic analysis revealed that this patient was homozygous for the Q141K SNP, while other patients were either heterozygous or wild-type (Mizuno et al. 2010). Login to comment
108 In addition, a clinical study on a Korean cohort revealed that the Q141K variant is associated with the risk of sunitinib-induced toxicity in metastatic renal cell carcinoma patients (Kim et al. 2013). Login to comment
109 A recent in vivo study using knockout mice confirmed that the Q141K variant is associated with an increase in the systemic exposure to sunitinib, as well as sunitinib-induced toxicity (Mizuno et al. 2012). Login to comment
110 An antibody-based flow cytometry assay for the quantitative determination of membrane proteins in human erythrocytes revealed significant differences in expression levels between BCRP wild-type and Q141K variants, suggesting that erythrocyte membrane protein levels may reflect genotype-dependent tissue expression patterns (Kasza et al. 2012). Login to comment
112 Detection of the Q141K polymorphism could also serve as a biomarker of poor prognosis in adult acute myeloid leukemia treated with idarubicin-based chemotherapy (Tiribelli et al. 2013). Login to comment
353 The c.421C>A (Q141K; rs2231142) variant with the highest allelic frequency displays lower stability (Furukawa et al. 2009; Basseville et al. 2012) leading to lower expression (Kondo et al. 2004), lower transport activity (Sparreboom et al. 2005), altered substrate specificity (Tamura et al. 2006), and impaired trafficking (Urquhart et al. 2008). Login to comment
472 But combination C421A wt with low protein expression DFS, OS are longer Regarding DFS, Q141K polymorphism per se did not affect survival, but stratifying patients in the three groups, patients with low ABCG2 and wt gene had a longer OS compared to patients with Q141K ABCG2 or with high ABCG2 expression. Login to comment

[hide] Heme in pathophysiology: a matter of scavenging, m... Front Pharmacol. 2014 Apr 8;5:61. doi: 10.3389/fphar.2014.00061. eCollection 2014. Chiabrando D, Vinchi F, Fiorito V, Mercurio S, Tolosano E
Heme in pathophysiology: a matter of scavenging, metabolism and trafficking across cell membranes.
Front Pharmacol. 2014 Apr 8;5:61. doi: 10.3389/fphar.2014.00061. eCollection 2014., [PMID:24782769]

Abstract [show]
Heme (iron-protoporphyrin IX) is an essential co-factor involved in multiple biological processes: oxygen transport and storage, electron transfer, drug and steroid metabolism, signal transduction, and micro RNA processing. However, excess free-heme is highly toxic due to its ability to promote oxidative stress and lipid peroxidation, thus leading to membrane injury and, ultimately, apoptosis. Thus, heme metabolism needs to be finely regulated. Intracellular heme amount is controlled at multiple levels: synthesis, utilization by hemoproteins, degradation and both intracellular and intercellular trafficking. This review focuses on recent findings highlighting the importance of controlling intracellular heme levels to counteract heme-induced oxidative stress. The contributions of heme scavenging from the extracellular environment, heme synthesis and incorporation into hemoproteins, heme catabolism and heme transport in maintaining adequate intracellular heme content are discussed. Particular attention is put on the recently described mechanisms of heme trafficking through the plasma membrane mediated by specific heme importers and exporters. Finally, the involvement of genes orchestrating heme metabolism in several pathological conditions is illustrated and new therapeutic approaches aimed at controlling heme metabolism are discussed.

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536 Genetic studies identified a SNP in the exon 5 of ABCG2 gene that leads to a glutamine-to-lysine amino acid substitution (Q141K) (Dehghan et al., 2008). Login to comment
537 The Q141K variant has been shown to have a significant reduced capacity to transport urate and, when expressed in mammalian cells, Q141K ABCG2 expression is significantly lower than that of wild-type protein (Imai et al., 2002; Mizuarai et al., 2004; Morisaki et al., 2005; Woodward et al., 2009). Login to comment

[hide] Metabolic syndrome, alcohol consumption and geneti... PLoS One. 2014 May 14;9(5):e97646. doi: 10.1371/journal.pone.0097646. eCollection 2014. Stiburkova B, Pavlikova M, Sokolova J, Kozich V
Metabolic syndrome, alcohol consumption and genetic factors are associated with serum uric acid concentration.
PLoS One. 2014 May 14;9(5):e97646. doi: 10.1371/journal.pone.0097646. eCollection 2014., [PMID:24827988]

Abstract [show]
OBJECTIVE: Uric acid is the end product of purine metabolism in humans, and increased serum uric acid concentrations lead to gout. The objective of the current study was to identify factors that are independently associated with serum uric acid concentrations in a cohort of Czech control individuals. METHODS: The cohort consisted of 589 healthy subjects aged 18-65 years. We studied the associations between the serum uric acid concentration and the following: (i) demographic, anthropometric and other variables previously reported to be associated with serum uric acid concentrations; (ii) the presence of metabolic syndrome and the levels of metabolic syndrome components; and (iii) selected genetic variants of the MTHFR (c.665C>T, c.1286A>C), SLC2A9 (c.844G>A, c.881G>A) and ABCG2 genes (c.421C>A). A backward model selection procedure was used to build two multiple linear regression models; in the second model, the number of metabolic syndrome criteria that were met replaced the metabolic syndrome-related variables. RESULTS: The models had coefficients of determination of 0.59 and 0.53. The serum uric acid concentration strongly correlated with conventional determinants including male sex, and with metabolic syndrome-related variables. In the simplified second model, the serum uric acid concentration positively correlated with the number of metabolic syndrome criteria that were met, and this model retained the explanatory power of the first model. Moderate wine drinking did not increase serum uric acid concentrations, and the urate transporter ABCG2, unlike MTHFR, was a genetic determinant of serum uric acid concentrations. CONCLUSION: Metabolic syndrome, moderate wine drinking and the c.421C>A variant in the ABCG gene are independently associated with the serum uric acid concentration. Our model indicates that uric acid should be clinically monitored in persons with metabolic syndrome.

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45 Variant c.421C.A (rs2231142, p.Q141K) results in a reduction of the urate transport rate by 53% compared with that for the wild-type ABCG2 and causes approximately 10% of the hyperuricemia and gout cases in Caucasians [28]. Login to comment
74 Genotype analysis The frequent genetic variants of genes encoding the urate transporter SLC2A9 (c.844G.A (rs16890979, p.V282I) and c.881G.A (rs3733591, p.R294H)), the urate transporter ABCG2 (c.421C.A (rs2231142, p.Q141K)) and the MTHFR enzyme (c.665C.T (known as c.677C.T; rs1801133, p.A222V) and c.1286A.C (known as c.1298A.C; rs1801131, p.E429A)) were selected for genotype analysis. Login to comment

[hide] Functional polymorphisms of the ABCG2 gene are ass... Int J Mol Sci. 2014 May 22;15(5):9149-59. doi: 10.3390/ijms15059149. Zhou D, Liu Y, Zhang X, Gu X, Wang H, Luo X, Zhang J, Zou H, Guan M
Functional polymorphisms of the ABCG2 gene are associated with gout disease in the Chinese Han male population.
Int J Mol Sci. 2014 May 22;15(5):9149-59. doi: 10.3390/ijms15059149., [PMID:24857923]

Abstract [show]
BACKGROUND: Gout is a common type of arthritis that is characterized by hyperuricemia, tophi and joint inflammation. Genetic variations in the ABCG2 gene have been reported to influence serum uric acid levels and to participate in the pathogenesis of gout, but no further data have been reported in the Han Chinese population. METHODS: Peripheral blood DNA was isolated from 352 male patients with gout and 350 gout-free normal male controls. High-resolution melting analysis and Sanger sequencing were performed to identify the genetic polymorphisms V12M, Q141K and Q126X in the ABCG2 gene. Genotype and haplotype analyses were utilized to determine the disease odds ratios (ORs). A prediction model for gout risk using ABCG2 protein function was established based on the genotype combination of Q126X and Q141K. RESULTS: For Q141K, the A allele frequency was 49.6% in the gout patients and 30.9% in the controls (OR 2.20, 95% confidence interval (CI): 1.77-2.74, p=8.99x10(-)(1)(3)). Regarding Q126X, the T allele frequency was 4.7% in the gout patients and 1.7% in the controls (OR 2.91, 95% CI: 1.49-5.68, p=1.57x10(-)(3)). The A allele frequency for V12M was lower (18.3%) in the gout patients than in the controls (29%) (OR 0.55, 95% CI 0.43-0.71, p=2.55x10(-)(6)). In the order of V12M, Q126X and Q141K, the GCA and GTC haplotypes indicated increased disease risk (OR=2.30 and 2.71, respectively). Patients with mild to severe ABCG2 dysfunction accounted for 78.4% of gout cases. CONCLUSION: The ABCG2 126X and 141K alleles are associated with an increased risk of gout, whereas 12M has a protective effect on gout susceptibility in the Han Chinese population. ABCG2 dysfunction can be used to evaluate gout risk.

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5 High-resolution melting analysis and Sanger sequencing were performed to identify the genetic polymorphisms V12M, Q141K and Q126X in the ABCG2 gene. Login to comment
7 A prediction model for gout risk using ABCG2 protein function was established based on the genotype combination of Q126X and Q141K. Login to comment
8 Results: For Q141K, the A allele OPEN ACCESS frequency was 49.6% in the gout patients and 30.9% in the controls (OR 2.20, 95% confidence interval (CI): 1.77-2.74, p = 8.99 &#d7; 10-13 ). Login to comment
11 In the order of V12M, Q126X and Q141K, the GCA and GTC haplotypes indicated increased disease risk (OR = 2.30 and 2.71, respectively). Login to comment
33 Furthermore, the SNPs Q141K and Q126X in the human ABCG2 gene have recently been recognized as clinical biomarkers to assess hyperuricemia and gout. Login to comment
37 In the present study, we developed an HRM assay to detect three functional SNPs (Q141K, V12M and Q126X) and then assessed the genetic association of those SNPs in the ABCG2 gene with gout to investigate the association between ABCG2 dysfunction and gout risk in a Han Chinese male population. Login to comment
42 The results obtained from the DNA sequencing analysis confirmed the reliability of the HRM assay. The genotype and allelic frequencies of the three SNPs (Q141K, V12M and Q126X) among the cases and controls were in Hardy-Weinberg equilibrium for all of the polymorphisms analyzed. Login to comment
43 For Q141K, the A allele was found on 49.6% of the chromosomes from the gout patients compared with 30.9% of the chromosomes from the controls (OR 2.20, 95% CI: 1.77-2.74, p = 8.99 &#d7; 10-13 ). Login to comment
48 The three groups are well distinguished: (A) V12M; (B) Q126X; and (C) Q141K. Login to comment
54 SNP Genotype * Allele Frequency Mode Case Control p-Value p-Value OR 95% CI 1/1 1/2 2/2 MAF 1/1 1/2 2/2 MAF Q141K 84 181 87 0.496 33 150 167 0.309 1.18 &#d7; 10-11 8.99 &#d7; 10-13 2.20 1.77-2.74 Q126X 0 33 319 0.047 0 12 338 0.017 1.31 &#d7; 10-3 1.57 &#d7; 10-3 2.91 1.49-5.68 V12M 16 97 239 0.183 35 133 182 0.290 3.67 &#d7; 10-5 2.55 &#d7; 10-6 0.55 0.43-0.71 * The minor allele was referred to as allele 1, and the major allele was referred to as allele 2. Login to comment
55 Allele 1 is A and allele 2 is C in Q141K. Allele 1 is T and allele 2 is C in Q126X. Login to comment
58 Haplotype Analysis We performed a 3-SNP haplotype analysis (in the order V12M, Q126X and Q141K). Login to comment
63 Haplotype frequency analysis of V12M, Q126X and Q141K. Allele Frequency p-Value OR 95% CI V12M Q126X Q141K Gout Control G C A 0.481 0.289 1.26 &#d7; 10-13 2.30 1.84-2.87 G T C 0.044 0.017 2.97 &#d7; 10-3 2.71 1.37-5.36 G C C 0.292 0.404 8.27 &#d7; 10-6 0.60 0.48-0.75 A C C 0.165 0.271 1.53 &#d7; 10-6 0.53 0.41-0.69 2.3. Login to comment
74 Estimated Function Genotype Combination Number (%) p-Value OR 95% CI Q141K Q126X Gout Control ࣘ1/4 function C/A T/C 22 (6.2) 8 (2.3) 8.47 &#d7; 10-6 5.90 2.56-13.58 1/2 function C/C T/C 95 (27.0) 37 (10.5) 1.12 &#d7; 10-13 5.51 3.46-8.77 A/A C/C 3/4 function C/A C/C 159 (45.2) 142 (40.6) 1.00 &#d7; 10-6 2.40 1.69-3.42 Full function C/C C/C 76 (21.6) 163 (46.6) ߟ ߟ p-Value, OR and 95% CI for each ABCG2 dysfunction were obtained via comparisons with full function. Login to comment
76 Discussion This study is the first to examine the possible role of ABCG2 variants, which have previously been found to be associated with gout, in terms of their genetic susceptibility to gout in the Han Chinese population. We found that the Q141K, Q126X and V12M alleles were strongly associated with gout in Chinese males. Login to comment
87 The Q141K SNP has been extensively studied; it has been found to impair protein-nucleotide binding stability [28], which has been linked to hyperuricemia in a variety of populations [29]. Login to comment
93 These findings reflect the diversity of the Q126X and Q141K distributions in different ethnic populations, which may explain the different prevalence of gout in Chinese and Caucasian populations. Login to comment
96 Matsuo et al. [16,18] reported that the genotype combination of Q126X and Q141K is a clinically important biomarker for predicting gout risk in the Japanese population. We analyzed the relationship between ABCG2 transport dysfunction and gout and found that dysfunctional ABCG2 is responsible for approximately 78.4% of gout cases. Login to comment
115 We selected three functional ABCG2 SNPs: V12M, Q126X and Q141K. Login to comment
124 SNP ID SNP Allele Sequence (5'-3') Size V12M A/G ATGGTATGGGCCATTCATTG 250 bp ATGCCTTCAGGTCATTGGAA Q141K A/C ATGTTGTGATGGGCACTCTG 158 bp CCACATTACCTTGGAGTCTG Q126X C/T GCTGCAAGGAAAGATCCAAG 163 bp CAGCCAAAGCACTTACCCAT 4.3. Login to comment
137 The recent findings on the roles of the ABCG2 Q141K and Q126X polymorphism in gout may pave the way for pharmacological chaperones targeting ABCG2 as a potential new therapeutic target for gout. Login to comment

[hide] Common dysfunctional variants of ABCG2 have strong... Sci Rep. 2014 Jun 9;4:5227. doi: 10.1038/srep05227. Nakayama A, Matsuo H, Nakaoka H, Nakamura T, Nakashima H, Takada Y, Oikawa Y, Takada T, Sakiyama M, Shimizu S, Kawamura Y, Chiba T, Abe J, Wakai K, Kawai S, Okada R, Tamura T, Shichijo Y, Akashi A, Suzuki H, Hosoya T, Sakurai Y, Ichida K, Shinomiya N
Common dysfunctional variants of ABCG2 have stronger impact on hyperuricemia progression than typical environmental risk factors.
Sci Rep. 2014 Jun 9;4:5227. doi: 10.1038/srep05227., [PMID:24909660]

Abstract [show]
Gout/hyperuricemia is a common multifactorial disease having typical environmental risks. Recently, common dysfunctional variants of ABCG2, a urate exporter gene also known as BCRP, are revealed to be a major cause of gout/hyperuricemia. Here, we compared the influence of ABCG2 dysfunction on serum uric acid (SUA) levels with other typical risk factors in a cohort of 5,005 Japanese participants. ABCG2 dysfunction was observed in 53.3% of the population investigated, and its population-attributable risk percent (PAR%) for hyperuricemia was 29.2%, much higher than those of the other typical environmental risks, i.e. overweight/obesity (BMI >/= 25.0; PAR% = 18.7%), heavy drinking (>196 g/week (male) or >98 g/week (female) of pure alcohol; PAR% = 15.4%), and aging (>/=60 years old; PAR% = 5.74%). SUA significantly increased as the ABCG2 function decreased (P = 5.99 x 10(-19)). A regression analysis revealed that ABCG2 dysfunction had a stronger effect than other factors; a 25% decrease in ABCG2 function was equivalent to "an increase of BMI by 1.97-point" or "552.1 g/week alcohol intake as pure ethanol" in terms of ability to increase SUA. Therefore, ABCG2 dysfunction originating from common genetic variants has a much stronger impact on the progression of hyperuricemia than other familiar risks. Our study provides a better understanding of common genetic factors for common diseases.

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18 Based on the previous studies8,11 , all of the participants were divided into four groups by the combination of common dysfunctional variants of ABCG2, non-functional Q126X (rs72552713) and half-functional Q141K (rs2231142), as follows: full function (normal function), 3/ 4 function (mild dysfunction), 1/2 function (moderate dysfunction) and #1/4 function (severe dysfunction) (see Supplementary Figure S1 and Table S2). Login to comment
37 Functional analyses revealed that Q126X is a nonfunctional variant and Q141K is a half-functional variant due to the halved ABCG2 expression on the membrane8 . Login to comment
38 Since haplotype frequency analyses demonstrated no simultaneous presence of the minor alleles of Q126X and Q141K in one haplotype, the combination of nonfunctional variant Q126X and half-functional variant Q141K makes it possible to estimate dysfunctional levels of ABCG28,10 (Supplementary Figure S1 and Table S2). Login to comment
45 As for the relationship between SNPs and the risk of hyperuricemia, Woodward et al. so far reported that the PAR% of Q141K for gout was 10% in Caucasians7 , and Yamagishi et al. indicated that PAR% for gout and/or hyperuricemia was 19% in Japanese16 . Login to comment
46 Ours is the first report to show the PAR% of ABCG2 dysfunction using the combination of Q126X and Q141K for functional evaluation. Login to comment
47 It is reasonable that the PAR% of Q141K in Caucasians would be lower than that in Japanese, because the minor allele frequency in Caucasians (0.11 according to Woodward et al.7 ) is lower than those of Japanese (0.31 by Yamagishi et al.16 and 0.29 in the present study). Login to comment
48 When we re-calculated PAR% according to the definition of hyperuricemia in Yamagishi et al.16 (SUA $ 7.0 mg/dl), the resulting PAR% of Q141K was 22.2%, which is comparable to that in Yamagishi et al.16 . Login to comment
49 In the present study, we defined hyperuricemia as SUA .7.0 mg/dl17 and obtained the PAR% of Q141K and Q126X as 23.5% and 2.6%, respectively. Login to comment
52 Subsequent regression analysis revealed that ABCG2 dysfunction defined by the combination of Q126X and Q141K significantly increased SUA, while previous studies showed the association of SUA and only Q141K7,8 . Login to comment
76 Genotyping of the two variants in ABCG2 gene, Q126X and Q141K, was performed with a LightCycler 480 (Roche Diagnostics) by high resolution melting (HRM) analysis22 . Login to comment
79 The MAFs of Q126X and Q141K were 0.025 and 0.294, respectively, and both variants were in Hardy-Weinberg equilibrium (P . Login to comment

[hide] Association between the low-dose irinotecan regime... Oncol Lett. 2014 Jun;7(6):2035-2040. Epub 2014 Apr 8. Moriya H, Saito K, Helsby N, Sugino S, Yamakage M, Sawaguchi T, Takasaki M, Kato H, Kurosawa N
Association between the low-dose irinotecan regimen-induced occurrence of grade 4 neutropenia and genetic variants of in patients with gynecological cancers.
Oncol Lett. 2014 Jun;7(6):2035-2040. Epub 2014 Apr 8., [PMID:24932285]

Abstract [show]
The occurrence of severe neutropenia during treatment with irinotecan (CPT-11) is associated with the *6 and *28 alleles of uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1). However, the correlation between these variants and the occurrence of severe neutropenia in a low-dose CPT-11 regimen for the treatment of gynecological cancers has not been extensively studied. There are also no studies regarding the association between the 421C>A mutation in ATP-binding cassette sub-family G member 2 (ABCG2) and the occurrence of severe neutropenia in CPT-11-treated patients with gynecological cancers. The present study was designed to determine the factors associated with the occurrence of grade 4 neutropenia during chemotherapy for gynecological cancers with combinations of CPT-11 and cisplatin or mitomycin C. In total, 44 patients with gynecological cancer were enrolled in the study. The association between the absolute neutrophil count (ANC) nadir values, the total dose of CPT-11 and the genotypes of UGT1A1 or ABCG2 was studied. No correlation was observed between the ANC nadir values and the total dose of CPT-11. The ANC nadir values in the UGT1A1*6/*28 and *6/*6 groups were significantly lower compared with those in the *1/*1 group (P<0.01). Univariate analysis showed no association between the occurrence of grade 4 neutropenia and the ABCG2 421C>A mutation. Subsequent to narrowing the factors by univariate analysis, multivariate logistic regression analysis only detected significant correlations between the occurrence of grade 4 neutropenia and the UGT1A1*6/*6 and *6/*28 groups (P=0.029; odds ratio, 6.90; 95% confidence interval, 1.22-38.99). No associations were detected between the occurrence of grade 4 neutropenia and the heterozygous variant (*1/*6 or *1/*28) genotype, type of regimen or age. In conclusion, the UGT1A1*6/*28 and *6/*6 genotypes were found to be associated with the occurrence of severe neutropenia in the low-dose CPT-11 regimen for gynecological cancers. This finding indicates that the determination of UGT1A1 variants may be as useful in CPT-11 chemotherapy for gynecological conditions as it is in colorectal and lung cancer patients treated with this drug.

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29 In addition, the 421C>A (Q141K) variant of ATPߛbinding cassette subߛfamily G member 2 (ABCG2), which encodes the breast cancer resistance protein (BCRP), a transporter known to target various anticancer drugs, including CPTߛ11, has been reported to reduce the expression of ABCG2 and cause resistance to CPTߛ11 in vitro (20). Login to comment

[hide] Determinants of the activity and substrate recogni... Drug Metab Rev. 2014 Nov;46(4):459-74. doi: 10.3109/03602532.2014.942037. Epub 2014 Jul 18. Szafraniec MJ, Szczygiel M, Urbanska K, Fiedor L
Determinants of the activity and substrate recognition of breast cancer resistance protein (ABCG2).
Drug Metab Rev. 2014 Nov;46(4):459-74. doi: 10.3109/03602532.2014.942037. Epub 2014 Jul 18., [PMID:25036722]

Abstract [show]
The xenobiotic transporters are among the most important constituents of detoxification system in living organisms. Breast cancer resistance protein (BCRP/ABCG2) is one of the major transporters involved in the efflux of xenobiotics. To understand its role in chemotherapeutic and multidrug resistance, it is crucial to establish the determinants of its substrate specificity, which obviously is of high relevance for successful therapy of many diseases. This article summarizes the current knowledge about the substrate preferences of BCRP. We overview the factors which determine its activity, inhibition and substrate recognition, focusing on the structural features of the transporter. BCRP substrate specificity is quite low as it interacts with a spectrum of substances with only a few common features: hydrophobic and aromatic regions, possibly a flat conformation and the metal ion-, oxygen- and nitrogen-containing functionalities, most of which may be the donors/acceptors of H-bonds. Several amino acid residues and structural motifs are responsible for BCRP activity and substrate recognition. Thus, the active form of BCRP, at least a dimer or a larger oligomer is maintained by intramolecular disulfide bridge that involves Cys(603) residues. The GXXXG motif in transmembrane helix 1, Cys residues, Arg(482) and Lys(86) are responsible for maintaining the protein structure, which confers transport activity, and the His(457) or Arg(456) residues are directly involved in substrate binding. Arg(482) does not directly bind substrates, but electrostatically interacts with charged molecules, which initiates the conformational changes that transmit the signal from the transmembrane regions to the ABC domain.

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201 To elucidate the significance of this polymorphism for porphyrin transport, a set of 18 variants of BCRP (Val12 Met, Gly51 Cys, Gln126 stop, Gln141 Lys, Thr153 Met, Gln166 Glu, Ile206 Leu, Phe208 Ser, Ser248 Pro, Glu334 stop, Phe431 Leu, Ser441 Asn, Arg482 Gly, Arg482 Thr, Phe489 Leu, Phe571 Ile, Asn590 Tyr and Asp620 Asn) have been expressed in insect cells. Login to comment

[hide] Role of the breast cancer resistance protein (BCRP... AAPS J. 2015 Jan;17(1):65-82. doi: 10.1208/s12248-014-9668-6. Epub 2014 Sep 19. Mao Q, Unadkat JD
Role of the breast cancer resistance protein (BCRP/ABCG2) in drug transport--an update.
AAPS J. 2015 Jan;17(1):65-82. doi: 10.1208/s12248-014-9668-6. Epub 2014 Sep 19., [PMID:25236865]

Abstract [show]
The human breast cancer resistance protein (BCRP, gene symbol ABCG2) is an ATP-binding cassette (ABC) efflux transporter. It was so named because it was initially cloned from a multidrug-resistant breast cancer cell line where it was found to confer resistance to chemotherapeutic agents such as mitoxantrone and topotecan. Since its discovery in 1998, the substrates of BCRP have been rapidly expanding to include not only therapeutic agents but also physiological substances such as estrone-3-sulfate, 17beta-estradiol 17-(beta-D-glucuronide) and uric acid. Likewise, at least hundreds of BCRP inhibitors have been identified. Among normal human tissues, BCRP is highly expressed on the apical membranes of the placental syncytiotrophoblasts, the intestinal epithelium, the liver hepatocytes, the endothelial cells of brain microvessels, and the renal proximal tubular cells, contributing to the absorption, distribution, and elimination of drugs and endogenous compounds as well as tissue protection against xenobiotic exposure. As a result, BCRP has now been recognized by the FDA to be one of the key drug transporters involved in clinically relevant drug disposition. We published a highly-accessed review article on BCRP in 2005, and much progress has been made since then. In this review, we provide an update of current knowledge on basic biochemistry and pharmacological functions of BCRP as well as its relevance to drug resistance and drug disposition.

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214 Of these SNPs, 34G>A (V12M) and 421C>A (Q141K) occur most frequently in East Asians (~30-60%) and with relatively low allele frequencies in Caucasians and African-American populations (~5-10%). Login to comment
217 In vitro expression and functional studies generally support the conclusion that the Q141K variant resulting from the 421C>A SNP has reduced cell surface expression in transfected cells and therefore cells expressing Q141K display lower efflux activities compared to those expressing wild-type BCRP (100). Login to comment
326 In vitro transport studies confirmed that uric acid is a BCRP substrate, and cells expressing the Q141K variant resulting from the ABCG2 421C>A SNP had lower uric acid efflux activity than cells expressing wild-type BCRP. Login to comment

[hide] Association of ABCG2 polymorphism with clinical ef... Cancer Chemother Pharmacol. 2015 Jan;75(1):173-82. doi: 10.1007/s00280-014-2630-6. Epub 2014 Nov 23. Koo DH, Ryu MH, Ryoo BY, Beck MY, Na YS, Shin JG, Lee SS, Kim EY, Kang YK
Association of ABCG2 polymorphism with clinical efficacy of imatinib in patients with gastrointestinal stromal tumor.
Cancer Chemother Pharmacol. 2015 Jan;75(1):173-82. doi: 10.1007/s00280-014-2630-6. Epub 2014 Nov 23., [PMID:25417047]

Abstract [show]
PURPOSE: Imatinib is a substrate of drug transporters and metabolizing enzymes, including members of the cytochrome P450 (CYP) system. Differences in imatinib pharmacokinetics among individuals might be influenced by genetic polymorphisms and be associated with variable clinical imatinib efficacy. This study sought to test how genetic polymorphisms can affect the clinical efficacy of imatinib and its blood levels in GIST patients. METHODS: A total of 209 GIST patients who had received imatinib 400 mg daily were genotyped for six single-nucleotide polymorphisms in three genes (CYP3A5 6986A>G; ABCB1 1236C>T, 2677G>A/T, and 3435C>T; and ABCG2 34G>A and 421C>A) via blood samples. Progression-free survival (PFS) and imatinib plasma trough levels were evaluated and compared according to genotypes. RESULTS: With a median follow-up of 39.6 months (range 16.7-97.5 months), the estimated 5-year PFS rate was 67.5 % (95 % CI 59.9-75.1). Among the CYP3A5, ABCB1, and ABCG2 genotypes, ABCG2 421C>A was associated with PFS. The 5-year PFS rate in patients with the AA variant of ABCG2 421C>A (92.3 %; 95 % CI 77.8-100.0) was significantly superior to that of patients with CC/CA genotypes (65.0 %; 95 % CI 56.9-73.1; p = 0.047). For the imatinib trough levels, there were no statistically significant differences when comparing polymorphisms among all genotypes, even after adjusting for clinical factors, including sex, age, body surface area, hemoglobin, albumin, and creatinine clearance. CONCLUSIONS: The ABCG2 421C>A genetic variation could influence clinical efficacy in terms of PFS in patients with advanced GIST undergoing imatinib therapy.

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103 The most studied polymorphism in ABCG2 is the non-synonymous SNP 421C>A in exon 5, which results in a glutamine to lysine substitution at codon 141 (Q141K), localized in the ATP-binding domain [24]. Login to comment
104 It is well known that ABCG2 harboring the Q141K polymorphism results in impaired expression, incomplete transport to the plasma membrane, and decreased efflux function of BCRP, and consequently results in reduced clearance of ABCG2 substrate drugs [25]. Login to comment
105 As cells expressing ABCG2 Q141K have a higher imatinib accumulation in vitro compared to cells wild type in ABCG2, this variant form of ABCG2 is associated with TableÊf;5ߒߙImatinib plasma trough level according to genotype SD standard deviation, BSA body surface area, Hb hemoglobin, WBC white blood cell, CCr creatinine clearance * p value for log-transformed imatinib level ** After adjusting for sex, age, BSA, Hb, Albumin, and CCr *** Patients who received imatinib for metastatic or recurrent GIST (N = 145) Genotype (N) Imatinib trough level (ng/mL) p value* p value** p value*** Mean SD CYP3A5 6986A>G AA (13) 1,342.8 450.3 0.918 0.764 0.656 AG (68) 1,375.6 633.3 GG (128) 1415.8 711.7 ABCB1 1236C>T CC (35) 1346.3 673.9 0.205 0.573 0.092 CT (94) 1347.2 678.3 TT (80) 1480.7 662.1 2677G>A/T GG (41) 1477.4 774.9 0.394 0.315 0.447 Hetero (135) 1350.2 620.5 TT (33) 1495.8 734.3 3435C>T CC (81) 1454.9 672.1 0.338 0.258 0.239 CT (98) 1343.4 708.1 TT (30) 1423.9 538.1 ABCG2 34G>A GG (119) 1360.5 486.8 0.930 0.940 0.959 GA (75) 1393.8 727.1 AA (15) 1405.7 659.2 421C>A CC (103) 1339.3 655.3 0.216 0.477 0.201 CA (88) 1477.4 693.5 AA (18) 1347.9 648.6 Fig.Êf;2ߒߙImatinib trough plasma levels according to genetic polymorphisms among metabolizer and efflux-transporter genes. Login to comment
107 This impaired functioning of the ABCG2 protein in cells transfected with ABCG2 Q141K is similar to the result of SNP-induced alterations in the function of the BCRP protein itself [28]. Login to comment

[hide] The molecular physiology of uric acid homeostasis. Annu Rev Physiol. 2015;77:323-45. doi: 10.1146/annurev-physiol-021113-170343. Epub 2014 Nov 12. Mandal AK, Mount DB
The molecular physiology of uric acid homeostasis.
Annu Rev Physiol. 2015;77:323-45. doi: 10.1146/annurev-physiol-021113-170343. Epub 2014 Nov 12., [PMID:25422986]

Abstract [show]
Uric acid, generated from the metabolism of purines, has proven and emerging roles in human disease. Serum uric acid is determined by production and the net balance of reabsorption or secretion by the kidney and intestine. A detailed understanding of epithelial absorption and secretion of uric acid has recently emerged, aided in particular by the results of genome-wide association studies of hyperuricemia. Novel genetic and regulatory networks with effects on uric acid homeostasis have also emerged. These developments promise to lead to a new understanding of the various diseases associated with hyperuricemia and to novel, targeted therapies for hyperuricemia.

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234 A common nonsynonymous SNP in ABCG, Q141K, reduced ABCG2 function by 53% in this system. Login to comment
235 Subsequent work in mammalian cells (38, 136) and Xenopus oocytes (136) revealed a temperature-dependent expression defect in Q141K-ABCG2, linked to instability of the nucleotide-binding domain (136). Login to comment
236 There are similarities in the biochemical phenotype of Q141K-ABCG2 and F508 CFTR proteins such that small-molecule therapies designed for cystic fibrosis can rescue the expression defect of Q141K-ABCG2 (136). Login to comment
722 Gout-causing Q141K mutation in ABCG2 leads to instability of the nucleotide-binding domain and can be corrected with small molecules. Login to comment

[hide] The role of the efflux carriers Abcg2 and Abcc2 fo... Chem Biol Interact. 2014 Oct 16;224C:36-41. doi: 10.1016/j.cbi.2014.10.009. Kranz J, Hessel S, Aretz J, Seidel A, Petzinger E, Geyer J, Lampen A
The role of the efflux carriers Abcg2 and Abcc2 for the hepatobiliary elimination of benzo[a]pyrene and its metabolites in mice.
Chem Biol Interact. 2014 Oct 16;224C:36-41. doi: 10.1016/j.cbi.2014.10.009., [PMID:25451572]

Abstract [show]
The ATP-binding cassette transporters Breast Cancer Resistance Protein (Abcg2) and Multidrug Resistance-associated Protein 2 (Abcc2) play an important role for the hepatobiliary elimination of drugs and toxins as well as their metabolites. Previous in vitro transport studies showed that both transporters are involved in the active efflux of phase II metabolites of carcinogenic benzo[a]pyrene (BP), however the role of these carriers in hepatobiliary elimination in vivo is still unknown. In the present study, Abcg2(-/-) and Abcc2(-/-) knockout mice were used to elucidate the role of Abcg2 and Abcc2 for the hepatobiliary excretion of BP and its metabolites. After intravenous application of [3H]BP the hepatobiliary excretion was significantly reduced in these mice: whereas wild type mice excreted on average 25.4% of the applied dose into the bile over 90min, Abcg2(-/-) knockout mice only excreted 10.7% and Abcc2(-/-) knockout mice 8.6%. As a consequence, [3H]BP concentrations were in general higher in the plasma and in most of the organs of the Abcg2 and Abcc2 knockout mice. Both transporters may have a protective function for BP-induced carcinogenesis in humans, due to its crucial importance for the hepatobiliary elimination of BP via bile. Subjects with reduced ABCG2 or ABCC2 expression might have higher oral bioavailability for BP due to a reduced excretion and so might be more susceptible to BP-induced carcinogenesis.

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172 One of these SNP, C421A generates an amino acid exchange (Gln141Lys) at the N-terminus of the protein and was shown to decrease the protein expression level and ATPase activity of the ABCG2 transporter [19,27]. Login to comment

[hide] Lack of efficacy of mitoxantrone in primary progre... J Neuroimmunol. 2015 Jan 15;278:277-9. doi: 10.1016/j.jneuroim.2014.11.017. Epub 2014 Nov 20. Grey Nee Cotte S, Salmen Nee Stroet A, von Ahsen N, Starck M, Winkelmann A, Zettl UK, Comabella M, Montalban X, Zipp F, Fleischer V, Kruse N, Gold R, Chan A
Lack of efficacy of mitoxantrone in primary progressive Multiple Sclerosis irrespective of pharmacogenetic factors: a multi-center, retrospective analysis.
J Neuroimmunol. 2015 Jan 15;278:277-9. doi: 10.1016/j.jneuroim.2014.11.017. Epub 2014 Nov 20., [PMID:25468777]

Abstract [show]
BACKGROUND: Mitoxantrone is used on an off-label basis in primary progressive MS (PPMS). ABC-transporter-genotypes are associated with therapeutic response in relapsing/secondary progressive MS (RP/SPMS). OBJECTIVE: To evaluate potential pharmacogenetic response markers for mitoxantrone in PPMS. METHODS: 41 mitoxantrone-treated PPMS-patients, 155 mitoxantrone-treated RP/SPMS-patients and 43 PPMS-controls were retrospectively assessed for clinical therapy-response and in correlation with four single-nucleotide-polymorphisms in ABCB1- and ABCG2-genes. RESULTS: 53.7% PPMS-patients were mitoxantrone-responders, in comparison to 78.1% of RP/SPMS-patients (p=0.039). There was no association between genotype and treatment response. CONCLUSION: Our data discourages the use of mitoxantrone in PPMS regardless of pharmacogenetic response markers previously described in RP/SPMS.

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16 Patients, material and methods After approval by local ethics committees and informed consent, genotyping for ABCG2 V12M (reference SNP rs2231137) and Q141K (rs2231142) and ABCB1 3435CNT (rs1045642) and 2677GNT (rs2032582) was performed using TaqManࡊ polymerase chain reaction Journal of Neuroimmunology 278 (2015) 277-279 Ìe; Corresponding author at: Department of Neurology, St. Josef-Hospital, Ruhr University Bochum, Gudrunstr. Login to comment

[hide] Investigation of the functional single-nucleotide ... Biomed Rep. 2015 Jan;3(1):105-109. Epub 2014 Nov 11. Sari FM, Yanar HT, Ozhan G
Investigation of the functional single-nucleotide polymorphisms in the transporter and susceptibility to colorectal cancer.
Biomed Rep. 2015 Jan;3(1):105-109. Epub 2014 Nov 11., [PMID:25469257]

Abstract [show]
Breast cancer resistance protein (BCRP) protects tissues by actively transporting xenobiotics and their metabolites out of the cells. BCRP is expressed in the apical membrane of normal intestinal and colonic epithelium. The BCRP substrates include a number of structurally unrelated compounds, such as drugs, pesticides, carcinogens and endogenous compounds. Although the functional and common BCRP alleles, 34G>A and 421C>A, are shown to vary by ethnicity, their potential mechanism has not been adequately described with regards to affecting the susceptibility to colorectal cancer. The present study aimed to evaluate the effects of the BCRP variants on the susceptibility to colorectal cancer and to predict the individual responses to xenobiotics transferred by BCRP. BCRP 421C>A was significantly associated with the colorectal cancer risk (odds ratio, 16.12; P=0.005). These findings are the first results of BCRP allele distributions in the Turkish population and provide an understanding of the correlation between therapeutic approaches and etiology of colorectal cancer.

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21 The BCRP 421C>A variant (rs2231142), causes a Gln141Lys (Q141K) change, and is associated with low levels of BCRP expression (13). Login to comment

[hide] Sunitinib-induced severe toxicities in a Japanese ... BMC Cancer. 2014 Dec 16;14:964. doi: 10.1186/1471-2407-14-964. Miura Y, Imamura CK, Fukunaga K, Katsuyama Y, Suyama K, Okaneya T, Mushiroda T, Ando Y, Takano T, Tanigawara Y
Sunitinib-induced severe toxicities in a Japanese patient with the ABCG2 421 AA genotype.
BMC Cancer. 2014 Dec 16;14:964. doi: 10.1186/1471-2407-14-964., [PMID:25515134]

Abstract [show]
BACKGROUND: Sunitinib is a multi-targeted receptor tyrosine kinase inhibitor that acts against receptors for vascular endothelial growth factor and platelet-derived growth factor. Common toxicities of sunitinib treatment include hypertension, hand-foot syndrome, vomiting, and diarrhea, and the proportion of grade 3 or 4 adverse events relating to sunitinib treatment range from 1 to 13% for all categories. It is reported that increased exposure to sunitinib is associated with improved clinical outcomes but also carries an increased risk of adverse effects. CASE PRESENTATION: A 73-year-old Japanese woman with metastatic renal cell carcinoma who received sunitinib at a dose of 50 mg once daily suffered a high-grade fever on day 11 of treatment. Sunitinib treatment was discontinued on day 12; however, severe thrombocytopenia and transaminase elevation occurred and persisted more than a week. Additionally, severe hypoxia due to pleural effusion and pulmonary edema developed despite immediate discontinuation of sunitinib. On day 14, three days after the discontinuation of sunitinib, the plasma concentrations of sunitinib and its major active metabolite N-desethyl sunitinib (SU12662) were extremely high (131.9 ng/mL and 28.4 ng/mL, respectively). By day 25, all toxicities had resolved, and a CT scan revealed marked tumor shrinkage. Genotyping of seven single-nucleotide polymorphisms that are potentially relevant to the pharmacokinetics of sunitinib was performed. The patient's genotype of ABCG2 (ATP-binding cassette, sub-family G (WHITE), member 2) 421C > A was homozygous for the variant allele (AA), which was reported to be associated with high exposure to sunitinib. Therefore, we speculated that the extremely high plasma concentrations of sunitinib and SU12662 caused by the ABCG2 421 AA genotype might have resulted in severe toxicities to the patient. CONCLUSION: The minor allele frequencies of ABCG2 421C > A are approximately three-fold higher in Asians than in Caucasians. Our report suggests that pharmacogenetic factors should be considered when severe and rapid-onset adverse drug reactions occur in Asian patients, including Japanese treated with sunitinib.

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85 Another population pharmacokinetics study also identified ethnic background as a significant covariate for the Table 1 Genotypes of seven SNPs in CYP3A5, ABCB1 and ABCG2 Gene SNP Allele Amino acid Genotype CYP3A5 rs776746 6986A > G Splice Site AG ABCB1 rs1128503 1236C > T G412G CT ABCB1 rs2032582 2677G > T/A A893S/T GT ABCB1 rs1045642 3435C > T I1145I CT ABCG2 rs2231137 34G > A V12M GG ABCG2 rs72552713 376G > A Q126X GG ABCG2 rs2231142 421C > A Q141K AA prediction of oral clearance [16]. Login to comment
137 Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, Tsuruo T, Miki Y, Sugimoto Y: C421A polymorphism in the human breast cancer resistance protein gene is associated with low expression of Q141K protein and low-level drug resistance. Login to comment

[hide] Rosuvastatin pharmacokinetics and pharmacogenetics... Eur J Clin Pharmacol. 2015 Mar;71(3):329-40. doi: 10.1007/s00228-014-1800-0. Epub 2015 Jan 30. Birmingham BK, Bujac SR, Elsby R, Azumaya CT, Zalikowski J, Chen Y, Kim K, Ambrose HJ
Rosuvastatin pharmacokinetics and pharmacogenetics in Caucasian and Asian subjects residing in the United States.
Eur J Clin Pharmacol. 2015 Mar;71(3):329-40. doi: 10.1007/s00228-014-1800-0. Epub 2015 Jan 30., [PMID:25630984]

Abstract [show]
PURPOSE: Systemic exposure to rosuvastatin in Asian subjects living in Japan or Singapore is approximately twice that observed in Caucasian subjects in Western countries or in Singapore. This study was conducted to determine whether pharmacokinetic differences exist among the most populous Asian subgroups and Caucasian subjects in the USA. METHOD: Rosuvastatin pharmacokinetics was studied in Chinese, Filipino, Asian-Indian, Korean, Vietnamese, Japanese and Caucasian subjects residing in California. Plasma concentrations of rosuvastatin and metabolites after a single 20-mg dose were determined by mass spectrometric detection. The influence of polymorphisms in SLCO1B1 (T521>C [Val174Ala] and A388>G [Asn130Asp]) and in ABCG2 (C421>A [Gln141Lys]) on exposure to rosuvastatin was also assessed. RESULTS: The average rosuvastatin area under the curve from time zero to time of last quantifiable concentration was between 64 and 84 % higher, and maximum drug concentration was between 70 and 98 % higher in East Asian subgroups compared with Caucasians. Data for Asian-Indians was intermediate to these two ethnic groups at 26 and 29 %, respectively. Similar increases in exposure to N-desmethyl rosuvastatin and rosuvastatin lactone were observed. Rosuvastatin exposure was higher in subjects carrying the SLCO1B1 521C allele compared with that in non-carriers of this allele. Similarly, exposure was higher in subjects carrying the ABCG2 421A allele compared with that in non-carriers. CONCLUSION: Plasma exposure to rosuvastatin and its metabolites was significantly higher in Asian populations residing in the USA compared with Caucasian subjects living in the same environment. This study suggests that polymorphisms in the SLCO1B1 and ABCG2 genes contribute to the variability in rosuvastatin exposure.

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4 The influence of polymorphisms in SLCO1B1 (T521>C [Val174Ala] and A388>G [Asn130Asp]) and in ABCG2 (C421>A [Gln141Lys]) on exposure to rosuvastatin was also assessed. Login to comment
37 In addition, an exploratory pharmacogenetic analysis was performed to examine the effects of three SNPs on rosuvastatin pharmacokinetics, including two polymorphisms in SLCO1B1 (T521>C [Val174Ala] and A388>G [Asn130Asp]) and one polymorphism in ABCG2 (C421>A [Gln141Lys]). Login to comment
62 All data were analyzed with Sequence Detection Systems (c) software v2.1: SLCO1B1 A388G (Asn130Asp) rs2306283 and ABI assay C__1901697_20 SLCO1B1 T521C (Val174Ala) rs4149056 and ABI assay C__30633906_10 ABCG2 C421A (Gln141Lys) rs2231142 and ABI assay C__15854163_70. Login to comment
133 The 421A allele, present at a higher frequency in the Asian populations (apart from the Asian Indians) than in the Caucasian population (Table 4), appeared to account Table 4 Genotype frequencies and HWE results by ethnic group Polymorphism Genotype Frequency (%) Pooled Asiana n=100 Chinese n=17 Filipino n=19 Asian-Indian n=16 Korean n=23 Vietnamese n=16 Japanese n=25 Caucasian n=22 SLCO1B1: T521>C (Val174Ala) T\T T\C C\C 78 (78 %) 22 (22 %) 0 15 (88 %) 2 (12 %) 0 14 (74 %) 5 (26 %) 0 16 (100 %) 0 0 18 (78 %) 5 (22 %) 0 12 (75 %) 4 (25 %) 0 19 (76 %) 6 (24 %) 0 15 (68 %) 6 (27 %) 1 (5 %) HWE p value - 1.00 1.00 1.00 1.00 1.00 1.00 0.54 SLCO1B1: A388>G (Asn130Asp) A\A A\G G\G 7 (7 %) 33 (33 %) 60 (60 %) 1 (6 %) 5 (29 %) 11 (65 %) 2 (11 %) 2 (11 %) 15 (79 %) 4 (25 %) 8 (50 %) 4 (25 %) 2 (9 %) 11 (48 %) 10 (43 %) 0 (0 %) 2 (12.5 %) 14 (87.5 %) 2 (8 %) 13 (52 %) 10 (40 %) 9 (41 %) 9 (41 %) 4 (18 %) HWE p value - 0.54 0.03 1.00 1.00 1.00 0.66 0.65 ABCG2: C421>A (Gln141Lys) C\C C\A A\A 49 (49 %) 46 (46 %) 5 (5 %) 7 (41 %) 10 (59 %) 0 10 (53 %) 7 (37 %) 2 (11 %) 14 (88 %) 1 (6 %) 1 (6 %) 13 (57 %) 10 (43 %) 0 7 (44 %) 8 (50 %) 1 (6 %) 12 (48 %) 11 (44 %) 2 (8 %) 15 (68 %) 7 (32 %) 0 HWE p value - 0.24 0.61 0.10 0.54 1.00 1.00 1.00 HWE Hardy-Weinberg equilibrium a The pooled Asian group comprised all Chinese, Filipino, Korean, Vietnamese and Japanese subjects for some but not all of the inter-ethnic group variability in exposure. Login to comment
199 A study in Chinese subjects investigating the effect of the ABCG2 C421>A (Gln141Lys) polymorphism on rosuvastatin exposure has suggested that the 421A allele is significantly associated with higher Cmax and AUC than the 421C allele [15]. Login to comment

[hide] Genome-wide association study of clinically define... Ann Rheum Dis. 2015 Feb 2. pii: annrheumdis-2014-206191. doi: 10.1136/annrheumdis-2014-206191. Matsuo H, Yamamoto K, Nakaoka H, Nakayama A, Sakiyama M, Chiba T, Takahashi A, Nakamura T, Nakashima H, Takada Y, Danjoh I, Shimizu S, Abe J, Kawamura Y, Terashige S, Ogata H, Tatsukawa S, Yin G, Okada R, Morita E, Naito M, Tokumasu A, Onoue H, Iwaya K, Ito T, Takada T, Inoue K, Kato Y, Nakamura Y, Sakurai Y, Suzuki H, Kanai Y, Hosoya T, Hamajima N, Inoue I, Kubo M, Ichida K, Ooyama H, Shimizu T, Shinomiya N
Genome-wide association study of clinically defined gout identifies multiple risk loci and its association with clinical subtypes.
Ann Rheum Dis. 2015 Feb 2. pii: annrheumdis-2014-206191. doi: 10.1136/annrheumdis-2014-206191., [PMID:25646370]

Abstract [show]
OBJECTIVE: Gout, caused by hyperuricaemia, is a multifactorial disease. Although genome-wide association studies (GWASs) of gout have been reported, they included self-reported gout cases in which clinical information was insufficient. Therefore, the relationship between genetic variation and clinical subtypes of gout remains unclear. Here, we first performed a GWAS of clinically defined gout cases only. METHODS: A GWAS was conducted with 945 patients with clinically defined gout and 1213 controls in a Japanese male population, followed by replication study of 1048 clinically defined cases and 1334 controls. RESULTS: Five gout susceptibility loci were identified at the genome-wide significance level (p<5.0x10-8), which contained well-known urate transporter genes (ABCG2 and SLC2A9) and additional genes: rs1260326 (p=1.9x10-12; OR=1.36) of GCKR (a gene for glucose and lipid metabolism), rs2188380 (p=1.6x10-23; OR=1.75) of MYL2-CUX2 (genes associated with cholesterol and diabetes mellitus) and rs4073582 (p=6.4x10-9; OR=1.66) of CNIH-2 (a gene for regulation of glutamate signalling). The latter two are identified as novel gout loci. Furthermore, among the identified single-nucleotide polymorphisms (SNPs), we demonstrated that the SNPs of ABCG2 and SLC2A9 were differentially associated with types of gout and clinical parameters underlying specific subtypes (renal underexcretion type and renal overload type). The effect of the risk allele of each SNP on clinical parameters showed significant linear relationships with the ratio of the case-control ORs for two distinct types of gout (r=0.96 [p=4.8x10-4] for urate clearance and r=0.96 [p=5.0x10-4] for urinary urate excretion). CONCLUSIONS: Our findings provide clues to better understand the pathogenesis of gout and will be useful for development of companion diagnostics.

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55 Two dysfunctional SNPs of ABCG2 We previously demonstrated that two dysfunctional SNPs of ABCG2, rs72552713 (Gln126Ter) and rs2231142 (Gln141Lys), were located on different haplotypes4 18 and strongly associated with hyperuricaemia and gout.4 18 32 Therefore, we additionally performed genotyping of these two SNPs by an allelic discrimination assay because SNPs are not on Illumina HumanOmniExpress V .1.0 (Illumina). Login to comment
86 ABCG2 is identified to have an association with SUA levels by recent GWASs.9-16 Subsequent genetic and functional analysis17 18 revealed that ABCG2 is a high-capacity urate exporter and shows the reduced transport of urate by a common half-functional variant, rs2231142 (Gln141Lys). Login to comment
109 &#a7;Non-synonymous SNPs (rs1260326, Leu446Pro; rs72552713, Gln126Ter; and rs2231142, Gln141Lys). Login to comment

[hide] ASSOCIATIONS BETWEEN BODY MASS INDEX AND SERUM URI... Nagoya J Med Sci. 2014 Aug;76(3-4):333-9. Suma S, Naito M, Okada R, Kawai S, Yin G, Morita E, Wakai K, Matsuo H, Hamajima N
ASSOCIATIONS BETWEEN BODY MASS INDEX AND SERUM URIC ACID LEVELS IN A JAPANESE POPULATION WERE SIGNIFICANTLY MODIFIED BY LRP2 rs2544390.
Nagoya J Med Sci. 2014 Aug;76(3-4):333-9., [PMID:25741042]

Abstract [show]
The genome-wide association study identified associations between the LRP2 polymorphism rs2544390 and serum uric acid (SUA) levels in a Japanese population. Our previous study on the LRP2 rs2544390 polymorphism identified an interaction between SUA and alcohol consumption. Here, we investigated an interaction with body mass index (BMI) using the same dataset. Subjects were 3,742 health checkup examinees (2,544 males and 1,198 females) aged 35-69 years. Those with the SLC22A12 258WW genotype, SLC2A9 rs11722228 C allele, and ABCG2 126QQ genotype and 141Q allele were selected for analysis to remove the strong influences of these genetic traits. In males, the odds ratio of BMI >/=25.0 relative to BMI <18.5 for hyperuricemia (SUA >/=7 mg/dL and/or under medication for hyperuricemia) was 6.58 (95% confidence interval [CI], 0.84-51.32) for CC, 10.08 (2.38-42.83) for CT, and 2.53 (0.54-11.78) for TT. The interaction was 0.59 (p=0.029) from the model including BMI (<25.0 and >/=25.0), genotype (CC/CT and TT), and the multiplicative interaction term between BMI >/=25.0 and the TT genotype. In females, the odds ratio of BMI >/=25.0 relative to BMI <18.5 for high SUA (>/=5 mg/dL and/or under medication for hyperuricemia) was 6.35 (95%CI, 1.68-24.08) for CC, 4.55 (1.85-11.18) for CT, and 5.93 (1.97-17.90) for TT. The interaction term was significant in the opposite direction for females (OR=2.75, p=0.011). The association between BMI and SUA was therefore modified by the LRP2 polymorphism in this Japanese population.

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17 Polymorphisms ABCG2 Q126X (rs72552713) and Q141K (rs2231142) have been shown to influence the risk of hyperuricemia through reduction of the uric acid transportation activity.16-18) Our previous studies have confirmed the associations with these polymorphisms,16,19,20) and we observed an interaction with alcohol consumption on SUA in our investigation of the LRP2 intron 1 polymorphism rs2544390.21) Because obesity is an important factor that determines SUA,6) the present study aimed to investigate the interaction between LRP2 rs2544390 and BMI on SUA levels using the same dataset as our previous work. Login to comment

[hide] Modulation of genetic associations with serum urat... PLoS One. 2015 Mar 26;10(3):e0119752. doi: 10.1371/journal.pone.0119752. eCollection 2015. Huffman JE, Albrecht E, Teumer A, Mangino M, Kapur K, Johnson T, Kutalik Z, Pirastu N, Pistis G, Lopez LM, Haller T, Salo P, Goel A, Li M, Tanaka T, Dehghan A, Ruggiero D, Malerba G, Smith AV, Nolte IM, Portas L, Phipps-Green A, Boteva L, Navarro P, Johansson A, Hicks AA, Polasek O, Esko T, Peden JF, Harris SE, Murgia F, Wild SH, Tenesa A, Tin A, Mihailov E, Grotevendt A, Gislason GK, Coresh J, D'Adamo P, Ulivi S, Vollenweider P, Waeber G, Campbell S, Kolcic I, Fisher K, Viigimaa M, Metter JE, Masciullo C,
Modulation of genetic associations with serum urate levels by body-mass-index in humans.
PLoS One. 2015 Mar 26;10(3):e0119752. doi: 10.1371/journal.pone.0119752. eCollection 2015., [PMID:25811787]

Abstract [show]
We tested for interactions between body mass index (BMI) and common genetic variants affecting serum urate levels, genome-wide, in up to 42569 participants. Both stratified genome-wide association (GWAS) analyses, in lean, overweight and obese individuals, and regression-type analyses in a non BMI-stratified overall sample were performed. The former did not uncover any novel locus with a major main effect, but supported modulation of effects for some known and potentially new urate loci. The latter highlighted a SNP at RBFOX3 reaching genome-wide significant level (effect size 0.014, 95% CI 0.008-0.02, Pinter= 2.6 x 10-8). Two top loci in interaction term analyses, RBFOX3 and ERO1LB-EDARADD, also displayed suggestive differences in main effect size between the lean and obese strata. All top ranking loci for urate effect differences between BMI categories were novel and most had small magnitude but opposite direction effects between strata. They include the locus RBMS1-TANK (men, Pdifflean-overweight= 4.7 x 10-8), a region that has been associated with several obesity related traits, and TSPYL5 (men, Pdifflean-overweight= 9.1 x 10-8), regulating adipocytes-produced estradiol. The top-ranking known urate loci was ABCG2, the strongest known gout risk locus, with an effect halved in obese compared to lean men (Pdifflean-obese= 2 x 10-4). Finally, pathway analysis suggested a role for N-glycan biosynthesis as a prominent urate-associated pathway in the lean stratum. These results illustrate a potentially powerful way to monitor changes occurring in obesogenic environment.

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252 The ATP-binding cassette transporter ABCG2 has been established as a high capacity urate transporter, is expressed in renal proximal tubules, liver and intestines, and the hyperuricemia causal Q141K mutation has been shown to reduce urate transport rates [39]. Login to comment
254 Interestingly, BMI-dependent effects of Q141K on urate response to acute fructose exposure have been recently reported [41]. Login to comment

[hide] An update on the genetic architecture of hyperuric... Arthritis Res Ther. 2015 Apr 10;17:98. doi: 10.1186/s13075-015-0609-2. Merriman TR
An update on the genetic architecture of hyperuricemia and gout.
Arthritis Res Ther. 2015 Apr 10;17:98. doi: 10.1186/s13075-015-0609-2., [PMID:25889045]

Abstract [show]
Genome-wide association studies that scan the genome for common genetic variants associated with phenotype have greatly advanced medical knowledge. Hyperuricemia is no exception, with 28 loci identified. However, genetic control of pathways determining gout in the presence of hyperuricemia is still poorly understood. Two important pathways determining hyperuricemia have been confirmed (renal and gut excretion of uric acid with glycolysis now firmly implicated). Major urate loci are SLC2A9 and ABCG2. Recent studies show that SLC2A9 is involved in renal and gut excretion of uric acid and is implicated in antioxidant defense. Although etiological variants at SLC2A9 are yet to be identified, it is clear that considerable genetic complexity exists at the SLC2A9 locus, with multiple statistically independent genetic variants and local epistatic interactions. The positions of implicated genetic variants within or near chromatin regions involved in transcriptional control suggest that this mechanism (rather than structural changes in SLC2A9) is important in regulating the activity of SLC2A9. ABCG2 is involved primarily in extra-renal uric acid under-excretion with the etiological variant influencing expression. At the other 26 loci, probable causal genes can be identified at three (PDZK1, SLC22A11, and INHBB) with strong candidates at a further 10 loci. Confirmation of the causal gene will require a combination of re-sequencing, trans-ancestral mapping, and correlation of genetic association data with expression data. As expected, the urate loci associate with gout, although inconsistent effect sizes for gout require investigation. Finally, there has been no genome-wide association study using clinically ascertained cases to investigate the causes of gout in the presence of hyperuricemia. In such a study, use of asymptomatic hyperurcemic controls would be expected to increase the ability to detect genetic associations with gout.

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132 The genetic basis is considerably simpler than that at SLC2A9, and the association signal is reported to be driven solely by the rs2231142 (Q141K) variant [36]. Login to comment
141 ABCG2 Q141K may also interact with extra-renal metabolic pathways to regulate serum urate (for example, via an influence on hepatic conversion of fructose to glucose) [39]. Login to comment
166 At ABCG2, the rs2231142 variant (Q141K), which is highly likely to be a causal variant at this locus [36], was associated with ABCG2 expression in the liver [4]. Login to comment

[hide] Endocytosis of ABCG2 drug transporter caused by bi... Biochim Biophys Acta. 2015 Aug;1853(8):1759-71. doi: 10.1016/j.bbamcr.2015.04.011. Epub 2015 Apr 24. Studzian M, Bartosz G, Pulaski L
Endocytosis of ABCG2 drug transporter caused by binding of 5D3 antibody: trafficking mechanisms and intracellular fate.
Biochim Biophys Acta. 2015 Aug;1853(8):1759-71. doi: 10.1016/j.bbamcr.2015.04.011. Epub 2015 Apr 24., [PMID:25918011]

Abstract [show]
ABCG2, a metabolite and xenobiotic transporter located at the plasma membrane (predominantly in barrier tissues and progenitor cells), undergoes a direct progressive endocytosis process from plasma membrane to intracellular compartments upon binding of 5D3 monoclonal antibody. This antibody is specific to an external epitope on the protein molecule and locks it in a discrete conformation within its activity cycle, presumably providing a structural trigger for the observed internalization phenomenon. Using routine and novel assays, we show that ABCG2 is endocytosed by a mixed mechanism: partially via a rapid, clathrin-dependent pathway and partially in a cholesterol-dependent, caveolin-independent manner. While the internalization process is entirely dynamin-dependent and converges initially at the early endosome, subsequent intracellular fate of ABCG2 is again twofold: endocytosis leads to only partial lysosomal degradation, while a significant fraction of the protein is retained in a post-endosomal compartment with the possibility of at least partial recycling back to the cell surface. This externally triggered, conformation-related trafficking pathway may serve as a general regulatory paradigm for membrane transporters, and its discovery was made possible thanks to consistent application of quantitative methods.

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14 Some sequence variants, e.g. the common human polymorphism Q141K, misdirects the molecule with partial folding defects to intracellular aggresomes [11]. Login to comment

[hide] Impact of ABCG2 polymorphisms on the clinical outc... Cancer Cell Int. 2015 Apr 19;15:43. doi: 10.1186/s12935-015-0191-3. eCollection 2015. Chen X, Chen D, Yang S, Ma R, Pan Y, Li X, Ma S
Impact of ABCG2 polymorphisms on the clinical outcome of TKIs therapy in Chinese advanced non-small-cell lung cancer patients.
Cancer Cell Int. 2015 Apr 19;15:43. doi: 10.1186/s12935-015-0191-3. eCollection 2015., [PMID:25960692]

Abstract [show]
OBJECTIVE: The primary purpose of this study was to investigate the correlation between single nucleotide polymorphisms (SNPs) of ATP binding cassette superfamily G member 2 (ABCG2) and outcome of tyrosine kinase inhibitions (TKIs) therapy in Chinese advanced non-small-cell lung cancer (NSCLC) patients. The secondary objective was to identify biomarkers to evaluate the response to treatment and outcome of the targeted therapy. METHODS: SNP genotyping (34 G/A, 421 C/A, 1143 C/T and -15622 C/T) of ABCG2 gene in 100 patients was performed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The clinical characteristics of 100 patients were collected. A total of 70 patients were treated with TKIs (gefitinib, erlotinib and icotinib). The association between ABCG2 polymorphisms and clinical characteristics was evaluated. Kaplan-Meier survival curves were plotted for overall survival (OS) and analyzed with the log-rank test. RESULTS: The three polymorphisms of the ABCG2 34 G/A, 421 C/A and 1143 C/T occurred more frequently compared with -15622 C/T in Chinese advanced NSCLC patients. There was no association between ABCG2 polymorphisms and clinical characteristics (p > 0.05). The median OS of patients with GG genotype at position 34 of the ABCG2 gene was significantly shorter than those with GA or AA genotype (p < 0.05). No significant difference of OS was found in 421 C/A and 1143 C/T polymorphisms (p > 0.05). CONCLUSION: ABCG2 34 G/A may be a possible predictor of the clinical outcome of TKIs therapy in NSCLC patients.

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254 Imai Y, Nakane M, Kage K, Tsukahara S, Ishikawa E, Tsuruo T, et al. C421A Polymorphism in the Human Breast Cancer Resistance Protein Gene Is Associated with Low Expression of Q141K Protein and Low-Level Drug Resistance 1 Supported in part by grants from the Ministry of Education, Culture, Sports, Science and Technology, the Ministry of Health, Labour and Welfare, Japan, and the Virtual Research Institute of Aging of Nippon Boehringer Ingelheim. Login to comment
258 Furukawa T, Wakabayashi K, Tamura A, Nakagawa H, Morishima Y, Osawa Y, et al. Major SNP (Q141K) variant of human ABC transporter ABCG2 undergoes lysosomal and proteasomal degradations. Login to comment

[hide] ABCG2: the molecular mechanisms of urate secretion... Am J Physiol Renal Physiol. 2015 Sep 15;309(6):F485-8. doi: 10.1152/ajprenal.00242.2015. Epub 2015 Jul 1. Woodward OM
ABCG2: the molecular mechanisms of urate secretion and gout.
Am J Physiol Renal Physiol. 2015 Sep 15;309(6):F485-8. doi: 10.1152/ajprenal.00242.2015. Epub 2015 Jul 1., [PMID:26136557]

Abstract [show]
The human propensity for high levels of serum uric acid (SUA) is a trait that has defied explanation. Is it beneficial? Is it pathogenic? Its role in the human diseases like gout and kidney stones was discovered over a century ago [Richette P, Bardin T. Lancet 375: 318-328, 2010; Rivard C, Thomas J, Lanaspa MA, Johnson RJ. Rheumatology (Oxford) 52: 421-426, 2013], but today emerging new genetic and epidemiological techniques have revived an age-old debate over whether high uric acid levels (hyperuricemia) independently increase risk for diseases like hypertension and chronic kidney disease [Feig DI. J Clin Hypertens (Greenwich) 14: 346-352, 2012; Feig DI, Madero M, Jalal DI, Sanchez-Lozada LG, Johnson RJ. J Pediatr 162: 896-902, 2013; Feig DI, Soletsky B, Johnson RJ. JAMA 300: 924-932, 2008; Wang J, Qin T, Chen J, Li Y, Wang L, Huang H, Li J. PLoS One 9: e114259, 2014; Zhu P, Liu Y, Han L, Xu G, Ran JM. PLoS One 9: e100801, 2014]. Part of the mystery of the role uric acid plays in human health stems from our lack of understanding of how humans regulate uric acid homeostasis, an understanding that could shed light on the historic role of uric acid in human adaptation and its present role in human pathogenesis. This review will highlight the recent work to identify the first important human uric acid secretory transporter, ABCG2, and the identification of a common causal ABCG2 variant, Q141K, for hyperuricemia and gout.

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12 This review will highlight the recent work to identify the first important human uric acid secretory transporter, ABCG2, and the identification of a common causal ABCG2 variant, Q141K, for hyperuricemia and gout. Login to comment
13 uric acid; kidney; ABCG2; Q141K; gout; hyperuricemia URIC ACID IS A TERMINAL METABOLITE of the purine metabolic pathway in humans and is only weakly soluble in water or blood. Login to comment
38 We found the Q141K variant had similar total and surface expression levels in the Xenopus oocytes but showed a 54% reduction in urate transport, marking Q141K as a loss-of-function mutation (17, 18, 30). Login to comment
39 A population-based study of 14,783 individuals supported rs2231142 (Q141K) as a causal variant for gout and increased SUA levels, marking the rs2231142 as a rare example in support of the common disease-common variant hypothesis (30). Login to comment
42 Matsuo et al. compared gouty and normal cohorts of Japanese males and found two ABCG2 mutations, Q141K (50% function) and Q126X (no function), and used these to correlate ABCG2 function with age of gout onset. Login to comment
50 Of the 28 loci, the ABCG2 loci (rs2231142/ Q141K mutation) resulted in the highest OR for gout risk (1.73) and contributed the largest increases in SUA (0.217 mg/dl) (12). Login to comment
51 Q141K Q141K / H155A 0.00 0.25 0.50 Q141K ABCG2 abundance relative to Wt Q141K ABCG2 H155A -- -- H155A GAPDH ABCG2 Wt ABCG2 Q141K H155 2 Q141K + H155A H155A ICL Q141K / Wt NBD B A C D ** Fig. 1. Login to comment
52 Q141K gout-causing mutation changes NBD structure in a model of ABCG2. Login to comment
53 A: model comparing Wt and Q141K NBDs as described in Woodward et al. (32) reveals a shift in an adjacent loop (black arrow, Q141K in blue, Wt in green). Login to comment
55 C: biochemical confirmation of the model shows the Q141K mutant expression levels can be rescued with the secondary H155A substitution. Login to comment
63 Recent work has focused on the molecular defect caused by the Q141K mutation of ABCG2 as both a potential therapeutic target and also as a model for understanding the basic structure/ function biology of ABC transporters. Login to comment
64 The Q141K mutation occurs in a residue of the nucleotide-binding domain, a position believed critical for interactions with the intracellular loops of the transmembrane portion of the protein. Login to comment
67 A comparative analysis between the Q141K ABCG2 mutant and the èc;F508 CFTR mutant reveals striking similarities. Login to comment
72 The Q141K ABCG2 mutant appears to only cause instability in the NBD domain. Login to comment
73 We recently demonstrated that artificially stabilizing the NBD domain of the Q141K mutant corrects the molecular defect. Login to comment
79 The specific source of the Q141K instability has remained unresolved. Login to comment
80 Recently, we have found that modeling the Q141K and Wt ABCG2 NBD domains (32) suggested a loop adjacent to the site of the Q141K substitution (see Fig. 1A) appears to be shifted outward, resulting from a clash between the mutant 141K residue and a histidine residue at the top of the loop. Login to comment
81 Replacing the histidine with an alanine appeared to resolve the shift and also significantly increased total Q141K (H155A) and mature, glycosylated protein abundance (Fig. 1, B-D) when expressed in HEK293 cells. Login to comment

[hide] Association of ABCB1 and FLT3 Polymorphisms with T... PLoS One. 2015 Aug 5;10(8):e0134102. doi: 10.1371/journal.pone.0134102. eCollection 2015. Chu YH, Li H, Tan HS, Koh V, Lai J, Phyo WM, Choudhury Y, Kanesvaran R, Chau NM, Toh CK, Ng QS, Tan PH, Chowbay B, Tan MH
Association of ABCB1 and FLT3 Polymorphisms with Toxicities and Survival in Asian Patients Receiving Sunitinib for Renal Cell Carcinoma.
PLoS One. 2015 Aug 5;10(8):e0134102. doi: 10.1371/journal.pone.0134102. eCollection 2015., [PMID:26244574]

Abstract [show]
Sunitinib is a tyrosine kinase inhibitor used as first-line treatment for metastatic renal cell carcinoma (mRCC). Asian ethnicity has been previously associated with lower clearance and greater toxicities for sunitinib treatment, relative to Caucasian ethnicity. Research focusing on identifying corresponding biomarkers of efficacy and toxicity has been hitherto conducted in Caucasian populations, and few of the reported associations have been externally validated. Our work thus aims to investigate candidate biomarkers in Asian patients receiving sunitinib, comparing the observed genotype effects with those reported in Caucasian populations. Using data from 97 Asian mRCC patients treated with sunitinib, we correlated 7 polymorphisms in FLT3, ABCB1, VEGFR2, ABCG2 and BIM with patient toxicities, response, and survival. We observed a stronger association of FLT3 738T genotype with leucopenia in our Asian dataset than that previously reported in Caucasian mRCC patients (odds ratio [OR]=8.0; P=0.03). We observed significant associations of FLT3 738T (OR=2.7), ABCB1 1236T (OR=0.3), ABCB1 3435T (OR=0.1), ABCB1 2677T (OR=0.4), ABCG2 421A (OR=0.3) alleles and ABCB1 3435, 1236, 2677 TTT haplotype (OR=0.1) on neutropenia. Primary resistance (OR=0.1, P=0.004) and inferior survival (progression-free: hazard ratio [HR]=5.5, P=0.001; overall: HR=5.0, P=0.005) were associated with the ABCB1 3435, 1236, 2677 TTT haplotype. In conclusion, ABCB1 and FLT3 polymorphisms may be helpful in predicting sunitinib toxicities, response and survival benefit in Asian mRCC patients. We have also validated the association between FLT3 738T and sunitinib-induced leucopenia previously reported in Caucasian populations, but have not validated other reported genetic associations.

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102 Genotype distribution Polymorphism rs number Variation na wt/wt wt/var var/var VAFe VEGFR2 1191 C/T rs2305948 V297I 94 64 30 0 0.160 (T) FLT3 738 T/C rs1933437 M227T 95 47 43 5 0.279 (C) ABCB1 1236 T/C rs1128503 G412G 93 35 49 9 0.360 (C) ABCB1 2677 G/TA rs2032582 A893S/T 96 25 44b 27c 0.375 (T); 0.135 (A) ABCB1 3435 C/T rs1045642 I1145I 96 34 50 12 0.385 (T) ABCG2 421 C/A rs2231142 Q141K 95 50 38 7 0.274 (A) BIM i2del d - i2del d 45 33 12 0 0.133 a Patients successfully genotyped. Login to comment

[hide] Concurrent effects of ABCB1 C3435T, ABCG2 C421A, a... Tumour Biol. 2015 Aug 7. Salimizand H, Amini S, Abdi M, Ghaderi B, Azadi NA
Concurrent effects of ABCB1 C3435T, ABCG2 C421A, and XRCC1 Arg194Trp genetic polymorphisms with risk of cancer, clinical output, and response to treatment with imatinib mesylate in patients with chronic myeloid leukemia.
Tumour Biol. 2015 Aug 7., [PMID:26250462]

Abstract [show]
There are a paucity and contradicted data about the impact of concurrent heredity of polymorphic genes and risk of chronic myeloid leukemia (CML). In the present study, the concurrent effects of three polymorphisms affecting the integrity of DNA consist of ABCB1 C3435T, ABCG2 C421A, and XRCC1 Arg194Trp on development of chronic myeloid leukemia were studied. Furthermore, the role of these polymorphisms in clinical and laboratory outcomes of patients was evaluated. In this case-control study, 70 CML patients and 140 healthy individuals were enrolled in the study. The clinical features of patients such as phase of disease and response to treatment and laboratory data before and after treatment with imatinib mesylate were collected. ABCB1 C3435T, ABCG2 C421A, and XRCC1 Arg194Trp single nucleotide polymorphisms were evaluated by restriction fragment length polymorphism-polymerase chain reaction. The T allele of ABCB1 C3435T, T allele of XRCC1 Arg194Trp, and C allele of ABCG2 C421A polymorphisms were significantly higher in patients than controls. TT genotype of ABCB1 and TT genotype of XRCC1 were associated with higher risk of chronic myeloid leukemia development. CC421 ABCG2/TT3435 ABCB1 and CC421 ABCG2/TT27157 XRCC1 were also correlated with a higher risk of CML. Patients with C allele of ABCB1 had poor cytogenetic response, and correlation of CC421 ABCG2/TT3435 ABCB1 diplotype with accelerated phase of CML was significant. Patients with CC421 ABCG2/TT3435 ABCB1 and CC421 ABCG2/TT27157 XRCC1 diplotypes might be at higher risk to rapid and severe development of CML and have weaker response to treatments with imatinib.

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29 Regarding ABCG2, one of the most important genetic variations is C421A (Q141K, rs2231142) which has been located on exon 5 and has a high association with BCRP activity [10-12]. Login to comment

[hide] Genetic variation in the ABCG2 gene is associated ... Clin Rheumatol. 2015 Oct 27. Jiri M, Zhang L, Lan B, He N, Feng T, Liu K, Jin T, Kang L
Genetic variation in the ABCG2 gene is associated with gout risk in the Chinese Han population.
Clin Rheumatol. 2015 Oct 27., [PMID:26506822]

Abstract [show]
Gout is a common type of arthritis that is characterized by hyperuricemia, tophi, and joint inflammation. Current evidence suggests that heredity contributes to the progression of gout. Previous studies have shown that regulation of the ATP-binding cassette subfamily G member 2 (ABCG2) pathways plays a role in gout occurrence. To investigate and validate potential genetic associations with the risk of gout, we conducted a case-control study. We conducted 143 cases and 310 controls and genotyped seven single-nucleotide polymorphisms (SNPs) in ABCG2 gene. ABCG2 SNP association analyses were performed using SPSS 17.0 Statistical Package, PLINK Software, HaploView software package, and SHEsis software platform. We identified that four susceptibility SNPs were potentially associated with occurrence of gout. Rs2622621 and rs3114018 in ABCG2 can actually increase the risk of gout in log-additive model (rs2622621, odds ratio (OR) = 1.90, 95 % confidence interval (CI) 1.39-2.61, p < 0.001; rs3114018, OR = 1.55, 95 % CI 1.13-2.13, p = 0.006). We found that rs17731799G/T-G/G and rs3114020 T/C-T/T in ABCG2 can actually increase the risk of gout in dominant model (rs17731799, OR = 1.67, 95 % CI 1.05-2.66, p = 0.028; rs3114020, OR = 1.58, 95 % CI 1.00-2.51, p = 0.048). The ABCG2 haplotype "GGCTCTC" (OR = 0.46, 95 % CI 0.28-0.75, p = 0.0019) decreased the gout risk. Our results, combined with those from previous studies, suggest that genetic variation in ABCG2 may influence gout susceptibility in the Han population.

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25 The most significant association at one of the loci was observed for a missense single-nucleotide polymorphism (SNP), rs2231142 (glutamine-to-lysine amino acid substitution (Q141K)) in the ABCG2 gene. Login to comment
74 ABCG2 is a high-capacity urate transporter which physiologically excretes urate Fig. 1 Haplotype block map for ABCG2 SNPs Table 3 ABCG2 haplotype frequencies and their association with gout (adjust by age and sex) Gene(s) Haplotype Frequency OR (95 % CI) p value ABCG2 GCCTAGT 0.2868 1 - GGCTCTC 0.2092 0.46 (0.28-0.75) 0.0019 AGACCTC 0.2014 0.71 (0.45-1.12) 0.14 GCCTCTC 0.0735 1.10 (0.57-2.13) 0.78 GGACCTC 0.0669 0.57 (0.27-1.18) 0.13 GCCTATC 0.0582 1.53 (0.75-3.14) 0.24 GGCTAGT 0.0378 0.37 (0.12-1.16) 0.09 GCCTAGC 0.0124 0.80 (0.19-3.46) 0.77 GCCTAGT 0.0538 0.47 (0.19-.13) 0.092 p value<0.05 indicates statistical significance OR odds ratio, CI confidence interval for the regulation of SUA. ABCG2 has the following two common dysfunctional variants: a non-sense variant Q126X and a missense variant Q141K [19, 22]. Login to comment

[hide] Genetic analysis of ABCG2 and SLC2A9 gene polymorp... Korean J Intern Med. 2015 Nov;30(6):913-20. doi: 10.3904/kjim.2015.30.6.913. Epub 2015 Oct 30. Kim YS, Kim Y, Park G, Kim SK, Choe JY, Park BL, Kim HS
Genetic analysis of ABCG2 and SLC2A9 gene polymorphisms in gouty arthritis in a Korean population.
Korean J Intern Med. 2015 Nov;30(6):913-20. doi: 10.3904/kjim.2015.30.6.913. Epub 2015 Oct 30., [PMID:26552468]

Abstract [show]
BACKGROUND/AIMS: Gout is a common inf lammatory arthritis triggered by the crystallization of uric acid in the joints. Serum uric acid levels are highly heritable, suggesting a strong genetic component. Independent studies to confirm the genetic associations with gout in various ethnic populations are warranted. We investigated the association of polymorphisms in the ABCG2 and SLC2A9 genes with gout in Korean patients and healthy individuals. METHODS: We consecutively enrolled 109 patients with gout and 102 healthy controls. The diagnosis of gout was based on the preliminary criteria of the America College of Rheumatology. Genomic DNA was extracted from whole blood samples. We identified single nucleotide polymorphism (SNP) changes in the ABCG2 and SLC2A9 genes using a direct sequencing technique. rs2231142 in ABCG2 and rs6449213 and rs16890979 in SLC2A9 and nearby regions were amplified by polymerase chain reaction. RESULTS: Patients with gout had significantly higher A/A genotype (29.3% vs. 4.9%, respectively) and A allele (52.8% vs. 26.5%, respectively) frequencies of rs2231142 in ABCG2 than did controls (chi(2) = 29.42, p < 0.001; odds ratio, 3.32; 95% confidence interval, 2.11 to 5.20). We found novel polymorphisms (c.881A>G and c.1002+78G>A) in the SLC2A9 gene. The univariate logistic regression analysis revealed that the c.881A>G and c.1002+78G>A SNPs were significantly higher in patients than in controls. CONCLUSIONS: We demonstrated a significant association between rs2231142 in the ABCG2 gene and gout and identified novel SNPs, c.881A>G and c.1002+78G>A, in the SLC2A9 gene that may be associated with gout in a Korean population.

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46 rs2231142, also known as Q141K and C421A, is an SNP in the ABCG2 gene and, thus, a missense variant. Login to comment
98 Furthermore, rs2231142, the missense SNP in ABCG2 (Q141K) has been associated with hyperuricemia and gout in Caucasian, African-American, Japanese, and Han Chinese samples [27-29]. Login to comment

[hide] ABCG2 in peptic ulcer: gene expression and mutatio... J Appl Genet. 2015 Nov 17. Salagacka-Kubiak A, Zebrowska M, Wosiak A, Balcerczak M, Mirowski M, Balcerczak E
ABCG2 in peptic ulcer: gene expression and mutation analysis.
J Appl Genet. 2015 Nov 17., [PMID:26578453]

Abstract [show]
The aim of this study was to evaluate the participation of polymorphism at position C421A and mRNA expression of the ABCG2 gene in the development of peptic ulcers, which is a very common and severe disease. ABCG2, encoded by the ABCG2 gene, has been found inter alia in the gastrointestinal tract, where it plays a protective role eliminating xenobiotics from cells into the extracellular environment. The materials for the study were biopsies of gastric mucosa taken during a routine endoscopy. For genotyping by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) at position C421A, DNA was isolated from 201 samples, while for the mRNA expression level by real-time PCR, RNA was isolated from 60 patients. The control group of healthy individuals consisted of 97 blood donors. The dominant genotype in the group of peptic ulcer patients and healthy individuals was homozygous CC. No statistically significant differences between healthy individuals and the whole group of peptic ulcer patients and, likewise, between the subgroups of peptic ulcer patients (infected and uninfected with Helicobacter pylori) were found. ABCG2 expression relative to GAPDH expression was found in 38 of the 60 gastric mucosa samples. The expression level of the gene varies greatly among cases. The statistically significant differences between the intensity (p = 0.0375) of H. pylori infection and ABCG2 gene expression have been shown. It was observed that the more intense the infection, the higher the level of ABCG2 expression.

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33 It is a non-synonymous polymorphism leading to a change of glutamine into lysine at position 141 in protein (Q141K) (Cusatis et al. 2006). Login to comment

[hide] The pharmacological impact of ATP-binding cassette... Acta Pharm Sin B. 2014 Apr;4(2):105-11. doi: 10.1016/j.apsb.2013.12.001. Epub 2014 Jan 13. Wu CP, V Ambudkar S
The pharmacological impact of ATP-binding cassette drug transporters on vemurafenib-based therapy.
Acta Pharm Sin B. 2014 Apr;4(2):105-11. doi: 10.1016/j.apsb.2013.12.001. Epub 2014 Jan 13., [PMID:26579371]

Abstract [show]
Melanoma is the most serious type of skin cancer and one of the most common cancers in the world. Advanced melanoma is often resistant to conventional therapies and has high potential for metastasis and low survival rates. Vemurafenib is a small molecule inhibitor of the BRAF serine-threonine kinase recently approved by the United States Food and Drug Administration to treat patients with metastatic and unresectable melanomas that carry an activating BRAF (V600E) mutation. Many clinical trials evaluating other therapeutic uses of vemurafenib are still ongoing. The ATP-binding cassette (ABC) transporters are membrane proteins with important physiological and pharmacological roles. Collectively, they transport and regulate levels of physiological substrates such as lipids, porphyrins and sterols. Some of them also remove xenobiotics and limit the oral bioavailability and distribution of many chemotherapeutics. The overexpression of three major ABC drug transporters is the most common mechanism for acquired resistance to anticancer drugs. In this review, we highlight some of the recent findings related to the effect of ABC drug transporters such as ABCB1 and ABCG2 on the oral bioavailability of vemurafenib, problems associated with treating melanoma brain metastases and the development of acquired resistance to vemurafenib in cancers harboring the BRAF (V600E) mutation.

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72 Recently, ABCG2 has been linked to the disease gout, as mutations (for example Q141K) in this transporter result in decreased efflux of urate from kidney epithelial cells54,55 . Login to comment

[hide] A meta-analysis of the associations between the Q1... Int J Clin Exp Pathol. 2015 Sep 1;8(9):9812-23. eCollection 2015. Li R, Miao L, Qin L, Xiang Y, Zhang X, Peng H, Mailamuguli, Sun Y, Yao H
A meta-analysis of the associations between the Q141K and Q126X ABCG2 gene variants and gout risk.
Int J Clin Exp Pathol. 2015 Sep 1;8(9):9812-23. eCollection 2015., [PMID:26617691]

Abstract [show]
BACKGROUND: Gout is an inflammatory disease in which genetic factors play a role. ABCG2 is a urate transporter, and the Q141K and Q126X variants of ABCG2 have been associated with a risk of developing gout, though previous studies of these associations have been inconsistent. Therefore, we conducted a meta-analysis to explore the relationship between these genetic variants and gout. METHODS: We examined 8 electronic literature databases. In total, 9 eligible articles on the associations between the Q141K (rs2231142) and Q126X (rs72552713) variants and gout risk, including 11 case-control studies were selected. We used odds ratios (OR) and 95% confidence intervals (CI) to assess the strength of these relationships in dominant, recessive, and co-dominant models. RESULTS: This study included 6652 participants (2499 gout patients and 4153 controls). The Q141K variant was found to significantly increase the risk of gout in Asians (dominant model: OR=2.64, 95% CI=2.04-3.43, P=0.02 for heterogeneity; recessive model: OR=3.19, 95% CI=2.56-3.97, P=0.28 for heterogeneity; co-dominant model: OR=1.37, 95% CI=1.18-1.59, P=0.09 for heterogeneity) and other populations (dominant model: OR=1.85, 95% CI=1.20-2.85, P<0.0001 for heterogeneity; recessive model: OR=3.78, 95% CI=2.28-6.27, P=0.19 for heterogeneity; co-dominant model: OR=1.48, 95% CI=1.26-1.74, P=0.19 for heterogeneity). The Q126X variant also significantly increased the risk of gout in Asians (dominant model: OR=3.87, 95% CI=2.07-7.24, P=0.06 for heterogeneity). CONCLUSIONS: These results suggest associations between the rs2231142 and rs72552713 ABCG2 gene polymorphisms and gout risk, which led to unfavorable outcomes. However, studies with larger sample sizes and homogeneous populations should be performed to confirm these results.

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3 ABCG2 is a urate transporter, and the Q141K and Q126X variants of ABCG2 have been associated with a risk of developing gout, though previous studies of these associations have been inconsistent. Login to comment
5 In total, 9 eligible articles on the associations between the Q141K (rs2231142) and Q126X (rs72552713) variants and gout risk, including 11 case-control studies were selected. Login to comment
8 The Q141K variant was found to significantly increase the risk of gout in Asians (dominant model: OR=2.64, 95% CI=2.04-3.43, P=0.02 for heterogeneity; recessive model: OR=3.19, 95% CI=2.56-3.97, P=0.28 for heterogeneity; co-dominant model: OR=1.37, 95% CI=1.18-1.59, P=0.09 for heterogeneity) and other populations (dominant model: OR=1.85, 95% CI=1.20-2.85, P<0.0001 for heterogeneity; recessive model: OR=3.78, 95% CI=2.28-6.27, P=0.19 for heterogeneity; co-dominant model: OR=1.48, 95% CI=1.26-1.74, P=0.19 for heterogeneity). Login to comment
12 Keywords: Gout, Q141K, Q126X, single nucleotide polymorphism, meta-analysis Introduction Gout is a recurrent relapsing inflammatory disease, caused by the precipitation of monosodium urate (MSU) crystals in the joints and soft tissues. Login to comment
26 Among these, Q141K and Q126X are the most commonly studied. Login to comment
27 SNP rs2231142, also referred to as C421A or Q141K in ABCG2, is located in exon 5 [10] and substitutes glutamic acid for diaminocaproic acid. Login to comment
33 The literature search was performed in English and Chinese using the following primary key words: gout, ABCG2, C421A, Q141K, rs2231142, C376T, Q126X, and rs72552713. Login to comment
37 Characteristics of the included studies (Q141K) First author (Ref.) Year Ethnicity/country Diagnostic standard Study design Sample size (case/control) HWE p-value Genotype distribution (case/control) Genotype frequency (case/control) CC CA AA CC (%) CA (%) AA (%) Wang Qiong [17] 2014 Asians China ACR preliminary diagnostic criteria for acute gout (1977) Case-control 185/311 Yes 64/157 86/126 35/28 34.6/50.5 46.5/40.5 18.9/9.0 Zhang Xin-Lei [28] 2014 Asians China ACR preliminary diagnostic criteria for acute gout (1977) Case-control 147/321 Yes 30/167 79/134 38/20 20.4/52.0 53.7/41.7 25.9/6.2 Li Fa-Gui [12] 2011 Asians China ACR preliminary diagnostic criteria for acute gout (1977) Case-control 200/235 Yes 64/103 91/112 45/20 32.0/43.8 45.5/47.7 22.5/8.5 Amanda 1 [25] 2010 Maori New Zealand ACR preliminary diagnostic criteria for acute gout (1977) Case-control 185/215 Yes 142/172 34/39 2/1 79.8/81.1 19.1/18.4 1.1/0.5 Amanda 2 [25] 2010 Pacific Islander New Zealand ACR preliminary diagnostic criteria for acute gout (1977) Case-control 173/109 Yes 58/69 78/36 37/4 33.5/63.3 45.1/33.0 21.4/3.7 Amanda 3 [25] 2010 Caucasian New Zealand ACR preliminary diagnostic criteria for acute gout (1977) Case-control 214/562 Yes 122/425 76/125 13/8 57.8/76.2 36.0/22.4 6.2/1.4 Danqiu Zhou [36] 2014 Asians China ACR preliminary diagnostic criteria for acute gout (1977) Case-control 352/350 Yes 87/167 181/150 84/33 24.7/47.7 51.4/42.9 23.9/9.4 Klaus [32] 2009 Caucasian German ACR preliminary diagnostic criteria for acute gout (1977) Case-control 677/1552 Yes 500/1241 168/299 9/12 73.9/80.0 24.8/19.2 1.3/0.8 Zhang Xiu-Juan [36] 2012 Asians China ARA diagnostic criteria for acute gout (1997) Case-control 110/236 Yes 35/120 55/96 20/20 31.8/50.8 50.0/40.7 18.2/8.5 You Yu-Quan [39] 2013 Asians China ACR preliminary diagnostic criteria for acute gout (1977) Case-control 154/160 Yes 48/98 78/49 28/13 31.2/60.3 50.6/30.6 18.2/8.1 Ye De-Shao [62] 2012 Asians China ACR preliminary diagnostic criteria for acute gout (1977) Case-control 102/102 Yes 23/53 42/40 37/9 22.5/52.0 41.2/39.2 36.3/8.8 Table 2. Login to comment
38 Characteristics of the included studies (Q126X) First author (Ref.) Year Ethnicity/ country Diagnostic standard Study design Sample size (case/control) HWE p-value Genotype distribution (case/control) Genotype frequency (case/control) CC CT TT CC (%) CT (%) TT (%) Zhang Xin-Lei [28] 2014 Asians China ACR preliminary diagnostic criteria for acute gout (1977) Case-control 136/321 Yes 127/320 9/1 0/0 93.4/99.7 6.6/0.3 0/0 Danqiu Zhou [36] 2014 Asians China ACR preliminary diagnostic criteria for acute gout (1977) Case-control 352/350 Yes 319/338 33/12 0/0 90.6/96.6 9.4/3.4 0/0 The following index terms were used: gout and ABCG2, gout and C421A, gout and Q141K, gout and rs2231142, gout and C376T, gout and Q126X, and gout and rs72552713. Login to comment
39 Selection criteria In this meta-analysis, we established the following inclusion criteria: (1) the publication Figure 2. Forest plot of the associations between the Q141K variant and gout risk using the A: Dominant model (CC compared to AC+AA), B: Recessive model (AA compared to CC+AC), and C: Co-dominant model (AC compared to CC+AA). Login to comment
40 Figure 3. Forest plot describing ethnicity in the dominant model (Q141K). Login to comment
48 Fortunately, the quality of the included studies was high, and we were able to collect the following data from each study: the first author`s name, publication year, country, ethnicity, diagnostic standard of gout, study design, total number of cases and controls, HWE P-value, genotype distribution and geno- Figure 4. Forest plot describing ethnicity in the recessive model (Q141K). Login to comment
53 Each relationship was assessed in a dominant model (Q141K: CC compared to AC+AA; Q126X: CC compared to CT+TT), a recessive model (Q141K: AA compared to CC+AC), and a co-dominant model (Q141K: AC compared to CC+AA). Login to comment
59 Figure 5. Forest plot describing ethnicity in the co-dominant model (Q141K). Login to comment
66 All of these 9 studies involved the Q141K variant, and only 2 studies referred to the Q126X variant. Login to comment
68 Of these 9 studies, of the Q141K variant, one study contained data for three different ethnicities; therefore, each of these studies was treated separately. Login to comment
77 Sensitivity analysis for the association between the Q141K variant and gout risk. Login to comment
79 Begg`s funnel plot examining the publication bias of studies using the recessive model (Q141K). Login to comment
87 Among the ABCG2 polymorphisms, Q141K and Q126X are the most commonly studied. Login to comment
88 Nevertheless, the role of the Q141K variant in gout risk and the potential for the Q126X variant to increase gout risk are controversial. Login to comment
90 Hirotaka et al. [25], Kazumasa et al. [11] and Xiaofei Lv [26] also showed that the Q141K variant increases the risk of gout. Login to comment
95 Our results suggested that the Q141K variant results in increased gout risk in dominant, recessive, and co-dominant models, and in subgroup analyses, Q126X also increased gout risk in a dominant model. Login to comment
101 Ethnic differences are always mentioned by researchers not only in discussions of gout [29] but also in the association between the Q141K polymorphism and gout risk [24, 30]. Login to comment
103 From this subgroup analysis, we found that regardless of race, the Q141K variant increases the risk of gout. Login to comment
118 Our evidence revealed that Q141K and Q126X are risk factors for the development of gout. Login to comment
182 Association of functional polymorphism rs2231142 (Q141K) in the ABCG2 gene with serum uric acid and gout in 4 US populations. Login to comment
195 Association between ABCG2 Q141K polymorphism and gout risk affected by ethnicity and gender: a systematic review and meta-analysis. Login to comment