DNA repair has long been implicated in colorectal cancer with the discovery that germ line mutations in MMR genes lead to HNPCC (29
) and mutations in the base excision repair gene, MUTYH
, lead to a familial polyposis syndrome. However, the etiological role of common genetic variation in DNA repair genes in colorectal adenoma and cancer has not been comprehensively studied in the context of epidemiological studies. Although some common SNPs in DNA repair genes have been reported to be associated with colorectal cancer and/or adenoma (18
), with the exception of the MHL1
-93G > A variant with microsatellite instable tumors (18
), most associations have not been replicated (30
). Furthermore, data on effect modifications by important environmental factors are sparse.
In our study, >3000 SNPs from 153 DNA repair genes were evaluated simultaneously among 2841 study subjects, which is the largest and most comprehensive study for colorectal adenoma risk focusing on DNA repair genes to date. Among the SNPs associated with risk, we found that genetic polymorphisms in EXO1 and ATM significantly modified the effect of cigarette smoking on risk, predisposing smokers to greater adenoma susceptibility. When stratified by genotype, smoking was only significantly associated with increased adenoma risk among individuals homozygous for the risk allele at EXO1 rs9350 (OR = 1.80, 95% CI = 1.47–2.19 for ever smokers with the CC genotype) and ATM rs17503908 (OR = 1.69, 95% CI = 1.40–2.03 for ever smokers with the GG genotype).
, located at chromosome 1q42–q43, encodes a protein with 5′→ 3′ and 3′→ 5′ double-stranded DNA exonuclease activity. It also exhibits some endonuclease activity correcting 5′-overhanging flap structures. EXO1
is involved in DNA MMR, recombination, replication and telomere stabilization (32
-mutant cells showed increased microsatellite instability and incomplete MMR capability (33
). Mice with EXO1
knockout were found to have lower survival rates and higher mutation rates as well as higher susceptibility to lymphomas (33
has been implicated in hereditary HNPCC due to its role in DNA MMR; however, studies investigating rare germ line variants in EXO1
have not shown consistent findings as reviewed by Liberti et.al.
We observed an increased colorectal adenoma risk among individuals carrying a C allele at rs9350 and a G allele at rs4658535 in EXO1
. The SNPs were highly correlated (r2
= 0.82), making it impossible to differentiate the associations of each statistically. Using the PolyPhen database, we found that rs9350 was predicted to be ‘probably damaging’ (position-specific independent counts score difference = 2.17) with the C to T substitution causing a non-synonymous amino acid change by replacing proline with leucine (35
), suggesting that rs9350 may be a causal variant. The substitution was also predicted to be ‘deleterious’ using the SIFT database. Several studies have examined the association between common polymorphisms in EXO1
and the risk of lung, oral, brain and colorectal cancer (36
); however, data are limited and inconclusive due to the small sample size and differences in the SNPs genotyped in the studies (36
). No studies have examined the association between this polymorphism and adenoma. Consistent with our findings, two case–control studies of colorectal cancer found a decreased cancer risk for individuals carrying T
allele at EXO1
rs9350 compared with C
). No association was observed with rs9350 in two studies of lung cancer (37
), one study of oral cancer (39
) and one study of breast cancer (44
), suggesting that the association of C allele at rs9350 may be organ/tissue specific. In addition, the previous studies in other cancers (37
) were relatively small (N
≤ 680 cases each) and conducted in Asian populations, where differences in environmental exposures may modify the association of rs9350 and cancer risk compared with Caucasians.
Smoking is an important risk factor for adenoma with current smokers having an 1.8-fold increased risk (95% CI = 1.5–2.1) in the full PLCO cohort (45
). We observed a stronger association between adenoma risk and rs9350 in EXO1
among smokers compared with non-smokers and hypothesize that the C allele at EXO1
rs9350 may increase risk among smokers by reducing the protein’s capacity or efficiency to repair the damage caused by smoking exposure. Tsai et al.
also reported an increased risk of oral cancer among smokers who carried the A allele at rs1047840 (r2
= 0.18 with rs9350 in our study) in EXO1
but not among non-smokers (39
). The haplotype results further confirmed the strong association between the EXO1
region encompassing rs9350 and adenoma risk.
We observed a significant decreased risk among never smokers for the T allele at rs17503908 in ATM
, located at chromosome 11q22.3, encodes a cell cycle checkpoint kinase which regulates many downstream proteins, including the tumor suppressor proteins p53 and BRCA1, the checkpoint kinase CHK2, checkpoint proteins RAD17 and RAD9 and the DNA repair protein NBS1. ATM is thought to be a master controller of the cell cycle checkpoint-signaling pathways and functions to repair DNA damage and maintain genome stability. Persons with ataxia telangiectasia, an autosomal recessive diseased caused by rare missense or truncating mutations in ATM
, have an increased sensitivity to ionizing radiation and an increased risk of cancer (46
). Heterozygous carriers of these rare ATM mutations have an increased risk of several cancers including colorectal cancer (47
Several lines of evidence have suggested the etiological role for ATM
in colorectal carcinogenesis (47
). Polymorphisms at rs1800056 and rs1800057 in ATM
have been associated with colorectal cancer risk (49
) and although the results were not replicated in a follow-up study (50
), rs1801516 has been associated with disease penetrance among HNPCC carriers (51
). In our study, the T allele at ATM
rs1801516 was also marginally associated with adenoma risk (P
= 0.011 for Caucasians and P
= 0.016 for all subjects), but no association was observed with rs1800056 (P
= 0.22). To date, no study has reported an association between ATM
rs17503908 and colorectal adenoma, which is located in an intronic region of ATM
. We observed an inverse association for the T allele at rs17503908. Differences in sensitivity to DNA damage or higher expression levels of ATM
could reduce risk for neoplastic transformation or subsequent proliferation by activating p53 (52
). In stratified analyses, this inverse association was restricted to never smokers. It is possible that smokers do not benefit from carrying this allele due to an antagonistic effect between the SNP and smoking exposure. An in vitro
study observed that smoking exposure activated ATM in human pulmonary adenocarcinoma cells through phosphorylation in a dose-dependent manner (53
). Similarly, benzo[a]pyrene diol epoxide, a polycyclic aromatic hydrocarbon found in tobacco smoke, has been shown to bind to ATM
) and induce ATM expression in esophageal cancer cell lines (55
), suggesting that ATM plays an active role in responding to tobacco smoke exposure. We speculate that the kinase encoded by the ATM
may be saturated by smoking exposure, which may prevent its protective effect.
Our study has several advantages and limitations. First, it is the largest study to date to investigate a broad range of DNA repair gene polymorphisms for colorectal adenoma risk. Although still somewhat underpowered to examine gene–environment interactions for the SNPs of moderate association, our findings provide promising leads for replication in future pooled analyses. Future analyses exploring these interactions with regard to colorectal cancer may lead to additional insight into colorectal neoplasm progression. Our study included only advanced adenoma cases and so the results may not be generalizable to non-advanced adenomatous polyps; however, advanced adenomas are more to progress to colorectal cancer and therefore clinically more relevant. Moreover, our study only included left-sided adenomas in the distal colon and rectum. Thus, the results of our study may not be generalizable to adenomas observed in the proximal colon, which may be more probably to occur as the result deficiencies in MMR. In addition, our study population came from a cancer prevention screening trial in which participants were generally more probably to be Caucasian, more educated, less probably to smoke and more physically active than the general population (56
). Thus, our results may not be broadly generalizable to the entire population or to other ethnicities. However, since our case–control study was nested within a randomized population-based colorectal cancer screening trial, this reduces the potential for selection bias often inherited in clinic-based case–control studies of adenoma, where persons may undergo endoscopy for reasons other than routine screening, such as gastrointestinal symptoms, blood in their stools, diagnostic follow-up or because they have a family history of colorectal cancer.
In summary, in this large comprehensive study of DNA repair gene polymorphisms and colorectal adenoma risk, we found that an SNP in EXO1 predicted to deleteriously alter function was associated with increased adenoma risk. The association was restricted to ever smokers and stronger in current smokers than former smokers. Although additional studies are needed to confirm our findings, this intriguing result suggests that genetic variation in EXO1 may modify susceptibility to colorectal adenoma, particularly among smokers.