Abstract 5367: Medroxyprogesterone acetate impedes 5α-dihydrotesterone induced androgen receptor signaling in normal and malignant human breast epithelial cells

Abstract

Abstract Combined hormone replacement therapy (cHRT) comprised of equine estrogens and the synthetic progestin medroxyprogesterone acetate (MPA) increases breast cancer risk in post-menopausal women [1], via as yet undetermined mechanisms. In addition to the progesterone receptor (PR), MPA binds with high affinity to the androgen receptor (AR), and therefore has the potential to elicit actions distinct to those of natural progesterone, which does not bind AR. We have proposed that MPA increases breast cancer risk by binding to the AR and interfering with the ability of endogenous AR ligands to inhibit proliferation of normal breast epithelial cells [2, 3]. To further investigate this proposal, we examined the effect of MPA on 5α-dihydrotestosterone (DHT)-induced AR signaling in normal human breast tissue and in a breast cancer cell line (ZR75-1) that expresses estrogen receptor alpha (ERα), PR, and AR. Normal human breast tissues was obtained from 12 post-menopausal women following surgery for benign or malignant disease. Normal histology was confirmed by a pathologist. Breast tissues (3mm3) were cultured in triplicate for 48 hours on gelatine sponges soaked in media containing 10% steroid deplete fetal calf serum (FCS) and DHT (1nM), MPA (1nM), or the AR antagonist, bicalutamide (Bic; 1uM), alone and in combination. Following culture, expression of AR, Ki67 (a marker of cell proliferation) and bcl-2 (a pro-survival factor) was assessed by immunohistochemistry. ZR75-1 cells were cultured for 24 and 48 hours in 10% steroid deplete FCS supplemented with DHT, MPA or the combination, followed by immunoblot analysis to detect AR and an androgen-regulated protein, FKBP5. Statistical analysis was preformed by Wilcoxon signed-rank test unless otherwise stated. Normal human breast tissues possessed significantly higher levels of immunoreactive AR than ERα or PR (p<0.0001; t-test for unequal variances). Treatment with DHT significantly increased AR expression (p=0.02) and decreased the percentage of Ki67 positive cells (p=0.03), but had no effect on expression of bcl-2. The effects of DHT on AR and Ki67 were reversed by co-incubation with Bic (p<0.05) or MPA (p<0.05). In ZR75 cells, DHT increased AR protein levels at 24 and 48 hours and increased FKBP5 protein levels at 48 hours. Co-incubation with MPA reversed the effects of DHT on AR and FKBP5 proteins. Collectively, these data support our hypothesis that MPA interferes with the action of endogenous AR ligands in normal and malignant human breast epithelial cells. This interference of androgen action corresponds to an increased proliferation index in normal human breast epithelial cells, which feasibly contributes to an increased risk of developing breast cancer. 1. Chlebowski., R.T., et al., Jama, 2003. 289: 3243-53 2. Birrell, S.N., et al., Faseb J, 2007. 21; 2285-93 3. Peters, A.A., et al., Cancer Res, 2009. 69: 6131-40 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5367.