Genetic Variations in NER Pathway, Smoking and 11 Cancer Sites: a Pooled Study Yi Ren Wang

Nucleotide Excision Repair Pathway

DNA Damage and Repair  Long-term effects of DNA damage may lead to irreversible mutations, contributing to oncogenesis  Reduced DNA repair capacity may be related to an increased risk of cancers

DNA Repair  Base excision repair  Nucleotide excision repair  Mismatch repair  Double-strand-break repair

NER Pathway  Particularly important mechanism by which the cell can prevent unwanted mutations by removing the vast majority of UV-induced DNA damage  The primary mechanism for the repair of bulky and helical distorting DNA adducts generated by cigarette smoke  Therefore, reduced efficiency of NER could increase susceptibility to DNA damage, and thus increase risk of cancer.

NER Pathway

Study Design

Data Source  Los Angeles Study: Lung: 611, UADT: 601 Controls: 1040  Memorial Sloan Kettering Cancer Center Study: Prostate: 154, Bladder: 245, Kidney: 34, Testes: 32 Controls: 205  TaiXing Study: Liver: 195, Esophagus: 205, Stomach: 196 Controls: 394

Study Design  SAS 9.1 software was used for data analysis.  ORs and 95% CLs was computed using unconditional logistic regression  Potential confounding factors adjusted in pooled analysis: age, gender, ethnicity, tobacco smoking  χ 2 test is performed to evaluate Hardy- Weinberg equilibrium.

11 Cancer Sites  Lung  Oropharynx  Nasopharynx  Larynx  Esophagus  Bladder  Kidney  Liver  Stomach  Prostate  Testes Established association with smoking Association with smoking unknown

Candidate SNPs in NER Pathway GeneSNPData SourceAmino Acid Change XPCrs SNPlex? XPCrs SNPlex&Taqm an ? XPD/ERCC 2 rs238406SNPlexR156R XPD/ERCC 2 rs SNPlexD711D XPD/ERCC 2 rs13181SNPlexK751Q XPD/ERCC 2 rs SNPlexS835S XPG/ERCC 5 rs SNPlexC529S XPG/ERCC 5 rs17655SNPlexD1104H XPG/ERCC 5 rs SNPlexH46H ERCC6rs SNPlex&Taqm an R1230P ERCC6rs SNPlex&Taqm an Q1413R ERCC1rs SNPlex? Overall Kappa between SNPlex & Tapman = , indicating almost perfect agreement. 95% CI = (0.9446, )

Results

No deviation from Hardy-Weinberg equilibrium in controls detected SNPGENOTYPEControl (%) P ERCC1 rs G/G1121 ( 96.6%) A/G38 ( 3.3%) A/A2 ( 0.2%) ERCC6 rs A/A991 ( 69.8%) A/G386 ( 27.2%) G/G42 ( 3.0%) ERCC6 rs G/G1276 ( 88.0%) C/G162 ( 11.2%) C/C12 ( 0.8%) XPC rs C/G55 ( 53.9%) C/C33 ( 32.4%) G/G14 ( 13.7%) XPC rs A/A929 ( 77.2%) A/T237 ( 19.7%) T/T38 ( 3.2%) XPD/ERCC2 rs C/C432 ( 51.2%) C/T368 ( 43.7%) T/T43 ( 5.1%) XPD/ERCC2 rs13181T/T460 ( 43.9%) G/T448 ( 42.7%) G/G140 ( 13.4%) XPD/ERCC2 rs T/T535 ( 50.2%) C/T441 ( 41.4%) C/C90 ( 8.4%) XPD/ERCC2 rs238406G/T471 ( 59.8%) G/G195 ( 24.7%) T/T122 ( 15.5%) XPG/ERCC5 rs C/T525 ( 47.0%) T/T312 ( 27.9%) C/C280 ( 25.1%) XPG/ERCC5 rs17655C/G420 ( 46.7%) C/C337 ( 37.5%) G/G142 ( 15.8%) XPG/ERCC5 rs G/G801 ( 79.0%) C/G203 ( 20.0%) C/C10 ( 1.0%)

Pooled association analysis of NER SNPs with 11 cancer sites All Population SNP GENO TYPECase (%)Control (%)COR (95% CI)AOR (95% CI)SNP GENO TYPECase (%)Control (%)COR (95% CI)AOR (95% CI) XPCC/G75 ( 46.6%)55 ( 53.9%)ERCC1G/G1164 ( 96.8%)1121 ( 96.6%) rs C/C61 ( 37.9%)33 ( 32.4%)1.36 ( 0.78, 2.35)1.34 ( 0.76, 2.35) rs A/G36 ( 3.0%)38 ( 3.3%)0.91 ( 0.57, 1.45)0.53 ( 0.31, 0.88) G/G25 ( 15.5%)14 ( 13.7%)1.31 ( 0.62, 2.75)1.41 ( 0.64, 3.10)A/A3 ( 0.2%)2 ( 0.2%)1.44 ( 0.24, 8.66)2.01 ( 0.26,15.82) XPCA/A1081 ( 78.9%)929 ( 77.2%)ERCC6A/A1178 ( 74.6%)991 ( 69.8%) rs A/T258 ( 18.8%)237 ( 19.7%)0.94 ( 0.77, 1.14)1.02 ( 0.81, 1.29) rs A/G342 ( 21.6%)386 ( 27.2%)0.75 ( 0.63, 0.88)0.89 ( 0.74, 1.07) T/T31 ( 2.3%)38 ( 3.2%)0.70 ( 0.43, 1.14)0.83 ( 0.49, 1.41)G/G60 ( 3.8%)42 ( 3.0%)1.20 ( 0.80, 1.80)1.45 ( 0.93, 2.24) XPD/ERCC2C/C526 ( 55.0%)432 ( 51.2%)ERCC6G/G1404 ( 86.6%)1276 ( 88.0%) rs C/T378 ( 39.5%)368 ( 43.7%)0.84 ( 0.70, 1.02)0.91 ( 0.73, 1.13) rs C/G199 ( 12.3%)162 ( 11.2%)1.12 ( 0.90, 1.39)1.03 ( 0.81, 1.31) T/T52 ( 5.4%)43 ( 5.1%)0.99 ( 0.65, 1.52)1.09 ( 0.68, 1.76)C/C18 ( 1.1%)12 ( 0.8%)1.36 ( 0.65, 2.84)1.12 ( 0.50, 2.50) XPD/ERCC2T/T412 ( 45.7%)460 ( 43.9%)XPG/ERCC5C/T557 ( 45.0%)525 ( 47.0%) rs13181G/T374 ( 41.5%)448 ( 42.7%)0.93 ( 0.77, 1.13)0.96 ( 0.77, 1.19) rs T/T430 ( 34.7%)312 ( 27.9%)1.30 ( 1.08, 1.57)1.11 ( 0.90, 1.37) G/G115 ( 12.8%)140 ( 13.4%)0.92 ( 0.69, 1.21)0.88 ( 0.64, 1.21)C/C251 ( 20.3%)280 ( 25.1%)0.84 ( 0.69, 1.04)0.87 ( 0.69, 1.09) XPD/ERCC2T/T449 ( 49.0%)535 ( 50.2%)XPG/ERCC5C/G507 ( 48.5%)420 ( 46.7%) rs C/T402 ( 43.8%)441 ( 41.4%)1.09 ( 0.90, 1.31)1.06 ( 0.86, 1.30) rs17655C/C368 ( 35.2%)337 ( 37.5%)0.90 ( 0.74, 1.10)0.98 ( 0.79, 1.22) C/C66 ( 7.2%)90 ( 8.4%)0.87 ( 0.62, 1.23)0.83 ( 0.56, 1.21)G/G171 ( 16.3%)142 ( 15.8%)1.00 ( 0.77, 1.29)1.01 ( 0.76, 1.33) XPD/ERCC2G/T539 ( 59.2%)471 ( 59.8%)XPG/ERCC5G/G931 ( 82.1%)801 ( 79.0%) rs238406G/G253 ( 27.8%)195 ( 24.7%)1.13 ( 0.91, 1.42)1.08 ( 0.85, 1.38) rs C/G158 ( 13.9%)203 ( 20.0%)0.67 ( 0.53, 0.84)0.66 ( 0.51, 0.84) T/T118 ( 13.0%)122 ( 15.5%)0.85 ( 0.64, 1.12)0.81 ( 0.60, 1.11)C/C45 ( 4.0%)10 ( 1.0%)3.87 ( 1.94, 7.73)3.10 ( 1.44, 6.66)

Pooled association analysis stratified by smoking: Ever Smoker Ever Smoker SNP GENO TYPECase (%)Control (%)COR (95% CI)AOR (95% CI)SNP GENO TYPECase (%)Control (%)COR (95% CI)AOR (95% CI) XPCC/G 39 ( 47.0%)27 ( 61.4%) ERCC1G/G 788 ( 95.7%)554 ( 96.5%) rs C/C 39 ( 47.0%)12 ( 27.3%)2.25 ( 1.00, 5.07)2.12 ( 0.93, 4.82) rs A/G 32 ( 3.9%)20 ( 3.5%)1.12 ( 0.64, 1.99)0.71 ( 0.38, 1.33) G/G 5 ( 6.0%)5 ( 11.4%)0.69 ( 0.18, 2.63)0.65 ( 0.17, 2.53) A/A 3 ( 0.4%)0 ( 0.0%) XPCA/A 735 ( 75.3%)458 ( 75.0%) ERCC6A/A 803 ( 72.9%)510 ( 70.3%) rs A/T 213 ( 21.8%)132 ( 21.6%)1.01 ( 0.79, 1.29)1.01 ( 0.76, 1.33) rs A/G 252 ( 22.9%)197 ( 27.2%)0.81 ( 0.65, 1.01)0.85 ( 0.67, 1.07) T/T 28 ( 2.9%)21 ( 3.4%)0.83 ( 0.47, 1.48)0.89 ( 0.48, 1.64) G/G 47 ( 4.3%)18 ( 2.5%)1.66 ( 0.95, 2.89)1.77 ( 0.99, 3.16) XPD/ERCC2C/C 338 ( 53.4%)209 ( 51.5%) ERCC6G/G 967 ( 85.4%)628 ( 85.6%) rs C/T 254 ( 40.1%)174 ( 42.9%)0.90 ( 0.70, 1.17)0.78 ( 0.59, 1.04) rs C/G 152 ( 13.4%)99 ( 13.5%)1.00 ( 0.76, 1.31)0.90 ( 0.68, 1.20) T/T 41 ( 6.5%)23 ( 5.7%)1.10 ( 0.64, 1.89)0.92 ( 0.51, 1.67) C/C 13 ( 1.1%)7 ( 1.0%)1.21 ( 0.48, 3.04)0.94 ( 0.34, 2.55) XPD/ERCC2T/T 320 ( 44.6%)230 ( 43.2%) XPG/ERCC5C/T 395 ( 47.1%)258 ( 46.7%) rs13181G/T 303 ( 42.2%)234 ( 43.9%)0.93 ( 0.73, 1.18)0.93 ( 0.72, 1.21) rs T/T 263 ( 31.4%)150 ( 27.2%)1.15 ( 0.89, 1.48)1.12 ( 0.86, 1.48) G/G 95 ( 13.2%)69 ( 12.9%)0.99 ( 0.70, 1.41)0.86 ( 0.59, 1.26) C/C 180 ( 21.5%)144 ( 26.1%)0.82 ( 0.62, 1.07)0.74 ( 0.56, 0.99) XPD/ERCC2T/T 358 ( 49.2%)258 ( 47.6%) XPG/ERCC5C/G 330 ( 47.2%)206 ( 46.7%) rs C/T 320 ( 44.0%)240 ( 44.3%)0.96 ( 0.76, 1.21)0.93 ( 0.73, 1.20) rs17655C/C 266 ( 38.1%)177 ( 40.1%)0.94 ( 0.72, 1.21)0.98 ( 0.75, 1.30) C/C 50 ( 6.9%)44 ( 8.1%)0.82 ( 0.53, 1.27)0.73 ( 0.46, 1.17) G/G 103 ( 14.7%)58 ( 13.2%)1.11 ( 0.77, 1.60)1.19 ( 0.81, 1.75) XPD/ERCC2G/T 348 ( 57.9%)225 ( 59.7%) XPG/ERCC5G/G 619 ( 81.3%)393 ( 79.2%) rs238406G/G 177 ( 29.5%)88 ( 23.3%)1.30 ( 0.96, 1.77)1.30 ( 0.94, 1.80) rs C/G 113 ( 14.8%)102 ( 20.6%)0.70 ( 0.52, 0.95)0.66 ( 0.48, 0.90) T/T 76 ( 12.6%)64 ( 17.0%)0.77 ( 0.53, 1.11)0.79 ( 0.54, 1.17) C/C 29 ( 3.8%)1 ( 0.2%)18.41 ( 2.50,. )9.62 ( 1.27,72.68)

Pooled association analysis stratified by smoking: Never Smoker Never Smoker SNP GENO TYPECase (%)Control (%)COR (95% CI)AOR (95% CI)SNP GENO TYPECase (%)Control (%)COR (95% CI)AOR (95% CI) XPCC/G 34 ( 47.9%)28 ( 49.1%) ERCC1G/G 352 ( 98.9%)565 ( 96.6%) rs C/C 19 ( 26.8%)21 ( 36.8%)0.75 ( 0.34, 1.65)0.79 ( 0.35, 1.78) rs A/G 4 ( 1.1%)18 ( 3.1%)0.36 ( 0.12, 1.06)0.22 ( 0.07, 0.68) G/G 18 ( 25.4%)8 ( 14.0%)1.85 ( 0.70, 4.89)1.70 ( 0.63, 4.60) A/A 0 ( 0.0%)2 ( 0.3%) XPCA/A 326 ( 87.2%)469 ( 79.6%) ERCC6A/A 352 ( 77.9%)478 ( 69.3%) rs A/T 45 ( 12.0%)103 ( 17.5%)0.63 ( 0.43, 0.92)1.06 ( 0.67, 1.68) rs A/G 88 ( 19.5%)188 ( 27.2%)0.64 ( 0.48, 0.85)0.98 ( 0.70, 1.37) T/T 3 ( 0.8%)17 ( 2.9%)0.25 ( 0.07, 0.87)0.49 ( 0.13, 1.87) G/G 12 ( 2.7%)24 ( 3.5%)0.68 ( 0.33, 1.38)1.01 ( 0.46, 2.22) XPD/ERCC2C/C 171 ( 57.0%)221 ( 51.0%) ERCC6G/G 415 ( 89.4%)645 ( 90.6%) rs C/T 118 ( 39.3%)192 ( 44.3%)0.79 ( 0.59, 1.08)1.06 ( 0.75, 1.52) rs C/G 44 ( 9.5%)62 ( 8.7%)1.10 ( 0.74, 1.65)1.36 ( 0.85, 2.18) T/T 11 ( 3.7%)20 ( 4.6%)0.71 ( 0.33, 1.52)1.50 ( 0.60, 3.76) C/C 5 ( 1.1%)5 ( 0.7%)1.55 ( 0.45, 5.40)1.99 ( 0.49, 8.06) XPD/ERCC2T/T 92 ( 50.5%)229 ( 44.7%) XPG/ERCC5C/T 154 ( 41.0%)265 ( 47.2%) rs13181G/T 70 ( 38.5%)212 ( 41.4%)0.82 ( 0.57, 1.18)0.98 ( 0.65, 1.47) rs T/T 155 ( 41.2%)161 ( 28.6%)1.66 ( 1.23, 2.23)1.05 ( 0.75, 1.47) G/G 20 ( 11.0%)71 ( 13.9%)0.70 ( 0.40, 1.22)0.79 ( 0.42, 1.48) C/C 67 ( 17.8%)136 ( 24.2%)0.85 ( 0.60, 1.21)1.17 ( 0.77, 1.76) XPD/ERCC2T/T 90 ( 47.9%)276 ( 53.0%) XPG/ERCC5C/G 164 ( 50.6%)212 ( 46.6%) rs C/T 82 ( 43.6%)200 ( 38.4%)1.26 ( 0.89, 1.78)1.42 ( 0.95, 2.11) rs17655C/C 97 ( 29.9%)159 ( 34.9%)0.79 ( 0.57, 1.09)1.04 ( 0.72, 1.50) C/C 16 ( 8.5%)45 ( 8.6%)1.09 ( 0.59, 2.02)1.07 ( 0.53, 2.16) G/G 63 ( 19.4%)84 ( 18.5%)0.97 ( 0.66, 1.42)0.78 ( 0.52, 1.19) XPD/ERCC2G/T 178 ( 62.5%)243 ( 59.7%) XPG/ERCC5G/G 292 ( 83.9%)405 ( 78.6%) rs238406G/G 70 ( 24.6%)107 ( 26.3%)0.89 ( 0.62, 1.28)0.75 ( 0.51, 1.12) rs C/G 41 ( 11.8%)101 ( 19.6%)0.56 ( 0.38, 0.83)0.66 ( 0.43, 1.02) T/T 37 ( 13.0%)57 ( 14.0%)0.89 ( 0.56, 1.40)0.90 ( 0.55, 1.49) C/C 15 ( 4.3%)9 ( 1.7%)2.31 ( 1.00, 5.35)2.39 ( 0.91, 6.24)

Selected association of NER SNPs with cancer, stratified by cancer site. Cancer SiteSNPGENOTYPECase (%)Control (%)Crude OR (95% CI)Adjusted OR (95% CI) Lung Cancer ERCC6 rs A/A362 ( 69.1%)580 ( 64.5%) A/G132 ( 25.2%)288 ( 32.0%)0.73 ( 0.58, 0.94)0.84 ( 0.64, 1.10) G/G30 ( 5.7%)31 ( 3.4%)1.55 ( 0.92, 2.61)1.93 ( 1.09, 3.41) XPG/ERCC5 rs C/T169 ( 51.2%)285 ( 47.0%) T/T62 ( 18.8%)103 ( 17.0%)1.02 ( 0.70, 1.47)1.00 ( 0.66, 1.53) C/C99 ( 30.0%)218 ( 36.0%)0.77 ( 0.56, 1.04)0.66 ( 0.47, 0.92) Bladder Cancer ERCC1 rs G/G104 ( 85.2%)86 ( 78.9%) A/G17 ( 13.9%)23 ( 21.1%)0.61 ( 0.31, 1.22)0.35 ( 0.13, 0.95) A/A1 ( 0.8%)0 ( 0.0%) XPD/ERCC2 rs238406G/T80 ( 57.6%)81 ( 62.3%) G/G43 ( 30.9%)30 ( 23.1%)1.45 ( 0.83, 2.54)2.70 ( 1.10, 6.65) T/T16 ( 11.5%)19 ( 14.6%)0.85 ( 0.41, 1.78)3.61 ( 0.94,13.88) XPG/ERCC5 rs17655C/G59 ( 41.3%)81 ( 62.8%) C/C76 ( 53.1%)41 ( 31.8%)2.54 ( 1.53, 4.22)4.56 ( 1.92,10.84) G/G8 ( 5.6%)7 ( 5.4%)1.57 ( 0.54, 4.57)2.67 ( 0.54,13.18) Kidney Cancer XPG/ERCC5 rs17655C/G7 ( 46.7%)81 ( 62.8%) C/C8 ( 53.3%)41 ( 31.8%)2.26 ( 0.77, 6.66)4.38 ( 1.08,17.79) G/G0 ( 0.0%)7 ( 5.4%) Esophagus Cancer XPD/ERCC2 rs13181T/T30 ( 54.5%)411 ( 45.1%) G/T24 ( 43.6%)383 ( 42.0%)0.86 ( 0.49, 1.49)0.72 ( 0.40, 1.28) G/G1 ( 1.8%)118 ( 12.9%)0.12 ( 0.02, 0.86)0.10 ( 0.01, 0.74) Stomach Cancer ERCC6 rs G/G181 ( 94.3%)342 ( 90.5%) C/G8 ( 4.2%)34 ( 9.0%)0.44 ( 0.20, 0.98)0.37 ( 0.16, 0.87) C/C3 ( 1.6%)2 ( 0.5%)2.83 ( 0.47,17.12)3.18 ( 0.51,19.92) XPG/ERCC5 rs G/G165 ( 93.2%)323 ( 84.3%) C/G12 ( 6.8%)57 ( 14.9%)0.41 ( 0.22, 0.79)0.36 ( 0.18, 0.73) C/C0 ( 0.0%)3 ( 0.8%)

Previous Published Results on XPG/ERCC5 rs VS. Stomach Cancer  Shehnaz K. Hussain, et al. Genetic variation in immune regulation and DNA repair pathways and stomach cancer in China

Pooled association analysis of XPG/ERCC5 rs WITHOUT stomach cancer SNPGENOTYPECase (%)Control (%)COR (95% CI)AOR (95% CI) XPG/ERCC5G/G 766 ( 80.0%)801 ( 79.0%) rs C/G 146 ( 15.3%)203 ( 20.0%)0.75 ( 0.59, 0.95)0.70 ( 0.54, 0.91) C/C 45 ( 4.7%)10 ( 1.0%)4.71 ( 2.35, 9.40)3.31 ( 1.54, 7.10)

Haplotype Analysis of NER SNPs with combined cancer sites GENESNPHAPLOTYPEFrequency (%)Crude OR (95% CI)Adjusted OR (95% CI) XPC rs C-A60.20% 0.80 ( 0.64, 0.99) 1.05 ( 0.82, 1.34) rs G-A39.80% 0.77 ( 0.57, 1.06) 1.10 ( 0.77, 1.56) XPD/ERCC2 rs238406G-C-T-C6.49% 1.26 ( 0.61, 2.59) 1.42 ( 0.65, 3.11) rs G-C-T-T19.91% 0.92 ( 0.65, 1.31) 1.11 ( 0.75, 1.63) rs13181G-T-G-C6.72% 1.61 ( 0.90, 2.88) 1.16 ( 0.61, 2.19) rs G-T-G-T13.59% 1.82 ( 1.25, 2.66) 1.28 ( 0.84, 1.95) T-C-T-C10.43% 1.66 ( 0.99, 2.81) 1.66 ( 0.94, 2.94) T-C-T-T25.73% 1.76 ( 1.27, 2.43) 1.87 ( 1.31, 2.67) XPG/ERCC5 rs C-G-C6.63% 2.88 ( 1.66, 5.01) 3.28 ( 1.80, 6.00) rs17655G-C-C33.52% 1.31 ( 1.05, 1.62) 1.20 ( 0.95, 1.52) rs G-C-T23.03% 0.98 ( 0.75, 1.28) 1.28 ( 0.95, 1.74) G-G-T28.16% 0.73 ( 0.58, 0.93) 0.95 ( 0.72, 1.26) ERCC6 rs A-C6.67% 0.77 ( 0.52, 1.14) 0.94 ( 0.61, 1.43) rs A-G77.38% 0.88 ( 0.73, 1.06) 1.01 ( 0.82, 1.26) G-G15.77% 1.29 ( 0.99, 1.69) 1.02 ( 0.75, 1.37)

Strengths  Large sample size  Multiple SNPs analyzed  Increased statistical power from pooled analysis to detect modest effects  Population-based study design  Detailed questionnaire data

Limitations  Recall bias  Misclassification of genotype data  Validity of the pooled estimate can be threatened by the heterogeneity of the studies (study populations, study design, genotyping methodology etc.)  Bias from population stratification or population mixing due to mixed ethnic ancestry

Comparison of esophagus cancer results between Los Angeles Study and TaiXing Study Los Angeles StudyTaiXing Study SNP GEN OTYP E Case (%)Control (%)Adjusted OR (95% CI)Case (%)Control (%)Adjusted OR (95% CI) ERCC6 rs A/G20 ( 33.9%)288 ( 32.0%)1.10 ( 0.62, 1.95)22 ( 11.2%)45 ( 12.1%)0.89 ( 0.51, 1.57) XPD/ERCC2 rs C/C13 ( 50.0%)128 ( 38.2%)1.56 ( 0.67, 3.59)40 ( 27.2%)109 ( 29.1%)0.98 ( 0.63, 1.54) XPG/ERCC5 rs T/T10 ( 25.0%)103 ( 17.0%)1.21 ( 0.54, 2.71)18 ( 9.6%)28 ( 7.5%)1.04 ( 0.53, 2.02) XPG/ERCC5 rs17655C/G12 ( 42.9%)150 ( 38.2%)1.85 ( 0.75, 4.57)43 ( 25.4%)93 ( 24.7%)1.03 ( 0.65, 1.64) XPG/ERCC5 rs17655G/G7 ( 25.0%)40 ( 10.2%)4.36 ( 1.34,14.22)48 ( 28.4%)95 ( 25.2%)1.32 ( 0.84, 2.10) XPG/ERCC5 rs C/G8 ( 21.1%)108 ( 21.5%)0.94 ( 0.41, 2.17)19 ( 10.6%)57 ( 14.9%)0.65 ( 0.36, 1.16)