Estrogens are clearly carcinogenic in humans and rodents, but the mechanisms by which these hormones induce cancer are only partially understood. Stimulation of cell proliferation and gene expression by binding to the estrogen receptor is one important mechanism in hormonal carcinogenesis; however, estrogenicity is not sufficient to explain the carcinogenic activity of all estrogens because some estrogens are not carcinogenic. Estrogens are nonmutagenic in many assays, but exhibit specific types of genotoxic activity under certain conditions. We have studied extensively the mechanisms by which estrogens induce neoplastic transformation in a model in vitro system. Diethylstilbestrol (DES) and 17β-estradiol (E2) and their metabolites induce morphological and neoplastic transformation of Syrian hamster embryo (SHE) cells that express no measurable levels of estrogen receptor. The estrogens induce DNA adduct formation that is detected by 32P-postlabeling. DNA adduct formation is detected in cells treated with DES, but not in cells treated with either α-or β-dienestrol. Similarly, morphological transformation of SHE calls is induced by treatment with DES, but not by treatment with α-or β-dienestrol. Exposure of SHE cells to E2 and its metabolites 2-hydroxyestradiol and 4-hydroxyestradiol leads to covalent DNA adduct formation, corresponding to the induction of cell transformation. Induction of morphological transformation of SHE cells by estrogens correlates well with the ability of estrogens to induce DNA adduct formation. DNA damage caused by DNA adduct formation may be important in hormonal carcinogenesis. It is clear that hormones affect carcinogenesis by epigenetic mechanisms such as stimulation of cell proliferation of estrogen-dependent target cells and reprogramming of cellular differentiation and gene expression. In addition, significant evidence exists that certain estrogens can also cause genetic alterations by mechanisms not involving the classic estrogen receptor. These findings indicate that hormonal carcinogenesis is most likely a result of the interplay of both genetic and epigenetic factors.