Single nucleotide polymorphisms (SNPs) of the IL-16 gene have been reported to influence the risk of several cancers, but their role in ovarian cancer (OC) has not been studied. Using the restriction fragment length polymorphism (PCR-RFLP) method, we examined four IL-16 SNPs: rs11556218 (T > G), rs4778889 (T > C), rs4072111 (C > T), and rs1131445 (T > C) in blood samples from 413 women of Central European descent, including 200 OC patients and 213 healthy controls. Among the patients, 62% were postmenopausal, 84.5% were diagnosed in late stages (FIGO IIb-IV), and 73.5% had high-grade serous OC (HGSOC). Minor allele frequencies in controls were 9.2% for rs11556218 (G allele), 13.7% for rs4778889 (C allele), 10.4% for rs4072111 (T allele), and 32.3% for rs1131445 (C allele). We found significant associations of rs11556218 (G vs. T allele: OR 2.76, 95% CI 1.84-4.14, < 0.0001) with elevated OC risk in the whole cohort ( < 0.001) and in both premenopausal ( < 0.001) and postmenopausal ( = 0.001) subgroups. These associations remained significant across heterozygote ( < 0.001), dominant ( < 0.001), and overdominant ( < 0.001) models. IL-16 rs4778889 was associated with OC risk predominantly in premenopausal women ( < 0.0001 in almost all models). In the whole cohort, the C allele was associated with OC risk (OR 1.54, CI 95% 1.06-2.23, = 0.024), and the association of rs4778889 was significant in dominant ( = 0.019), overdominant ( = 0.033), and heterozygote ( = 0.027) models. Furthermore, rs4778889 was linked with HGSOC ( = 0.036) and endometriosis-related OC subtypes ( = 0.002). No significant associations were found for rs4072111 or rs1131445 ( = 0.81 or 0.47, respectively). In conclusion, rs11556218 and rs4778889 SNPs are associated with OC risk, especially in premenopausal women.