Specific estrogen metabolites may initiate and promote hormone-related cancers. In epidemiological studies, significantly lower excretion of urinary estradiol (E2) and lower ratio of urinary 2-hydroxy estrogens to 16α-hydroxyestrone (2:16 OH-E1) have been reported in prostate cancer cases compared to controls. Although soy supplementation has been shown to increase the ratio 2:16 OH-E1 in women, no studies to our knowledge have investigated the effects of soy supplementation on estrogen metabolism in men. The objective of this randomized controlled trial was to determine the effects of soy protein isolate consumption on estrogen metabolism in men at high risk for developing advanced prostate cancer. Fifty-eight men supplemented their habitual diets with 1 of 3 protein isolates: 1) isoflavone-rich soy protein isolate (SPI1) (107 mg isoflavones/d); 2) alcohol-washed soy protein isolate (SPI-2) (<6 mg isoflavones/d); or 3) milk protein isolate (MPI), each providing 40 g protein/d. At 0, 3, and 6 mo of supplementation, the urinary estrogen metabolite profile was measured by GC-MS. Both soy groups had higher E2 excretion than the MPI group at 3 and 6 mo. After 6 mo of supplementation, the SPI+ group had a significantly higher urinary 2:16 OH-E1 ratio than the MPI group. Increased urinary E2 excretion and 2:16 OH-E1 ratio in men consuming soy protein isolate are consistent with studies in postmenopausal women and suggest that soy consumption may be beneficial in men at high risk of progressing to advanced prostate cancer as a result of effects on endogenous estrogen metabolism.