Stat5 acts independently of androgen receptor to drive advanced prostate cancer

描述（由申请人提供）：Stat5独立于雄激素受体驱动晚期前列腺癌：去势抵抗性前列腺癌（CRPC）是前列腺癌（PCa）的致命形式，缺乏有效的治疗选择。进展到CRPC通常发生在开始雄激素剥夺治疗的三年内。CRPC中雄激素受体（AR）转录活性的恢复归因于多种分子机制，但没有一种分子机制能够完全解释从激素响应性PCa到CRPC的转变。在肿瘤发生的癌症干细胞理论的支持下，最近的研究表明前列腺癌干细胞（PCSC）亚群的扩增可能是驱动CRPC发展的机制。对于AR表达阴性的PCSCs，其生存能力是通过独立于AR的信号通路维持的，这预示着对雄激素治疗缺乏反应性。我们的初步数据表明，转录因子Stat5是一种经过验证的PCa治疗靶蛋白，对ar阴性和ar缺失的PCa细胞的存活至关重要。在缺乏AR的情况下，Stat5缺失可显著下调抗凋亡蛋白Survivin的表达，Stat5激活在预期分离的PCSCs中增加。该建议的中心假设是，活性Stat5通过ar独立的信号通路促进PCa细胞的活力和致死性CRPC的发展。总体目标是：1)确定抗凋亡蛋白Survivin是否是ar不依赖、stat5驱动的PCa细胞存活的中介；2)确定Stat5是否在前列腺癌干细胞诱导的CRPC发育和生长中起关键作用。第一个具体目标是通过qPCR、Western blotting和Survivin在Stat5激活或抑制后的染色质免疫沉淀，确定Stat5在ar阴性和ar缺失的PCa细胞中上调Survivin的机制。在ar阴性和ar缺失的PCa细胞中，通过诱导Stat5/Survivin的表达，将采用机制研究来确定Survivin的表达是否能挽救Stat5抑制诱导的凋亡。第二个具体目标是确定从PCa细胞、异种移植肿瘤和临床PCa中分离的PCSCs中Stat5的激活水平，以及Stat5抑制剂是否在体外诱导PCSCs凋亡。最重要的是，我们将使用独特的CWR22Pc PCa异种移植模型系统来确定Stat5驱动的PCSCs是否在体内诱导CRPC生长，以及Stat5抑制剂是否阻断pcsc诱导的CRPC生长。除了提供CRPC发展的机制见解外，拟议的研究还探索了Stat5抑制作为CRPC的新的一线/辅助治疗方法。这些研究具有重要意义和创新性，因为它们提出了新的和前所未有的Stat5抑制概念，作为一种双重策略，能够靶向AR依赖和AR独立的生存途径，无论AR状态如何，都可以消除CRPC的生长。对于有意将基础科学发现转化为治疗进步以改善患者预后的医学博士/博士生而言，该提案代表了前列腺癌研究方面的杰出培训经验。

DESCRIPTION (provided by applicant): Stat5 Acts Independently of Androgen Receptor to Drive Advanced Prostate Cancer: Castrate-resistant prostate cancer (CRPC) is the lethal form of prostate cancer (PCa) and lacks effective therapeutic options. Progression to CRPC typically occurs within three years of initiation of androgen deprivation therapy. Recovery of androgen receptor (AR) transcriptional activity in CRPC has been attributed to various molecular mechanisms, but none have completely accounted for the transition from hormone-responsive PCa to CRPC. Recent studies, supported by the cancer stem cell theory of tumorigenesis, have implicated expansion of the prostate cancer stem cell (PCSC) subpopulation as a mechanism which may drive development of CRPC. Viability of PCSCs, which are negative for expression of AR, is sustained through signaling pathways independent of AR, predicting lack of responsiveness to androgen-based therapies. Our preliminary data demonstrate that the transcription factor Stat5, a validated therapeutic target protein in PCa, is crucial for survival of AR-negative and AR-depleted PCa cells. Stat5 depletion robustly down-regulates expression of the anti-apoptotic protein Survivin in the absence of AR, and Stat5 activation is increased in prospectively isolated PCSCs. The central hypothesis of this proposal is that active Stat5 promotes viability of PCa cells and development of lethal CRPC through AR-independent signaling pathways. The overall objectives are to 1) determine if the anti-apoptotic protein Survivin is a mediator of AR-independent, Stat5-driven survival of PCa cells; 2) determine if Stat5 is critical for prostate cancer stem cell-induced development and growth of CRPC. The first specific aim will determine mechanisms by which Stat5 up-regulates Survivin in AR-negative and AR-depleted PCa cells by qPCR, Western blotting and chromatin immunoprecipitation of Survivin following Stat5 activation or inhibition. Mechanistic studies will be employed to determine if Survivin expression rescues Stat5 inhibition-induced apoptosis in AR-negative and AR-depleted PCa cells using inducible expression of Stat5/Survivin following Stat5/Survivin depletion. The second specific aim will determine Stat5 activation levels in PCSCs prospectively isolated from PCa cells, xenograft tumors and clinical PCas, and if Stat5 inhibitors induce apoptosis of PCSCs in vitro. Most importantly, we will determine if Stat5-driven PCSCs induce development of CRPC growth in vivo using the unique CWR22Pc PCa xenograft model system, and if Stat5 inhibitors block PCSC-induced development of CRPC growth in this model system. In addition to providing mechanistic insight on the development of CRPC, the proposed studies explore Stat5 inhibition as a new first-line/adjuvant therapy for CRPC. These studies are significant and innovative because they propose the novel and unprecedented concept of Stat5 inhibition as a dual strategy capable of targeting both AR-dependent and AR-independent survival pathways, eliminating growth of CRPC regardless of AR status. This proposal represents an outstanding training experience in prostate cancer research for an MD/PhD student interested in translating basic science discoveries into therapeutic advances for improved patient outcomes.