ABCMdb

PMID: 25492507

Nakano E, Masamune A, Niihori T, Kume K, Hamada S, Aoki Y, Matsubara Y, Shimosegawa T
Targeted next-generation sequencing effectively analyzed the cystic fibrosis transmembrane conductance regulator gene in pancreatitis.
Dig Dis Sci. 2015 May;60(5):1297-307. doi: 10.1007/s10620-014-3476-9. Epub 2014 Dec 10., [PubMed]

Sentences

No. Mutations Sentence Comment

6 ABCC7 p.Glu585* ABCC7 p.Lys411Glu We could identify 12 non-synonymous variants including three novel ones [c.A1231G (p.K411E), c.1753G[T (p.E585X) and c.2869delC (p.L957fs)] and seven synonymous variants including three novel ones in the exonic regions. Login to comment 7 ABCC7 p.Gln1352His ABCC7 p.Leu1156Phe The frequencies of the c.4056G[C (p.Q1352H) and the c.3468G[T (p.L1156F) variants were higher in patients with chronic pancreatitis than those in controls. Login to comment 85 ABCC7 p.Glu585* ABCC7 p.Lys411Glu Based on the presence in dbSNP137, Exome Variant Server (NHLBI GO Exome Sequencing Project, Seattle, WA, USA; URL: http://evs.gs.washington.edu/ EVS/), and the Human Genetic Variation Database, three non-synonymous variants [c.1231A[G (p.K411E), c.1753G[T (p.E585X) and c.2869delC (p.L957fs)] and three synonymous variants (c.372C[T, c.3975A[G and c.4254G[A) were novel. Login to comment 88 ABCC7 p.Gln1352His The patient also had the c.4056G[C (p.Q1352H) variant in a Fig. 1 Graph of the mean depth, median depth, and sequencing coverage for all the 27 exons in the CFTR gene. Login to comment 90 ABCC7 p.Glu585* ABCC7 p.Arg31Cys ABCC7 p.Arg1453Trp ABCC7 p.Val470Met ABCC7 p.Ile556Val ABCC7 p.Ile125Thr ABCC7 p.Gln1352His ABCC7 p.Gly1349Ser ABCC7 p.Lys411Glu ABCC7 p.Leu1156Phe ABCC7 p.Arg31His On average, 90.3 % of the coding region was successfully covered by C20 reads Table 2 Non-synonymous CFTR variants detected in this study Exon Non-synonymous variant Amino acid change dbSNP135 Genotype SIFT (score) PolyPhen-2 (score) Alcoholic CP (%) Idiopathic CP (%) Hereditary/ familial CP (%) 2 c.91C[T p.R31C rs1800073 CT D (0.012) PD (0.989) 0/46 (0) 3/121 (2.5) 0/26 (0) 2 c.92G[A p.R31H rs149353983 GA T (0.183) B (0.003) 0/46 (0) 1/121 (0.8) 0/26 (0) 4 c.374T[C p.I125T rs141723617 TC D (0.005) B (0.17) 0/46 (0) 2/121 (1.6) 1/26 (3.8) 10 c.1231A[G p.K411E - AG D (0.015) B (0.233) 0/46 (0) 1/121 (0.8) 0/26 (0) 11 c.1408G[A p.V470M rs213950 GA T (1) B (0) 21/46 (45.7) 65/121 (53.7) 11/26 (42.3) AA 5/46 (10.9) 19/121 (15.7) 1/26 (3.8) 12 c.1666A[G p.I556V rs75789129 AG T (0.536) B (0.334) 2/46 (4.3) 8/121 (6.6) 0/26 (0) GG 0/46 (0) 0/121 (0) 0/26 (0) 13 c.1753G[T p.E585X - GT - - 1/46 (2.2) 0/121 (0) 0/26 (0) 17 c.2869delC p.L957fs - - - 0/46 (0) 1/121 (0.8) 0/26 (0) 21 c.3468G[T p.L1156F rs139729994 GT T (0.163) PD (0.994) 2/46 (4.3) 10/121 (8.3) 2/26 (7.7) TT 1/46 (2.2) 0/121 (0) 0/26 (0) 25 c.4045G[A p.G1349S rs201686600 GA D (0) PD (1) 1/46 (2.2) 0/121 (0) 0/26 (0) 25 c.4056G[C p.Q1352H rs113857788 GC D (0) PD (1) 5/46 (10.9) 11/121 (9.1) 4/26 (15.4) CC 0/46 (0) 0/121 (0) 0/26 (0) 27 c.4357C[T p.R1453W rs4148725 CT D (0) PD (0.999) 3/46 (6.5) 6/121 (5.0) 1/26 (3.8) B benign, CP chronic pancreatitis, D damaging, PD probably damaging, T tolerated, SIFT Sorting Intolerant From Tolerant heterozygous form (Table 6). Login to comment 91 ABCC7 p.Glu585* The nonsense variant c.1753G[T (p.E585X) was found in a patient with alcoholic CP. He was diagnosed as having alcoholic CP at 28 years old. The c.1231A[G (p.411E) variant was found in a 19-year-old male with idiopathic CP. He had suffered from pancreatitis attacks since 12 years old. Login to comment 94 ABCC7 p.Leu1156Phe The patient also had the c.3468G[T (p.L1156F) variant in a heterozygous form. Login to comment 97 ABCC7 p.Gln1352His The frequency of the c.4056G[C (p.Q1352H) variant was higher in all patients with CP than that in controls (P = 0.009; Table 3). Login to comment 99 ABCC7 p.Leu1156Phe The frequency of the c.3468G[T (p.L1156F) variant was also higher in patients with CP than that in controls (P = 0.04). Login to comment 100 ABCC7 p.Glu585* ABCC7 p.Arg31Cys ABCC7 p.Arg1453Trp ABCC7 p.Val470Met ABCC7 p.Ile556Val ABCC7 p.Gln1352His ABCC7 p.Gly1349Ser ABCC7 p.Lys411Glu ABCC7 p.Leu1156Phe ABCC7 p.Arg31His There were no significant difference for any other non-synonymous or synonymous variants detected in the exons Table 3 Comparison of the non-synonymous variant frequencies between the patients with CP and controls Amino acid change Genotype All CP (%) HGVD (%) P value (vs. HGVD) All CP Alcoholic CP Nonalcoholic CP Idiopathic CP Hereditary/ familial CP p.R31C CT 3/193 (1.6) 12/1102 (1.1) 0.48 [0.99 0.41 0.18 [0.99 p.R31H GA 1/193 (0.5) 0 - - - - - p.I125T TC 3/193 (1.6) 5/1102 (0.5) 0.11 [0.99 0.057 0.15 0.13 p.K411E AG 1/193 (0.5) 0 - - - - - p.V470M GA 97/193 (50.3) 573/1199 (47.8) 0.66 0.57 0.68 0.38 0.12 AA 25/193 (13.0) 185/1199 (15.4) p.I556V AG 10/193 (5.2) 78/1150 (6.8) 0.70 0.79 0.81 [0.99 0.45 GG 0/193 (0) 3/1150 (0.3) p.E585X GT 1/193 (0.5) 0 - - - - - p.L957fs 1/193 (0.5) 0 - - - - - p.L1156F GT 14/193 (7.3) 45/1136 (4.0) 0.04 0.06 0.07 0.11 0.30 TT 1/193 (0.5) 1/1136 (0.1) p.G1349S GA 1/193 (0.5) 4/1094 (0.4) 0.56 0.19 [0.99 [0.99 [0.99 p.Q1352H GC 20/193 (10.4) 57/1153 (4.9) 0.009 0.12 0.037 0.17 0.062 CC 0/193 (0) 1/1153 (0.1) p.R1453W CT 10/193 (5.2) 42/1144 (3.7) 0.32 0.25 0.49 0.45 [0.99 CP chronic pancreatitis, HGVB Human Genetic Variation Database P values were determined versus HGVD by the Fisher`s exact test Table 4 Synonymous variants in the exons of the CFTR gene detected in this study Exon Synonymous variant Amino acid change dbSNP135 Genotype Alcoholic CP (%) Idiopathic CP (%) Hereditary/ familial CP (%) 4 c.372C[T p.G124= - CT 0/46 (0) 1/121 (0.8) 0/26 (0) 13 c.1731C[T p.Y577= rs55928397 CT 0/46 (0) 1/121 (0.8) 0/26 (0) 15 c.2562T[G p.T854= rs1042077 TG 20/46 (43.5) 69/121 (57.0) 12/26 (46.2) GG 6/46 (13.0) 18/121 (14.9) 0/26 (0) 23 c.3723C[A p.G1241= rs185065886 CA 1/46 (2.2) 0/121 (0) 0/26 (0) 25 c.3975A[G p.R1325= - AG 0/46 (0) 1/121 (0.8) 0/26 (0) 27 c.4254G[A p.E1418= - GA 0/46 (0) 1/121 (0.8) 0/26 (0) 27 c.4389G[A p.Q1463= rs1800136 GA 1/46 (2.2) 3/121 (2.5) 0/26 (0) CP chronic pancreatitis between all patients with CP and controls (Tables 3, 5). Login to comment 114 ABCC7 p.Glu585* ABCC7 p.Arg31Cys ABCC7 p.Arg31Cys ABCC7 p.Arg31Cys ABCC7 p.Arg1453Trp ABCC7 p.Arg1453Trp ABCC7 p.Ile556Val ABCC7 p.Gln1352His ABCC7 p.Gly1349Ser ABCC7 p.Lys411Glu ABCC7 p.Leu1156Phe ABCC7 p.Leu1156Phe ABCC7 p.Arg31His Comprehensive analysis by targeted NGS enabled us to identify novel and Table 5 Comparison of the synonymous variant frequencies between the patients with CP and controls Synonymous variant Genotype All CP (%) HGVD (%) P value (vs. HGVD) All CP Alcoholic CP Nonalcoholic CP Idiopathic CP Hereditary/ familial CP c.C372T CT 1/193 (0.5) 0 - - - - - c.1731C[T CT 1/193 (0.5) 0 - - - - - c.2562T[G TG 101/193 (52.3) 528/1154 (45.8) 0.22 0.81 0.11 0.045 0.033 GG 24/193 (12.4) 181/1154 (15.7) c.3723C[A CA 1/193 (0.5) 3/671 (4.5) [0.99 0.23 [0.99 [0.99 [0.99 c.3975A[G AG 1/193 (0.5) 0 - - - - - c.4254G[A GA 1/193 (0.5) 0 - - - - - c.4389G[A GA 4/193 (2.1) 40/1112 (3.6) 0.48 [0.99 0.53 0.81 [0.99 AA 0/193 (0) 1/1112 (0.1) CP chronic pancreatitis, HGVD Human Genetic Variation Database P values were determined against HGVD by the Fisher`s exact test Table 6 Total CFTR sequencing results of patients carrying rare non-synonymous CFTR variants a Pancreatitis-associated mutations in the PRSS1, SPINK1, CTRC, and CPA1 genes Case# Etiology Age at onset Rare variant Additional non-synonymous variants c.1210-34TG(9_13) c.1210-12T(5_9) Mutation in other pancreatitis susceptibility genesa A1 Idiopathic 34 p.R31C/- p.R1453W/- TG11/TG11, 7T/7T - A2 Idiopathic 8 p.R31C/- - TG11/TG12, 7T/7T - A3 Idiopathic 16 p.R31C/- - TG11/TG12, 7T/7T - A4 Idiopathic 10 p.R31H/- - TG11/TG12, 7T/7T - A5 Idiopathic 16 p.I125T/- p.L1156F/- TG11/TG12, 7T/7T CTRC p.R29Q/- A6 Idiopathic 2 p.I125T/- - TG11/TG12, 7T/7T - A7 Hereditary 28 p.I125T/- p.R1453W/- TG11/TG12, 7T/7T - A8 Idiopathic 19 p.K411E/- p/L1156F/- TG11/TG12, 7T/7T - A9 Alcoholic 28 p.E585X/- p.I556V/- TG11/TG11, 7T/7T - A10 Idiopathic 21 p.L957fs/- p.Q1352H/- TG11/TG12, 7T/7T - A11 Alcoholic 40 p.G1349S/- - TG11/TG11, 7T/7T - rare variants in the CFTR gene. Login to comment 115 ABCC7 p.Glu585* The c.1753G[T (p.E585X) variant is a nonsense variant, and the c.2869delC (p.L957fs) variant leads to a stop codon afterward at amino acid 967. Login to comment 116 ABCC7 p.Glu585* These variants result in a heavily truncated protein missing nearly two-thirds (p.E585X) or more than one-third (p.L957fs) of its amino acids. Login to comment 119 ABCC7 p.Lys411Glu The pathogenic potential of another novel variant, c.1231A[G (p.K411E), is currently unknown, but the in silico analyses suggest that this variant is deleterious. Login to comment 120 ABCC7 p.Leu1156Phe Importantly, the clinical phenotype of this patient might be complicated by the presence of another variant, p.L1156F. Login to comment 122 ABCC7 p.Ile125Thr This is also the case with the c.374T[C (p.I125T) variant. Login to comment 123 ABCC7 p.Arg1453Trp ABCC7 p.Ile556Val Two of the three patients carrying this variant had other non-synonymous variants (p.I556V and p.R1453W). Login to comment 124 ABCC7 p.Ile125Thr This p.I125T variant was originally reported in Chinese patients with idiopathic bronchiectasis and considered to be associated with CFTR-RD [35]. Login to comment 134 ABCC7 p.Gln1352His We found a significant association between the p.Q1352H variant and CP. Login to comment 138 ABCC7 p.Gln1352His Glutamine at 1,352 is located in the second nucleotide-binding fold of CFTR, and its change to histidine (p.Q1352H) causes reductions in both the protein expression and channel activity of CFTR [40]. Login to comment 139 ABCC7 p.Leu1156Phe Similarly, we found that the p.L1156F variant was overexpressed in patients with CP. Login to comment 140 ABCC7 p.Leu1156Phe A functional study reported reduced Cl- / HCO3 - permeability in the presence of the p.L1156F variant [41]. Login to comment 142 ABCC7 p.Gln1352His ABCC7 p.Leu1156Phe Indeed, seven out of 25 patients carrying the SPINK1 variant(s) had the CFTR p.Q1352H and/or p.L1156F variants. Login to comment 143 ABCC7 p.Ile125Thr ABCC7 p.Leu1156Phe One patient was trans-heterozygous for the CTRC p.R29Q and CFTR p.I125T/p.L1156F variants. Login to comment 145 ABCC7 p.Arg75Gln Trans-heterozygosity of the SPINK1 p.N34S with the Table 7 Distribution of the c.1210-34TG(9_13) and c.1210-12T(5_9) variants in patients with CP CP chronic pancreatitis c.1210-34TG(9_13), c.1210-12T(5_9) All CP (%) Alcoholic CP (%) Idiopathic CP (%) Hereditary/ familial CP (%) TG10/TG11, 7T/9T 1/193 (0.5) 0/46 (0) 1/121 (0.8) 0/26 (0) TG11/TG11, 7T/7T 46/193 (23.8) 15/46 (32.6) 23/121 (19.0) 8/26 (30.8) TG11/TG11, 7T/9T 4/193 (2.1) 1/46 (2.2) 3/121 (2.5) 0/26 (0) TG11/TG12, 5T/7T 5/193 (2.6) 0/46 (0) 4/121 (3.3) 1/26 (3.8) TG11/TG12, 6T/7T 1/193 (0.5) 0/46 (0) 1/121 (0.8) 0/26 (0) TG11/TG12, 7T/7T 124/193 (64.2) 27/46 (58.7) 81/121 (66.9) 16/26 (61.5) TG11/TG13, 6T/7T 1/193 (0.5) 1/46 (2.2) 0/121 (0) 0/26 (0) TG11/TG13, 7T/7T 1/193 (0.5) 0/46 (0) 1/121 (0.8) 0/26 (0) TG12/TG12, 7T/7T 6/193 (3.1) 0/46 (0) 6/121 (5.0) 0/26 (0) TG12/TG13, 5T/7T 4/193 (2.1) 2/46 (4.3) 1/121 (0.8) 1/26 (3.8) CFTR p.R75Q wasreportedtoincreaseCPrisk[31].6.5 % of the patients with idiopathic or hereditary CP carried variants in at least two pancreatitis susceptibility genes [32]. Login to comment 151 ABCC7 p.Arg1453Trp ABCC7 p.Val470Met ABCC7 p.Ile556Val ABCC7 p.Ile556Val ABCC7 p.Gln1352His ABCC7 p.Gln1352His ABCC7 p.Gln1352His ABCC7 p.Gln1352His ABCC7 p.Gln1352His ABCC7 p.Leu1156Phe ABCC7 p.Leu1156Phe ABCC7 p.Leu1156Phe ABCC7 p.Leu1156Phe Sequence capture eliminates the necessity of setting up hundreds of PCR, instead allowing for parallel Table 8 Total CFTR sequencing results of patients with SPINK1, PRSS1, CTRC, or CPA1 mutations Case# Etiology CFTR variantsa c.1210-34TG(9_13) c.1210-12T(5_9) SPINK1 PRSS1 CTRC CPA1 B1 Familial p.Q1352H/- TG11/TG12, 7T/7T p.N34S/p.N34S B2 Idiopathic - TG12/TG12, 7T/7T p.N34S/p.N34S B3 Idiopathic - TG11/TG12, 7T/7T p.N34S/p.N34S B4 Idiopathic p.L1156F/-, p.Q1352H/- TG11/TG11, 7T/7T p.N34S/- B5 Idiopathic p.Q1352H/- TG11/TG12, 7T/7T p.N34S/- B6 Idiopathic p.Q1352H/- TG11/TG12, 7T/7T p.N34S/- B7 Idiopathic - TG11/TG12, 7T/7T p.N34S/- B8 Idiopathic - TG11/TG12, 7T/7T p.N34S/- B9 Idiopathic - TG11/TG12, 7T/7T p.N34S/- B10 Idiopathic - TG11/TG12, 7T/7T p.N34S/- B11 Idiopathic - TG11/TG12, 7T/7T p.N34S/- B12 Idiopathic - TG11/TG12, 7T/7T p.N34S/- B13 Idiopathic - TG11/TG12, 7T/7T p.N34S/- B14 Alcoholic - TG12/TG13, 5T/7T p.N34S/- B15 Idiopathic - TG11/TG12, 7T/7T p.N34S/IVS3?2T[C B16 Idiopathic p.R1453W/- TG11/TG11, 7T/7T p.N34S/IVS3?2T[C B17 Idiopathic - TG11/TG12, 7T/7T IVS3?2T[C/IVS3?2T[C B18 Idiopathic - TG11/TG12, 7T/7T IVS3?2T[C/IVS3?2T[C B19 Hereditary p.I125T/-, p.L1156F/- TG11/TG12, 5T/7T IVS3?2T[C/- B20 Familial p.L1156F/- TG11/TG12, 7T/7T IVS3?2T[C/- B21 Idiopathic - TG11/TG12, 7T/7T IVS3?2T[C/- B22 Alcoholic p.Q1352H/- TG11/TG12, 7T/7T IVS3?2T[C/- B23 Alcoholic - TG11/TG12, 7T/7T IVS3?2T[C/- B24 Idiopathic - TG11/TG12, 7T/7T p.P45S/- B25 Idiopathic - TG12/TG12, 7T/7T IVS3?2T[C/- p.R122H/- B26 Hereditary TG11/TG12, 7T/7T p.R122H/- B27 Idiopathic p.I556V/- TG11/TG12, 7T/7T p.N29I/- B28 Idiopathic p.I125T/-, p.L1156F/- TG11/TG12, 7T/7T p.R29Q/- B29 Idiopathic - TG11/TG12, 7T/7T p.T368_Y369ins20/- Nine patients had the non-synonymous CFTR variants, which are probably damaging based on the SIFT or the PolyPhen-2 prediction The p.I556V variant appeared to be benign based on the SIFT or the PolyPhen-2 prediction Case B28 is the same as A5 in Table 6 a We excluded the p.V470M variant from the list because of its similar frequencies in patients and controls enrichment of target regions in a single experiment. 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