PR Rapid and Genomic Effects Participate in the Response of Breast Cancer Cells to Tamoxifen

GALIGNIANA, NATALIA M; DÍAZ FLAQUÉ, MARÍA C.; VENTURUTTI, LEANDRO; MERCOGLIANO, MARÍA FLORENCIA; DE MARTINO, MARA; IZZO, FRANCO; PROIETTI, CECILIA J.; ELIZALDE, PATRICIA V.

Chicago

Congreso; 16th International Congress of Endocrinology & the Endocrine Society?s 96th Annual Meeting (ICE-ENDO 2014); 2014

The progesterone receptor (PR) is known to participate in breast cancer cell proliferation and to cross-talk with the oestrogen receptor alpha (ERα), yet its roles in resistance to anti-oestrogen therapies are not fully understood. In the present work, we studied the effects of the synthetic progestin medroxyprogesterone acetate (MPA) alone or in combination with tamoxifen (Tam), a commonly used selective ER modulator, in Tam-sensitive and -resistant breast cancer cell lines. We used Tam at a concentration (1 μM) in which it acts as an antagonist of ERα-mediated effects and therefore mimics its expected role in patients. We found that Tam inhibited MPA-induced activation of the c-Src/ERK1/2 signaling pathway in BT474-HR cells, which are sensitive to the antiproliferative effects of Tam. As a result, two synergic downstream effector mechanisms were disrupted. On the one hand, Western blot and chromatin immunoprecipitation assays revealed that MPA-induced c-Src/ERK1/2 phosphorylation and recruitment of the transcription factor AP-1 to the cyclin D1 promoter, along with ERα and PR itself, were inhibited by co-treating BT474-HR cells with MPA+Tam. This prevented MPA from inducing cyclin D1 transcription and cell proliferation. Interestingly, Tam failed to block MPA-activation of c-Src/ERK1/2 and assembly of the said complex in the Tam-resistant cell line BT474 (1). On the other hand, we found that PR-dependent activation of c-Src/ERK1/2 leads to the activation of c-Myc and subsequent down-regulation of miR-16, a microRNA we have previously shown acts as a tumor suppressor in progestin-induced breast cancer growth (2). This decrease in miR-16 levels resulted in increased expression of its target, the anti-apoptotic protein Bcl-2. Co-treatment of BT474-HR cells with MPA+Tam disrupted MPA activation of c-Src/ERK1/2, restoring miR-16 levels to control levels and keeping Bcl-2 expression repressed. Such inhibitory effects of Tam were not evidenced in BT474 cells. Together, these results suggest that Tam effects on PR-dependent c-Src/ERK1/2 signaling can inhibit cell proliferation by regulating the formation of transcription factor complexes whilst inducing apoptosis through control of microRNA levels only in Tam-sensitive cells. Our results provide insight into complex synergic mechanisms underlying Tam resistance involving the PR, a receptor not classically studied in the context of ER-targeted therapies shown here to be a possible therapeutic target to prevent its unrestrained functions in Tam-resistant cells. (1) Díaz Flaqué et al., Breast Cancer Res. 2013; 15(6):R118. (2) Rivas et al., Breast Cancer Res. 2012; 14(3):R77