Background: The manganese superoxide dismutase ( MnSOD ) gene encodes an antioxidant enzyme (SOD2) that may protect cells from oxidative damage. The MnSOD allele with Val as amino acid 16 encodes a protein that has 30–40% lower activity compared with the MnSOD Ala variant, hence possibly increasing susceptibility to oxidative stress. On the other hand, some epidemiologic studies suggest that the Ala allele is associated with a higher risk of cancer, including prostate cancer. Methods: We conducted a nested case–control study in the Health Professionals Follow-up Study with 612 incident prostate cancer cases and 612 matched controls to investigate the role of the MnSOD gene Ala16Val polymorphism and its joint association with plasma carotenoid concentrations in relation to risk of total prostate cancer and aggressive prostate cancer (advanced stage or Gleason sum ≥7). Results: The allele frequencies in the controls were 49.8% for Ala and 50.2% for Val . No association was found between the MnSOD genotype and risk of total and aggressive prostate cancer. Furthermore, no statistically significant interaction was observed between the MnSOD genotype and any of the plasma carotenoids in relation to risk of total and aggressive prostate cancer. In analyses in which we combined data from plasma and dietary carotenoids and created a quintile score to reflect long-term carotenoid status, a 3-fold [95% confidence interval: 1.37–7.02] increased risk of aggressive prostate cancer was observed among men with the Ala/Ala genotype in the presence of low long-term lycopene status ( P -value, test for interaction = 0.02) as compared with men with the Ala/Val + Val/Val genotypes with low long-term lycopene status. Conclusion: In this cohort of mainly white men, the MnSOD gene Ala16Val polymorphism was not associated with total or aggressive prostate cancer risk. However, men with the MnSODAla/Ala genotype who had low long-term lycopene status had a higher risk of aggressive prostate cancer compared with individuals with the other genotypes. These results are consistent with findings from earlier studies that reported when antioxidant status is low, the MnSODAla/Ala genotype may be associated with an increased risk of aggressive prostate cancer.