Regulation of estrogen receptor (ER) levels in MCF-7 cells by progesterone metabolites

Estradiol-17beta (E2) may participate in carcinoma of mammary cells containing estradiol receptors (ER) at sufficient levels. Hence, the regulation of ER levels may be important for the progression of estrogen-dependent mammary carcinomas. Our previous findings that the progesterone metabolite, 5alpha-pregnane-3,20-dione (5alphaP), exhibits marked mitogenic and metastatic properties, whereas the progesterone metabolites, 4-pregnen-3alpha-ol-20-one (3alphaHP) and 4-pregnen-20alpha-ol-3-one (20alphaHP), oppose these actions, prompted examination of the possible effects of these progesterone metabolites on ER concentration in MCF-7 breast cancer cells. Cells were exposed for 24h to 0 (control) or 10(-10) to 10(-6)M E2, 5alphaP, 3alphaHP, 20alphaHP or combinations of these steroids, and ER concentrations were determined for intracellular estrogen receptors by specific binding of [(3)H]E2. The total ER number (nuclear plus cytosolic) in control samples was 2551+/-164 per cell. E2 and 5alphaP resulted in significant dose-dependent increases in total ER numbers ( approximately 1.6-fold and approximately 2.2-fold at 10(-6)M, respectively). In combination, E2+5alphaP resulted in additive increases in ER numbers. Individually, 3alphaHP and 20alphaHP each resulted in dose-dependent decreases (43% and 54% at 10(-6)M, respectively) in total ER numbers and inhibited the E2- or 5alphaP-induced increases in ER levels. In combination, 3alphaHP+20alphaHP resulted in dose-dependent additive suppression of ER levels. Treatment with cycloheximide or actinomycin D indicated that both transcription and translation are involved in 5alphaP and 3alphaHP action on ER numbers. Real time RT-PCR showed increases in expression of ERalpha transcripts due to 5alphaP and increases in expression of ERbeta due to 3alphaHP; expression levels of either ERalpha or ERbeta were not significantly altered when cells were treated with 5alphaP+3alphaHP. The results are the first to show that the pro- and anti-cancer progesterone metabolites also have marked selective (up or down) regulatory effects on ER levels in MCF-7 breast cancer cells.