Abstract 5626: EBP50: Novel biomarker for resistance to endocrine and HER-2 targeted therapies in breast cancer. .

Backgroud: Na+/H+ Exchanger Regulatory Factor (NHERF1) also named ERM Binding Protein 50 (EBP50) functions as a molecular scaffold to coordinate ion transport and second messenger signal transduciton. Recently, a number of studies suggest a role for EBP50 expression in cancer progression. Methods: Microarray profiling. Stable transfection. Soft agar and MTT growth assays. Immunoblot and immunoprecipitation assays. Confocal microscopy. Duolink in situ. Results: The mean levels of EBP50 were reduced in tamoxifen (Tam)-resistant, metastatic breast tumors compared to Tam-sensitive tumors. Knockdown of EBP50 decreased Tam sensitivity in MCF-7 and ZR-75B cells. EBP50 knockdown cells showed higher levels of phosphorylated HER2 and EGFR, as well as their downstream signalling pathways. PI3K/AKT inhibitors were able to restore Tam sensitivity in knockdown cells. Similarly, knockdown EBP50 in ERα/HER2 positive BT-474 cells increased constitutive pHER2 and these cells were resistant not only to Tam, but also to Trastuzumab (Tras) treatment. An EGFR inhibitor or Lapatinib were able to restore Tras sensitivity. Overexpression of EBP50 restored Tam sensitivity in BT-474 cells. We demonstrate that EBP50 is capable of binding to HER2 and ERα using immunoprecipitation and PLA assay, suggesting that EBP50 interacts directly with both. Overexpression of EBP50 decreased pHER2 and pEGFR levels suggesting that EBP50 can downregulate its activity. EBP50 overexpression upregulated ERα transcriptional activity. ERα transcriptional activity in cells overexpressing EBP50 was costitutively higher and more sensitive to Tam. We previously published that Dicer overexpression and loss of RhoGDIα were associated with Tam resistance. Interestingly, EBP50 knockdown was associated with higher Dicer and lower RhoGDIα levels suggesting that Tam resistance is associated with inter-related mechanisms. We performed microarray analysis and identified kinase signiture in EBP50 knockdown cells. This kinase signature predicts Tam resistance. Our data suggest that EBP50 is a novel negative regulator of HER2 signaling, and its loss confers resistance to both Tam and Tras. We hypothesize that EBP50 levels might be a new predictive biomarker for targeted therapy. Citation Format: Guowei Gu, Arnoldo Corona-Rodriguez, Kyle Covington, Natalie Fernandez, Jianling Zhou, Weiyan Cai, Sebastiano Ando’, Suzanne A.W. Fuqua. EBP50: Novel biomarker for resistance to endocrine and HER-2 targeted therapies in breast cancer. . [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 5626. doi:10.1158/1538-7445.AM2013-5626