Identification of novel indications for commonly prescribed drugs could accelerate translation of therapies. We investigated whether any clinically used drugs might be useful in treating prostate cancer by coupling an efficient, high-throughput laboratory- based screen and a large prospective cohort study. In stage one, we conducted an in vitro prostate cancer cell cytotoxicity screen of 3,187 compounds. Digoxin emerged as the leading candidate, given its potency in inhibiting proliferation in vitro (the concentration of the drug at which proliferation was inhibited by 50%: mean of 163 nM) and its common use. In stage two, we evaluated the association between the leading candidate drug from stage one and prostate cancer risk in 47,884 men followed up from 1986 through 2006. Regular digoxin users [vs nonusers: relative risk (RR) = 0.76; 95% confidence interval (CI), 0.61-0.95], especially users for ≥10 years (RR = 0.54; 95% CI, 0.37-0.79; P trend < 0.001), had a lower prostate cancer risk. Digoxin was highly potent in inhibiting prostate cancer cell growth in vitro, and its use was associated with a 25% lower prostate cancer risk. siGnificance: Our two-stage transdisciplinary approach for drug repositioning provides compelling justification for further mechanistic and possibly clinical testing of the leading nonchemotherapy candidate, digoxin, a cardiac glycoside, as a drug for prostate cancer treatment. Perhaps of equal importance, our study illustrates the power of the transdisciplinary approach in translational cancer research. By coupling laboratory and epidemiologic methods and thinking, we reduced the probability of identifying false-positive candidate drugs for the next steps in testing. © 2011 American Association for Cancer Research.