The progesterone receptor (PR) plays an important role in diverse biological processes. A major function of this nuclear hormone receptor is to bind to DNA and regulate the expression of progesterone-responsive genes. Recent research suggests PR can also act indirectly by antagonizing the actions of estrogen receptor (ER). Estrogen has been shown to increase expression of PR in normal human breast tissue and progesterone has been implicated in blocking estrogen-induced cell proliferation. However, the role of PR in the growth of estrogen-dependent breast cancer cell lines is controversial, with progesterone stimulating and inhibiting growth in different situations. Though molecular studies have suggested various mechanisms of crosstalk between estrogen and progesterone signaling pathways, it is difficult to distinguish the effect of these interactions from the independent activities of the two hormones. Through the use of the fluorescence anisotropy microplate assay (FAMA) as a novel high throughput screening method, we identified small molecule inhibitors of PR binding to its DNA binding site, the progesterone response element. Inhibitor specifcity was evaluated by comparing progesterone, estrogen, and glucocorticoid gene activity in cell-based reporter assays using the same T47D breast cancer cell line background. Identifying the sites and modes of action of small molecule inhibitors of PR will be useful in dissecting the mechanisms of PR action and in testing its novel role in the inhibition of ER action. This project aims to (1) Determine the site and mode of action of small molecule inhibitors of PR action, and (2) Test whether small molecule inhibitors of PR action can enhance ER action by relieving PR inhibition of ER. Relevance: The roles of progesterone receptor, based on its direct effects and its indirect effects mediated by inhibiton of estrogen receptor, remain controversial and are not well understand. Using a novel high- throughput screen and the diverse steroid hormone systems available in our laboratory, this research aims to characterize small molecules that specifically inhibit PR binding to DNA. Characteriztion of these inhibitors will begin to elucidate the direct and indirect effects of PR and PR inhibitors in normal cells, breast cancer, and multiple sclerosis.
