Due to its lipophilicity and persistence, an organochlorine compound, beta-hexachlorocyclohexane (beta-HCH), is known to frequently accumulate in human adipose and breast tissues. An epidemiological study has indicated that exposure to beta-HCH could be one of the significant environmental risk factors for the development of human breast cancers. Additionally, beta-HCH has recently been identified as an environmental estrogen capable of activating estrogen receptor (ER) through a ligand-independent pathway. In the present investigation, we examined the impact of long-term in vitro exposure to beta-HCH on cell transformation and the metastatic potentials of MCF-7 cells. We found that continuous exposure of MCF-7 cells to beta-HCH at 100 nM and 1 microM or to 17beta-estradiol (E(2)) at 1 nM for up to 13 months (33 passages) not only enhanced their transformation tendencies but also promoted their invasiveness. Western blot analysis revealed that beta-HCH induced transformation-related biochemical changes in MCF-7 cells, such as a decline in the levels of ERalpha and p44/42 MAP kinase and a significant increase in expression of c-ErbB2 and MMP-9 levels. In contrast, long-term E(2) treatment resulted in the downregulation of ERalpha and p44/42 MAP kinase and upregulation of MMP-9 only, but no changes in c-ErbB2. Together, these results indicate that these biochemical changes induced by beta-HCH are consistent with the events taking place in these cells to promote the phenotypical expression of transformed cells. Our results provide the in vitro mechanistic basis supporting the hypothesis that beta-HCH is one of the epigenetic risk factors assisting the progression of breast cancer cells to an advanced state of malignancy.