Reprogrammingof theERR a andER a TargetGene Landscape Triggers Tamoxifen Resistance in Breast Cancer

Endocrine treatment regimens for breast cancer that target the estrogen receptor-a (ERa) are effective, but acquired resistance remains a limiting drawback. One mechanism of acquired resistance that has been hypothesized is functional substitution of the orphan receptor estrogen-related receptor-a (ERRa) for ERa. To examine this hypothesis, we analyzed ERRa and ERa in recurrent tamoxifen-resistant breast tumors and conducted a genome-wide target gene profiling analysis of MCF-7 breast cancer cell populations that were sensitive or resistant to tamoxifen treatment. This analysis uncovered a global redirection in the target genes controlled by ERa, ERRa, and their coactivator AIB1, defining a novel set of target genes in tamoxifen-resistant cells. Beyond differences in the ERa and ERRa target gene repertoires, both factors were engaged in similar pathobiologic processes relevant to acquired resistance. Functional analyses confirmed a requirement for ERRa in tamoxifenand fulvestrant-resistant MCF-7 cells, with pharmacologic inhibition of ERRa sufficient to partly restore sensitivity to antiestrogens. In clinical specimens (n 1⁄4 1041), increased expression of ERRa was associated with enhanced proliferation and aggressive disease parameters, including increased levels of p53 in ERa-positive cases. In addition, increased ERRa expression was linked to reduced overall survival in independent tamoxifen-treated patient cohorts. Taken together, our results suggest that ERa and ERRa cooperate to promote endocrine resistance, and they provide a rationale for the exploration of ERRa as a candidate drug target to treat endocrine-resistant breast cancer. Cancer Res; 75(4); 720–31. 2015 AACR. Introduction The ligand-activated transcription factor ERa is a key driver of the breast cancer phenotype in around 70% of patients (1). Accordingly, endocrine treatment modalities targeting ERa, such as the selective estrogen receptormodulator (SERM) tamoxifen or downregulator (SERD) fulvestrant (Fulv; SERM) constitute the basis for therapeutic intervention in ERa-positive tumors (2). Fulvestrant as a pure ERa antagonist and tamoxifen as partial antagonist counteract the pro–proliferative and antiapoptotic stimuli classically induced by estrogens (3). Beyond the widespread improvements by these treatment regimens, the frequent onset of antiestrogen resistance remains a major limitation (4), underlining the clinical need for alternative drug targets. A compelling body of evidence suggests involvement of another nuclear hormone receptor, estrogen-related receptor-a (ERRa), in the pathogenesis of breast cancer. Increased ERRa expression was found in mammary tumors and correlated with an impaired disease-free and overall patient survival (5, 6). The pathophysiologic relevance of ERRa has been further demonstrated in vivo, as ERRa inhibition in xenograft systems reduces breast tumor growth, and in HER-2/neu–driven breast cancer mouse models, ERRa knockout delays tumor formation (7, 8). Despite the structural relationship, ERa and ERRa share only 33% homology in their ligand-binding domains, resulting in the insensitivity of ERRa to classical ERa ligands such as estrogen and tamoxifen (9–11). Because of the lack of known natural ligands, ERRa is classified as orphan receptor, whereas its transcriptional activity can be abrogated by small-molecule inhibitors (i.e., XCT790; ref. 12). ERRa and ERa possess a high sequence homology in their central DNA-binding domains (68%), and therefore each recognize the others cognate-binding motif (9, 13). Identification of a subset of common target genes (e.g., pS2; refs. 14, 15), raised the hypothesis that ERRa bypasses the requirement for ERa in endocrine-resistant breast cancers and fuels resistance. This concept has been corroborated by the interplay of ERRa and known determinants of antiestrogen resistance such as HER-2/neu and the coactivator AIB1 (6, 8, 16). As a versatile nuclear hormone Division of Molecular Genetics, German Cancer Research Center (DKFZ), Heidelberg,Germany. InstituteofPathology,UniversityMedical Center Hamburg-Eppendorf, Hamburg, Germany. Foundation PATH—Patients' Tumor Bank of Hope, Munich, Germany. Institute of Pathology, University Hospital Cologne, Cologne, Germany. Genomics Core Facility, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany. Molecular Biology of Breast Cancer, University Women's Clinic, Heidelberg, Germany. Breast Cancer Group, Cardiff University,Cardiff, UnitedKingdom. Institute of Pathology, University of Heidelberg, Heidelberg, Germany. Gynecologic Oncology, National Center for Tumor Diseases, University of Heidelberg, Heidelberg, Germany. Phenex Pharmaceuticals AG, Heidelberg, Germany. Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). Corresponding Author: Peter Lichter, Division of Molecular Genetics, German Cancer Research Center (DKFZ), Im Neuenheimer Feld 580, Heidelberg 69120, Germany. Phone: 0049-6221-424619; Fax: 0049-6221-424639; E-mail: peter.lichter@dkfz.de doi: 10.1158/0008-5472.CAN-14-0652 2015 American Association for Cancer Research. Cancer Research Cancer Res; 75(4) February 15, 2015 720 on May 4, 2017. © 2015 American Association for Cancer Research. cancerres.aacrjournals. org Downloaded from Published OnlineFirst February 2, 2015; DOI: 10.1158/0008-5472.CAN-14-0652