Abstract 474: Estrogen-responsive miRNAs as modulators of E2-induced apoptosis in AI-resistant breast cancer

Abstract

Abstract Long-term estrogen deprivation with tamoxifen or aromatase inhibitors (AI) is the basic principle of endocrine treatment of ER-positive breast cancer. Acquired resistance is however a major obstacle in treatment success. Based on clinical observations and in vitro as well as animal in vivo experiments it has been suggested that tumor cell clones evolve over time and become vulnerable to E2-induced apoptosis, thereby potentially providing a promising second line treatment option. In vitro, this vulnerability is mimicked in the AI-resistant breast cancer models MCF-7:5C and MCF-7:2A, which during long-term E2-deprivation reconfigure their survival signaling including endoplasmic reticulum, oxidative and inflammatory stress related pathways. E2-stimulation of these cells initiates an unfolded protein response which in turn triggers apoptosis through the intrinsic and subsequently extrinsic pathway (Jordan VC 2015). Recently, we identified miRNA profiles matching the biology of AI resistance and vulnerability to E2-induced apoptosis (Hoppe R et al. 2016). Here we investigate the modulatory role of miRNAs in E2-induced apoptosis through the identification of their global expression changes in E2-stimulated 5C and 2A models compared to MCF-7:WS8 reference. Each cell line was treated with 10-9 M E2 or vehicle over a 72h time course (6, 12, 24, 72 h). miRNA profiles were generated using Affymetrix GeneChip miRNA2.0 arrays. At each time point relative miRNA expression changes (E2/control) were evaluated resulting in a total of 72 (5C), 104 (2A) and 94 (WS8) differentially expressed miRNAs (FC > 1.5 or < 1/1.5, Pt-test < 0.05). Differential expression analyses between consecutive time points revealed 16 (5C), 43 (2A) and 27 (WS8) miRNAs (maximum absolute FC difference > 1.5, PF-test < 0.05). Differential area under the curve (dAUC) analyses at 6-72, 6-24, and 24-72 h mined the overall, early, and late-responding miRNAs of the different phenotypes (Ppermutation test < 0.05). Representative early down-regulated miRNA candidates potentially modulating E2-induced apoptosis in 5C cells are miR-543 and miR-432 of the DLK1-DIO3 locus on Chr. 14q32.31. Late responding up-regulated miRNA candidates are miR-150* and miR-149* the low expression of which in tumor tissues registered in The Cancer Genome Atlas (miRNA-Seq v. 3.1.17.0) was associated with worse outcome in all PAM50 breast cancer subtypes and Lum B, respectively (HR = 2.1, 95% CI: 1.4-3.2; P = 0.00017; HR = 6.8, 95% CI: 1.5-3.0; P = 0.0036). We are currently in the process to subject respective miRNA sets to functional enrichment analyses (KEGG, GO) and to correlate E2-responsive miRNA candidates with respective transcriptome signatures. In summary, we will present miRNAs matching to the biological processes inherent to E2-induced apoptosis. These may serve as potential targets for the amplification of the apoptotic trigger upon E2-treatment. Citation Format: Reiner Hoppe, Ping Fan, Stefan Winter, Florian Büttner, V. Craig Jordan, Hiltrud B. Brauch. Estrogen-responsive miRNAs as modulators of E2-induced apoptosis in AI-resistant breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 474. doi:10.1158/1538-7445.AM2017-474