In this case-control study, we evaluated associations of 240 common SNPs in genes involved in NER, BER or cell cycle control with risk of smoking-related lung cancer. Thirty-eight SNPs, 24 in NER, 10 in BER and four in cell cycle control genes, were associated with lung cancer risk at P<0.05 in our study population. The strongest associations with risk of lung cancer were observed for rs2074508 in GTF2H4, rs10500298 in LIG1, rs747658 and rs3219073 in PARP1, and rs1799782 and rs3213255 in XRCC1. In addition, haplotypes in six different genes (ERCC2, GTF2H1, GTF2H4, PARP1, XRCC1 and CCND1) were found associated with lung cancer risk.
codes for the p52 subunit of NER transcription factor IIH (TFIIH). The TFIIH complex has ATPase and helicase activities and opens DNA at sites of DNA distorting damage. The p52 subunit may regulate the ATPase activity of the TFIIH subunit (XPB), a protein coded by ERCC3
]. In our study, single SNP analyses found the minor allele of rs2074508 in GTF2H4
significantly associated with a reduced risk of lung cancer (additive model). The common GTF2H4
haplotype associated with reduced lung cancer risk captured this minor allele. Wang et al
] recently reported associations between cervical dysplasia and two SNPs near GTF2H4
, rs2894054, located 3.7 kb upstream, and rs6926723, located 0.9 kb downstream (within valyl-tRNA synthetase, VARSL
) from GTF2H4
. Published studies of GTF2H4
genetic variation and cancer risk appear otherwise not to exist [19
]. Providing a first comprehensive survey of common variation in GTF2H4
and lung cancer susceptibility, the GTF2H4
SNPs evaluated in our study captured 9 (90%) of the 10 HapMap phase 1 & 2 common-variant (MAF≥0.05) CEU SNPs.
DNA ligase I (encoded by LIG1
) joins Okazaki fragments in the lagging strand during DNA synthesis and completes BER and NER [20
]. In our study, case and control genotype distributions differed significantly for one of the 21 examined LIG1
rs10500298 minor allele homozygotes were more common among lung cancer cases than controls and more common among adenocarcinomas than squamous cell carcinomas. A literature search identified five published studies on lung cancer risk and LIG1
genetic variability [study #1: one SNP, 530 and 570 non-Hispanic white cases and controls [21
]; study #2, 34 SNPs, 143 and 172 French Caucasian cases and controls [22
]; study #3: four SNPs, ~440 and ~790 mixed race (~60% Caucasian) cases and controls [23
]; study #4: five SNPs, 113 and 299 Latino American cases and controls [24
], and study #5: five SNPs, 255 and 280 African American cases and controls [24
]]. The French Caucasian, mixed race, and African American studies reported significant lung cancer associations involving either single LIG1
SNPs and/or haplotypes. Our SNP panel included two SNPs (rs20581, rs20579) associated with risk of lung cancer in the mixed race study. However, neither SNP was found to be associated with lung cancer risk in our study. SNP rs10500298 was not in strong linkage disequilibrium with a third mixed race lung cancer risk-associated SNP (rs20580) nor with three French Caucasian lung cancer risk-associated SNPs (rs3730994, rs3786763, rs3730912).
Upon detection of DNA strand breaks, poly (ADP-ribose) polymerase 1 (PARP1) adds poly(ADP-ribose) to nuclear proteins, recruits XRCC1, and initiates BER [25
]. Interestingly, inhibition of PARP1 creates a state of synthetic lethality in cells that are unable to complete homologous recombination as a result of BRCA1
loss and PARP1 inhibitors show promise in treating BRCA-deficient breast and ovarian cancer [25
]. In our study, the minor alleles of two PARP1
SNPs, rs747658 and rs3219073 (in high linkage disequilibrium), reduced lung cancer risk, per minor allele, by one third. Consistent with this, the relatively common PARP1
haplotype that contained the minor alleles from these two SNPs was strongly statistically significantly associated with a lower risk of lung cancer as well. Thus far, only a few studies on PARP1
variability and lung cancer risk have been published. Among Japanese, having at least one minor allele of SNP rs3219145 (Lys940Arg), a SNP absent in HapMap CEU whites and not evaluated by us, was associated with increased lung cancer risk (OR 1.40, 95% CI 1.04–1.90) [27
]. Lockett et al
] reported that poly (ADP-ribose) polymerase activity is lower in lymphocytes collected from individuals with at least one 762Ala allele (Val762Ala; rs1136410) and in vitro experiments confirmed lower enzymatic activity of PARP1-Ala762 [29
]. Consistent with this, among Han Chinese, having at least one minor allele of SNP rs1136410 was associated with an increased lung cancer risk (OR 1.26, 95% CI 1.05–1.52) [30
]. However, no significant association was observed in Korean [31
] and Japanese study populations [27
], and in our study population the association was not statistically significant either (OR 1.18, 95% CI 0.96–1.46). The PARP1
SNPs evaluated in our study captured 46 (96%) of 48 phase 1 & 2 common-variant HapMap CEU SNPs.
Commonly described as a scaffold protein, XRCC1 may coordinate BER through interactions not only with PARP1, but also with DNA ligase III (LIG3), DNA polymerase β (POLB), apurinic/apyrimidinic (AP) nuclease (APEX1), and polynucleotide kinase 3′-phosphatase (PNKP) [32
]. In our study population, two SNPs in XRCC1
, rs1799782 (Arg194Trp) and rs3213255, were strongly associated with risk of lung cancer. Many published studies on XRCC1
variability and lung cancer risk have evaluated rs1799782 and two other non-synonymous SNPs, rs25489 (Arg280His) and rs25487 (Arg399Gln) (for example, [33
]). However, four recent meta-analyses, Kiyohara et al
], Wang et al
], Zheng et al
], and Vineis et al
], did not find any one of these three commonly studied non-synonymous coding SNPs associated with lung cancer risk in whites. In analyses adjusted for age and restricted to smokers with high pack-year exposures, a large white-only case-control study [34
] also observed significantly lower lung cancer risk in association with Trp194 [34
]. Therefore, our observation with respect to rs1799782 may reflect a study population enriched with heavy smokers combined with a Trp194 lung cancer protective effect that depends on a history of heavy smoking. The XRCC1
SNPs evaluated in our study captured 18 (75%) of 24 phase 1 & 2 common-variant (MAF≥0.05) HapMap CEU SNPs.
The effect of genetic variation in DNA repair and cell cycle control genes on lung cancer risk may become detectable only in the presence of certain environmental factors that cause DNA damage such as cigarette smoking. Our study population consisted of former and current smokers only and different results may be observed among never smokers. It should also be noted that multiple testing may have led to chance findings and that associations will need to be confirmed in additional studies. However, SNPs rs747658 and rs3219073 in PARP1 did remain associated with lung cancer risk at P<0.05 after applying the conservative Bonferroni correction.
Although our study population was relatively large, it is possible that some associations were not detected due to insufficient power. Specifically, testing under a log-additive model at a 0.05 two-sided significance level, our case-control study (722 cases and 929 controls) had under 90% power for very common variants (population MAF≥0.25) with lung cancer effects (OR) less than 1.3 and for common variants (population MAF=0.05) with lung cancer effects (OR) less than 1.6 [40
]. Haplotype-based analyses may have improved our power for detecting genes with lung cancer association involving uncommon or multiple susceptibility alleles [41
]. Evaluations of gene-environment interactions and case-case comparisons were limited to the 38 SNPs associated with lung cancer risk at P
<0.05 and it is possible that important interactions and/or associations were not identified due to the limited number of SNPs investigated.
Unfortunately, our study population contained only a relatively small number of non-white subjects which limited our ability to evaluate SNP-lung cancer risk associations in non-white race groups. However, using 45 case and 58 control subjects with African genetic ancestry, we observed that four SNPs (ERCC3 rs4150407, LIG1 rs175628, RPA2 rs7356, and RPA3 rs13246995) were statistically significantly (P<0.05) associated with lung cancer risk, and replicated directional associations observed in subjects with Caucasian genetic ancestry.
Contributing to an expanding knowledge base about the role of genetic variation in DNA repair and cell cycle pathway genes in lung carcinogenesis, our findings may help identify biological processes behind lung cancer susceptibility. However, knowing the genotype of a single common SNP can not be used usefully to assign smokers to very high or low risk groups. This limitation motivates the search for multi-SNP indices capable of stratifying smokers into more meaningful risk groups [42
]. As an illustration of one uncomplicated approach [42
], we formed a 31-SNP genotype risk summary score that identified second, third, and fourth quartile groups at 1.04-fold, 2.21-fold, and 3.44-fold increased risk, respectively, relative to a first, lowest risk, quartile reference group.
To conclude, our data suggests that common variation in DNA repair and cell cycle control pathway genes is associated with smoking-related risk of lung cancer. Specifically, we observed strong associations with lung cancer risk for SNPs in GTF2H4, LIG1, PARP1 and XRCC1. An illustrative genotype risk summary score that combined genotype information for 31 SNPs in 15 genes risk-stratified current and ex-cigarette smokers over a 3.4-fold range. If confirmed in additional studies, combining genotype information for SNPs known to be associated with lung cancer risk may assist in classifying current and former smokers according to risk.