Gallbladder cancer (GBC) involves interplay of sex steroids, including estrogen and progesterone. Since CYP17 is a key enzyme involved in estrogen and testosterone hormone biosynthesis as well as in xenobiotic metabolism, it may be a potential candidate gene in the carcinogenesis of the gallbladder. Hence, the present study aimed to investigate the association of CYP17 (rs2486758, and rs743572) polymorphisms with GBC susceptibility. The present study included a total of 414 histologically confirmed GBC and 230 healthy controls. The CYP17 (rs2486758 and rs743572) polymorphisms were genotyped by TaqMan-Allele discrimination assays. Statistical analysis was performed by using SPSS ver. 16. Overall, both the CYP17 SNPs did not indicate any association with GBC risk at genotype, haplotype, or at the genotypic interaction levels. However, in the case-only analysis, CYP rs743572 showed association with increased risk of GBC in tobacco users at hetero genotype and dominant models, as compared to non-user GBC patients. The TCrs2486758-AGrs743572 genotypic combination was also associated with increased GBC susceptibility in tobacco users. CYP17 rs743572 is associated with increased risk of GBC in tobacco users in the North Indian population. However, the study requires confirmation in other populations.