Progesterone reverses the mesenchymal phenotypes of basal phenotype breast cancer cells via a membrane progesterone receptor mediated pathway

Introduction: Basal phenotype breast cancers (BPBC) are often associated with apparent epithelial to mesenchymal transition (EMT). The role of progesterone (P4) in regulating EMT of BPBC has not been reported. Methods: The EMT relevant biology was investigated in vitro using human BPBC cell models (MDA-MB468 and MDA-MB231) with P4, PR agonist (RU486), and PR antagonist (R5020) treatments. The essential role of membrane progesterone receptor alpha (mPR alpha) in the P4-regulated EMT was demonstrated by knocking down the endogenous gene and/or stably transfecting exogenous mPR alpha gene in the BPBC cell models. Results: The expression of snail and down-stream EMT proteins such as occludin, fibronectin, and E-cadherin was significantly regulated by P4 incubation, which was accompanied by cell morphological reversion from mesenchymal to epithelial phenotypes. In searching for the cell mediator of P4' action in the MDA-MB468 (MB468) cells, it was found that mPR alpha but not the nuclear PR has an essential role in the P4 mediated EMT inhibition. Knocking down the expression of mPR alpha with specific siRNA blocked the P4's effects on expression of the EMT proteins. In another BPBC cell line - MDA-MB231 (MB231), which is mPR alpha negative by Western blotting, P4 treatment did not alter cell proliferation and EMT protein expressions. Introduction of the exogenous mPR alpha cDNA into these cells caused cell proliferation, but not EMT, to become responsive to P4 treatment. In further studies, it was found that activation of the PI3K/Akt pathway is necessary for the P4-induced EMT reversion. To define the potential inter-mediate steps between mPR alpha and PI3K, we demonstrated that mPR alpha, caveolin-1 (Cav-1), and epidermal growth factor receptor (EGFR) are colocalized in the membrane of caveolar vesicle and the P4-repressed EMT in MB468 cells can be blocked by EGFR inhibitor (AG1478) and PI3K inhibitor (wortmannin). Conclusions: Our data suggest that the signaling cascade of P4 induced mesenchymal repression is mediated through mPR alpha and other caveolae bound signaling molecules namely Cav-1, EGFR, and PI3K. This novel finding may have great impact on fully understanding the pathogenesis of BPBC and provide an essential clue for developing a targeted therapeutic strategy for treatment of BPBC.