Copy number polymorphism of glutathione-S-transferase genes (GSTM1 & GSTT1) in susceptibility to lung cancer in a high-risk population from north-east India

Lung cancer is the leading cause of cancer-related deaths worldwide1. In India, it is the most frequent cancer amongst men2. In north eastern (NE) part of India lung cancer is among the ten leading sites of cancer, with the highest age-adjusted incidence rate (AAR) in Mizoram State (24.5 in males and 26.3 in females). Aizwal district alone showed an AAR of 36.0 in males and 38.7 in females which is almost three to ten times higher than Delhi3. High incidence rates suggest role of both genetic as well as environmental factors such as smoking, tobacco use and dietary carcinogen consumption. Tobacco smoking remains the primary aetiological factor associated with the development of lung cancer accounting for nearly 80-90 per cent of the disease. Polycyclic aromatic hydrocarbons (PAHs) particularly benzo[a]pyrene (BaP) and nitrogen containing nitrosamines and aromatic amines are main carcinogens present in tobacco smoke that are implicated in lung carcinogenesis. Non-smoking tobacco and betel quid have also been implicated in lung carcinogenesis probably due to their accompanied consumption with smoking4. Deleterious effects of tobacco carcinogens are primarily mediated through DNA adduct formations following their activation in the detoxifying pathways. Activated PAHs and N-nitroso compounds produced by phase I xenobiotic metabolizing enzymes are substrates for the glutathione-S-transferase M1 and T1 (GSTM1 and GSTT1) phase II enzymes. Evidences are available in literature on association of GSTM1 and GSTT1 genotypes with lung cancer5,6. Most of these studies have compared the “null” genotype with the “non-null” genotype and thus do not distinguish between one and two copy number of the genes. However, studies have reported a trimodal phenotype distribution for both GSTM1 and GSTT1 identifying homozygous wild type (+/+), hemizygous (+/-) and null (-/-) genotypes of the genes7,8. These studies suggest a gene dosage effect with three alleles corresponding to fast, intermediate and slow enzyme activity. Enzymatic activity of GSTT1 has been reported to be varying with the copy number of the gene9. Sprenger et al7 in their genotype-phenotype comparison showed correlation of significantly increased enzyme activity in individuals with two copy number of the GSTT1 compared to those with one copy number. Roodi et al10 showed that the relative risk of breast cancer increased with the present allele (+/- and +/+ genotypes) compared with -/- genotype, however, this trend was not significant. Several methods (standard and long-range PCR) in the past have been used for distinguishing GSTM110,11 and GSTT17,11,12 alleles into three genotypes. These methods were primarily based on the fact that the two genes are flanked by highly homologous regions. Recent studies have used Taqman based real-time PCR assays to discriminate between the wild-type, hemizygous deletion, and homozygous deletion of the GSTM1 and GSTT1 genes13,14. In our previous report on association of GST polymorphisms, comparing the null genotype (-/-) with combined non-null genotype (+/- and +/+) using traditional multiplex PCR-gel electrophoresis method in high risk north-east Indian population, we showed a significant protective effect of GSTM1 and GSTT1 null genotypes in lung cancer15. The present study was conducted to examine the relationship between GSTM1 and GSTT1 genes and lung cancer risk by assessing potential gene dosage effects and gene-environment interactions.