We have investigated the ability of tamoxifen to regulate members of the transforming growth factor β (TGF-β) family in human breast cancers in vivo. Using immunohistochemical techniques, we find that 3 months of tamoxifen treatment causes a consistent induction of extracellular TGF-β1 in breast cancer biopsies, compared with matched pretreatment samples from the same patient. The induced TGF-β is locaized between and around stromal fibroblasts and appears to be derived from these cells. Lower levels of TGF-β1,-β2, and -β3 seen in epithelial cells were not altered by tamoxifen treatment. The increased stromal staining of TGF-β1 occurred in estrogen receptor-negative as well as estrogen receptor-positive tumors. These results provide in vivo evidence for a novel, estrogen receptor-independent mechanism of action for tamoxifen, involving the stromal induction of a potent growth inhibitor for epithelial cells.