Abstract 3341: Cross-talk between PI3K/AKT and ERα signaling leads to STAT3/6-modulated epigenetic transcription in aggressive breast cancer

The phosphatidylinositol 3-kinase (PI3K)/AKT pathway is a critical survival cascade in tumor progression. Our previous study demonstrated that combinational treatment of a potent PI3K inhibitor, LY294002 (LY), and DNA methyltransferase inhibitor, 5’-Aza-deoxycytidine (DAC), compounds effectively reduce tumor formation in the immunodeficiency mouse model, suggesting that PI3K/AKT signaling suppresses tumorigenesis through the epigenetic modulation. While tumor size of the LY-treated xenografts is dramatically decreased compared to the control group, the tumor growth cannot be completely attenuated by either LY or LY/DAC treatment, indicating that there is an alternative route for tumor survival. Here, we performed a microarray analysis to identify LY-mediated expression profiling on an ERα-positive breast cancer cell line, T-47D, the cell line applied in the previous xenograft experiments. A subset of 24 LY-activated genes associated with better clinical outcome are found to be not only direct targets of PI3K/AKT but involved in estrogen (E2)/ERα signaling by ingenuity pathway analysis, suggesting the involvement of ERα signaling in PI3K/AKT-mediated tumor growth. To clarify their relationship, pharmacological experiments using an ERα antagonist, ICI 182, 780 (ICI) and LY coupled with E2 was conducted. We observed that E2 stimulation induces both AKT and ERα activation in a time-dependent manner. Either ICI or LY treatment cannot repress the activation levels of AKT or ERα, respectively. Though, either single or combined inhibitor treatment can effectively restore E2-suppressed gene expression. These findings indicate that cross-talk between PI3K/AKT and ERα signaling cascades may cooperatively regulate cell growth and associated gene expression. In addition, transcription factor motif analysis showed that signal transducer and activator of transcription 3 and 6 (STAT3/6) binding motifs are enriched with highest prediction score on the promoter regions of the examined AKT-directed targets. Inhibitors prohibit E2-mediated assembly of STAT3/6 with ERα and STAT3/6 recruitment onto the promoter regions of target loci using co-immunoprecipitation (co-IP) and chromatin IP-qPCR. Moreover, repressive histone marks, H3K27me3 and H3K9me3, are found to occupy on the same regions as STAT3/6 upon E2 treatment. DAC treatment can effectively restore E2-induced suppression of AKT-directed target genes. Further association studies are ongoing to interrogate whether STAT3/6 is part of repressive transcription complex with H3K27me3 and H3K9me3. In summary, we propose that cross-talk of ERα with PI3K/AKT signaling represents the potential function switch of ERα from genomic binding to no-genomic binding. Combined treatment of PI3K/AKT inhibitors with hormone therapy may shed a light of therapeutic strategy to patients with aggressive or metastatic breast tumors. Citation Format: Hang-Kai Hsu, Pei-Yin Hsu, Yao Wang, Zelton D. Sharp, Tim H.-M. Huang, Victor X. Jin. Cross-talk between PI3K/AKT and ERα signaling leads to STAT3/6-modulated epigenetic transcription in aggressive breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3341. doi:10.1158/1538-7445.AM2014-3341