We conducted a prospective study to assess the association between cardiorespiratory fitness and prostate cancer. The subjects were men, aged 20-80 yr, who received a preventive medical exam at the Cooper Clinic in Dallas, TX, during 1970-1989 and provided information on cardiorespiratory fitness and prostate cancer (N = 12,975). Cardiorespiratory fitness was assessed at a baseline examination between 1971 and 1989 using a maximal exercise treadmill test. Questionnaires were mailed to the men in 1982 and 1990 to ascertain incident cases of prostate cancer. Ninety-four cases of incident prostate cancer were identified. Higher cardiorespiratory fitness levels were inversely associated with the probability of development of incident prostate cancer controlling for age, body mass index, and smoking habits; adjusted estimates of the incidence rate ratio declined from 1.1 (95% CI 0.63-1.77) to 0.73 (95% CI 0.41-1.29) to 0.26 (95% CI 0.10-0.63) across increasing quartiles of fitness (P for trend <0.004). This protective effect was limited to participants <60 yr old. Also, an inverse association was observed between physical activity and prostate cancer. Compared with expending <1000 kcal·wk-1, participants who expended 1000-<2000, 2000-<3000, or ≥3000 kcal·wk-1 had adjusted incidence rate ratios of 0.37 (95% CI 0.17-0.79), 0.62 (95% CI 0.27-1.41), and 0.37 (95% CI 0.14-0.98), respectively. The results suggest that cardiorespiratory fitness and physical activity levels may protect against the development of incident prostate cancer.