[CANCER RESEARCH 64, 4378 – 4384, June 15, 2004] Identification of Genetic Variants in Base Excision Repair Pathway and Their Associations with Risk of Esophageal Squamous Cell Carcinoma Bingtao Hao, 1,4 Haijian Wang, 1,4 Kaixin Zhou, 1,4 Yi Li, 1 Xiaoping Chen, 1,4 Gangqiao Zhou, 1,4 Yunping Zhu, 1 Xiaoping Miao, 2 Wen Tan, 2 Qingyi Wei, 3 Dongxin Lin, 2 and Fuchu He 1,4 1 Laboratory of Systems Biology, Beijing Institute of Radiation Medicine, Beijing, People’s Republic of China; 2 Department of Etiology and Carcinogenesis, Cancer Institute, Chinese Academy of Medical Sciences/Peking Union Medical College, Beijing, People’s Republic of China; 3 Department of Epidemiology, University of Texas M. D. Anderson Cancer Center, Houston, Texas; and 4 Chinese National Human Genome Center at Beijing, Beijing, People’s Republic of China ABSTRACT The etiology of esophageal squamous cell carcinoma (ESCC) has been shown to be associated with genetic and certain environmental factors that produce DNA damage. Base excision repair (BER) genes are responsible for repair of DNA damage caused by reactive oxygen species and other electrophiles and therefore are good candidate susceptibility genes for ESCC. We first screened eight BER genes for new and potential functional polymorphisms by resequencing 27 DNA samples. We then identified and genotyped for important tagging single nucleotide polymorphisms (SNPs) in a case-control study of 419 patients with newly diagnosed esophageal cancer and 480 healthy controls by frequency matching on age and sex. The association between genotypes and ESCC risk was estimated by unconditional multivariate logistic regression analysis, and stepwise re- gression procedure was used for constructing the final logistic regression model. We identified 129 SNPs in the eight BER genes, including 18 SNPs that cause amino acid changes. In the final model, 4 SNPs, including 2 in the coding regions (ADPRT Val762Ala and MBD4 Glu346Lys) and others in noncoding regions (LIG3 A3704G and XRCC1 T-77C), remained as significant predictors for the risk of ESCC. The adjusted odd ratios were 1.25 [95% confidence interval (CI) 1.02–1.53] for the ADPRT 762Ala allele, 1.25 (95% CI 1.02–1.53) for the MBD4 346 Lys allele, 0.78 (95% CI 0.63– 0.97) for the LIG3 3704G allele, and 1.38 (95% CI 1.01–1.89) for the XRCC1–77C allele. In addition, we observed a significant gene-gene in- teraction between XRCC1 Gln399Arg and ADPRT Val762Ala. The results suggest that the polymorphisms in five BER genes may be associated with the susceptibility to ESCC in a Chinese population. INTRODUCTION Cancer of the esophagus ranks among 10 most frequent cancers in the world, with a marked regional variation in incidence and mortality worldwide (1). Northern China is one of the regions where esophageal squamous cell carcinoma (ESCC) is common, and there are 250,000 patients newly diagnosed with ESCC each year in China, accounting for more than half of the world’s cases (2). Although the integrated etiology of ESCC remains to be fully elucidated, accumulative epi- demiological evidence suggests that tobacco smoking, heavy alcohol drinking, micronutrient deficiency, and dietary carcinogen exposure may cause the disease (3– 6). All these factors can induce or enhance DNA damage mediated by either oxidative stress or DNA-binding electrophiles, which in turn may initiate and/or promote carcinogen- esis. However, to safeguard the integrity of genome and prevent the detrimental consequences of DNA damage, humans have developed a complex set of DNA repair systems. Defects in DNA repair have been demonstrated to be a critical mechanism in human carcinogenesis (7). In addition, accumulating evidence suggests that suboptimal DNA repair capacity caused by genetic polymorphism is associated with increased cancer risk (8 –11). Thus, ESCC is probably associated with not only environmental factors but also individuals’ suboptimal DNA repair capacity. Base excision repair (BER) is one of the important DNA repair pathways against DNA damage resulted from many insults, including altered metabolism, reactive oxygen species, and methylating and deaminating agents (7, 12–14). BER has two major two steps: exci- sion of damaged base residues and core BER reaction (7, 14). Briefly, a battery of glycosylases, each dealing with a relatively narrow, partially overlapping spectrum of lesions, feeds into a core reaction by releasing the modified base and creating abasic sites; the core BER reaction is initiated by strand incision at the abasic site by the APEX endonuclease; and DNA polymerase  performs a one-nucleotide gap-filling reaction and removes the 5'-terminal baseless sugar resi- due via its lyase activity. This is then followed by sealing of the remaining nick by the XRCC1-ligase3 complex (14). It is the so- called short-patch repair that performs a one-nucleotide gap-filling reaction, which is the dominant mode of the repair activity in mam- mals. The gap of 2–10 bases was filled by the long-patch repair mode, which involves DNA pol, pol/, and proliferating cell nuclear antigen in repair synthesis, FEN1 endonuclease in removal of the displaced DNA flap, and DNA ligase 1 in gap sealing. Enzymes from the BER pathway can also rectify single-strand interruptions in DNA. ADP-ribosyltransferase (ADPRT), as well as XRCC1, temporarily binds to single-strand interruptions in DNA and may act to recruit repair proteins (7, 14). Knockout mouse models showed inactivation of BER core proteins (i.e., APEX1, POLB, and XRCC1) induces embryonic lethality, highlighting the pivotal roles of these genes (7). Recently, a knockout mouse model of MBD4, a glycosylase, revealed that although MBD4 inactivation does not by itself cause cancer predisposition in mice, it may alter the mutation spectrum in cancer cells and therefore contribute to the cancer-predisposition phenotype (15, 16). Until last year, inherited deficiencies in the BER pathway had not been causally linked to any human genetic disorders (7). It has recently been found that the bi-allelic mutations in a DNA glycosylase gene, MUTYH, may lead to an autosomal recessive syndrome of adenomatous colorectal polyposis with a high colorectal cancer risk (17–20). Thus, common polymorphisms of BER genes are plausible candidates that may contribute to susceptibility to cancer, including ESCC according to the Common Disease-Common Variant hypoth- esis (21). Several studies have in fact found associations between genetic polymorphisms in some BER genes such as OGG1 and XRCC1 and risk of certain cancers, including ESCC, and the results are encour- aging (Refs. 10, 11; reviewed in Ref. 22). However, most of the previous studies were designed to analyze individual gene, which obviously has limitations to elucidate the effect of the entire BER pathway. To comprehensively investigate the roles of the polymor- phisms in the BER genes in the development of ESCC and identify potential genetic markers for ESCC in Chinese populations, we con- ducted this study by using a candidate-gene association approach. We Received 2/4/04; revised 3/22/04; accepted 3/30/04. Grant support: National “863” High Technology Project Grants 2001AA224011 (F. He) and 2002BA711A06 (D. Lin) and National Natural Science Foundation Grant 39990570 (D. Lin). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. Requests for reprints: Fuchu He, Laboratory of Systems Biology, Beijing Institute of Radiation Medicine, Beijing 100850, China. Phone: 86-10-681-71208; Fax: 86-10-682- 14653; E-mail: hefc@nic.bmi.ac.cn. 4378 Downloaded from http://aacrjournals.org/cancerres/article-pdf/64/12/4378/2515650/zch01204004378.pdf by guest on 01 July 2022