A population-based association study of SNPs of GSTP1, MnSOD, GPX2 and Barrett's esophagus and esophageal adenocarcinoma

Carcinogenesis vol.28 no.6 pp.1323–1328, 2007 doi:10.1093/carcin/bgm007 Advance Access publication February 2, 2007 A population-based association study of SNPs of GSTP1, MnSOD, GPX2 and Barrett’s esophagus and esophageal adenocarcinoma Seamus J.Murphy1,2,
, Anne E.Hughes1, Chris C.Patterson3, Lesley A.Anderson2, R.G.Peter Watson4, Brian T.Johnston4, Harry Comber5, Jim McGuigan6, John V.Reynolds7 and Liam J.Murray2 1 Department of Medical Genetics, Queen’s University Belfast, Royal Group of Hospitals, Grosvenor Road, Belfast BT12 6BA, 2Centre for Clinical and Population Sciences, Queen’s University Belfast, Mulhouse Building, Grosvenor Road, Belfast BT12 6BJ, 3Department of Medical Statistics, Queen’s University Belfast, Mulhouse Building, Grosvenor Road, Belfast, BT12 6BJ, 4Department of Gastroenterology, Royal Group of Hospitals, Grosvenor Road, Belfast, BT12 6BA, 5National Cancer Registry Ireland, Elm Court, Boreenmonna Road, Cork, Ireland, 6Department of Thoracic Surgery, Royal Group of Hospitals, Grosvenor Road, Belfast, BT12 6BA and 7Department of Thoracic Surgery, St James’s Hospital, James’s Street, Dublin 8, Ireland Oxidative stress appears to be important in the pathogenesis of Barrett’s esophagus (BE) and esophageal adenocarcinoma (EAC). Single-nucleotide polymorphisms (SNPs) of antioxidant enzyme genes may play a part in determining individual susceptibility to these diseases. The Factors Influencing the Barrett’s Adenocarcinoma Relationship (FINBAR) study is a population-based, case– control study of BE and EAC in Ireland. DNA from EAC (n 5 207), BE (3 cm BE at endoscopy with specialized intestinal metaplasia on biopsy, n 5 189) and normal population controls (n 5 223) were analyzed. Several SNPs spanning the genes for glutathione S-transferase P1 (GSTP1), manganese superoxide dismutase (MnSOD) and glutathione peroxidase 2 (GPX2) were genotyped using multiplex polymerase chain reaction and SNaPshotä. The x2 test was used to compare genotype and allele frequencies between case and control subjects. Linkage disequilibrium between SNPs was quantified using Lewontin’s D# value and haplotype frequency estimates obtained using Haploview. Eleven SNPs were genotyped (six for GSTP1, three for MnSOD and two for GPX2); all were in Hardy–Weinberg equilibrium. None was significantly associated with EAC or BE even before Bonferroni correction. Odds ratios for EAC for individual SNPs ranged from 0.68 [95% confidence interval (CI) 0.43–1.08] to 1.25 (95% CI 0.73– 2.16), and for BE from 0.84 (95% CI 0.52–1.30) to 1.30 (95% CI 0.85–1.97). SNPs in all three genes were in strong linkage disequilibrium (D# . 0.887) but haplotype analysis did not show any significant association with EAC or BE. SNPs involving the GSTP1, MnSOD and GPX2 genes were not associated with BE or EAC. Further studies aimed at identifying susceptibility genes should focus on different antioxidant genes or different pathways. Methods Introduction Barrett’s esophagus (BE) is a condition in which the native squamous mucosa of the lower esophagus is replaced by columnar mucosa, defined histologically as specialized intestinal metaplasia (SIM). It is an Abbreviations: BE, Barrett’s esophagus; CI, confidence interval; EAC, esophageal adenocarcinoma; FINBAR, Factors Influencing the Barrett’s Adenocarcinoma Relationship; GI, gastrointestinal; GPX2, glutathione peroxidase 2; GSTP1, glutathione S-transferase P1; MnSOD, manganese superoxide dismutase; OR, odds ratio; PCR, polymerase chain reaction; ROS, reactive oxygen species; SIM, specialized intestinal metaplasia; SNP, single-nucleotide polymorphism. Study subjects and recruitment The FINBAR study is an Irish population-based case–control study of BE and EAC. The study methods have been described in detail elsewhere (23). Three groups of subjects were recruited: subjects with EAC, subjects with longsegment BE and normal population controls. All subjects were white Caucasians aged 35–85 years. Cases of EAC were subjects with a histological confirmation of adenocarcinoma within the esophagus. Cases from Northern Ireland were identified from electronic pathology records from all pathology laboratories within the province. Cases from the Republic of Ireland were identified from the main hospitals involved in the diagnosis and treatment of esophageal cancer and through the National Cancer Registry in Ireland, Cork. Subjects with BE were identified in the following manner: In Northern Ireland, Ó The Author 2007. Published by Oxford University Press. All rights reserved. For Permissions, please email: 1323
To whom correspondence should be addressed at Division of Gastroenterology, Box 1069, Mount Sinai Medical Center, One Gustave Levy Place, New York, NY 10029-6574, USA. Tel: þ1 212 659 9393; Fax: þ1 212 659 9853; Email: important risk factor for the development of esophageal adenocarcinoma (EAC) (1). Most subjects with BE never develop EAC, and both environmental and genetic susceptibility factors are believed to be important in determining individual risk. Antioxidant status has emerged as an important determinant of risk. Reactive oxygen species (ROS), such as superoxide anion (O2.), are produced continuously in all cells (2). However, excess levels of ROS are harmful to cells, causing damage to cellular lipids, proteins and DNA. Antioxidant enzymes [e.g. superoxide dismutase and glutathione S-transferase (GST)] protect against the adverse effects of ROS by preventing their formation, scavenging them or promoting their decomposition. There is substantial evidence that ROS plays an important role in the pathogenesis of BE and EAC in both animal models (3–5) and human studies (6–8); there is a positive correlation between the severity of esophagitis and ROS levels, with high levels found in BE (6,7) and EAC (9). Glutathione S-transferase P1 (GSTP1) is quantitatively the most important GST isoform in normal esophageal epithelium (10). GSTP1 expression, GSTP1 mRNA levels, glutathione content and GST enzyme activities are all reduced in BE compared with normal esophageal epithelium (10–14). Glutathione peroxidase 2 (GPX2) is detected mainly in gastrointestinal (GI) tissues and liver in humans (15). Messenger RNA levels of GPX2 are significantly increased in BE tissue as compared with normal squamous mucosa (16). Manganese superoxide dismutase (MnSOD, SOD2) is a major cellular defense mechanism against oxidative damage (17,18). In humans, tissue MnSOD levels are reduced in patients with reflux esophagitis and BE compared with normal esophagus (7). Whereas expression of MnSOD is increased in EAC (19), superoxide dismutase activity is decreased, possibly due to enzyme inactivation (20). Further, exogenous administration of superoxide dismutase reduces the risk of developing SIM and EAC in a rat model (20). Previous studies have looked for an association between singlenucleotide polymorphisms (SNPs) of GSTP1 and risk of BE or EAC. One study found that a func (...truncated)