Background: Exposure to beta-Hexachlorocyclohexane (beta-HCH), a contaminant of the hexachlorohexane pesticide lindane, has been implicated as a risk factor in the development of breast cancers in epidemiological studies. Previous studies in our laboratory have demonstrated the ability of beta-HCH to elicit its actions via a ligand-independent activation of the estrogen receptor through increased c-Neu (= erbB2 or HER-2) expression and kinase activation in both the BG-1 and MCF-7 cell lines. In addition, long term exposure (33 passages) to beta-HCH was shown to promote the selection of MCF-7 cells which exhibit a more metastatic phenotype.
Methods: In this current study, we decided to investigate the long-term effects of beta-HCH in both the MCF10AT1 cell line which was derived from a normal epithelial cell line by stably transfecting a mutated c-Ha-ras and a MMTV-Neu mouse model for mammary cancer in vivo. MCF10AT1 cells were exposed for 20 passages with beta-HCH, 4-OH-Tamoxifen (Tam), or 17-beta-estradiol (E2) after which cells were analyzed for proliferation rates and mRNA expression by RT-PCR. In our in vivo studies, MMTV-Neu mice were injected with beta-HCH and observed for tumor formation over a 70 week period.
Results: beta-HCH and Tam selected MCF10AT1 cells demonstrated increased mRNA expression of MMP-13 (collagenase-3) a marker of increased invasiveness. beta-HCH treatment was also seen to increase the expression in a number of proto-oncogenes (c-Neu, Cyclin D1, p27), cell status markers (Met-1, CK19), and the inflammatory marker NFkappaB. Previous studies, have demonstrated the role of these markers as evidence of malignant transformations, and further illustrate the ability of beta-HCH to be carcinogenic. To demonstrate beta-HCH's tumorigenic properties in an in vivo system, we used an MMTV-Neu mouse model.MMTV-Neu is a c-Neu overexpressing strain which has been shown to spontaneously develop mammary tumors at later stages of aging. In this experiment, beta-HCH exposure was shown to both accelerate the appearance (~8 weeks for median tumor-free period) and incidence (~25% increase at the end of the test period) of tumors when compared to control mice receiving only the corn-oil vehicle.
Conclusion: Based upon these results, it was concluded that beta-HCH does act as a breast cancer promoter which exerts its tumorigenic activity via increased c-Neu expression.