Modulation of the estrogen receptor (ER) has been one of the most successful treatment strategies in women with breast cancer. The two most commonly used methods of ER inhibition are either competitive binding of the ER with anti-estrogens (e.g. tamoxifen) or to decrease the local production of estrogen in target tissues using an aromatase inhibitor. However, resistance to hormonal therapy emerges in more than 50% of patients whose tumors initially express ER, and a high percentage of tumors are a priori resistant. An extensive effort to introduce novel anti-estrogens, such as the selective estrogen receptor down-regulator (SERD) fulvestrant, or the selective estrogen modulator (SERM) raloxifene, has had limited success beyond that achieved with tamoxifen. An emerging strategy to enhance the efficacy of hormone therapy involves epigenetic modulation of ER signaling. Preclinical studies from our and other laboratories have shown that the addition of an HDAC inhibitor to tamoxifen treatment reverses tamoxifen resistance and synergistically induces cell death. Furthermore, in a recently completed clinical trial by our group, a subset of patients with hormone therapy resistant tumors had durable tumor regression when treated with this combination, providing clinical evidence for HDAC inhibition as an approach for reversing hormone therapy resistance. Correlative studies accompanying this trial in peripheral blood mononuclear cells suggest that histone acetylation and HDAC2 expression was higher in patients with a response than those without treatment benefit. In an effort to determine the relevant HDAC targets for synergy with tamoxifen, we demonstrated that siRNA depletion of HDAC2 was sufficient to potentiate the cytotoxic effects of tamoxifen and modulate estrogen signaling in ER- positive breast cancer cells. HDAC2 represses ER regulated pro-apoptotic genes and thus its inhibition allows SERMs to exert their anti-agonistic effects even in settings of ER hypersensitivity and high endogenous estrogen levels. Therefore, we hypothesize that HDAC2 plays an important role in hormone therapy resistance and its selective depletion may offer a novel strategy to reverse resistance to anti-estrogen therapy. In this application, we seek to determine the relevance of HDAC2 as a therapeutic target and predictive marker of response for the treatment of hormone sensitive and hormone resistant breast cancer. In SPECIFIC AIM 1 we will define the optimal therapeutic setting for combined HDAC2 inhibition-anti-estrogen therapy for the treatment of breast cancer in in vitro models. In SPECIFIC AIM 2, we will evaluate HDAC expression in patient samples and determine their potential as a predictive marker for response to this combined therapy. In SPECIFIC AIM 3 we will evaluate the ability of HDAC2 depletion either alone or in combination with anti- estrogen therapy to induce the regression of primary tumors and to reduce metastases using in vivo breast cancer xenograft models. PUBLIC HEALTH RELEVANCE: Patients with metastatic breast cancer have a median survival of approximately 24 months and account for over 40,000 deaths per year in the United States. Hormonal therapy is one of the most successful and least toxic modalities for patients with breast cancer. However more than 50% of the patients are a priori resistant to hormone therapy, or become therapy resistant over time. Our preclinical and clinical data suggest that epigenetic modulation of the estrogen receptor by HDAC inhibitors may introduce a novel strategy to reverse hormone therapy resistance. This application will determine the optimal clinical setting to integrate HDAC inhibitors into hormonal therapy.