Abstract 504: ERα36 as a potential target in breast cancer that promotes an aggressive tumor phenotype in vitro and predicts outcome in vivo.

Steroid hormones exhibit dynamic effects on cancer progression. 17β-estradiol (E2), in particular, enhances tumor growth and development through a non-classical estrogen receptor (ER) found in the plasma membrane of triple negative breast cancer (TNBC) cells. Our lab has previously found that E2 rapidly increases protein kinase C (PKC) activity, which is associated with increased tumorigenicity, and promotes cancer cell survival and metastatic potential by acting through the ERα variant, ERα36. The aims of this study were to elucidate the mechanism by which ERα36 promotes cancer cell survivability and to determine if ERα36 levels in human cancer tissue can predict clinical outcome. Using specific inhibitors against signaling molecules we hypothesized to as part of the proposed anti-apoptotic pathway, we determined the effect of pathway inhibition on the protective effect of E2 against taxol-induced apoptosis in HCC38 TNBC cells. In order to evaluate these effects, we used MTT to measure cell viability, TUNEL to measure cell death, and caspase-3 activity as a marker of apoptosis. Our results show that the anti-apoptotic effect of E2 against taxol-induced apoptosis is mediated through ERα36 at the cell membrane, which activates a signaling cascade including phosphatidylcholine specific phospholipase D, lysophosphatidic acid, and phosphoinositide-3 kinase. In order to determine if ERα36 is a potential target for either diagnosis or treatment in breast cancer, tissue microarrays containing breast cancer patient tumor tissue were assayed for presence of ERα36 by immunohistochemistry (IHC) and blindly scored by three independent observers to quantify the number of positive cells and intensity. We also performed IHC for VEGF, which is associated with aggressive tumor formation. We performed categorical analysis using either chi-square test or fisher9s exact test to obtain p-values for correlations of several variables with either ERα36 or VEGF, as well as correlation of ERα36 to VEGF. Our analysis included TNM classification, stage, age, receptor status(ER, PR, HER2), p53 status, and VEGF. Our analyses indicate ERα36 intensity was significantly correlated to age (p=.0450 for patients 50) and VEGF (p=.0226) in our breast cancer cohort. Interestingly, receptor status was not correlated to ERα36, suggesting that ERα36 may be a possible target regardless of clinical ER status and classification of the cancer. The results of this study indicate that ERα36 mediates the anti-apoptotic effect of E2 in breast cancer and that it is also correlated to variables that may be associated with adverse outcome, such as tumor angiogenesis. This study therefore suggests that ERα36 may serve as a potential biomarker for further classification of breast cancer patients, as well as a potential target for therapeutic intervention in the development of adjuvant therapies. Citation Format: Reyhaan A. Chaudhri, Agreen Hadadi, Zvi Schwartz, Ruth O9Regan, Barbara D. Boyan. ERα36 as a potential target in breast cancer that promotes an aggressive tumor phenotype in vitro and predicts outcome in vivo. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 504. doi:10.1158/1538-7445.AM2013-504