The combined actions of DHT and MPA lead to altered AR signaling in normal and malignant post-menopausal breast epithelial cells

Abstract Consistent with several observational studies examining combined hormone replacement therapy (cHRT: conjugated equine estrogen in conjunction with the synthetic progestin medroxyprogesterone acetate, MPA) in postmenopausal women, a re-analysis of the placebo-controlled randomized Women's Health Initiative clinical trial demonstrated a markedly increased breast cancer risk in newly menopausal women following ≥ 5 years of cHRT (Hazard Ratio, 3.05; 95% Confidence Interval, 1.62-5.70) [1]. We have previously demonstrated that androgen receptor (AR)-mediated effects of MPA impede 5α-dihydrotestosterone (DHT)-induced AR signaling in normal and malignant breast epithelial cells (AACR abstract 2010). The current study aimed to further investigate the biological actions of DHT and/or MPA on steroid receptor expression and cancer-related intracellular signaling pathways. Immunohistochemical analysis of estrogen receptor alpha (ERα), progesterone receptor (PR) and AR expression was performed on histologically normal human post-menopausal breast tissues and measured by image analysis in tissues cultured ex vivo with vehicle (0.1% ethanol control), DHT (1nM), MPA (1nM) or the AR antagonist, bicalutamide (Bic;1uM) for 48 hr, either alone or in combination. Microarray analysis and qRT-PCR validation were performed using the ERα positive breast cancer cell line, ZR-75-1 to determine changes in gene expression in key intracellular signaling pathways. The microarray data was analyzed with Ingenuity Gene Pathway Analysis and Gene Ontology software. Statistical tests included both Wilcoxon matched pairs test and one-way ANOVA (p<0.05). DHT treatment increased AR expression in cultured breast tissues compared to vehicle control (p<0.05), and co-treatment with either MPA or Bic impeded this effect. No change in ERα or PR protein levels was induced by the hormone treatments. Microarray studies revealed that DHT or MPA treatment for 6 hr altered the expression of 439 and 858 genes, respectively, whereas co-treatment altered 1494 genes (p<0.05). Only 114 genes were uniquely regulated by DHT, and the expression of 32% (41% induced and 27% repressed) of these genes was abrogated by MPA. Similarly, the expression of 38% (51% induced and 24% repressed) of the 690 genes uniquely regulated by co-treatment with DHT and MPA was altered compared to DHT alone (p<0.05). Examples of genes that were regulated by DHT (p<0.05) and this effect of DHT was antagonised by co-treatment with DHT and MPA (p<0.05) are FGFR2, OLR1 and C1ORF116. Co-treatment with DHT and MPA altered the expression of genes involved in cell growth, cell cycle, cell death, cancer and intracellular signaling pathways compared to individual treatments (p<0.05). Collectively, these findings suggest an AR-mediated mechanism for the action of MPA in breast cancer. 1.Prentice, R.L., et al. Am J Epidemiol, 2009 170(1): 12-23. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 274. doi:1538-7445.AM2012-274Aleksandra M. Ochnik, Nicole L. Moore, Stephen N. Birrell, Lisa M. Butler, Shalini Jindal, Luke Selth, Wayne D. Tilley and Theresa E. Hicke

Medroxyprogesterone acetate impedes 5 alpha-dihydrotesterone induced androgen receptor signaling in normal and malignant human breast epithelial cells

Abstract not availableAleksandra M. Ochnik, Tanja Jankovic-Karasoulos, Elisa J. Cops, Stephen N. Birrell, Lisa M. Butler, Shalini Jindall, Tina Bianco-Miotto, Mervyn Thomas, Wayne D. Tilley and Theresa E. Hicke