ABCMdb

ABCC1 p.Arg433Ser

Predicted by SNAP2:	A: D (91%), C: D (85%), D: D (95%), E: D (95%), F: D (95%), G: D (95%), H: D (85%), I: D (91%), K: D (71%), L: D (91%), M: D (91%), N: D (91%), P: D (95%), Q: D (85%), S: D (91%), T: D (91%), V: D (91%), W: D (95%), Y: D (91%),
Predicted by PROVEAN:	A: D, C: D, D: D, E: D, F: D, G: D, H: D, I: D, K: N, L: D, M: D, N: D, P: D, Q: D, S: D, T: D, V: D, W: D, Y: D,

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[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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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

[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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103 Another report showed that a naturally occurring mutation of R433S in MRP1 caused decreased organic anion transport and J. Cervenak et al. / Cancer Letters 234 (2006) 62-72 65 increased doxorubicin resistance [73]. 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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852 Table 5 Frequency of ABCC1 genetic variants in different populations, position on DNA, putative effect, and frequencies (according to Le Saux et al., 2000; Ito et al., 2001; Moriya et al., 2002; Conrad et al., 2002; Oselin et al., 2003b; Wang et al., 2004) Position/ Nucleotide Aminoacid or effect Orientals Caucasians Function 128G>C C43S 0.01 - elevateda 218C>T T73I 0.00-0.04 - 257C>T S92F 0.00 0.00 decreaseda 350C>T T117M - 0.02 (decreased)a 689G>A R230N 0.00 0.00 (decreased)a 816G>A synonymous - 0.04 825T>C synonymous - 0.30 1057G>A V353M 0.00 0.005 elevateda 1299G>T R433S - 0.01 elevated Vmax of doxorubicin, decreased transport of LTC4 a,b 1684T>C synonymous - 0.80 1898G>A R633Q - 0.01 (decreased)a 2012G>T G671V - 0.03 doxorubicine-induced cardiomyopathyc 2168G>A R723Q 0.01-0.07 - decreaseda 2965G>A A989T 0.00 0.005 (decreased)a 3140G>C C1047S 0.00 0.00 3173G>A R1058Q 0.01 - 4002G>A synonymous - 0.28 4535C>T S1512L - 0.03 decreaseda a Letourneau et al. (2005). 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

[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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351 The G1299T polymorphism (exon 10) results in the change R433S in a cytoplasmic loop of MRP1, and was observed to increase doxorubicin resistance, but decrease transport of several organic anions [167]. 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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73 In contrast, the decrease in LTC4 and estrone-3-sulfate transport due to the Arg433Ser substitution appeared to result in a lower Vmax (Conrad et al., 2002) rather than changes in Km. Login to comment
74 Interestingly, the same Arg433Ser variant was associated with increased doxorubicin resistance in transfected cells, demonstrating that a single amino acid substitution can result in markedly different substrate-dependent functional consequences. Login to comment
81 MRP1 (ABCC1) NH2 NBD NBD in out Membrane Cys43Ser Ser92Phe Thr117Met Arg230Gln Val353Met Arg633Gln Gly671Val Arg723Gln Arg433Ser Ala989Thr Cys1047Ser Val1146Ile Arg1058Gln Thr1401Met Ser1512Leu Thr73Ile COOH NBD NBD COOH NBD COOH NBD NBD Table1MRP1(ABCC1)singlenucleotidepolymorphisms.Location,allelefrequencyandfunctionaleffects. Positionin codingsequence Aminoacid exchangeLocation Allelefrequency EffectNCBIIDReferenceAfCaJpothers 128G>CCys43SerExon2--1[1]-Decreaseinvincristineresistance[2]rs41395947 Disruptedplasmamembranetraffickingin transfectedcells[2] 218C>TThr73IleExon2--1[1]3.7Chinese[3]Noinfluenceonexpressionandtransportin membranevesicles[4] rs41494447 257C>TSer92PheExon30a 0a 0a 0Chinese[3]Noinfluenceonexpressionandtransportin membranevesicles[4] 350C>TThr117MetExon3-100[5]--Noinfluenceonexpressionandtransportin membranevesicles[4] 689G>AArg230GlnExon70a 0a 0a 0Chinese[3]Noinfluenceonexpressionandtransportin membranevesicles[4] 1057G>AVal353MetExon90a 0.5a 0a -- 1299G>TArg433SerExon10-1.4[6]--Changesintransportandresistance[7] 1898G>AArg633GlnExon13-[8]--Noinfluenceonexpressionandtransportin membranevesicles[4] 2012G>TGly671ValExon16-2.8[6]--Noinfluenceonexpressionandtransportin membranevesicles[6] Associatedwithanthracycline-induced cardiotoxicity[9] 2168G>AArg723GlnExon17--7.3[1]5.6Chinese[3]Noinfluenceonexpressionandtransportin membranevesicles[4]noinfluenceonmRNA expressioninenterocytes(n=1)[10] rs4148356 2965G>AAla989ThrExon220a 0.5a 0a -Noinfluenceonexpressionandtransportin membranevesicles(non-significantreduction inE17βGtransport)[4] 323 3140G>CCys1047SerExon234.5a 0a 0a -Noinfluenceonexpressionandtransportin membranevesicles[4] rs13337489 3173G>AArg1058GlnExon23--1[1]-Noinfluenceonexpressionandtransportin membranevesicles[4] rs41410450 3436G>AVal1146IleExon24-----rs28706727 4102C>TThr1401MetExon29-----rs8057331 4535C>TSer1512LeuExon31-[5]--Noinfluenceonexpressionandtransportin membranevesicles[4] ReferencewithoutfrequencymeansthatSNPwasdetectedbutnofrequencydetermined. 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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155 ABCC2 (Multidrug Resistance-Associated Protein 2) Table 6.5 Frequency of ABCC1 genetic variants in different populations, position on DNA, putative effect, and frequencies (according to (33, 77-80, 136)) Position Amino acid or effect Orientals Caucasians Function c.128G>C C43S 0.01 - Elevateda c. 218C>T T73I 0.00-0.04 - c. 257C>T S92F 0.00 0.00 Decreaseda c. 350C>T T117M - 0.02 (Decreased)a c. 689G>A R230N 0.00 0.00 (Decreased)a c. 816G>A Synonymous - 0.04 c. 825T>C Synonymous - 0.30 c. 1057G>A V353M 0.00 0.005 Elevateda c. 1299G>T R433S - 0.01 Elevated vmax of doxorubicin, decreased transport of LTC4 a,b c. 1684T>C Synonymous - 0.80 c. 1898G>A R633Q - 0.01 (Decreased)a c. 2012G>T G671V - 0.03 Doxorubicine-induced cardiomyopathyc c. 2168G>A R723Q 0.01-0.07 - Decreaseda c. 2965G>A A989T 0.00 0.005 (Decreased)a c. 3140G>C C1047S 0.00 0.00 c. 3173G>A R1058Q 0.01 - c. 4002G>A Synonymous - 0.28 c. 4535C>T S1512L - 0.03 Decreaseda References: a [81], b [77], c [84] an inducible expression of ABCC2, which contributes also to the phenomenon of drug resistance. 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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67 Those studied include C43S, T73I, S92F, T117M, R230Q, V353M, R433S, R633Q, G671V, R723Q, A989T, C1047S, R1058Q, A1337T, and S1512L. Login to comment
69 However, it has been noted that C43S, R433S, and A989T result in decreased ABCB1 function [43]. Login to comment

[hide] Identification of human multidrug resistance prote... J Hum Genet. 2001;46(11):656-63. Conrad S, Kauffmann HM, Ito K, Deeley RG, Cole SP, Schrenk D
Identification of human multidrug resistance protein 1 (MRP1) mutations and characterization of a G671V substitution.
J Hum Genet. 2001;46(11):656-63., [PMID:11721885]

Abstract [show]
The multidrug resistance protein 1 (MRP1) belonging to the ATP-binding cassette (ABC) superfamily of transport proteins can confer resistance to multiple natural product drugs and methotrexate in human tumor cells. In addition, MRP1 is expressed in normal tissues acting as an efflux pump for glutathione, glucuronate, and sulfate conjugates and may thus influence the pharmacokinetic properties of many drugs. Using polymerase chain reaction-single-strand conformation polymorphism analysis, we screened 36 Caucasian volunteers for mutations in the coding exons of the MRP1 gene, including the adjacent intron sequences. Among several mutations found, two are expected to cause amino acid substitutions. One of these mutations (G671V) was of special interest because it is located near the first nucleotide binding domain. To determine whether this mutation caused a change in the MRP1 phenotype, a mutant MRP1 expression vector was constructed and transfected into SV40-transformed human embryonic kidney cells (HEKSV293T) and the transport properties of the mutant protein were examined. Transport of the MRP1 substrates leukotriene C4, 17beta-estradiol 17beta-(D)-glucuronide, and estrone sulfate by membrane vesicles prepared from transiently transfected HEKSV293T cells was comparable to that of wild-type MRP1.

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78 Mutations in the MRP1 gene and median mRNA expression levels of samples with different MRP1 genotypes Position Location SNPs Change FRE EXPa Intron 7 809ϩ54 C/A Intronic 2.8 101 Exon 8b 825 T/C Silent 2.8 51 Intron 8 1040ϩ13 T/C Intronic 16.7 95 Exon 10 1299 G/T Arg433Ser 1.4 156 Intron 11b 1474-48 C/T Intronic 2.8 146 Intron 11 1474-8 T/C Intronic 9.7 72 Intron 12 1678-9 DEL T Intronic 11.1 106 Exon 13b 1684 T/C Silent 8.3 106 Exon 13 1704 C/T Silent 2.8 59 Exon 16 2012 G/T Gly671Val 2.8 43 Intron 20 2736-36 T/C Intronic 2.8 132 Intron 20 2736-18 CC/TT Intronic 2.8 132 Intron 20 2736-6 T/C Intronic 2.8 132 Intron 21 2871ϩ17 G/A Intronic 1.4 132 Intron 26 3819ϩ7 G/A Intronic 1.4 124 Exon 28b 4002 G/A Silent 2.8 95 Intron 30 4487ϩ5 A/T Intronic 2.8 137 Intron 30 4487ϩ18 A/G Intronic 1.4 142 Intron 30 4487ϩ28 C/G Intronic 2.8 137 Densitometrical analysis was performed with TINA software (Raytest, Straubenhardt, Germany), average relative MRP1 expression was set as 100% FRE, allelic frequency in %; EXP, expression level; SNP, single-nucleotide polymorphism a The average expression level of all samples was 100 Ϯ 55 b Recently described by Ito et al. 2001b Fig. 2. Login to comment
118 They are located in the second transmembrane spanning domain (R433S) and in the vicinity of the first ATP-binding site (G671V). Login to comment

[hide] A naturally occurring mutation in MRP1 results in ... Pharmacogenetics. 2002 Jun;12(4):321-30. Conrad S, Kauffmann HM, Ito K, Leslie EM, Deeley RG, Schrenk D, Cole SP
A naturally occurring mutation in MRP1 results in a selective decrease in organic anion transport and in increased doxorubicin resistance.
Pharmacogenetics. 2002 Jun;12(4):321-30., [PMID:12042670]

Abstract [show]
The human 190 kDa multidrug resistance protein, MRP1, is a polytopic membrane glycoprotein that confers resistance to a wide range of chemotherapeutic agents. It also transports structurally diverse conjugated organic anions, as well as certain unconjugated and conjugated compounds, in a reduced glutathione-stimulated manner. In this study, we characterized a low-frequency (<1%) naturally occurring mutation in MRP1 expected to cause the substitution of a conserved arginine with serine at position 433 in a predicted cytoplasmic loop of the protein. Transport experiments with membrane vesicles prepared from transfected human embryonic kidney cells and HeLa cells revealed a two-fold reduction in the ATP-dependent transport of the MRP1 substrates, leukotriene C4 (LTC4) and oestrone sulphate. Kinetic analysis showed that this reduction was due to a decrease in Vmax for both substrates but Km was unchanged. In contrast, 17beta-oestradiol-17beta-(D-glucuronide) transport by the Arg433Ser mutant MRP1 was similar to that by wild-type MRP1. Fluorescence confocal microscopy showed that the mutant MRP1 was routed correctly to the plasma membrane. In contrast to the reduced LTC4 and oestrone sulphate transport, stably transfected HeLa cells expressing Arg433Ser mutant MRP1 were 2.1-fold more resistant to doxorubicin than cells expressing wild-type MRP1, while resistance to VP-16 and vincristine was unchanged. These results provide the first example of a naturally occurring mutation predicted to result in an amino acid substitution in a cytoplasmic region of MRP1 that shows an altered phenotype with respect to both conjugated organic anion transport and drug resistance.

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2 In this study, we characterized a low-frequency (<1%) naturally occurring mutation in MRP1 expected to cause the substitution of a conserved arginine with serine at position 433 in a predicted cytoplasmic loop of the protein. Login to comment
5 In contrast, 17â- oestradiol-17â-(D-glucuronide) transport by the Arg433 Ser mutant MRP1 was similar to that by wild-type MRP1. Login to comment
7 In contrast to the reduced LTC4 and oestrone sulphate transport, stably transfected HeLa cells expressing Arg433 Ser mutant MRP1 were 2.1-fold more resistant to doxorubicin than cells expressing wild-type MRP1, while resistance to VP-16 and vincristine was unchanged. Login to comment
31 In addition, we have recreated the Arg433 Ser substitution in a MRP1 mammalian expression vector and analysed its transport and drug resistance properties after transfection into human cells. Login to comment
38 Site-directed mutagenesis The mutation Arg433 Ser was generated using the following sense primer 59P-GTGGACGCTCAGTC GTTCATGGACTTGGCC-39 (substituted nucleotides are underlined) with the USE Mutagenesis Kit (Amersham Pharmacia Biotech, PQ, Canada), according to the manufacturer`s instructions. Login to comment
79 Results A heterozygous Arg433 Ser substitution in MRP1 does not result in any overt clinical symptoms A mutation in exon 10 of the human MRP1 gene expected to result in substitution of Arg433 to Ser was found in a single individual among 91 healthy Caucasian volunteers. Login to comment
81 An Arg433 Ser substitution causes a reduction in MRP1-mediated LTC4 and oestrone sulphate transport activity, but leaves E217âG transport activity intact To determine whether MRP1-mediated transport of [3 H]LTC4, [3 H]oestrone sulphate and [3 H]E217âG was affected by a Arg433 Ser substitution, this mutation was generated in a mammalian MRP1 expression vector and transiently transfected into HEKSV293T cells. Login to comment
82 Stably transfected clonal HeLa cell lines expressing the Arg433 Ser MRP1 mutant were also generated in order to confirm the results obtained with the transient transfectants and to be able to test drug resistance in longer term assays. Login to comment
85 In contrast, levels of [3 H]E217âG uptake by the Arg433 Ser mutant MRP1 and wild-type MRP1 were similar and proportional to their relative protein expression levels (Fig. 2). Login to comment
87 Arg433 Ser mutant MRP1 is correctly routed to the plasma membrane To determine whether the decrease in LTC4 and oestrone sulphate transport caused by the Arg433 Ser substitution might be associated with a change in the trafficking of MRP1 to the plasma membrane, the subcellular localization of the mutant and wild-type MRP1 was determined by indirect immunofluorescent confocal microscopy. Login to comment
88 As shown in Fig. 4, both the wild-type and mutant proteins expressed in HeLa cells showed a pattern of strong plasma membrane staining, indicating that MRP1 trafficking in the transfected cells was unaffected by the Arg433 Ser substitution. Login to comment
89 Kinetic analysis of ATP-dependent [3 H]LTC4 and [3 H]oestrone sulphate transport by Arg433 Ser mutant MRP1 Since the initial time courses of LTC4 and oestrone sulphate uptake indicated a two-fold reduction in transport activity of the Arg433 Ser mutant MRP1, the effect of this mutation on the transport kinetics of these substrates was examined. Login to comment
91 A non-linear regression analysis of the data obtained showed small but not significant differences in the apparent Km values of the wild-type and Arg433 Ser mutant proteins for LTC4 and oestrone sulphate. Login to comment
92 In contrast, the apparent Vmax values of the Arg433 Ser mutant for both substrates were approximately two-fold lower than for wild-type MRP1 when normalized for relative levels of protein expression. Login to comment
93 Drug resistance profile of cells expressing Arg433 Ser MRP1 In addition to its ability to transport conjugated organic anions, MRP1 can also confer resistance to a variety of anticancer drugs by reducing intracellular drug accumulation [21,22]. Login to comment
94 To determine whether the Arg433 Ser substitution affected the drug resistance properties of MRP1, the drug resistance profile of Arg433 Ser MRP1 Fig. 1 Expression and ATP-dependent [3 H]LTC4 transport activity of wild-type and mutant Arg433 .Ser MRP1 in membrane vesicles prepared from transfected cells. Login to comment
95 (a) MRP1 expression levels in membrane vesicles of human embryonic kidney cells transfected with pcDNA3.1(-) as control, pcDNA3.1(-)wt-MRP1K (wtMRP1) and pcDNA3.1(-) MRP1K-Arg433 Ser (R433S) cDNA expression vectors were determined by dot blot analysis with MAb QCRL-1. Login to comment
97 (b) Time course of [3 H]LTC4 uptake in membrane vesicles shown in (a) prepared from HEKSV293T cells transfected with wild-type MRP1 (wtMRP1, j), mutant (R433S, m) and control (pcDNA3.1(-), h) cDNA expression vectors. Results shown are means (Æ SD) of triplicate determinations in a single experiment. Similar results were obtained in two additional independent experiments with vesicles from transfected HEKSV293T cells and in two additional experiments with vesicles from transfected HeLa cells. Login to comment
99 The Arg433 Ser mutation increased resistance to the cationic anthracycline agent, doxorubicin by approximately two-fold (P , 0.01) relative to wild-type MRP1 (Fig. 6a). Login to comment
100 In contrast, the sensitivity of the Arg433 Ser mutant to the Vinca alkaloid, vincristine, and the electroneutral epipodophyllotoxin, VP-16 (Fig. 6b,c) was comparable to that conferred by wild-type MRP1. Login to comment
108 Thus, it seems likely that this transport protein contains more than one Fig. 2 Expression and ATP-dependent [3 H]E217âG transport activity of wild-type and mutant Arg433 Ser MRP1 in membrane vesicles prepared from transfected cells. Login to comment
110 (b) Time course of [3 H]E217âG uptake in membrane vesicles shown in (a) (wtMRP1, j; mutant R433S, m; control pcDNA3.1(-), h). Login to comment
114 (b) Time course of [3 H]oestrone sulphate uptake in the presence of 1 mmol GSH in the membrane vesicles shown in (a) prepared from cells transfected with wild-type MRP1 (wtMRP1, j), mutant (R433S, m) and empty vector control (pcDNA3.1(-), h) cDNA expression vectors. Results shown are means (Æ SD) of triplicate determinations in a single experiment. Similar results were obtained in three additional independent experiments with vesicles from transfected HeLa cells and in two experiments with vesicles from transfected HEKSV293T cells. Login to comment
118 Of all the mutations detected, two are predicted to cause amino acid substitutions in the protein, Gly671 Val and Arg433 Ser. Login to comment
121 According to most topological models of MRP1, the second mutation, an Arg433 Ser substitution, is located close to the COOH-proximal end of the cytoplasmic loop connecting the predicted transmembrane (TM)7 to TM8 in the second membrane spanning domain (MSD2) of MRP1 [41-43]. Login to comment
123 When the properties of a recombinant Arg433 Ser MRP1 mutant expressed in transfected human cells were examined, a two-fold reduction in LTC4 and GSH-stimulated oestrone sulphate transport activity was observed even though the expression levels and membrane localization of the mutant MRP1 and wild-type MRP1 were comparable. Login to comment
125 Thus, the Vmax values for LTC4 and GSH-stimulated oestrone sulphate transport by the Arg433 Ser mutant MRP1 were approximately two-fold lower than Fig. 4 Confocal microscopy of transfected HeLa cells expressing wild-type and Arg433 Ser mutant MRP1 proteins. Login to comment
130 (b) Transfected HeLa cells expressing Arg433 Ser mutant MRP1. Login to comment
132 (a) ATP-dependent uptake of [3 H]LTC4 by the membrane vesicles shown in Fig. 1(a) was measured at various LTC4 concentrations (0.01-1 ìM) for 45 s at 23 8C [wild-type MRP1 (wtMRP1, j; and Arg433 Ser mutant MRP1 (R433S, m)]. Login to comment
135 (b) ATP-dependent uptake of [3 H]oestrone sulphate by the membrane vesicles shown in Fig. 3(a) [wild-type MRP1 (wtMRP1, j) and Arg433 Ser mutant MRP1 (R433S, m)]. Login to comment
141 In contrast, the levels of E217âG uptake by the Arg433 Ser mutant MRP1 were unchanged. Login to comment
155 HeLa cells expressing the Arg433 Ser mutant protein were observed to be two-fold more resistant to the anthracycline doxorubicin in addition to displaying a two-fold reduction in LTC4 and GSH-stimulated oestrone sulphate transport activity. Enhanced resistance was specific for this anthracycline as the resistance of cells expressing the Arg433 Ser mutant to the Vinca alkaloid, vincristine, and to the epipodophyllotoxin, VP-16, remained comparable to cells expressing wild-type MRP1. Login to comment
160 Furthermore, TM segments corresponding to those identified in P-glycoprotein as being important for substrate binding and/or transport have also been Table 1 Kinetic parameters of [3 H]LTC4 and GSH-stimulated [3 H]oestrone sulphate uptake by membrane vesicles from cells transfected with expression vectors encoding wild-type and Arg433 Ser mutant MRP1 Km (ìM) Vmax (pmol/mg/min) Normalized Vmax (pmol/mg/min) Substrate wtMRP1 R433S wtMRP1 R433S wtMRP1 R433S LTC4 0.18 Æ 0.04 0.14 Æ 0.06 121 Æ 8 71 Æ 10 121 Æ 8 59 Æ 8 Oestrone sulphate 0.89 Æ 0.07 1.10 Æ 0.34 72 Æ 2 41 Æ 4 72 Æ 2 34 Æ 3 Apparent Km and Vmax values for LTC4 and GSH-stimulated oestrone sulphate uptake were determined as described in Materials and Methods and the legend to Fig. 5. Login to comment
169 Thus, our data, demonstrating that the doxorubicin resistance of cells expressing the Arg433 Ser mutation is increased while vincristine and VP-16 resistance are unaffected, indicate that amino acid changes in this cytoplasmic loop of MRP1, similar to the comparable loop in P-glycoprotein, can selectively affect the drug resistance profile conferred by this transporter. Login to comment
171 Cells stably transfected with wild-type MRP1 (wtMRP1, j), mutant (R433S, m) and control (pcDNA3.1(-), h) cDNA expression vectors were exposed to (a) doxorubicin, (b) vincristine and (c) etoposide at the concentrations indicated for 72 h at 37 8C. Login to comment
174 100 75 50 25 0.1 1 10 100 Vincristine (nM) %Controlabsorbance 100 75 50 25 1 10 100 Etoposide (µM) %Controlabsorbance Table 2 Relative drug resistance of HeLa cells expressing wild-type and Arg433 Ser mutant MRP1 Relative resistancea Drug wtMRP1 Arg433 Ser MRP1 Ratio Doxorubicin 2.5 Æ 0.6 (n ¼ 5) 5.3 Æ 1.0 (n ¼ 8)b 2.1 Vincristine 7.0 Æ 4.1 (n ¼ 5) 10.3 Æ 5.0 (n ¼ 10) 1.5 VP-16 3.4 Æ 1.4 (n ¼ 6) 3.8 Æ 1.6 (n ¼ 6) 1.1 a The resistance of stably transfected HeLa cells was determined using a tetrazolium-based microtitre plate assay. The relative resistance factors were obtained by dividing the IC50 values for wild-type or Arg433 Ser mutant MRP1 transfected cells by the IC50 values for empty vector control transfected cells and are normalized for differences in protein expression levels as appropriate. Login to comment
177 Thus, the Arg433 Ser substitution results in an additional potential phosphorylation site which, if utilized, might in some direct or indirect way influence the activity of the protein. Login to comment
179 The MRP1 mutation predicted to cause the Arg433 Ser substitution described here was present in a cohort of 91 healthy Caucasian volunteers with an estimated allelic frequency of ,1%. Login to comment
181 However, the mutation was heterozygous in the case found and the alterations in Arg433 Ser MRP1 transport activity observed in the current study suggest that homozygous carriers should be examined before ruling out possible physiological consequences associated with this mutation in vivo. Login to comment
182 Thus, the two-fold decrease in LTC4 and GSH-stimulated oestrone sulphate transport by the Arg433 Ser mutant MRP1 in vitro raises the possibility that individuals bearing this polymorphism might be subject to a variety of biological effects, such as an impaired response to an inflammatory stimulus [30-32]. Login to comment

[hide] Charged amino acids in the sixth transmembrane hel... J Biol Chem. 2002 Nov 1;277(44):41326-33. Epub 2002 Aug 18. Haimeur A, Deeley RG, Cole SP
Charged amino acids in the sixth transmembrane helix of multidrug resistance protein 1 (MRP1/ABCC1) are critical determinants of transport activity.
J Biol Chem. 2002 Nov 1;277(44):41326-33. Epub 2002 Aug 18., 2002-11-01 [PMID:12186871]

Abstract [show]
The multidrug resistance protein, MRP1 (ABCC1), is an ATP-binding cassette transporter that confers resistance to chemotherapeutic agents. MRP1 also mediates transport of organic anions such as leukotriene C(4) (LTC(4)), 17beta-estradiol 17-(beta-d-glucuronide) (E(2)17betaG), estrone 3-sulfate, methotrexate (MTX), and GSH. We replaced three charged amino acids, Lys(332), His(335), and Asp(336), predicted to be in the sixth transmembrane (TM6) helix of MRP1 with neutral and oppositely charged amino acids and determined the effect on substrate specificity and transport activity. All mutants were expressed in transfected human embryonic kidney cells at levels comparable with wild-type MRP1, and confocal microscopy showed that they were correctly routed to the plasma membrane. Vesicular transport studies revealed that the MRP1-Lys(332) mutants had lost the ability to transport LTC(4), and GSH transport was reduced; whereas E(2)17betaG, estrone 3-sulfate, and MTX transport were unaffected. E(2)17betaG transport was not inhibited by LTC(4) and could not be photolabeled with [(3)H]LTC(4), indicating that the MRP1-Lys(332) mutants no longer bound this substrate. Substitutions of MRP1-His(335) also selectively diminished LTC(4) transport and photolabeling but to a lesser extent. Kinetic analyses showed that V(max) (LTC(4)) of these mutants was decreased but K(m) was unchanged. In contrast to the selective loss of LTC(4) transport in the Lys(332) and His(335) mutants, the MRP1-Asp(336) mutants no longer transported LTC(4), E(2)17betaG, estrone 3-sulfate, or GSH, and transport of MTX was reduced by >50%. Lys(332), His(335), and Asp(336) of TM6 are predicted to be in the outer leaflet of the membrane and are all capable of forming intrahelical and interhelical ion pairs and hydrogen bonds. The importance of Lys(332) and His(335) in determining substrate specificity and of Asp(336) in overall transport activity suggests that such interactions are critical for the binding and transport of LTC(4) and other substrates of MRP1.

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238 We previously showed that the naturally occurring mutation of MRP1-Arg433 to Ser predicted to be located in the fourth cytoplasmic loop in close proximity to the membrane interface of TM8 results in a 2-fold decrease in LTC4 and estrone 3-sulfate transport efficiency. 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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285 Thus, substitution of Arg433 with Ser in an ICL predicted to be close to TM8 of MRP1 caused by the low frequency (<1%) G1299T MRP1 polymorphism in exon 10 results in a substrate selective alteration in organic anion transport activity and drug resistance when the mutant protein is expressed in stably transfected HeLa cells [190]. Login to comment
286 E217βG transport was unaffected by the Arg433Ser mutation A fourth mutant phenotype is that where mutants exhibit a substrate-selective loss or gain of transport activity. Login to comment
290 In contrast, cells expressing the mutant MRP1-Arg433Ser were 2.5-fold more resistant to doxorubicin while resistance to VP-16 and vincristine remained comparable to cells expressing wild-type MRP1. Login to comment

[hide] Polymorphisms of MRP1 (ABCC1) and related ATP-depe... Pharmacogenet Genomics. 2005 Aug;15(8):523-33. Conseil G, Deeley RG, Cole SP
Polymorphisms of MRP1 (ABCC1) and related ATP-dependent drug transporters.
Pharmacogenet Genomics. 2005 Aug;15(8):523-33., [PMID:16006996]

Abstract [show]
Genetic variations in drug metabolizing enzymes and targets are established determinants of adverse drug reactions and interactions, but less is known about the role of genetic polymorphisms in membrane transport proteins. MRP1 (ABCC1) is one of 13 polytopic membrane proteins that comprise the 'C' subfamily of the ATP-binding cassette (ABC) superfamily of transport proteins. MRP1 and related ABCC family members, including MRP2, 3, 4 and 5 (ABCC2, 3, 4 and 5), each have a distinctive pattern of tissue expression and substrate specificity. Together, these five transporters play important roles in the disposition and elimination of drugs and other organic anions, and in maintenance of blood-tissue barriers, as confirmed by enhanced chemosensitivity of respective knockout mice. Moreover, Mrp2 (Abcc2) deficient animals display mild conjugated hyperbilirubinemia, corresponding to a human condition known as Dubin-Johnson syndrome (DJS). Naturally occurring mutations in MRP/ABCC-related drug transporters have been reported, some of which are non-synonymous single nucleotide polymorphisms. The consequences of the resulting amino acid changes can sometimes be predicted from in vitro site-directed mutagenesis studies or from knowledge of mutations of analogous (conserved) residues in ABCC proteins that cause DJS, Pseudoxanthoma elasticum (ABCC6), cystic fibrosis (CFTR/ABCC7) or persistent hyperinsulinemic hypoglycemia of infancy (SUR1/ABCC8). Continual updating of databases of sequence variants and haplotype analysis, together with in vitro biochemical validation assays and pharmacological studies in knockout animals, should make it possible to determine how genetic variation in the MRP-related transporters contributes to the range of responses to drugs and chemicals observed in different human populations.

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148 Fig. 3 Exon 1 2 3 MSDMSD NBD1 MSD NBD2 C4535T(S1512L) G3173A (R1058Q) G3140C (C1047S) G2965A (A989T) G2168A (R723Q) G2012T(G671V) G1898A (R633Q) G1299T(R433S) G1057A (V353M) G689A (R230Q) C350T(T117M) C257T(S92F) C218T(T73I) C128C (C43S) (TM1-5) (TM6-11) (TM12-17) 4 5 6 7 8 9101112 1314 151617 1819 20 21 22 23 242526272829 30 31 Location of non-synonymous SNPs in the coding regions of the genes in the MRP1/ABCC1 gene. 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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70 2650C > T exon 21 synonymous (p.L884L) Kim et al., 2001 md-v-110 rs9282563 c.2677G > T exon 21 p.A893S Kim et al., 2001 md-v-031 rs2032582 c.2677G > A exon 21 p.A893T Kim et al., 2001 md-v-109 IVS 21 + 14 - 17 delAATA intron 21 Epidauros md-v-092 IVS 21 + 49 T > C intron 21 Epidauros md-v-042 rs2032583 IVS 21 + 66 T > C intron 21 Epidauros md-v-108 IVS 26 - 156 T > C intron 25 Epidauros md-v-095 IVS 26 - 68 A > G intron 25 Epidauros md-v-164 c.3320A > C exon 26 p.Q1107P Cascorbi et al., 2002 md-v-033 c.3322T > C exon 26 p.W1108R Kroetz et al., 2003 md-v-225 c.3325C > T exon 26 p.L1109F Epidauros md-v-165 c 3364C > T exon 26 synonymous (p.A1132A) Hoffmeyer et al., 2000 md-v-034 c.3321T > A exon 26 p.S1141T Kim et al., 2001 md-v-035 c.3435C > T (Tag8) exon 26 synonymous (p.I1145I) Hoffmeyer et al., 2000 md-v-036 rs1045642 IVS 26 + 59 T > G intron 26 Epidauros md-v-097 rs2235047 IVS 26 + 80 T > C intron 26 Epidauros md-v-040 rs2235048 IVS 26 + 123_24 insCATG intron 26 Epidauros md-v-096 Tag 11 intron 27 Soranzo et al., 2004 rs1186746 Tag 12 intron 27 Soranzo et al., 2004 rs1186745 MRP1 (ABCC1) c.816G > A exon 8 synonymous (p.P272P) Epidauros mr-v-014 c.825T > C exon 8 synonymous (p.V275V) Saito et al., 2002 mr-v-015 rs246221 c.1062T > C exon 9 synonymous (p.N354N) Saito et al., 2002 mr-v-016 rs35587 c.1068G > A exon 9 synonymous (p.T356T) Epidauros mr-v-057 rs8187852 IVS 9 + 8 A > G intron 9 Saito et al., 2002 mr-v-017 rs35588 c.1303G > T exon 10 p.R433S Conrad et al., 2002 mr-v-018 IVS 10 + 64 C > T intron 10 Epidauros mr-v-019 MRP2 (ABCC2) g. Login to comment

[hide] Functional characterization of non-synonymous sing... Pharmacogenet Genomics. 2005 Sep;15(9):647-57. Letourneau IJ, Deeley RG, Cole SP
Functional characterization of non-synonymous single nucleotide polymorphisms in the gene encoding human multidrug resistance protein 1 (MRP1/ABCC1).
Pharmacogenet Genomics. 2005 Sep;15(9):647-57., [PMID:16041243]

Abstract [show]
The 190-kDa ATP-binding cassette (ABC) multidrug resistance protein 1 (MRP1) encoded by the MRP1/ABCC1 gene mediates the active cellular efflux of glucuronide, glutathione and sulfate conjugates. It can also confer resistance to a diverse spectrum of chemotherapeutic agents and transport a variety of toxicants. In the present study, we examined 10 MRP1/ABCC1 missense genetic variants [non-synonymous single nucleotide polymorphisms (SNPs)] to determine whether or not they affect expression or function of the transporter. Variants 218C>T (Thr73Ile), 257C>T (Ser92Phe), 350C>T (Thr117Met), 689G>A (Arg230Gln), 1898G>A (Arg633Gln), 2168G>A (Arg723Gln), 2965G>A (Ala989Thr), 3140G>C (Cys1047Ser), 3173G>A (Arg1058Gln) and 4535C>T (Ser1512Leu) were recreated using site-directed mutagenesis and transfected into human embryonic kidney cells. Immunoblotting experiments showed that all mutant proteins were expressed at levels comparable to wild-type MRP1. Vesicular transport assays revealed that the Ala989Thr mutation caused a significant decrease in estradiol 17beta-glucuronide transport due to a decrease in apparent affinity (Km) for this organic anion. The transport properties of the other mutants were comparable to wild-type MRP1. When the MRP1/ABCC1 non-synonymous SNPs were evaluated by the SIFT algorithm using subsets of homologs and orthologs of MRP1/ABCC1, Arg230Gln, Val353Met, Arg433Ser, Gly671Val and Arg1058 mutations were predicted to be deleterious, whereas the PolyPhen algorithm predicted Ser92Phe and Gly671Val to be potentially damaging. Thus most predictions of these algorithms were not in accordance with our experimental results. In conclusion, our data suggest that none of the MRP1/ABCC1 variants studied are likely by themselves to have major deleterious effects in healthy individuals, and the SIFT and PolyPhen algorithms appear to be poor predictors of the phenotypic consequences of these MRP1 mutations at least in vitro.

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7 When the MRP1/ABCC1 non-synonymous SNPs were evaluated by the SIFT algorithm using subsets of homologs and orthologs of MRP1/ABCC1, Arg230Gln, Val353Met, Arg433Ser, Gly671Val and Arg1058 mutations were predicted to be deleterious, whereas the PolyPhen algorithm predicted Ser92Phe and Gly671Val to be potentially damaging. Login to comment
28 Of these mutations, the Fig. 1 128G >C (C43S) 128G >T(T73I) 689G >A (R230Q)1057G >A (V353M) 1299G >T(R433S) 1898G >A (R633Q) 2012G >T(G671V) 2168G >A (R723Q) 3173G >A (R1058Q) 4535C >T(S1512L) 3140G >C (C1047S) 2965G >A (A989T) 350C >T(T117M) 257C >T(S92F) 313029282726252423222120181716151413121110987654321 19 MSD1 MSD1 MSD2 MSD3 MSD2 NBD1 MSD3 NBD2 TM 1 2 3 4 5 6 7 8 Val353Met Ala989Thr Cys1047Ser Arg1058Gln NBD2NBD1 Ser1512Leu Arg633Gln Arg433Ser Arg723Gln Thr73lle Thr117Met Arg230Gln Cys43Ser Ser92Phe Gly671Val 9 10 11 12 13 14 15 16 17 (a) (b) Location of reported non-synonymous single nucleotide polymorphisms (SNPs) in MRP1/ABCC1. Login to comment
35 A second mutation, Arg433Ser (1299G > T), located in cytoplasmic loop 4 (CL4) proximal to transmembrane (TM) helix 8 in the second MSD, decreased both LTC4 and estrone-3-sulfate (E13SO4) transport but increased resistance to doxorubicin [19]. Login to comment
46 The template for generating Table 1 Frequencies of non-synonymous single nucleotide polymorphisms in MRP1/ABCC1 Variant Amino acid substitution Allelic frequency Population References 128G > C Cys43Ser 0% (0/26) Japanese [16] 1% (1/96) Japanese [17] 218C > T Thr73Ile 0% (0/26) Japanese [16] 1% (1/96) Japanese [17] 3.7% (2/54) Chinese [37] 257C > T Ser92Phe 0% (0/220) Caucasian www.pharmGKB.org 0.5% (1/200) African-American 0% (0/60) Japanese 0% (0/14) Pacific-Islander 350C > T Thr117Met 1.6% (1/64) Caucasian [28] 689G > A Arg230Gln 0% (0/220) Caucasian www.pharmGKB.org 0.5% (1/200) African-American 0% (0/60) Japanese 0% (0/14) Pacific-Islander 1057G > A Val353Met 0.5% (1/220) Caucasian www.pharmGKB.org 0% (0/200) African-American 0% (0/60) Japanese 0% (0/14) Pacific-Islander 1299G > T Arg433Ser 1.4% (1/72) Caucasian [20] 0% (0/110) Caucasian [19] 1898G > A Arg633Gln 0.8% (2/234) Caucasian [29] 2012G > T Gly671Val 2.8% (2/72) Caucasian [20] 2.6% (6/234) Caucasian [29] 2168G > A Arg723Gln 3.8% (1/26) Japanese [16] 1% (1/96) Japanese [30] 7.3% (7/96) Japanese [17] 5.6% (3/54) Chinese [37] 2965G > A Ala989Thr 0.5% (1/220) Caucasian www.pharmGKB.org 0% (0/200) African-American 0% (0/60) Japanese 0% (0/14) Pacific-Islander 3140G > C Cys1047Ser 0% (0/220) Caucasian www.pharmGKB.org 4.5% (9/200) African-American 0% (0/60) Japanese 0% (0/14) Pacific-Islander 3173G > A Arg1058Gln 0% (0./26) Japanese [16] 1% (1/96) Japanese [17] 4535C > T Ser1512Leu 3.1% (2/24) Caucasian [28] Characterization of MRP1/ABCC1 variants in vitro Le´tourneau et al. 649 the Arg633Gln and Arg723Gln mutants was created by subcloning a HindIII fragment (1329 bp) encoding amino acids 517-959 into pGEM-3z [20]. Login to comment
94 Results from previously characterized non-synonymous SNPs, Cys43Ser, Arg433Ser and Gly671Val, were included where available for comparison [18-20]. Login to comment
123 Previous mutagenesis and inhibition studies have Fig. 3 Cys43Ser Thr73lle Ser92Phe Thr117Met Arg230Gln Arg433Ser Arg633Gln Gly671Val Arg723Gln Ala989Thr Cys1047Ser Arg1058Gln Ser1512Leu Cys43Ser Thr73lle Ser92Phe Thr117Met Arg230Gln Arg433Ser Arg633Gln Gly671Val Arg723Gln Ala989Thr Cys1047Ser Arg1058Gln Ser1512Leu Thr73lle Ser92Phe Thr117Met Arg230Gln Arg633Gln Arg723Gln Ala989Thr Cys1047Ser Arg1058Gln Ser1512Leu LTC4 % WT-MRP1 uptake 0 25 50 75 100 125 E217βG % WT-MRP1 uptake 0 25 50 75 100 125 150 MTX % WT-MRP1 uptake 0 25 50 75 100 125 (b) (c) (a) ATP-dependent vesicular transport of organic anions by mutant MRP1 proteins. Login to comment
128 Hatched bars represent previously published data on non-synonymous SNPs, Cys43Ser, Arg433Ser and Gly671Val, using membrane vesicles from stably transfected HeLa cells and are included for comparison [18-20]. Login to comment
134 Thus, when the subset included the human homologs MRP2, MRP3 and MRP6, the SIFT analysis predicted that the Arg433Ser, Gly671Val and Arg1058Gln substitutions would be deleterious for MRP1 function. Login to comment
135 However, when the analysis was expanded to include the six known mammalian orthologs of MRP1, the Arg230Gln, Val353Met, Arg433Ser and Gly671Val mutations were also predicted to be deleterious. Login to comment
136 The Val353Met mutation has not yet been tested in vitro; however, of the remaining five, only Arg433Ser has been found to adversely affect MRP1 function [19]. Login to comment
158 This observation may be construed as being Table 2 Conservation of the amino acids substituted by non-synonymous SNP of human MRP1/ABCC1a Protein Speciesb C43S T73I S92F T117Mc R230Q V353M R433S R633Qc G671V R723Q A989T C1047S R1058Q S1512L MRP1 Human C T S T R V R R G R A C R S Monkey C T S M R V R R G Q A C R S Dog C T S M R V R R G R A R R S Cow C A S M Q V R R G R A R R S Rat C A S M Q V R W G R A R R S Mouse C T S M H V R R G R A R R S MRP2 Human L A V T K A K R G K A I R E Monkey L A V T K A K R G K A I R E Dog L A V T K A K R G K A I Q Q Rat L A A T K V K R G K A A R E Mouse L A A T K V K V G K A T R E Rabbit L A V T K V K R G K A I R E MRP3 Human C L S M Y I R K G Q A V R A Rat C L S M L L R K G Q A L R V MRP4 Human - - - - I F K R G R Y T K Y MRP5 Human - - - - V T R S G R T R R S MRP6 Human P A A M R I R S G V A L R A CFTR Human - - - - R Y K A G K L I Q Q SUR1 Human V L L A T V Q R G E L R L E SUR2 Human V L H T Q V Q R G E I N L P Pgp Human - - - - - E K S G A G R R Q YCF1 Saccharomyces cervisiae A I L V T V K L G K S Y R G Mrp1 Caenorhabditis elegans T L D F L I R T G R G L R K Mrp2 Caenorhabditis elegans T F D I L I K T G R G I R K AtMRP2 Arabidopsis thaliana Q L R W L M S P G R R K R E AtMRP1 Arabidopsis thaliana H T A V L M S P G R R K R E a Aligned using Clustal W (http://pbil.univ-lyon1.fr/). Login to comment

[hide] Functional importance of three basic residues clus... J Biol Chem. 2006 Jan 6;281(1):43-50. Epub 2005 Oct 17. Conseil G, Deeley RG, Cole SP
Functional importance of three basic residues clustered at the cytosolic interface of transmembrane helix 15 in the multidrug and organic anion transporter MRP1 (ABCC1).
J Biol Chem. 2006 Jan 6;281(1):43-50. Epub 2005 Oct 17., 2006-01-06 [PMID:16230346]

Abstract [show]
The multidrug resistance protein 1 (MRP1) mediates drug and organic anion efflux across the plasma membrane. The 17 transmembrane (TM) helices of MRP1 are linked by extracellular and cytoplasmic (CL) loops of various lengths and two cytoplasmic nucleotide binding domains. In this study, three basic residues clustered at the predicted TM15/CL7 interface were investigated for their role in MRP1 expression and activity. Thus, Arg1138, Lys1141, and Arg1142 were replaced with residues of the same or opposite charge, expressed in human embryonic kidney cells, and the properties of the mutant proteins were assessed. Neither Glu nor Lys substitutions of Arg1138 and Arg1142 affected MRP1 expression; however, all four mutants showed a decrease in organic anion transport with a relatively greater decrease in leukotriene C4 and glutathione transport. These mutations also modulated MRP1 ATPase activity as reflected by a decreased vanadate-induced trapping of 8-azido-[32P]ADP. Mutation of Lys1141 to either Glu or Arg reduced MRP1 expression, and routing to the plasma membrane was impaired. However, only the Glu-substituted Lys1141 mutant showed a decrease in organic anion transport, and this was associated with decreased substrate binding and vanadate-induced trapping of 8-azido-ADP. These studies identified a cluster of basic amino acids likely at the TM15/CL7 interface as a region important for both MRP1 expression and activity and demonstrated that each of the three residues plays a distinct role in the substrate specificity and catalytic activity of the transporter.

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23 Furthermore, the naturally occurring substitution of Arg433 by a Ser at the interface of CL4 and TM8 causes a selective decrease in LTC4 and estrone sulfate transport efficiency but increases drug resistance (23). Login to comment

[hide] NAD(P)H oxidase and multidrug resistance protein g... Circulation. 2005 Dec 13;112(24):3754-62. Epub 2005 Dec 5. Wojnowski L, Kulle B, Schirmer M, Schluter G, Schmidt A, Rosenberger A, Vonhof S, Bickeboller H, Toliat MR, Suk EK, Tzvetkov M, Kruger A, Seifert S, Kloess M, Hahn H, Loeffler M, Nurnberg P, Pfreundschuh M, Trumper L, Brockmoller J, Hasenfuss G
NAD(P)H oxidase and multidrug resistance protein genetic polymorphisms are associated with doxorubicin-induced cardiotoxicity.
Circulation. 2005 Dec 13;112(24):3754-62. Epub 2005 Dec 5., 2005-12-13 [PMID:16330681]

Abstract [show]
BACKGROUND: A significant number of patients treated with anthracyclines develop cardiotoxicity (anthracycline-induced cardiotoxicity [ACT]), mainly presenting as arrhythmias (acute ACT) or congestive heart failure (chronic ACT). There are no data on pharmacogenomic predictors of ACT. METHODS AND RESULTS: We genotyped participants of the German non-Hodgkin lymphoma study (NHL-B) who were followed up for the development of heart failure for a median of >3 years. Single-nucleotide polymorphisms (SNPs) were selected from 82 genes with conceivable relevance to ACT. Of 1697 patients, 55 developed acute and 54 developed chronic ACT (cumulative incidence of either form, 3.2%). We detected 5 significant associations with polymorphisms of the NAD(P)H oxidase and doxorubicin efflux transporters. Chronic ACT was associated with a variant of the NAD(P)H oxidase subunit NCF4 (rs1883112, -212A-->G; symbols with right-pointing arrows, as edited?' odds ratio [OR], 2.5; 95% CI, 1.3 to 5.0). Acute ACT was associated with the His72Tyr polymorphism in the p22phox subunit (rs4673; OR, 2.0; 95% CI, 1.0 to 3.9) and with the variant 7508T-->A (rs13058338; OR, 2.6; 95% CI, 1.3 to 5.1) of the RAC2 subunit of the same enzyme. In agreement with these results, mice deficient in NAD(P)H oxidase activity, unlike wild-type mice, were resistant to chronic doxorubicin treatment. In addition, acute ACT was associated with the Gly671Val variant of the doxorubicin efflux transporter multidrug resistance protein 1 (MRP1) (OR, 3.6; 95% CI, 1.6 to 8.4) and with the Val1188Glu-Cys1515Tyr (rs8187694-rs8187710) haplotype of the functionally similar MRP2 (OR, 2.3; 95% CI, 1.0 to 5.4). Polymorphisms in adrenergic receptors previously demonstrated to be predictive of heart failure were not associated with ACT. CONCLUSIONS: Genetic variants in doxorubicin transport and free radical metabolism may modulate the individual risk to develop ACT.

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194 As opposed to polarized cells, in the cardiomyocytes MRP1 is found in the cytoplasm in addition to plasma membrane.37 It has been postulated that such an expression may permit sequestration of doxorubicin in lysosomes, ie, away from its target the nucleus.40 A low-frequency protein variant of MRP1 (Arg433Ser) has been shown to affect resistance to anthracyclines in vitro but has not been found in our study.41 The effect on doxorubicin resistance or transport of the variant 671Val, which shows an association with acute ACT in this study, has not been investigated.42 MRP2 has a substrate spectrum similar to that of MRP1 and increases the resistance to doxorubicin when overexpressed in HEK-293 cells.43 Physiologically, the MRP2 protein is expressed in the apical membrane of polarized cells in the liver and in the kidney, and there is no evidence for its expression in the heart. Login to comment
199 As opposed to polarized cells, in the cardiomyocytes MRP1 is found in the cytoplasm in addition to plasma membrane.37 It has been postulated that such an expression may permit sequestration of doxorubicin in lysosomes, ie, away from its target the nucleus.40 A low-frequency protein variant of MRP1 (Arg433Ser) has been shown to affect resistance to anthracyclines in vitro but has not been found in our study.41 The effect on doxorubicin resistance or transport of the variant 671Val, which shows an association with acute ACT in this study, has not been investigated.42 MRP2 has a substrate spectrum similar to that of MRP1 and increases the resistance to doxorubicin when overexpressed in HEK-293 cells.43 Physiologically, the MRP2 protein is expressed in the apical membrane of polarized cells in the liver and in the kidney, and there is no evidence for its expression in the heart. Login to comment

[hide] Metabolism and transport of oxazaphosphorines and ... Drug Metab Rev. 2005;37(4):611-703. Zhang J, Tian Q, Yung Chan S, Chuen Li S, Zhou S, Duan W, Zhu YZ
Metabolism and transport of oxazaphosphorines and the clinical implications.
Drug Metab Rev. 2005;37(4):611-703., [PMID:16393888]

Abstract [show]
The oxazaphosphorines including cyclophosphamide (CPA), ifosfamide (IFO), and trofosfamide represent an important group of therapeutic agents due to their substantial antitumor and immuno-modulating activity. CPA is widely used as an anticancer drug, an immunosuppressant, and for the mobilization of hematopoetic progenitor cells from the bone marrow into peripheral blood prior to bone marrow transplantation for aplastic anemia, leukemia, and other malignancies. New oxazaphosphorines derivatives have been developed in an attempt to improve selectivity and response with reduced toxicity. These derivatives include mafosfamide (NSC 345842), glufosfamide (D19575, beta-D-glucosylisophosphoramide mustard), NSC 612567 (aldophosphamide perhydrothiazine), and NSC 613060 (aldophosphamide thiazolidine). This review highlights the metabolism and transport of these oxazaphosphorines (mainly CPA and IFO, as these two oxazaphosphorine drugs are the most widely used alkylating agents) and the clinical implications. Both CPA and IFO are prodrugs that require activation by hepatic cytochrome P450 (CYP)-catalyzed 4-hydroxylation, yielding cytotoxic nitrogen mustards capable of reacting with DNA molecules to form crosslinks and lead to cell apoptosis and/or necrosis. Such prodrug activation can be enhanced within tumor cells by the CYP-based gene directed-enzyme prodrug therapy (GDEPT) approach. However, those newly synthesized oxazaphosphorine derivatives such as glufosfamide, NSC 612567 and NSC 613060, do not need hepatic activation. They are activated through other enzymatic and/or non-enzymatic pathways. For example, both NSC 612567 and NSC 613060 can be activated by plain phosphodiesterase (PDEs) in plasma and other tissues or by the high-affinity nuclear 3'-5' exonucleases associated with DNA polymerases, such as DNA polymerases and epsilon. The alternative CYP-catalyzed inactivation pathway by N-dechloroethylation generates the neurotoxic and nephrotoxic byproduct chloroacetaldehyde (CAA). Various aldehyde dehydrogenases (ALDHs) and glutathione S-transferases (GSTs) are involved in the detoxification of oxazaphosphorine metabolites. The metabolism of oxazaphosphorines is auto-inducible, with the activation of the orphan nuclear receptor pregnane X receptor (PXR) being the major mechanism. Oxazaphosphorine metabolism is affected by a number of factors associated with the drugs (e.g., dosage, route of administration, chirality, and drug combination) and patients (e.g., age, gender, renal and hepatic function). Several drug transporters, such as breast cancer resistance protein (BCRP), multidrug resistance associated proteins (MRP1, MRP2, and MRP4) are involved in the active uptake and efflux of parental oxazaphosphorines, their cytotoxic mustards and conjugates in hepatocytes and tumor cells. Oxazaphosphorine metabolism and transport have a major impact on pharmacokinetic variability, pharmacokinetic-pharmacodynamic relationship, toxicity, resistance, and drug interactions since the drug-metabolizing enzymes and drug transporters involved are key determinants of the pharmacokinetics and pharmacodynamics of oxazaphosphorines. A better understanding of the factors that affect the metabolism and transport of oxazaphosphorines is important for their optional use in cancer chemotherapy.

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627 Substitution of Arg433 with Ser predicted to be close to TM8 of MRP1 caused by the low frequency G1299T polymorphism in exon 10 leads to a substrate selective change in organic anion transport activity and drug resistance using MRP1-expressing HeLa cells (Conrad et al., 2002) or human leukemia CEM-7A cells (Assaraf et al., 2003). Login to comment

[hide] Nucleotide sequence analyses of the MRP1 gene in f... BMC Genomics. 2006 May 10;7:111. Wang Z, Sew PH, Ambrose H, Ryan S, Chong SS, Lee EJ, Lee CG
Nucleotide sequence analyses of the MRP1 gene in four populations suggest negative selection on its coding region.
BMC Genomics. 2006 May 10;7:111., [PMID:16684361]

Abstract [show]
BACKGROUND: The MRP1 gene encodes the 190 kDa multidrug resistance-associated protein 1 (MRP1/ABCC1) and effluxes diverse drugs and xenobiotics. Sequence variations within this gene might account for differences in drug response in different individuals. To facilitate association studies of this gene with diseases and/or drug response, exons and flanking introns of MRP1 were screened for polymorphisms in 142 DNA samples from four different populations. RESULTS: Seventy-one polymorphisms, including 60 biallelic single nucleotide polymorphisms (SNPs), ten insertions/deletions (indel) and one short tandem repeat (STR) were identified. Thirty-four of these polymorphisms have not been previously reported. Interestingly, the STR polymorphism at the 5' untranslated region (5'UTR) occurs at high but different frequencies in the different populations. Frequencies of common polymorphisms in our populations were comparable to those of similar populations in HAPMAP or Perlegen. Nucleotide diversity indices indicated that the coding region of MRP1 may have undergone negative selection or recent population expansion. SNPs E10/1299 G>T (R433S) and E16/2012 G>T (G671V) which occur at low frequency in only one or two of four populations examined were predicted to be functionally deleterious and hence are likely to be under negative selection. CONCLUSION: Through in silico approaches, we identified two rare SNPs that are potentially negatively selected. These SNPs may be useful for studies associating this gene with rare events including adverse drug reactions.

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8 SNPs E10/1299 G>T (R433S) and E16/2012 G>T (G671V) which occur at low frequency in only one or two of four populations examined were predicted to be functionally deleterious and hence are likely to be under negative selection. Login to comment
45 We found that SNPs E10/1299G>T, which resulted in arginine-serine substitution at amino acid position 433 (R433S) and E16/2012G>T, which resulted in glycine-valine substitution at amino acid position 671 (G671V), may potentially adversely affect the function of MRP1. Login to comment
123 2000 SNPe6* E10/1299 G>T R433S 0.12 benign -3.59003 0 0 0 1.39 Conrad et al . Login to comment

[hide] Genetic variations and haplotype structures of the... Drug Metab Pharmacokinet. 2007 Feb 25;22(1):48-60. Fukushima-Uesaka H, Saito Y, Tohkin M, Maekawa K, Hasegawa R, Kawamoto M, Kamatani N, Suzuki K, Yanagawa T, Kajio H, Kuzuya N, Yasuda K, Sawada J
Genetic variations and haplotype structures of the ABC transporter gene ABCC1 in a Japanese population.
Drug Metab Pharmacokinet. 2007 Feb 25;22(1):48-60., 2007-02-25 [PMID:17329911]

Abstract [show]
Multidrug resistance-related protein 1 (MRP1), an ATP-binding cassette transporter encoded by the ABCC1 gene, is expressed in many tissues, and functions as an efflux transporter for glutathione-, glucuronate- and sulfate-conjugates as well as unconjugated substrates. In this study, the 31 exons and their flanking introns of ABCC1 were comprehensively screened for genetic variations in 153 Japanese subjects to elucidate the linkage disequilibrium (LD) profiles and haplotype structures of ABCC1 that is necessary for pharmacogenetic studies of the substrate drugs. Eighty-six genetic variations including 31 novel ones were found: 1 in the 5'-flanking region, 1 in the 5'-untranslated region (UTR), 20 in the coding exons (9 synonymous and 11 nonsynonymous variations), 4 in the 3'-UTR, and 60 in the introns. Of these, eight novel nonsynonymous variations, 726G>T (Trp242Cys), 1199T>C (Ile400Thr), 1967G>C (Ser656Thr), 2530G>A (Gly844Ser), 3490G>A (Val1164Ile), 3550G>A (Glu1184Lys), 3901C>T (Arg1301Cys), and 4502A>G (Asp1501Gly), were detected with an allele frequency of 0.003. Based on the LD profiles, the analyzed regions of the gene were divided into five LD blocks (Blocks -1 and 1 to 4). The multiallelic repeat polymorphism in the 5'-UTR was defined as Block -1. For Blocks 1, 2, 3 and 4, 32, 23, 23 and 13 haplotypes were inferred, and 9, 7, 7 and 6 haplotypes commonly found on > or = 10 chromosomes accounted for > or = 91% of the inferred haplotypes in each block. Haplotype-tagging single nucleotide polymorphisms for each block were identified to capture the common haplotypes. This study would provide fundamental and useful information for the pharmacogenetic studies of MRP1-dependently effluxed drugs in Japanese.

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26 In Caucasian populations, 1299GÀT (Arg433Ser), 1898GÀA (Arg633Gln), 2012GÀT (Gly671Val), and 4535CÀT (Ser1512Leu) have been reported.79) In addition, Arg433Ser decreases the transport activity for LTC4 and estrone sulfate, but not for estradiol 17b-glucuronide, in vitro.10) Ito et al. found 16 genetic polymorphisms, including 4 nonsynonymous and 8 synonymous ones, in 48 Japanese subjects.11) An in vitro functional study showed that one of the non-synonymous variations, 2168GÀA (Arg723Gln), leads to reduced transport activity for LTC4, estradiol 17b-glucuronide and methotrexate.12) However, no haplotype analysis has been reported for the Japanese population. Login to comment

[hide] Multidrug resistance associated proteins as determ... Curr Drug Metab. 2007 Dec;8(8):787-802. Yu XQ, Xue CC, Wang G, Zhou SF
Multidrug resistance associated proteins as determining factors of pharmacokinetics and pharmacodynamics of drugs.
Curr Drug Metab. 2007 Dec;8(8):787-802., [PMID:18220559]

Abstract [show]
The multidrug resistance associated proteins (MRP1, MRP2, MRP3, MRP4, MRP5, MRP6, MRP7, MRP8 and MRP9) belong to the ATP-binding cassette superfamily (ABCC family) of transporters. They are expressed differentially in the liver, kidney, intestine, brain and other tissues. These transporters are localized to the apical and/or basolateral membrane of the hepatocytes, enterocytes, renal proximal tubule cells and endothelial cells of the blood-brain barrier. Several MRPs (mainly MRP1-3) are associated with tumor resistance which is often caused by an increased efflux and decreased intracellular accumulation of natural product anticancer drugs and other anticancer agents. MRPs transport a structurally diverse array of important endogenous substances and xenobiotics and their metabolites (in particular conjugates) with different substrate specificity and transport kinetics. Most MRPs are subject to induction and inhibition by a variety of compounds. Several nuclear receptors, including pregnane X receptor (PXR), liver X receptor (LXR), and farnesoid receptor (FXR) participate in the regulation of MRPs. MRPs play an important role in the absorption, distribution and elimination of various drugs in the body and thus may affect their efficacy and toxicity and cause drug-drug interactions. MRPs located in the blood-brain barrier can restrict the penetration of compounds into the central nervous system. Mutation of MRP2 causes Dubin-Johnson syndrome, while mutations in MRP6 are responsible for pseudoxanthoma elasticum. More recently, mutations in mouse Mrp6/Abcc6 gene is associated with dystrophic cardiac calcification (DCC), a disease characterized by hydroxyapatite deposition in necrotic myocytes. A single nucleotide polymorphism, 538G>A in the MRP8/ABCC11 gene, is responsible for determination of earwax type. A better understanding of the function and regulating mechanism of MRPs can help minimize and avoid drug toxicity, unfavourable drug-drug interactions, and to overcome drug resistance.

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405 Important Single Nucleotide Polymorphisms (SNPs) of MRP Genes MRP Chromosomal location Amino acid variation Nucleotide variation Location References Cys43Ser Thr73Ile G128C C218T Exon2 Exon2 [239] Arg433Ser G1299T Exon10 [258] Gly671Val G2012T Exon16 [259] Arg723Gln G2168A Exon17 [239] MRP1 16p13.11-p13.12 Arg1058Gln G3173A Exon23 [239] C-24T Promoter [100, 239] Val417Ile G1249A Exon10 [100, 238, 239] Gly676Arg G2026C Exon16 [237] Try709Arg T2125C Exon17 [236] Arg768Trp Ser789Phe C2302T C2366T Exon18 Exon18 [100, 238, 239] I1173F R1150H A3517T G3449A Exon25 Exon25 [240] Ile1324Ile C3972T Exon28 [100, 239] MRP2 10q23-24 Ala1450Thr G4348A Exon31 [100, 238, 239] (Table 2) contd…. Login to comment

[hide] Characterization and analyses of multidrug resista... Pharmacogenet Genomics. 2009 Mar;19(3):206-16. Yin JY, Huang Q, Yang Y, Zhang JT, Zhong MZ, Zhou HH, Liu ZQ
Characterization and analyses of multidrug resistance-associated protein 1 (MRP1/ABCC1) polymorphisms in Chinese population.
Pharmacogenet Genomics. 2009 Mar;19(3):206-16., [PMID:19214144]

Abstract [show]
OBJECTIVE: To explore the distribution frequencies of four common single nucleotide polymorphisms (SNPs) of MRP1/ABCC1 in a mainland Chinese population and investigate whether these SNPs affect the expression and function of the MRP1/ABCC1. METHODS: The genotype of 208 healthy volunteers was determined using PCR-restriction fragment length polymorphism. The four candidated SNPs were recreated by site-directed mutagenesis and tested for their effect on MRP1/ABCC1 expression and multidrug resistance function in stable transfected HEK293 and CHO-K1 cell lines. Real-time PCR, western blot and confocal microscopy were used to determine the mRNA, protein expression, and protein trafficking. At last, the effect of mutations on MRP1/ABCC1-mediate drug resistance was determined using methyl thiazolyl tetrazolium assay. RESULTS: The allelic frequencies of Cys43Ser (128G>C), Thr73Ile (218C>T), Arg723Gln (2168G>A), and Arg1058Gln (3173G>A) in mainland Chinese were 0.5, 1.4, 5.8, and 0.5%, respectively. None of these mutations had any effect on MRP1/ABCC1 expression and trafficking, but that Arg723Gln mutation significantly reduced MRP1/ABCC1-mediated resistance to daunorubicin, doxorubicin, etoposide, vinblastine, and vincristine. The Cys43Ser mutation did not affect all tested drug resistance. In contrast, the Thr73Ile mutation reduced resistance to methotrexate and etoposide, whereas the Arg1058Gln mutation increased the response of two anthracycline drugs and etoposide in HEK293 and CHO-K1 cells as well as vinblastine and methotrexate in CHO-K1 cells. CONCLUSION: The allelic frequency of the Arg723Gln mutation is relatively higher than other SNPs in mainland Chinese population and therefore this mutation significantly reduces MRP1/ABCC1 activity in multidrug resistance.

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25 To date, only three naturally occurring mutations of MRP1/ABCC1 (Gly671Val, Arg433Ser, and Cys43Ser) have been fully investigated for their effects on MRP1/ ABCC1-mediated MDR [18-20]. Login to comment
28 For example, the substitution of highly conserved Arg433 by Ser significantly decreased the transport of leukotriene C4 and estrone sulfate, but increased doxorubicin resistance [18]. Login to comment
157 It has also been reported that Gly671Val and Arg433Ser mutations were found in Caucasian, but not in Asian populations [15,18,19]. Login to comment

[hide] ABCC1 polymorphisms contribute to level and declin... Pharmacogenet Genomics. 2009 Sep;19(9):675-84. Siedlinski M, Boezen HM, Boer JM, Smit HA, Postma DS
ABCC1 polymorphisms contribute to level and decline of lung function in two population-based cohorts.
Pharmacogenet Genomics. 2009 Sep;19(9):675-84., [PMID:19687781]

Abstract [show]
OBJECTIVE: The ATP-binding cassette transporter ABCC1 [i.e. multidrug resistance-associated protein 1 (MRP1)] is a membrane-bound pump excreting a variety of xenobiotics from the cell, and thus ABCC1 may play an important role in smoking-related lung function loss and development of chronic obstructive pulmonary disease (COPD). We earlier showed that bronchial epithelium of COPD patients have lower ABCC1 expression than that of healthy controls, with even further decrements in more severe COPD stages. In line with these results, we now aimed to assess effects of ABCC1 single nucleotide polymorphisms (SNPs) on both the level and the longitudinal course of lung function in the general population. METHODS: All 51 prevalent (minor allele frequency >5%) and noncorrelated (r<0.8) ABCC1 SNPs were analyzed in two independent, prospective, population-based cohorts, that is, Doetinchem (n = 1152) and Vlagtwedde-Vlaardingen (n = 1390) studies (three and seven median lung function measurements, respectively, per patient), using linear regression and linear mixed-effect models. RESULTS: SNPs rs4148382 and rs212093 in the 3'-ABCC1 region were significantly associated with a higher and lower forced expiratory volume in 1 s (FEV1), respectively, in both the cohorts. Another rs35621 SNP (intron 14) was significantly associated with a highly excessive FEV1 decline in both cohorts. All replicated associations were additionally confirmed by permutation testing. CONCLUSION: This is the first study showing a significant relationship between ABCC1 SNPs and lung function in two independent cohorts. These SNPs are therefore putative candidates for studies aiming to prevent COPD and investigating pharmacogenetics in established COPD.

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72 Table 2 Additive effects of 53 ABCC1 SNPs on the level and change of FEV1 in the Doetinchem cohort (n = 1152) Additive effect on the level of FEV1 (ml)a Additive effect on the change of FEV1 (ml/year)b SNP Position MAF (%) HWE P value B SE P B SE P rs4148330 50 region 34.2 0.96 - 63.2 20.4 0.002 - 1.8 1.4 0.20 rs504348 [28] 50 region 19.3 0.61 - 54.9 24.7 0.03 - 2.4 1.7 0.15 rs152022 Intron 1 20.3 0.12 9.0 24.0 0.71 0.4 1.6 0.83 rs215049 Intron 1 31.9 0.72 - 32.5 21.1 0.12 - 1.5 1.5 0.31 rs215068 Intron 1 30.9 0.21 22.7 21.5 0.29 - 0.7 1.5 0.64 rs215079 Intron 1 21.0 0.24 3.2 23.9 0.89 - 1.5 1.7 0.37 rs215100 Intron 1 32.1 0.77 - 48.8 20.9 0.02 - 1.0 1.4 0.50 rs246220 Intron 1 12.6 0.34 - 19.0 30.1 0.53 1.2 2.1 0.55 rs4781699 Intron 1 31.4 0.63 - 56.6 20.9 0.01 - 2.2 1.4 0.13 rs8060750 Intron 1 12.7 0.52 24.4 29.2 0.40 - 1.1 2.0 0.59 rs8045000 Intron 1 32.8 - - 51.2 20.8 0.01 - 1.8 1.4 0.20 rs7190484 Intron 1 38.1 - - 48.8 20.1 0.02 - 1.8 1.4 0.19 rs7196970 Intron 1 40.6 - - 28.6 20.1 0.15 - 1.0 1.4 0.49 rs215101 Intron 1 10.5 - 36.2 33.1 0.27 - 0.9 2.2 0.70 rs4781711 Intron 1 6.9 - - 22.0 41.5 0.60 0.3 2.9 0.91 rs215059 Intron 1 14.2 - 15.6 28.8 0.59 0.8 2.0 0.69 rs4148337 Intron 3 31.9 0.72 - 25.4 21.0 0.23 - 1.0 1.4 0.51 rs4781718 Intron 3 14.3 0.39 - 0.1 27.8 0.99 - 2.1 1.9 0.28 rs246213 Intron 5 8.8 0.97 - 46.9 34.9 0.18 - 2.9 2.4 0.24 rs246240 Intron 5 18.0 0.28 3.1 25.3 0.90 - 0.7 1.7 0.71 rs7188937 Intron 5 46.4 - - 22.0 19.2 0.25 - 1.9 1.3 0.14 rs246233 Intron 6 9.0 - - 8.8 37.7 0.82 1.8 2.6 0.49 rs246230 Intron 7 14.6 0.42 - 5.2 27.5 0.85 - 0.1 1.9 0.97 rs193537 Intron 7 29.5 - - 14.0 22.3 0.53 - 0.2 1.5 0.89 rs60782127 (Arg433Ser) [29,30] Exon 10 2.2 1.00 - 25.2 68.0 0.71 3.3 4.7 0.48 rs12445600 Intron 10 27.8 0.57 - 22.8 22.0 0.30 - 0.5 1.5 0.74 rs4148344 Intron 10 15.7 - 12.5 28.4 0.66 0.5 1.9 0.80 rs35596 Intron 12 23.1 0.44 - 11.0 23.7 0.64 0.9 1.6 0.57 rs35597 Intron 12 46.7 0.33 32.9 20.1 0.10 3.1 1.4 0.02 rs35610 Intron 13 15.8 0.30 - 38.0 27.5 0.17 - 1.6 1.9 0.41 rs35621 Intron 14 10.7 0.61 -7.8 31.4 0.80 - 3.0 2.1 0.17 rs4148353 Intron 15 14.8 0.46 - 4.5 28.3 0.87 0.4 2.0 0.84 rs45511401 (Gly671Val) [30,31] Exon 16 4.9 1.00 22.5 45.4 0.62 - 0.7 3.1 0.81 rs2074086 Intron 18 31.9 0.43 11.7 21.0 0.58 - 1.0 1.4 0.49 rs2074087 Intron 18 14.2 - 42.1 30.6 0.17 - 3.6 2.1 0.08 rs4148358 Intron 19 23.5 0.94 11.1 23.2 0.63 - 0.1 1.6 0.93 rs2283515 Intron 19 46.6 - - 17.5 19.8 0.38 - 2.8 1.3 0.04 rs12449239 Intron 19 14.9 - - 20.4 29.8 0.49 - 0.7 2.0 0.74 rs11864374 Intron 21 22.6 0.62 - 27.4 24.2 0.26 - 2.9 1.6 0.08 rs9933511 Intron 21 27.1 0.09 9.5 22.7 0.68 - 0.5 1.5 0.75 rs3819552 Intron 21 49.0 0.57 6.6 19.9 0.74 2.7 1.4 0.04 rs2238475 Intron 23 4.6 0.16 - 10.8 46.0 0.82 1.2 3.2 0.71 rs2238476 Intron 23 5.9 0.74 28.0 41.5 0.50 - 1.5 2.8 0.59 rs12599429 Intron 26 31.8 0.85 17.7 21.5 0.41 2.0 1.5 0.17 rs212079 Intron 26 4.1 0.85 - 20.3 50.0 0.69 - 5.9 3.4 0.09 rs212083 Intron 27 19.5 0.46 - 10.6 24.6 0.67 - 3.9 1.7 0.02 rs2230671 (Ser1334Ser) Exon 28 29.8 0.98 5.7 21.6 0.79 1.9 1.5 0.19 rs3743527 30 UTR 22.5 0.09 - 58.6 23.5 0.01 - 1.3 1.6 0.44 rs212090 30 UTR 45.5 - - 11.2 19.4 0.56 4.3 1.3 0.001 rs212093 30 region 45.3 0.10 - 42.5 19.3 0.03 - 2.9 1.3 0.03 rs4148382 30 region 9.7 1.00 94.2 33.4 0.005 - 3.6 2.3 0.11 rs212094 30 region 18.0 - 15.9 25.8 0.54 - 5.3 1.8 0.003 rs12448760 30 region 29.1 - -7.5 21.6 0.73 2.7 1.5 0.07 The lack of HWE P values indicates haplotype-tagged SNPs. Login to comment

[hide] Structural and functional properties of human mult... Curr Med Chem. 2011;18(3):439-81. He SM, Li R, Kanwar JR, Zhou SF
Structural and functional properties of human multidrug resistance protein 1 (MRP1/ABCC1).
Curr Med Chem. 2011;18(3):439-81., [PMID:21143116]

Abstract [show]
Multidrug ABC transporters such as P-glycoprotein (P-gp/MDR1/ABCB1) and multidrug resistance protein 1 (MRP1/ABCC1) play an important role in the extrusion of drugs from the cell and their overexpression can be a cause of failure of anticancer and antimicrobial chemotherapy. Recently, the mouse P-gp/Abcb1a structure has been determined and this has significantly enhanced our understanding of the structure-activity relationship (SAR) of mammalian ABC transporters. This paper highlights our current knowledge on the structural and functional properties and the SAR of human MRP1/ABCC1. Although the crystal structure of MRP1/ABCC1 has yet to be resolved, the current topological model of MRP1/ABCC1 contains two transmembrane domains (TMD1 and TMD2) each followed by a nucleotide binding domain (NBD) plus a third NH2-terminal TMD0. MRP1/ABCC1 is expressed in the liver, kidney, intestine, brain and other tissues. MRP1/ABCC1 transports a structurally diverse array of important endogenous substances (e.g. leukotrienes and estrogen conjugates) and xenobiotics and their metabolites, including various conjugates, anticancer drugs, heavy metals, organic anions and lipids. Cells that highly express MRP1/ABCC1 confer resistance to a variety of natural product anticancer drugs such as vinca alkaloids (e.g. vincristine), anthracyclines (e.g. etoposide) and epipodophyllotoxins (e.g. doxorubicin and mitoxantrone). MRP1/ABCC1 is associated with tumor resistance which is often caused by an increased efflux and decreased intracellular accumulation of natural product anticancer drugs and other anticancer agents. However, most compounds that efficiently reverse P-gp/ABCB1-mediated multidrug resistance have only low affinity for MRP1/ABCC1 and there are only a few effective and relatively specific MRP1/ABCC1 inhibitors available. A number of site-directed mutagenesis studies, biophysical and photolabeling studies, SAR and QSAR, molecular docking and homology modeling studies have documented the role of multiple residues in determining the substrate specificity and inhibitor selectivity of MRP1/ABCC1. Most of these residues are located in the TMs of TMD1 and TMD2, in particular TMs 4, 6, 7, 8, 10, 11, 14, 16, and 17, or in close proximity to the membrane/cytosol interface of MRP1/ABCC1. The exact transporting mechanism of MRP1/ABCC1 is unclear. MRP1/ABCC1 and other multidrug transporters are front-line mediators of drug resistance in cancers and represent important therapeutic targets in future chemotherapy. The crystal structure of human MRP1/ABCC1 is expected to be resolved in the near future and this will provide an insight into the SAR of MRP1/ABCC1 and allow for rational design of anticancer drugs and potent and selective MRP1/ABCC1 inhibitors.

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735 The naturally occurring replacement of Arg433 by Ser (neutral) at the interface of CL4 and TM8 caused a 2-fold decrease in LTC4 and estrone sulfate transport but were 2.1-fold more resistant to doxorubicin while while resistance to etoposide (VP-16) and vincristine remained unchanged [363]. Login to comment
816 There are at least 15 naturally occurring mutations identified in MRP1/ABCC1, including Cys43Ser in TM1, Thr73Ile in CL1, Ser92Phe in TM2, Arg230Asn in L0, Val353Met at TM6/TM7 interface, Arg433Ser in TM8, Gly671Val in TM11, Arg723Gln located between the Walker A and Walker B motifs of NBD1, Ala861Thr at NBD1/TM12 interface, Ala989Thr in TM12, Cys1047Ser in TM13, Arg1058Gln in CL7, Val1146Ile in CL7, Thr1337Ala between the Walker A and Walker B motifs of NBD2, and Thr1401Met, and many of them have been found to affect its transport activity [171, 362, 363, 366, 367, 377-384]. Login to comment
819 A naturally-occurring mutation of 1299G>T in exon 10 causing Arg433Ser in TM8 of TMD1 resulted in decreased transport of LTC4 and increased resistance to doxorubicin [363]. Login to comment

[hide] Promoter polymorphism of MRP1 associated with redu... World J Gastroenterol. 2010 Dec 28;16(48):6104-10. Zhao J, Yu BY, Wang DY, Yang JE
Promoter polymorphism of MRP1 associated with reduced survival in hepatocellular carcinoma.
World J Gastroenterol. 2010 Dec 28;16(48):6104-10., 2010-12-28 [PMID:21182225]

Abstract [show]
AIM: to investigate the effect of the G-1666A polymorphism in the multidrug resistance related protein-1 (MRP1) on outcome of hepatocellular carcinoma (HCC). METHODS: a cohort of 162 patients with surgically resected HCC who received no postsurgical treatment until relapse was studied. Genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism analysis. Electrophoretic mobility shift assay (EMSA) was used to evaluate the influence of the G-1666A polymorphism on the binding affinity of the MRP1 promoter with its putative transcription factors. RESULTS: Kaplan-Meier analysis showed that patients with GG homologues had a reduced 4-year disease-free survival compared with those carrying at least one A allele (P = 0.011). Multivariate Cox regression analysis indicated that the -1666GG genotype represented an independent predictor of poorer disease-free survival [hazard ratio (HR) = 3.067, 95% confidence interval (CI): 1.587-5.952, P = 0.001], and this trend became worse in men (HR = 3.154, 95% CI: 1.604-6.201, P = 0.001). A similar association was also observed between 4-year overall survival and the polymorphism in men (HR = 3.342, 95% CI: 1.474-7.576, P = 0.004). Moreover, EMSA suggested that the G allele had a stronger binding affinity to nuclear proteins. CONCLUSION: the MRP1 -1666GG genotype predicted a worse outcome and was an independent predictor of poor survival in patients with HCC from Southeast China.

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46 G1299T (Arg433Ser) confers resistance to doxorubicin by reducing intracellular drug accumulation in HeLa cells that stably express mutant MRP1, whereas the G3173A (Arg1058Gln) variation increases the response to etoposide in HEK293 and CHO-K1 cells[12,13] . Login to comment

[hide] ABCC1 polymorphism Arg723Gln (2168G > A) is associ... Clin Exp Pharmacol Physiol. 2011 Sep;38(9):632-7. doi: 10.1111/j.1440-1681.2011.05571.x. Yin JY, Han LF, Huang Q, Xu XJ, Zhou HH, Liu ZQ
ABCC1 polymorphism Arg723Gln (2168G > A) is associated with lung cancer susceptibility in a Chinese population.
Clin Exp Pharmacol Physiol. 2011 Sep;38(9):632-7. doi: 10.1111/j.1440-1681.2011.05571.x., [PMID:21736601]

Abstract [show]
1. In a previous in vitro study, we showed that the Arg723Gln (2168G > A) polymorphism significantly ABCC1-induced multidrug resistance. The aim of the present study was to further investigate the association of this polymorphism with lung cancer susceptibility and chemotherapy response in a Chinese population. 2. A total of 77 lung cancer patients (54 men, 23 women) and 71 cancer-free controls (49 men, 22 women) were enrolled in the study. Genomic DNA was extracted from peripheral blood and all samples were genotyped using polymerase chain reaction-restriction fragment length polymorphism. 3. Individuals carrying the 723Gln (A) allele have a 3.4-fold increased risk (adjusted odds ratio (OR) 3.42; 95% confidence interval (CI) 1.29-9.06; P = 0.013) of lung cancer compared with wild-type individuals. Further stratified analysis indicated that older individuals (> 50 years) carrying the 723Gln (A) allele have the highest susceptibility to lung cancer (adjusted OR 4.10; 95% CI 1.25-13.48; P = 0.020). However, no substantial association was found between the Arg723Gln (2168G > A) polymorphism and chemotherapy response in Chinese lung cancer patients. 4. In conclusion, the Arg723Gln (2168G > A) polymorphism of ABCC1 appears to be a potential susceptibility marker for lung cancer in the Chinese population, especially in older people.

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24 For example, Cys43Ser (128G>C), which is located in the NH2 proximal region, was found to be important for the maintenance of MRP1/ABCC1-induced drug resistance.6 Another polymorphism, Arg433Ser (1299G>A), has the potential to increase resistance to doxorubicin in transfected Hela cells.7 Our previous investigation also showed that Arg723Gln (2168G>A) could significantly reduce MRP1/ABCC1 induced MDR.8 Furthermore, a number of in vivo studies have provided the clinical data to support the important role of MRP1/ABCC1 polymorphisms in disease susceptibility, drug metabolism and toxicity. Login to comment

[hide] Impact of genetic polymorphisms in transmembrane c... Naunyn Schmiedebergs Arch Pharmacol. 2004 Jan;369(1):69-77. Epub 2003 Nov 4. Gerloff T
Impact of genetic polymorphisms in transmembrane carrier-systems on drug and xenobiotic distribution.
Naunyn Schmiedebergs Arch Pharmacol. 2004 Jan;369(1):69-77. Epub 2003 Nov 4., [PMID:14598019]

Abstract [show]
Active transport across biological membranes has become a noticeable factor in the absorption, distribution, and excretion of an increasing number of drugs. Different transmembrane transport systems including organic anion transporters (OATP, solute carrier family SLC21A), organic cation transporters (OCT, SLC22A), dipeptide transporters (PEPT, SLC15A), nucleoside transporters (CNT, SLC28A), monocarboxylate carriers (MCT, SLC2A), and members of the large ATP-binding cassette family (ABC, SLC3A) are involved in drug disposition. Genetic polymorphisms in transport proteins frequently occur and contribute to interindividual differences in the efficacy and safety of pharmatherapy. Currently, the most advanced research has been done on P-glycoprotein (ABCB1, SLC3A1.201.1). Knowledge of this transporter indicates that haplotype analysis rather than association with single nucleotide polymorphisms (SNPs) provides the most appropriate interpretation of pharmacogenetic data from drug transporters. This review gives an overview and update on the pharmacological impact of genetic variants in transmembrane transporters.

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166 Digoxin plasma levels are inversely correlated with Pgp expression levels The functional relevance of the MRP1 SNPs G671 V, positioned within NBD1, and of Arg433Ser, located in a putative cytoplasmic loop, was tested using in vitro transport assays (Conrad et al. 2002). Login to comment
167 While the first did not change MRP1 transport activity for conjugated organic anions, the Arg433Ser polymorphism reduced transport activity of the substrates leucotriene C4 and estrone sulfate by half due to a decrease in Vmax for both compounds, whereas Km remained unchanged. Login to comment
168 In addition, doxorubicin-resistance of cells overexpressing the mutant Arg433Ser MRP1 was 2-fold higher compared to the wild-type. 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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7118 Nucleotide Change Amino Acid Change In Vitro Function Protein Expression/Localization ABCC1 MRP1 G128C C43S 1↔ Intracellular C218T T73I 1↔ Normal C257T S92F 2↔ Normal C350T T117M 2↔ Normal G689A R230Q ↔ Normal G1057A V353M N.D. N.D. G1299T R433S 2↔ Normal G1898A R633Q 2↔ Normal G2012T G671V ↔ Normal G2168A R723Q 2 Normal G2965A A989T 2↔ Normal G3140C C1047S 1↔ Normal G3173A R1058Q ↔ Normal C4535T S1512L ↔ Normal ABCC2 MRP2 C-24T N.D. N.D. G1058A R353H N.D. N.D. G1249A V417I ↔ Normal C2366T S789F 12 Intracellular T2780G L927R N.D. N.D. C3298T R1100C N.D. N.D. G3299A R1100H N.D. N.D. T3563A V1188E N.D. N.D. G4348A A1450T ↔ Normal/Intracellular G4544A C1515Y N.D. N.D. ABCC3 MRP3 G32A G11D ↔ Normal C202T H68Y N.D. N.D. G296A R99Q N.D. Normal C1037T S346F 2 Normal C1537A Q513K N.D. N.D. T1643A L548Q N.D. N.D. G1820A S607N 2 Normal C2221T Gln741STOP N.D. N.D. G2293C V765L ↔ Normal G2395A V799M N.D. N.D. C2758T P920S 1 Normal G2768A R923Q 1 Normal C3657A S1219R N.D. N.D. C3856G R1286G ↔ Normal G3890A R1297H N.D. N.D. C4042T R1348C 1 Normal A4094G Q1365R ↔ Normal C4141A R1381S ↔ Intracellular C4217T T1406M N.D. N.D. G4267A G1423R N.D. N.D. ABCC4 MRP4 C52A L18I N.D. N.D. C232G P78A 2↔ Normal T551C M184T N.D. N.D. G559T G187W 2 Reduced A877G K293E ↔ Normal G912T K304N ↔ Normal C1067T T356M N.D. N.D. C1208T P403L 2↔ Normal G1460A G487E 2 Normal A1492G K498E ↔ Normal A1875G I625M N.D. N.D. C2000T P667L N.D. N.D. A2230G M744V ↔ Normal G2269A E757K N.D. Intracellular G2459T R820I N.D. N.D. G2560T V854F N.D. N.D. G2698T V900L N.D. N.D. G2867C C956S 1↔ Normal G3211A V1071I ↔ Normal C3425T T1142M N.D. N.D. G3659A R1220Q N.D. N.D. A3941G Q1314R N.D. N.D. 2, reduced function; 1, increased function; ↔, no change in function; N.D. not determined. Login to comment
7115 Nucleotide Change Amino Acid Change In Vitro Function Protein Expression/Localization ABCC1 MRP1 G128C C43S 1࢒ Intracellular C218T T73I 1࢒ Normal C257T S92F 2࢒ Normal C350T T117M 2࢒ Normal G689A R230Q ࢒ Normal G1057A V353M N.D. N.D. G1299T R433S 2࢒ Normal G1898A R633Q 2࢒ Normal G2012T G671V ࢒ Normal G2168A R723Q 2 Normal G2965A A989T 2࢒ Normal G3140C C1047S 1࢒ Normal G3173A R1058Q ࢒ Normal C4535T S1512L ࢒ Normal ABCC2 MRP2 C-24T N.D. N.D. G1058A R353H N.D. N.D. G1249A V417I ࢒ Normal C2366T S789F 12 Intracellular T2780G L927R N.D. N.D. C3298T R1100C N.D. N.D. G3299A R1100H N.D. N.D. T3563A V1188E N.D. N.D. G4348A A1450T ࢒ Normal/Intracellular G4544A C1515Y N.D. N.D. ABCC3 MRP3 G32A G11D ࢒ Normal C202T H68Y N.D. N.D. G296A R99Q N.D. Normal C1037T S346F 2 Normal C1537A Q513K N.D. N.D. T1643A L548Q N.D. N.D. G1820A S607N 2 Normal C2221T Gln741STOP N.D. N.D. G2293C V765L ࢒ Normal G2395A V799M N.D. N.D. C2758T P920S 1 Normal G2768A R923Q 1 Normal C3657A S1219R N.D. N.D. C3856G R1286G ࢒ Normal G3890A R1297H N.D. N.D. C4042T R1348C 1 Normal A4094G Q1365R ࢒ Normal C4141A R1381S ࢒ Intracellular C4217T T1406M N.D. N.D. G4267A G1423R N.D. N.D. ABCC4 MRP4 C52A L18I N.D. N.D. C232G P78A 2࢒ Normal T551C M184T N.D. N.D. G559T G187W 2 Reduced A877G K293E ࢒ Normal G912T K304N ࢒ Normal C1067T T356M N.D. N.D. C1208T P403L 2࢒ Normal G1460A G487E 2 Normal A1492G K498E ࢒ Normal A1875G I625M N.D. N.D. C2000T P667L N.D. N.D. A2230G M744V ࢒ Normal G2269A E757K N.D. Intracellular G2459T R820I N.D. N.D. G2560T V854F N.D. N.D. G2698T V900L N.D. N.D. G2867C C956S 1࢒ Normal G3211A V1071I ࢒ Normal C3425T T1142M N.D. N.D. G3659A R1220Q N.D. N.D. A3941G Q1314R N.D. N.D. 2, reduced function; 1, increased function; ࢒, no change in function; N.D. not determined. Login to comment

[hide] Genetic association analysis of transporters ident... Pharmacogenet Genomics. 2012 Jun;22(6):447-65. Grover S, Gourie-Devi M, Bala K, Sharma S, Kukreti R
Genetic association analysis of transporters identifies ABCC2 loci for seizure control in women with epilepsy on first-line antiepileptic drugs.
Pharmacogenet Genomics. 2012 Jun;22(6):447-65., [PMID:22565165]

Abstract [show]
OBJECTIVE: The ATP-binding cassette (ABC) superfamily of transporters is known to efflux antiepileptic drugs (AEDs) primarily in the brain, gastrointestinal tract, liver, and kidneys. In addition, they are also known to be involved in estrogen disposition and may modulate seizure susceptibility and drug response. The objective of the present study was to investigate the role of genetic variants from ABC transporters in seizure control in epilepsy patients treated with monotherapy of first-line AEDs for 12 months. METHODS: On the basis of gene coverage and functional significance, a total of 98 single nucleotide polymorphisms from ABCB1, ABCC1, and ABCC2 were genotyped in 400 patients from North India. Of these, 216 patients were eligible for therapeutic assessment. Genetic variants were compared between the 'no-seizures' and the 'recurrent-seizures' groups. Bonferroni corrections for multiple comparisons and adjustment for covariates were performed before assessment of associations. RESULTS: Functionally relevant promoter polymorphisms from ABCC2: c.-1549G>A and c.-1019A>G either considered alone or in haplotype and diplotype combinations were observed for a significant association with seizure control in women (odds ratio>3.5, P<10, power>95%). Further, low protein-expressing CGT and TGT (c.-24C>T, c.1249G>A, c.3972C>T) haplotypes were always observed to be present in combination with the AG (c.-1549G>A, c.-1019A>G) haplotype that was over-represented in women with 'no seizures'. CONCLUSION: The distribution of the associated variants supports the involvement of ABCC2 in controlling seizures in women possibly by lowering of its expression. The biological basis of this finding could be an altered interaction of ABCC2 with AEDs and estrogens. These results necessitate replication in a larger pool of patients.

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87 - 330-21247T > C Intron 1 0.005 6 rs4148731 chr7:87239329 c.-330 - 8935C > T Intron 1 0.000 7 rs9282564 chr7:87229440 c.61A > G Exon 3 (Asn21Asp) 0.000 8 rs9282565 chr7:87214875 c.239C > A Exon 5 (Ala80Glu) 0.000 9 rs28381826 chr7:87214531 c.286 + 297G > A Intron 5 0.000 10 rs1989830 chr7:87205663 c.287 - 6124C > T Intron 5 0.135 11 rs2520464 chr7:87201086 c.287 - 1547A > G Intron 5 0.409 12 rs2235023 chr7:87190452 c.827+ 127G > A Intron 9 0.000 13 rs10276036 chr7:87180198 c.1000 - 44C > T Intron 10 0.401 14 rs2229109 chr7:87179809 c.1199G > A Exon 12 (Ser400Asn) 0.000 15 rs1128503 chr7:87179601 c.1236T > C Exon 13 (Gly412Gly) 0.390 16 rs2235036 chr7:87175271 c.1795G > A Exon 16 (Ala599Thr) 0.000 17 rs2235039 chr7:87165854 c.2401G > A Exon 21 (Val801Met) 0.000 18 rs2235040 chr7:87165750 c.2481 + 24G > A Intron 21 0.155 19 rs2032581 chr7:87160810 c.2485A > G Exon 22 (Ile829Val) 0.000 20 rs2032582 chr7:87160618 c.2677T/A > G Exon 22 (Ser/Thr893Ala) 0.318 21 rs7779562 chr7:87144816 c.3085 -72G > C Intron 25 0.043 22 rs2707944 chr7:87144641 c.3188C > G Exon 26 (Ala1063Gly) 0.000 23 rs2229107 chr7:87138659 c.3421A > T Exon 27 (Thr1141Ser) 0.000 24 rs1045642 chr7:87138645 c.3435T > C Exon 27 (Ile1145Ile) m Expression and activity [28] m mRNA expression [29] Altered substrate specificity [30] 0.375 25 rs2235048 chr7:87138511 c.3489 + 80C > T Intron 27 0.381 26 rs17064 chr7:87133470 c.3932A > T 30 UTR 0.000 ABCC1 1 rs504348 chr16:16043174 rs50438C > G Near gene region k Promoter activity [31] 0.135 2 rs215106 chr16:16047542 c.48 + 3886A > G Intron 1 0.210 3 rs215049 chr16:16070768 c.48 + 27112G > C Intron 1 0.245 4 rs246220 chr16:16082128 c.49 - 19545C > G Intron 1 0.118 5 rs119774 chr16:16086833 c.49 - 14840G > A Intron 1 0.089 6 rs246217 chr16:16090354 c.49 - 11319C > A Intron 1 0.118 7 rs2014800 chr16:16099966 c.49 - 1707C > T Intron 1 0.398 8 rs41494447 chr16:16101842 c.218C > T Exon 2 (Thr73Ile) 0.000 9 rs4781712 chr16:16103232 c.226 - 401A > G Intron 2 0.355 10 rs246240 chr16:16119024 c.616 -7942A > G Intron 5 0.114 11 rs924135 chr16:16123459 c.616 - 3507A > T Intron 5 0.412 12 rs903880 chr16:16130514 c.809 + 54C > A Intron 7 0.147 13 rs8187852 chr16:16139709 c.1057G > A Exon 9 (Met353Val) 0.000 14 rs35587 chr16:16139714 c.1062T > C Exon 9 (Asn354Asn) 0.182 15 rs35592 chr16:16141823 c.1219 - 176T > C Intron 9 0.172 16 rs60782127 chr16:16142079 c.1299G > T Exon 10 (Arg433Ser) k Transport of leukotriene C4 and estrone sulfate [32] 0.008 17 rs3765129 chr16:16149901 c.1474 - 48C > T Intron 11 0.032 18 rs35597 chr16:16158034 c.1678 - 3979G > A Intron 12 0.320 19 rs35621 chr16:16168608 c.1913 - 1575C > T Intron 14 0.103 20 rs45511401 chr16:16173232 c.2012G > T Exon 16 (Gly671Val) 0.024 21 rs4148356 chr16:16177275 c.2168G > A Exon 17 (Arg723Gln) 0.000 22 rs3851713 chr16:16184873 c.2644 + 428A > T Intron 19 0.340 23 rs2239995 chr16:16192565 c.2645 - 3919G > A Intron 19 0.324 24 rs11864374 chr16:16201885 c.2871 + 1155G > A Intron 21 0.338 25 rs35529209 chr16:16205325 c.2965G > A Exon 22 (Thr989Ala) k Transport of estradiol 17b-glucuronide [32] 0.000 26 rs3887893 chr16:16205501 c.3079 + 62G > A Intron 22 0.448 27 rs13337489 chr16:16208683 c.3140G > C Exon 23 (Ser1047Cys) 0.000 28 rs2299670 chr16:16220858 c.3819 + 1090A > G Intron 26 0.399 29 rs8057331 chr16:16230411 c.4202C > T Exon 29 (Thr1401Met) 0.000 30 rs212090 chr16:16236004 c.5462T > A 30 UTR 0.357 31 rs212093 chr16:16237754 rs212093G > A Near gene region 0.429 32 rs4148382 chr16:16238494 rs4148382G > A Near gene region 0.034 ABCC2 1 g.-1774G > delG chr10:101535688 g.-1774G > delG Near gene region k Promoter activity [33] 0.000 2 rs1885301 chr10:101541053 c.-1549G > A Near gene region k Promoter activity [haplotype containing (- 1549A)-(- 24T)] [33] k Clearance of irinotecan (ABCC2*2 containing the G allele) [34] 0.379 450 Pharmacogenetics and Genomics 2012, Vol 22 No 6 Table 2 (continued) N dbSNP ida Positionb Allelesc Gene location (effect) Function MAF 3 rs2804402 chr10:101541583 c. Login to comment

[hide] Identification and functional characterization of ... Pharmacogenetics. 2004 Apr;14(4):213-23. Lee YM, Cui Y, Konig J, Risch A, Jager B, Drings P, Bartsch H, Keppler D, Nies AT
Identification and functional characterization of the natural variant MRP3-Arg1297His of human multidrug resistance protein 3 (MRP3/ABCC3).
Pharmacogenetics. 2004 Apr;14(4):213-23., [PMID:15083066]

Abstract [show]
The human multidrug resistance protein 3 (MRP3, symbol ABCC3) is an ATP-binding cassette transporter that mediates the efflux of organic anions, including lipophilic substances conjugated with glucuronate, sulphate or glutathione, across the basolateral membrane of polarized cells (e.g. hepatocytes) into blood. Genetic variants of MRP3 may affect the transport of these substances out of cells. The aims of this study were: (i) to identify MRP3 polymorphisms; (ii) to functionally characterize one relatively frequent MRP3 polymorphism; and (iii) to establish whether MRP3 transports bilirubin glucuronosides. Exonic nucleotide variants in the ABCC3 gene were identified by single-strand conformation polymorphism analysis. The 3890G>A mutation, resulting in MRP3-ArgHis, was introduced into the ABCC3 cDNA which was stably transfected into MDCKII cells. For the functional characterization of MRP3-ArgHis in comparison with MRP3, ATP-dependent transport was analysed in isolated membrane vesicles. Two non-synonymous MRP3 variants were identified with an allele frequency of 0.003 for 1643T>A (MRP3-LeuGln) and 0.08 for 3890G>A (MRP3-ArgHis). Because of the high frequency of the 3890G>A mutation, and because of the close proximity of Arg to the second nucleotide-binding domain, we pursued the functional characterization of the MRP3-ArgHis polymorphic variant. MRP3-ArgHis was correctly localized to the basolateral membrane of polarized MDCKII cells. We identified monoglucuronosyl bilirubin, bisglucuronosyl bilirubin and leukotriene C4 as substrates for both MRP3 and MRP3-ArgHis. Dehydroepiandrosterone-3-sulphate and 17beta-glucuronosyl oestradiol were transported with similar kinetics by MRP3 and MRP3-ArgHis. This experimental setup provides a useful tool to analyse the functional consequences of polymorphic variants of MRP3.

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209 However, a naturally occurring variant (MRP1-Arg433 Ser) has been described, which was correctly routed to the plasma membrane but showed a selective decrease in organic anion transport [45]. 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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184 Nonetheless, two very low frequency (,2%) non-synonymous ABCC1 SNPs, e22/G2965A (Ala989Thr) and e10/G1299GT (Arg433Ser), were reported experimentally to result in decreased estradiol 17b-glucoronide (53) and organic anion transport but increased doxorubicin resistance (54), respectively. Login to comment
183 Nonetheless, two very low frequency (,2%) non-synonymous ABCC1 SNPs, e22/G2965A (Ala989Thr) and e10/G1299GT (Arg433Ser), were reported experimentally to result in decreased estradiol 17b-glucoronide (53) and organic anion transport but increased doxorubicin resistance (54), respectively. Login to comment

[hide] Influence of ABCB1 and ABCG2 polymorphisms on doxo... Cancer Sci. 2008 Apr;99(4):816-23. Lal S, Wong ZW, Sandanaraj E, Xiang X, Ang PC, Lee EJ, Chowbay B
Influence of ABCB1 and ABCG2 polymorphisms on doxorubicin disposition in Asian breast cancer patients.
Cancer Sci. 2008 Apr;99(4):816-23., [PMID:18377430]

Abstract [show]
The influence of three high frequency ABCB1 polymorphisms (c.1236C>T, c.2677G>A/T, and c.3435C>T) and the ABCG2 c.421C>A polymorphism on the disposition of doxorubicin in Asian breast cancer patients receiving adjuvant chemotherapy was investigated in the present study. The allelic frequency of the ABCB1 c.1236T, c.2677T, c.2677A, and c.3435T variants were 60%, 38%, 7%, and 22%, respectively, and the frequency of the ABCG2 c.421A allele was 23%. Pairwise analysis showed increased exposure levels to doxorubicin in patients harboring at least one ABCB1 c.1236T allele (P = 0.03). Patients homozygous for the CC-GG-CC genotype had significantly lower doxorubicin exposure levels compared to the patients who had CT-GT-CT (P = 0.02) and TT-TT-TT genotypes (P = 0.03). Significantly increased clearance of doxorubicin was also observed in patients harboring CC-GG-CC genotypes when compared to patients harboring the CT-GT-CT genotype (P = 0.01). Patients harboring the CC-GG-CC genotypes had significantly lower peak plasma concentrations of doxorubicinol compared to patients who had TT-TT-TT genotypes (P = 0.03). No significant influences on doxorubicin pharmacokinetic parameters were observed in relation to the ABCG2 c.421C>A polymorphism. In conclusion, the present exploratory study suggests that the three high frequency linked polymorphisms in the ABCB1 gene might be functionally important with regards to the altered pharmacokinetics of doxorubicin in Asian breast cancer patients, resulting in significantly increased exposure levels and reduced clearance.

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161 (44) Naturally occurring non-synonymous mutations (Arg433Ser) in the MRP1 cytoplasmic domain, leading to doxorubicin resistance,(45) also needs further investigation in terms of its role in altering doxorubicin disposition in cancer patients. 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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311 Another SNP G2012T in exon 16 (Gly671Val) was found to have no functional consequences, whereas exon 10 SNP G1299T (Arg433Ser; in the cytosolic interface of transmembrane segment 8) resulted in significant decrease in the transport ability of many organic anions but increased doxorubicin resistance (Conseil, Deeley, and Cole 2005). Login to comment

[hide] The G671V variant of MRP1/ABCC1 links doxorubicin-... Pharmacogenet Genomics. 2012 Apr;22(4):273-84. Jungsuwadee P, Zhao T, Stolarczyk EI, Paumi CM, Butterfield DA, St Clair DK, Vore M
The G671V variant of MRP1/ABCC1 links doxorubicin-induced acute cardiac toxicity to disposition of the glutathione conjugate of 4-hydroxy-2-trans-nonenal.
Pharmacogenet Genomics. 2012 Apr;22(4):273-84., [PMID:22293538]

Abstract [show]
OBJECTIVE: Doxorubicin-induced acute cardiotoxicity is associated with the Gly671Val (G671V; rs45511401) variant of multidrug resistance-associated protein 1 (MRP1). Doxorubicin redox cycling causes lipid peroxidation and generation of the reactive electrophile, 4-hydroxy-2-trans-nonenal (HNE). Glutathione forms conjugates with HNE, yielding an MRP1 substrate, GS-HNE, whose intracellular accumulation can cause toxicity. METHODS: We established stable HEK293 cell lines overexpressing wild-type MRP1 (HEKMRP1), G671V (HEKG671V), and R433S (HEKR433S), a variant not associated with doxorubicin-induced cardiotoxicity and investigated the sensitivity of HEKG671V cells to doxorubicin and transport capacity of G671V toward GS-HNE. RESULTS: In ATP-dependent transport studies using plasma membrane-derived vesicles, the Vmax (pmol/min/mg) for GS-HNE transport was the lowest for G671V (69+/-4) and the highest for R433S (972+/-213) compared with wild-type MRP1 (416+/-22), whereas the Km values were 2.8+/-0.4, 6.0 or more, and 1.7+/-0.2 micromol/l, respectively. In cells, the doxorubicin IC50 (48 h) was not different in HEKMRP1 (463 nmol/l) versus HEKR433S (645 nmol/l), but this parameter was significantly lower in HEKG671V (181 nmol/l). HEKG671V retained significantly (approximately 20%) more, whereas HEKR433S retained significantly less intracellular doxorubicin than HEKMRP1. Similarly, HEKG671V cells treated with 1.5 micromol/l of doxorubicin for 24 h retained significantly more GS-HNE. In cells treated with 0.5 micromol/l of doxorubicin for 48 , glutathione and glutathione disulfide levels and the glutathione/glutathione disulfide ratio were significantly decreased in HEKG671V versus HEKMRP1; these values were similar in HEKR433S versus HEKMRP1. CONCLUSION: These data suggest that decreased MRP1-dependent GS-HNE efflux contributes to increased doxorubicin toxicity in HEKG671V and potentially in individuals carrying the G671V variant.

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3 Methods We established stable HEK293 cell lines overexpressing wild-type MRP1 (HEKMRP1), G671V (HEKG671V), and R433S (HEKR433S), a variant not associated with doxorubicin-induced cardiotoxicity and investigated the sensitivity of HEKG671V cells to doxorubicin and transport capacity of G671V toward GS-HNE. Login to comment
4 Results In ATP-dependent transport studies using plasma membrane-derived vesicles, the Vmax (pmol/min/mg) for GS-HNE transport was the lowest for G671V (69 ± 4) and the highest for R433S (972 ± 213) compared with wild-type MRP1 (416 ± 22), whereas the Km values were 2.8 ± 0.4, 6.0 or more, and 1.7 ± 0.2 lmol/l, respectively. Login to comment
20 Among several variants, they identified Arg433Ser (R433S), Original article 273 1744-6872 c 2012 Wolters Kluwer Health | Lippincott Williams & Wilkins DOI: 10.1097/FPC.b013e328350e270 located in the second transmembrane spanning domain, and Gly671Val (G671V; rs45511401), near the Walker A motif in the first nucleotide-binding domain. Login to comment
21 The R433S MRP1 variant showed a 50% decreased transport maximum for leukotriene C4 (LTC4) [8]; however, cells expressing this variant were more resistant to doxorubicin than those expressing wild-type (WT) MRP1 [8]. Login to comment
37 In this study, we examined whether cells expressing the G671V and R433S variants versus WT MRP1 were more sensitive to doxorubicin, and characterized the transport properties of WT MRP1 relative to the G671V and R433S MRP1 variants, specifically with respect to GS-HNE transport activity. Login to comment
46 MRP1 variants of G671Vand R433S were generated using the QuickChange II XL site-directed mutagenesis kit (Stratagene, La Jolla, California, USA) according to the manufacturer`s instructions with the following mutagenic primers (for G671V, forward primer: 50 -TCCATCCCCGAAGTTGCTTTGGTG 274 GCCGTG-30 and reverse primer: 50 -CACGGCCACCAAAG CAACTTCGGGGATGGA-30 ; for R433S, forward primer: 50 -GTGGACGCTCAGAGCTTCATGGACTTGGC-30 and reverse primer: 50 -GCCAAGTCCATGAAGCTCTGAGCGT CCAC-30 ). Login to comment
114 In an attempt to understand this phenomenon, we developed HEK293 cell lines that stably expressed human MRP1 SNPs G671V, and three different control cells: HEK293 cells transfected with pUSEamp(+) vector (HEKpUSE), WT MRP1 (HEKMRP1), or the R433S variant (HEKR433S), which is not associated with doxorubicin-induced acute cardiac toxicity. Login to comment
141 MRP1 G671V decreases GS-HNE transport Jungsuwadee et al. 277 Fig. 1 (b) 3000 MRP1pUSE G671V R433S *** *** *** 2000 Meanfluorescenceintensity 1000 0 pUSE MRP1 G671V R433S (a) (c) 10.0 * * * 7.5 5.0 2.5 MRP1/18S 0.0 pUSE MRP1 G671V R433S Expression in stable cell lines of multidrug resistance-associated protein 1 (MRP1) and its variants. Login to comment
143 Plasmids containing wild-type MRP1 or the MRP1 variants G761V and R433S were transfected into HEK293 cells to generate stable cell lines. Login to comment
149 Fig. 2 (a) (b) Genotype Doxorubicin IC50 nmol/l (95% CI) pUSE 58.0 (51.4-65.5) MRP1 463.0 (332.7-644.2) G671V 181.4 (137.4-239.5) R433S 645.2 (158.3-2630.0) pUSE125 100 75 50 Percentsurvival 25 0 MRP1 G671V R433S 10 Doxorubicin (μmol/l) -1-2-3 Percent survival of HEKpUSE, HEKMRP1, HEKR433S, and HEKG671V cells in the presence of doxorubicin. Login to comment
156 The G671V variant was comparable with WT MRP1 with respect to LTC4 transport, whereas LTC4 transport was decreased by 75% in the R433S variant compared with WT MRP1 (Fig. 6b), consistent with previous reports [7,8]. Login to comment
160 In contrast, the Vmax of the R433S variant was increased over two-fold relative to WT MRP1. Login to comment
162 The estimate of the Km for the R433S variant was higher (Z 6 mmol/l), consistent with its increased Vmax (Fig. 6c). Login to comment
165 To investigate whether the cardiac sarcolemma can transport GS-HNE, and the Fig. 3 (a) (b) (c) (d) 1.5 ** ** **** ** ** ** *** ** 1.0 Fractionofcontrol doxorubicinaccumulation Fractionofcontrol doxorubicinaccumulation 0.5 0.0 8 250 200 150 100 GS-HNE(%control) 50 0 pUSE MRP1 G671V R433S 6 4 GS-HNE(nmol/mg) 2 0 Doxorubicin (1.5 μmol/l) - + - + - + - + pUSE MRP1 G671V R433S 1.5 1.0 0.5 0.0 pUSE 1 h M RP1 h G 671V 1 h R433S 1 h pUSE 24 h M RP1 24 h G 671V 24 h R433S 24 h Doxorubicin and glutathione-conjugated 4-hydroxy-2-trans-nonenal (GS-HNE) retention in HEKpUSE, HEKMRP1, HEKR433S, and HEKG671V cells. Login to comment
182 These data are in contrast with an earlier report [8] showing that HeLa cells expressing R433S are two-fold more resistant than cells expressing WT MRP1. Login to comment
185 We next examined cellular GS-HNE levels after treatment of cells with 1.5 mmol/l of doxorubicin for 24 h and Fig. 4 20(a) (b) (c) 15 12.5 10.0 7.5 5.0Glutathione (nmol/mgprotein) GSSG(nmol/mgprotein) 2.5 0.0 15 10 5 Glutathione/GSSGratio 0 pUSE pUSE 0.8 *** *** *** ** *** ***0.6 0.4 0.2 0.0 M RP1 G 671V R433S M RP1 G 671V R433S pUSE M RP1 G 671V R433S 10 Glutathione(nmol/mg) Glutathione/GSSGratio 5 0 0 16 14 12 10 8 0 10 20 30 40 50 Time (h) 25 50 Time (h) 1.5 1.0 0.5 0.0 0 10 20 30 40 50 Time (h) GSSG(nmol/mg) Time course of intracellular glutathione and GSSG in HEKpUSE, HEKMRP1, HEKR433S, and HEKG671V cells. Login to comment
205 It is also interesting to note that the R433S variant showed significantly increased transport of GS-HNE, despite decreased transport of LTC4 (Fig. 6b) and estrone sulfate, and unaltered transport of E217G [8]. Login to comment
219 Although increased retention of doxorubicin could also contribute to the increased oxidative stress in cells expressing the G671V variant, because amino acids in the third membrane spanning domain, especially between amino acids 959 and 1187, are considered most critical for doxorubicin transport [35], it seems less Fig. 6 Genotype Km ± SE (μmol/l) Vmax± SE (pmol/min/mg) Vmax/Km (mg/l/min) pUSE (95% CI) 6.3 ±12.9 (0.0-33.5) 28.4± 36.4 (0.0-104.8) 4.5 MRP1 (95% CI) 1.7 ±0.2 (1.2-2.1) 416.1±21.7 (371.1-461.2) 244.7 G671V (95% CI) 2.8± 0.4 (2.1-3.6) 69.1 ±4.1 (60.5-77.7) 24.7 R433S (95% CI) ≥6 972.1± 212.7 (531.0-1413) Approximately 160 pUSE MRP1 600 500 Protein(pmol/min/mg) 400 300 200 100 0 0 1 2 3 GS-HNE (μmol/l) 4 5 6 7 G671V R433S 125 *** *** 100 75 50 25 0 PercentLTC4efflux relativetoWT M RP1G 671VR433S M RP1+glutathione 5 G 671V+glutathione 5 R433S+glutathione 5 M RP1+glutathione 0.5 G 671V+glutathione 0.5 R433S+glutathione 0.5 2.5 (a) (b) (c) MRP1 pUSE/293 MRP1 G671V R433S Na/K-ATPase MRP1/NaK-ATPase banddensity (arbitraryunit) 2.0 1.5 1.0 0.5 0.0 MRP1 G671V R433S Multidrug resistance-associated protein 1 (MRP1) expression and transport activities of HEKMRP1, HEKR433S, and HEKG671V. 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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179 3.1.2.1 ABCC1/MRP1 In 2001, Conrad et al. described two non-synonymous polymorphisms, R433S and G671V. Login to comment

[hide] Two polymorphic variants of ABCC1 selectively alte... Drug Metab Dispos. 2013 Dec;41(12):2187-96. doi: 10.1124/dmd.113.054213. Epub 2013 Sep 30. Conseil G, Cole SP
Two polymorphic variants of ABCC1 selectively alter drug resistance and inhibitor sensitivity of the multidrug and organic anion transporter multidrug resistance protein 1.
Drug Metab Dispos. 2013 Dec;41(12):2187-96. doi: 10.1124/dmd.113.054213. Epub 2013 Sep 30., [PMID:24080162]

Abstract [show]
In this study we compared the in silico predictions of the effect of ABCC1 nonsynonymous single nucleotide polymorphisms (nsSNPs) with experimental data on MRP1 transport function and response to chemotherapeutics and multidrug resistance protein 1 (MRP1) inhibitors. Vectors encoding seven ABCC1 nsSNPs were stably expressed in human embryonic kidney (HEK) cells, and levels and localization of the mutant MRP1 proteins were determined by confocal microscopy and immunoblotting. The function of five of the mutant proteins was determined using cell-based drug and inhibitor sensitivity and efflux assays, and membrane-based organic anion transport assays. Predicted consequences of the mutations were determined by multiple bioinformatic methods. Mutants C43S and S92F were correctly routed to the HEK cell plasma membrane, but the levels were too low to permit functional characterization. In contrast, levels and membrane trafficking of R633Q, G671V, R723Q, A989T, and C1047S were similar to wild-type MRP1. In cell-based assays, all five mutants were equally effective at effluxing calcein, but only two exhibited reduced resistance to etoposide (C1047S) and vincristine (A989T; C1047S). The GSH-dependent inhibitor LY465803 (LY465803 [N-[3-(9-chloro-3-methyl-4-oxo-4H-isoxazolo-[4,3-c]quinolin-5-yl)-cyclohexylmethy l]-benzamide)] was less effective at blocking calcein efflux by A989T, but in a membrane-based assay, organic anion transport by A989T and C1047S was inhibited by MRP1 modulators as well as wild-type MRP1. GSH accumulation assays suggest cellular GSH efflux by A989T and C1047S may be impaired. In conclusion, although six in silico analyses consistently predict deleterious consequences of ABCC1 nsSNPs G671V, changes in drug resistance and inhibitor sensitivity were only observed for A989T and C1047S, which may relate to GSH transport differences.

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27 Previously, we generated and partially characterized recombinant forms of ABCC1 nsSNPs: rs45511401 (2012G.T; G671V), rs60782127 (1299G.T; R433S), and rs41395947 (128G.C; C43S) (Conrad et al., 2001, 2002; Leslie et al., 2003). Login to comment
29 In contrast, the R433S mutant exhibited reduced LTC4 and estrone sulfate transport levels (Conrad et al., 2002). Login to comment
30 Cells expressing MPR1-R433S also showed increased doxorubicin resistance, suggesting R433S can better efflux this drug despite its lower organic anion transport activity. Login to comment
34 In contrast to our experimental data, the predictive algorithms SIFT (Sorting Tolerant From Intolerant) and PolyPhen indicated that G671V would adversely affect MRP1 function but C43S and A989T were less likely to do so, whereas predictions for R433S were mixed (L&#e9;tourneau et al., 2005). Login to comment

[hide] Importance of ABCC1 for cancer therapy and prognos... Drug Metab Rev. 2014 Aug;46(3):325-42. doi: 10.3109/03602532.2014.901348. Epub 2014 Mar 26. Kunicka T, Soucek P
Importance of ABCC1 for cancer therapy and prognosis.
Drug Metab Rev. 2014 Aug;46(3):325-42. doi: 10.3109/03602532.2014.901348. Epub 2014 Mar 26., [PMID:24670052]

Abstract [show]
Multidrug resistance presents one of the most important causes of cancer treatment failure. Numerous in vitro and in vivo data have made it clear that multidrug resistance is frequently caused by enhanced expression of ATP-binding cassette (ABC) transporters. ABC transporters are membrane-bound proteins involved in cellular defense mechanisms, namely, in outward transport of xenobiotics and physiological substrates. Their function thus prevents toxicity as carcinogenesis on one hand but may contribute to the resistance of tumor cells to a number of drugs including chemotherapeutics on the other. Within 48 members of the human ABC superfamily there are several multidrug resistance-associated transporters. Due to the well documented susceptibility of numerous drugs to efflux via ABC transporters it is highly desirable to assess the status of ABC transporters for individualization of treatment by their substrates. The multidrug resistance associated protein 1 (MRP1) encoded by ABCC1 gene is one of the most studied ABC transporters. Despite the fact that its structure and functions have already been explored in detail, there are significant gaps in knowledge which preclude clinical applications. Tissue-specific patterns of expression and broad genetic variability make ABCC1/MRP1 an optimal candidate for use as a marker or member of multi-marker panel for prediction of chemotherapy resistance. The purpose of this review was to summarize investigations about associations of gene and protein expression and genetic variability with prognosis and therapy outcome of major cancers. Major advances in the knowledge have been identified and future research directions are highlighted.

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148 However, substitution of highly conserved Arg433 with serine (G1299T, rs60782127) significantly reduces transport of estrone-3-sulfate and LTC4 and increases resistance to doxorubicin (Conrad et al., 2002). Login to comment
150 Interestingly, this Arg433Ser substitution was associated with increased doxorubicin resistance in transfected cells. Login to comment
159 NCBI ID Reference Amino acid exchange Nucleotide exchange Location Function MAFa rs41395947 Cys43Ser G128C Exon 2 Non-synonymous Unknown rs41494447 Thr73Ile C218T Exon 2 Non-synonymous T &#bc; 0.003 rs8187844 Ser92Phe C257T Exon 3 Non-synonymous T &#bc; 0.004 rs8187848 Arg230Gln G689A Exon 7 Non-synonymous A &#bc; 0.009 rs2230669 Pro272Pro G816A Exon 8 Synonymous A &#bc; 0.037 rs246221 Val275Val T825C Exon 8 Synonymous C &#bc; 0.301 rs35592 non-coding T-176C Intron 9 Non-coding C &#bc; 0.257 rs60782127 Arg433Ser G1299T Exon 10 Non-synonymous T &#bc; 0.004 rs35605 Leu562Leu T1684C Exon 13 Synonymous T &#bc; 0.173 rs112282109 Arg633Gln G1898A Exon 14 Non-synonymous A &#bc; 0.004 rs45511401 Gly671Val G2012T Exon 16 Non-synonymous T &#bc; 0.050 rs4148356 Arg723Gln G2168A Exon17 Non-synonymous A &#bc; 0.027 rs35529209 Ala989Thr G2965A Exon 22 Non-synonymous Unknown rs13337489 Cys1047Ser G3140C Exon 23 Non-synonymous C &#bc; 0.000 rs41410450 Arg1058Gln G3173A Exon 23 Non-synonymous Unknown rs2238476 non-coding G-1960A Intron 23 Non-coding T &#bc; 0.062 rs2230671 Ser1334Ser G4002A Exon 28 Synonymous T &#bc; 0.208 rs28364006 Thr1337Ala A4009G Exon 28 Non-synonymous Unknown rs369410659 Ser1512Leu C4535T Exon 31 Non-synonymous Unknown a Minor allele frequencies for Caucasinans in dbSNP based on HapMap-CEU population or 1000 genomes. Login to comment

[hide] Genetic variation of the ABC transporter gene ABCC... BMC Genet. 2015 Sep 23;16(1):114. doi: 10.1186/s12863-015-0271-3. Slomka M, Sobalska-Kwapis M, Korycka-Machala M, Bartosz G, Dziadek J, Strapagiel D
Genetic variation of the ABC transporter gene ABCC1 (Multidrug resistance protein 1-MRP1) in the Polish population.
BMC Genet. 2015 Sep 23;16(1):114. doi: 10.1186/s12863-015-0271-3., [PMID:26395522]

Abstract [show]
BACKGROUND: Multidrug resistance-associated protein 1 (MRP1), encoded by the ABCC1 gene, is an ATP-binding cassette transporter mediating efflux of organic anions and xenobiotics; its overexpression leads to multidrug resistance. In this study, 30 exons (from 31 in total) of the ABCC1 gene as well as and their flanking intron sequences were screened for genetic variation, using the High Resolution Melting (HRM) method, for 190 healthy volunteers representing the Polish population. Polymorphism screening is an indispensable step in personalized patient therapy. An additional targeted SNP verification study for ten variants was performed to verify sensitivity of the scanning method. RESULTS: During scanning, 46 polymorphisms, including seven novel ones, were found: one in 3' UTR, 21 in exons (11 of them non-synonymous) and 24 in introns, including one deletion variant. These results revealed some ethnic differences in frequency of several polymorphisms when compared to literature data for other populations. Based on linkage disequilibrium analysis, 4 haplotype blocks were determined for 9 detected polymorphisms and 12 haplotypes were defined. To capture the common haplotypes, haplotype-tagging single nucleotide polymorphisms were identified. CONCLUSIONS: Targeted genotyping results correlated well with scanning results; thus, HRM is a suitable method to study genetic variation in this model. HRM is an efficient and sensitive method for scanning and genotyping polymorphic variants. Ethnic differences were found for frequency of some variants in the Polish population compared to others. Thus, this study may be useful for pharmacogenetics of drugs affected by MRP1-mediated efflux.

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136 Variants c.596C > T (p.Ser199Leu) and c.814C > T (p.Pro272Ser) were located in the third intracellular loop (between TM5 and TM6), variant c.1299G > T (p.Arg433Ser) in the fourth intracellular loop, variant c.3196C > T (p.Arg1066Trp) in the seventh intracellular loop (between TM7 and TM8). Login to comment
142 The novel variant c.596C > T (p.Ser199Leu) was estimated as a probably damaging substitution, likewise as four others: c.1299G > T (Arg433 Ser), c.2012G > T (p.Gly671Val), c.3886C > T (p.Arg 1296Trp) and c.3901C > T (p.Arg1301Cys). Login to comment
143 On the other hand, analysis for HumVar-trained model indicated that three polymorphisms: c.1299G > T (Arg433Ser), c.2012G > T (p.Gly671Val), c.3901C > T (p.Arg1301Cys), lead to probably damaging substitutions and two others, c.596C > T (p.Ser199Leu) and c.3886C > T (p.Arg1296Cys), are possibly damaging variants. Login to comment
144 Table 2 Summary of ABCC1 variants detected during scanning by HRM Exon scanned by HRM dbSNP ID Variant position NM_004996.3: Intron/amino acid residue NP_004987.2: Observed genotypesa, b (n) HWE exact test P-valuec MAFd R/R R/V V/V 2 rs8187843 c.225 + 26G > A Intron 164 25 0 1 (A) 0.066 4 rs587783373* c.352-79G > A Intron 185 1 0 1 (A) 0.003 4 rs4148337 c.352-66 T > C Intron 15 80 91 0.727 (T) 0.296 5 rs483352860* c.596C > T p.Ser199Leu 186 1 0 1 (T) 0.003 6 rs8187846 c.677 + 17C > T Intron 188 1 0 1 (T) 0.003 7 rs483352864* c.809 + 16C > T Intron 188 1 0 1 (T) 0.003 7 rs45609533 c.809 + 31G > T Intron 183 5 0 1 (T) 0.013 7 rs903880 c.809 + 54C > A Intron 112 65 11 0.684 (A) 0.231 7 rs246232 c.809 + 64C > G Intron 84 90 14 0.174 (G) 0.314 8 rs546943313 c.810-73C > T Intron 187 1 0 1 (T) 0.003 8 rs200194736 c.814C > T p.Pro272Ser 187 1 0 1 (T) 0.003 8 rs2230669 c.816G > A p.Pro272= 172 16 0 1 (A) 0.043 8 rs246221 c.825 T > C p.Val275= 84 92 12 0.059 (C) 0.309 8 rs587783372* c.855G > A p.Pro285= 187 1 0 1 (A) 0.003 9 rs35587 c.1062 T > C p.Asn354= 78 91 16 0.185 (C) 0.332 9 rs35588 c.1218 + 8A > G Intron 82 91 16 0.245 (G) 0.327 9 rs483352877* c.1218 + 9C > T Intron 188 1 0 1 (T) 0.003 10 rs60782127 c.1299G > T p.Arg433Ser 186 2 0 1 (T) 0.005 12 rs17265551 c.1677 + 56C > T Intron 162 27 0 0.604 (T) 0.072 13 rs35604 c.1678-37G > A Intron 2 45 142 0.745 (G) 0.130 13 rs483352863* c.1678-34G > A Intron 188 1 0 1 (A) 0.003 13 rs35605 c.1684 T > C p.Leu562= 2 45 142 0.745 (T) 0.130 13 rs8187858 c.1704C > T p.Tyr568= 157 31 1 1 (T) 0.088 14 rs112282109 c.1898G > A p.Arg633Gln 187 1 0 1 (A) 0.003 16 rs8187863 c.2001C > T p.Ser667= 187 1 0 1 (T) 0.003 16 rs45511401 c.2012G > T p.Gly671Val 161 25 2 0.296 (T) 0.077 17 rs4148356 c.2168G > A p.Arg723Gln 181 9 0 1 (A) 0.024 19 rs45607032 c.2461-39_2461-38delAT Intron 179 9 0 1 (delAT) 0.024 19 rs2074087 c.2461-30C > G Intron 0 44 144 0.083 (C) 0.117 19 rs45492500 c.2461-27G > A Intron 172 14 2 0.056 (A) 0.048 21 rs11075296 c.2871 + 26C > T Intron 0 0 189 1 - 22 rs768191257 c.2876A > G p.Lys959Arg 187 1 0 1 (G) 0.003 22 rs3851716 c.3079 + 10G > A Intron 0 0 188 1 - 22 rs34794353 c.3079 + 24C > T Intron 187 1 0 1 (T) 0.003 22 rs3887893 c.3079 + 62 T > C Intron 67 96 25 0.358 (C) 0.388 23 rs191017838 c.3171G > A p.Leu1057= 187 2 0 1 (A) 0.005 23 rs199773531 c.3196C > T p.Arg1066Trp 188 1 0 1 (T) 0.003 25 rs41278168 c.3591-5C > T Intron 187 1 0 1 (T) 0.003 27 rs200922662 c.3886C > T p.Arg1296Trp 187 1 0 1 (T) 0.003 27 rs201533167 c.3901C > T p.Arg1301Cys 187 1 0 1 (T) 0.003 Linkage disequilibrium analysis Based on full genotype sets of 44 polymorphic variants confirmed by Hardy-Weinberg equilibrium exact test (Table 2), linkage disequilibrium analysis using r2 and |D`| statistics was performed (Additional file 4). Login to comment
154 Bold variants signifies the ones which were validated by genotyping results Table 3 Summary of ABCC1 selected SNPs genotyping by HRM and comparing them with scanning results dbSNP ID Variant residue NM_004996.3: Intron/amino acid residue NP_004987.2: Observed genotypesa (n) HWE exact test P-valueb MAFc (genotyping) MAFc (scanning) Chi-square test P-valued R/R R/V V/V rs41395947 c.128G > C p.Cys43Se 380 0 0 1 - - - rs2230669 c.816G > A p.Pro272= 362 18 0 1 (A) 0.024 (A) 0.043 0.079 rs246221 c.825 T > C p.Val275 197 160 23 0.243 (C) 0.271 (C) 0.309 0.187 rs8187852 c.1057G > A p.Val353Met 379 0 0 1 - - - rs35587 c.1062 T > C p.Asn354= 204 142 33 0.247 (C) 0.274 (C) 0.332 0.044 rs35588 c.1218 + 8A > G Intron 190 160 30 0.709 (G) 0.289 (G) 0.325 0.214 rs60782127 c.1299G > T p.Arg433Ser 373 6 0 1 (T) 0.008 (T) 0.005 0.623 rs35605 c.1684 T > C p.Leu562= 13 105 262 0.588 (T) 0.172 (T) 0.130 0.063 rs8187858 c.1704C > T p.Tyr568= 325 55 0 0.242 (T) 0.072 (T) 0.087 0.374 rs45511401 c.2012G > T p.Gly671Val 346 28 3 0.007 (T) 0.045 (T) 0.077 0.038 rs4148356 c.2168G > A p.Arg723Gln 360 19 0 1 (A) 0.025 (A) 0.024 0.888 rs45517537 c.2581G > A p.Ala861Thr 380 0 0 1 - - - rs35529209 c.2965G > A p.Ala989Thr 378 0 0 1 - - - rs13337489 c.3140G > C p.Cys1047Ser 380 0 0 1 - - - rs28706727 c.3436G > A p.Val1146Ile 380 0 0 1 - - - rs2230671 c.4002G > A p.Ser1334= 204 140 32 0.296 (A) 0.271 (A) 0.277 0.850 rs28364006 c.4009A > G p.Thr1337Ala 380 0 0 1 - - - a Number of genotypes detected during this study, R - reference allele, V - variant allele. b P-value is consistent with Hardy-Weinberg equilibrium if P > 0.001. c Minor allele shown in brackets with its frequency. Login to comment