Polymorphisms in DNA repair genes as risk factors for asbestos-related malignant mesothelioma in a general population study

Abstract

Differences in response to carcinogenic agents are due to the allelic variants of the genes that control it. Key genes are those involved in the repair of the DNA damage caused by such agents. This paper describes the results of a case-control epidemiological study designed to determine the genotypes of four of these genes in persons exposed to a single genotoxic factor, i.e. asbestos, who had or had not developed malignant mesothelioma (MM). Our working hypothesis was that an imperfect DNA repair, as revealed by subtle polymorphic variants, could reduce protection against the chronic DNA insult provoked by asbestos and eventually result in mutagenesis and cancer. Seven variants (i.e. XRCC1-R399Q-NCBI SNP, XRCC1-R194W, XRCC3-T241M, XRCC3-IVS6-14, XPD-K751Q, XPD-D312N, OGG1-S326C) were investigated in 81 patients and 110 age and sex-matched controls, all residents at Casale Monferrato, a Piedmontese town highly exposed to asbestos pollution.

Unconditional multivariable logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs). When considered as a categorical variable, XRCC1-399Q showed an increased OR both in heterozygotes (OR = 2.08; 95% CI = 1.00–4.33) and homozygotes (2.38; 95% CI = 0.82–6.94), although individual ORs were not significant. When it was considered as a continuous variable OR was significant (OR = 1.68; 95% CI: 1.02–2.75).

When genotypes were divided into “non-risk” and “risk” genotypes, i.e. those thought to be associated with increased risk in the light of the functional significance of the variants, XRCC1-399Q (Q homozygotes + Q/R heterozygotes versus R homozygotes) had an OR = 2.147 (95% CI: 1.08–4.28), whereas that of XRCC3-241T (T homozygotes + M/T heterozygotes versus M homozygotes) was 4.09 (95% CI: 1.26–13.21) and that of OGG1-326C was increased, though not significantly. None of the haplotypes showed a significantly different frequency between patients and controls.

This is the first report of an association between polymorphisms in DNA repair genes and asbestos-associated MM. Our data indicate that genetic factors are involved in MM development.

Introduction

The association of asbestos exposure and malignant mesothelioma (MM) is well documented, but the mechanism of action has not been completely clarified [1]. Asbestos fibers chronically retained in the lung can be carcinogenic as the result of: (1) mechanical effects, such as interference with mitotic spindle formation and the segregation of chromosomes, leading to breaks and aberrations [2]; (2) generation of reactive oxygen species (ROS) either by reactions involving catalytic iron or by frustrated fagocytes [3], [4]; (3) local and systemic immunosuppression. Asbestos induced oxidative damage has been clearly demonstrated, both in vitro and in vivo: its consequences include DNA single-strand breaks and DNA base modification [5]. Concentration of 8-hydroxy-2′-deoxyguanosine (8OHdG) in DNA was significantly higher in leukocytes from asbestos workers than from unexposed controls [6]. 8OHdG is one of the most specific forms of damage induced by ROS and is also mutagenic, since at high levels it increases the probability of G to T transversion during cell division.

About 80% of MM patients have a history of asbestos exposure [1], but only 5–10% of individuals exposed to high levels actually develop MM and its onset is usually 30–40 years after the first exposure [7]. The combined role of genetics and environment in the etiology of MM has also been suggested by its familial aggregation [8], [9], though the traits that may predispose to MM have not been extensively studied. Two studies of MM risk associated to glutathione-S-transferase (GSTM1) and the N-acetyltransferase 2 (NAT2) produced contradictory results. Hirvonen et al. [10] showed an increased risk of MM in subjects occupationally exposed to asbestos and carrying a homozygous deletion of the GSTM1 gene or the NAT2 slow acetylator genotype, whereas Neri et al. [11] did not observe any association with the GSTM1 gene and found an increased risk for NAT2 fast acetylators. These workers also reported an association of MM with microsomal Epoxide Hydrolase (mEH) and found that mEH interacted with both NAT2 and GSTM1 genes according to a multiplicative model. The same genotypes are associated with other cancer types, such as lung and bladder cancer [12]. Other studies observed an increased number of micronuclei in lymphocytes from MM patients as compared to controls, coupled with indirect evidence of an increased susceptibility to chromosomal aberration [13], [14].

Association of defective DNA repair with several cancer types has been described by many papers [15], [16], [17], [18].

The postulated association of gene polymorphisms leading to defective DNA repair with an increased risk of MM is examined in this paper. We focused on DNA repair after oxidative damage and studied seven single-nucleotide polymorphisms (SNP) for four genes: XRCC1, XRCC3, XPD, OGG1. SNPs were chosen because they had a significant effect on the transcript (amino acid substitution or possible splice defect) and have been associated with certain cancer types and/or deficient DNA repair [15], [16], [17], [18], [19], [20], [21], [22], [23], [24], [25]. XRCC1, XRCC3 and XPD were studied each for two SNPs reported to be in linkage disequilibrium so that haplotypes could be drawn.

Our working hypothesis was that imperfect DNA repair, as revealed by subtle polymorphic variants which in normal conditions are not associated with a disease phenotype, could reduce protection against the chronic DNA insult provoked by asbestos fibers, especially oxidative damage, and eventually result in mutagenesis and cancer. The laboratory investigation was part of a population-based case-control study on MM, with all cases and controls from the same well-defined geographical area, concurrently recruited and examined for asbestos exposure and SNPs.