We investigated dietary supplement use and prostate cancer risk in the Carotene and Retinol Efficacy Trial (CARET). CARET was a randomized, double-blinded, placebo-controlled trial testing a daily dose of 30 mg beta-carotene + 25,000 IU retinyl palmitate for lung cancer prevention (1985-1996; active follow-up occurred through 2005). Secondary outcomes, including prostate cancer, were also assessed. Participants were queried about dietary supplements, health history, family history of cancer, smoking, and lifestyle habits. Cox proportional hazards regression estimated multivariate-adjusted relative risk [and 95% confidence intervals (95% CI)] of prostate cancer for dietary supplement users and nonusers with or without the high-dose CARET vitamins during the intervention and postintervention phases. After an average of 11 years of follow-up, 890 prostate cancer cases were reported. Neither the CARET nor other supplements were associated with total prostate cancer risk. For aggressive prostate cancer, men in the CARET intervention arm who used additional supplements had a relative risk for aggressive prostate cancer (Gleason >or=7 or stage III/IV) of 1.52 (95% CI, 1.03-2.24; P < 0.05), relative to all others. These associations disappeared in the postintervention period (0.75; 95% CI, 0.51-1.09). Conversely, there was no association of CARET + other supplements with nonaggressive disease, relative to all others. There was no effect modification by smoking or time on CARET intervention in any analyses. CARET only included smokers, so findings reported here may not apply to nonsmokers. Our results are consistent with other studies suggesting that dietary supplements may influence prostate cancer risk.