The aim of the study was to evaluate the role of N-acetyltransferase 2 (NAT2) gene polymorphism in the development of cervical cancer by comparing patients having invasive cervical squamous cell carcinoma (SCC) with healthy control subjects. The study group consisted of 42 women with invasive cervical SCC and 50 control subjects. All of the patients were primarily treated with surgical intervention. Blood samples (5 mL) were obtained from the patients before surgery or during follow-up to 2 years after surgery. DNA was extracted from the leukocytes by a high pure PCR template preparation kit (catalog No. 1 796 828; Roche Diagnostics GmbH, Mannheim, Germany). NAT2*5A, NAT2*6A, and NAT2*7A/B polymorphisms of NAT2 were detected by using a LightCycler-NAT2 mutation detection kit in real-time PCR (catalog No. 3113914, LightCycler instrument; Roche Diagnostics GmbH, Mannheim, Germany). We found that the risk of cervical SCC was 9.045-fold higher in individuals with NAT2*5A mutant allele (95% confidence interval, 1.448-56.524; P = 0.018). The frequency of the NAT2*5A slow genotypes in the patients with cervical cancer (23.8%) was significantly higher compared with that in the control group (6%). Individuals with the NAT2*5A slow genotype had a significantly higher risk of cervical cancer compared with individuals with the NAT2*5A fast genotype (odds ratio, 7.469; 95% confidence interval, 1.673-33.350; P = 0.008). However, there was no significant association between the NAT2*6A and NAT2*7A/B fast or slow acetylator status and the development of cervical cancer. In conclusion, NAT2*5A slow acetylator genotype was found to be significantly higher in patients with cervical cancer. These results suggest that NAT2*5A gene polymorphisms in patients may be associated with genetic susceptibility to cervical cancer.