P2-02-07: Regulation of mTOR Signaling by Proto-Oncogene PELP1.
Authors: VK Gonugunta, V Cortez, C Rethman, BC Nair, GR Sareddy, RK Vadlamudi
Journal: Cancer Research (2011)

Abstract BACKGROUND: Despite positive effects of hormonal therapy, initial or acquired resistance to endocrine therapies frequently occurs. Emerging evidence suggests that ER action is complex and requires functional interactions with coregulators. In addition, ER also participates in extra-nuclear signaling events in the cytoplasm and growth factor cross talk with ER is implicated in the development of therapy resistance. The mammalian target of rapamycin (mTOR) is a serine/threonine kinase that plays an important role on cell growth and proliferation. Proline, Glutamic-acid and Leucine-rich Protein 1 (PELP1) is an ER coregulator that functions in nuclear as well as extra-nuclear actions. PELP1 functions as a proto-oncogene and is a prognostic indicator of shorter breast cancer specific survival. The objective if this study is to examine whether PELP1 participates in ER-growth factor signaling cross talk via mTOR pathway. METHODS: To test this hypothesis, we have used ER-positive model cells (MCF7, ZR75) along with PELP1 over expressing (MCF7-PELP1, ZR75-PELP1) and PELP1 knock down cells (MCF7-PELP1 shRNA, ZR-75-PELP1 shRNA). Estrogen and Heregulin were used as signal inducers. Rapamycin that inhibit mTORC1 and AZD8055, an inhibitor of both mTORC1 and mTORC2 activities were used as pharmacological inhibitors to block mTOR pathway. The affect of blocking mTOR axis on PELP1 mediated oncogenic functions was determined using ERE and E2F reporter gene assays, cell proliferation (MTT) and anchorage independent assays. Cell cycle progression was monitored by flow cytometry. PELP1 modulation of mTOR signaling was validated using siRNA approach in MCF7, ZR75 cells and/or by PELP1 overexpression in PELP1-shRNA clones and evaluated the role of identified components for changes in the mTOR signaling using Western analysis. IHC analysis of mTOR signaling components was performed using tissues collected from PELP1 overexpressing and PELP1 siRNA liposome treated tumors. RESULTS: Over expression of PELP1 enhanced the estrogen and heregulin mediated cell proliferation in breast cancer cells. Knock down of PELP1 with siRNA significantly reduced the activation of mTOR signaling components (p70S6K and 4E-BP1). Accordingly, over expression of PELP1 in the breast cancer cells correlated with increased mTOR signaling. Pharmacological inhibition of mTOR substantially reduced PELP1 mediated ER coactivation functions in ERE reporter gene assays. Rapamycin (10−7M) or AZD8055 (10−8M) abolished PELP1 mediated growth advantage in estrogen and heregulin stimulated breast cancer cells. Further, combinatorial treatment of mTOR inhibitors sensitized PELP1 overexpressing cells to tamoxifen therapy. IHC studies using xenograft tissues with PELP1 overexpression or down regulation, showed correlation of PELP1 expression with the activation of mTOR signaling components. CONCLUSIONS: Our results suggest that PELP1 driven oncogenic functions involve PELP1 modulation of mTOR signaling and blockage of mTOR signaling render PELP1 driven tumors highly sensitive to therapeutic inhibition. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-02-07.