Abstract 5628: Novel cell permeable peptide inhibitors of PELP1 oncogenic functions
Estrogen receptor (ERα) is the fundamental transcription factor in breast cancer progression and a majority of breast cancers start as ERα positive. Therapies targeting ERα are effective; however, many eventually develop resistance and recur as metastases. ERα signaling is mediated by coregulator proteins that can have altered expression and/or function that promotes therapy resistance and metastasis by enhancing hormone-independent ERα signaling and by epigenetic changes. An alternative strategy for inhibiting ERα signaling in therapy resistant and metastatic tumors is to inhibit the function of ERα coregulators. PELP1 is an ERα coactivator and proto-oncogene which is overexpressed in breast cancer, promotes breast cancer progression by epigenetic changes at ERα target genes and is an independent marker of poor prognosis and breast cancer survival. The objective of this study is to develop an inhibitor of PELP1-mediated ERα coactivator functions. Since PELP1 lacks any known enzymatic activities, we targeted PELP1 protein-protein interactions to inhibit its oncogenic functions. We used a yeast two-hybrid system to screen a library of random peptides (∼10-7) fused to a transcriptional activation domain and identified 23 peptides with high affinity for PELP1. Two of these PELP1-inhibiting peptides (PIP-1 and PIP-2) significantly block PELP1 coactivator functions and we used a TAT-peptide fusion of PIPs to deliver the peptides into cells. Cellular uptake of the TAT- PIP was confirmed by fluorescent microscopy and biotin-avidin peptide pull-down assays confirmed PELP1-PIP binding. Both PIP- 1 and -2 significantly inhibited PELP1-mediated proliferation in two ERα-positive breast cancer cells (MCF7 and ZR75). A GenBank search revealed homology of PIPs with the methyltransferase G9a (Bat8/KMT1C/EHMT2) that is commonly overexpressed in aggressive breast cancer, contributes to epigenetic silencing of tumor suppressors and promotes cancer invasion and metastasis. In vivo co-immunoprecipitation analysis and an in vitro binding assay confirmed PELP1 interaction with G9a. Peptide competition assays indicated that the TAT-fused peptides could interrupt PELP1-G9a interaction. Both PIP-1 and -2 substantially inhibited PELP1-mediated cell invasion/migration in Boyden chamber assays. Interestingly, disrupting PELP1-G9a interaction with PIPs sensitized resistant cells to hormonal therapy. Mechanistic studies revealed that PELP1-G9a interactions play a critical role in epigenetic changes in therapy resistant cells. In conclusion, our studies identified a novel inhibitor of PELP1 that can be used for therapeutic targeting of ERα-coregulator driven cancer progression and therapy resistance. This work was supported by the NIH grant CA095681 and CPRIT pre-doctoral fellowship. 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 5628. doi:1538-7445.AM2012-5628