Methylenetetrahydrofolate reductase (MTHFR) catalyzes the synthesis of 5-methyltetrahydrofolate, which is involved in the methylation of homocysteine to methionine. Genetic polymorphisms that decrease MTHFR activity result in an altered cancer risk depending on folic acid intake. In this study we examined the C677T and A1298C polymorphisms of the MTHFR gene in specimens from 81 patients with prostate cancer and 42 controls selected from patients with benign prostatic hypertrophy (BPH). Genomic DNA was isolated from archived formaldehyde-fixed and paraffin-embedded tissue blocks. MTHFR genotypes were determined by restriction-fragment-length-polymorphism polymerase chain reaction. The MTHFR polymorphism frequencies in the prostate-cancer and BPH specimens were, respectively, 60% and 48% for 677CC, 31% and 48% for 677CT, 9% and 5% for 677TT, 36% and 43% for 1298AA, 53% and 40% for 1298AC, and 11% and 17% for 1298CC. Although such differences fall within the realm of chance variation (P>0.05), the data suggest that the 677CT genotype may be associated with a reduced risk of prostate cancer: the age-adjusted odds ratio (aOR) was 0.6 [95% confidence interval (CI): 0.3-1.4]; the odds-ratio reduction was similar in both blacks and whites (aOR=0.4 in blacks, and 0.6 in whites); and when polymorphisms at the 677 and 1298 loci were analyzed in conjunction, a lower frequency of the 677CT-1298AA genotype was observed in the patients with prostate cancer (aOR=0.3, 95% CI: 0.1-1.1). This particular genotype, moreover, was associated with lower Gleason score tumors (aOR=0.1 for Gleason-score 7 versus 6 tumors, 95% CI: 0.0-0.7) and earlier stage disease (aOR=0.3 for stage III versus II, 95% CI: 0.3-2.6). These findings suggest that polymorphisms of the MTHFR gene may alter the risk of developing prostate cancer.