The elevated incidence of breast cancer in women has been associated with prolonged exposure to high levels of estrogens. Our laboratory has demonstrated that treatment of the immortalized human breast epithelial cells MCF-10F with 17beta-estradiol (E2) or its metabolites 4-hydroxy-estradiol (4-OH-E2) and 2-hydroxy-estradiol (2-OH-E2) induces phenotypical changes indicative of neoplastic transformation. The MCF-10F cells treated with E2, 2-OH-E2 and 4-OH-E2 form colonies in agar methocel and lost their ductulogenic capacity in collagen matrix, expressing phenotypes similar to those induced by the carcinogen benz(a)pyrene (BP). To investigate whether these phenotypic changes were associated with genomic changes, MCF-10F cells treated with either E2, 2-OH-E2, or 4-OH-E2 at different doses (0.007 nM, 70 nM and 3.6 microM) were analyzed using a combination of standard G-banding and comparative genomic hybridization (CGH). Whereas no aneuploidy was observed in any of the transformed cells, the CGH revealed instead that only cells treated with 4-OH-E2 at the highest concentration (3.6 microM) exhibited DNA gains at 8q24, 9q34 and 20q13 and losses at 13q21.