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Background and Aims: Antituberculosis drug-induced hepatotoxicity (anti-TB-DIH) is a growing problem of serious concern in many countries around the world. Drug metabolizing enzymes have been proposed to play an influential role in the pathogenesis of anti-TB-DIH. Therefore, the main aim of this study was to investigate the possible association of GSTM1, GSTT1 and CYP2E1 gene polymorphisms with anti-TB-DIH. Methods: A prospective case-control study was conducted in 244 North Indian TB patients receiving anti-TB treatment. These patients were prospectively followed up both clinically and biochemically, before and during the treatment period, for the diagnosis of anti-TB-DIH. The frequencies of GSTM1, GSTT1 and CYP2E1 gene polymorphisms were determined from genomic DNA, utilizing multiplex polymerase chain reaction and restriction fragment length polymorphism methods respectively. Logistic regression analysis was used to determine the association between gene polymorphisms and anti-TB-DIH. Results: Anti-TB-DIH developed in 35 patients (cases). Out of 209 patients (without anti-TB-DIH), only 100 were taken as controls. As a result of genotype analysis, the frequencies of GSTM1 and GSTT1 null genotypes were found to be higher in cases as compared to controls. However, statistically significant difference was observed only for GSTT1 (Adjusted-OR= 2.39; 95% CI= 1.06-5.39; P= 0.04). Further, combined effects of these two gene polymorphisms on the risk of anti-TB-DIH were analysed. However, the effects were insignificant. Also, no significant difference was reported in the frequencies of three genotypes (C1/C1 …