BACKGROUND. Chemoprevention utilizing dietary agents is an effective means to slow the development of prostate cancer. We evaluated the potential additive and synergistic effects of genistein and resveratrol for suppressing prostate cancer in the Simian Virus-40 T-antigen (SV-40 Tag) targeted probasin promoter rat model, a transgenic model of spontaneously developing prostate cancer. METHODS. Rats were fed genistein or resveratrol (250 mg/kg AIN-76A diet) alone and in combination, and a low-dose combination (83 mg genistein + 83 mg resveratrol/kg diet). Histopathology and mechanisms of action studies were conducted at 30 and 12 weeks of age, respectively. RESULTS. Genistein, resveratrol, and the high-dose combination treatments suppressed prostate cancer. The low-dose combination did not elicit protection against prostate cancer and was most likely below the effective dose for causing significant histopathological changes. Total genistein and resveratrol concentrations in the blood reached 2,160 and 211 nM, respectively in rats exposed to the single treatments. Polyphenol treatments decreased cell proliferation and insulin-like growth factor-1 (IGF-1) protein expression in the prostate. In addition, genistein as a single agent induced apoptosis and decreased steroid receptor coactivator-3 (SRC-3) in the ventral prostate (VP). CONCLUSIONS. Genistein and resveratrol, alone and in combination, suppress prostate cancer development in the SV-40 Tag model. Regulation of SRC-3 and growth factor signaling proteins are consistent with these nutritional polyphenols reducing cell proliferation and increasing apoptosis in the prostate. © 2009 Wiley-Liss, Inc.