Stat5 acts independently of androgen receptor to drive advanced prostate cancer

描述(申请人提供)：STAT5独立作用于雄激素受体，驱动晚期前列腺癌：去势耐药前列腺癌(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通常发生在雄激素剥夺治疗开始后的三年内。雄激素受体(AR)转录活性的恢复被归因于多种分子机制，但没有一种机制完全解释了从激素反应性PCa到CRPC的转变。最近的研究表明，在肿瘤发生的肿瘤干细胞理论的支持下，前列腺癌干细胞(PCSC)亚群的扩张可能是推动CRPC发生的机制之一。不表达AR的PCSCs的生存能力是通过独立于AR的信号通路维持的，这预示着对基于雄激素的治疗缺乏反应性。我们的初步数据表明，转录因子Stat5是PCa中有效的治疗靶蛋白，对于AR阴性和AR耗尽的PCa细胞的生存至关重要。在缺乏AR的情况下，Stat5的缺失强烈下调了抗凋亡蛋白Survivin的表达，而在预期分离的PCSCs中，Stat5的激活增加了。这一建议的中心假设是，激活的Stat5通过AR不依赖的信号通路促进PCa细胞的活性和致死性CRPC的发展。总体目标是：1)确定抗凋亡蛋白Survivin是否是AR非依赖性、Stat5驱动的前列腺癌细胞存活的介质；2)确定Stat5是否在前列腺癌干细胞诱导的CRPC的发育和生长中起关键作用。第一个特定目标将确定Stat5在AR阴性和AR耗竭的PCa细胞中上调Survivin的机制，通过qPCR、Western blotting和Stat5激活或抑制后Survivin的染色质免疫沉淀。机制研究将通过在Stat5/Survivin缺失后通过诱导Stat5/Survivin的表达来确定Survivin的表达是否挽救了AR阴性和AR耗竭的PCa细胞中Stat5的抑制诱导的凋亡。第二个特定目的是确定从PCSC、异种移植瘤和临床PCAS中预期分离的PCSCs中Stat5的激活水平，以及Stat5抑制剂是否在体外诱导PCSCs的凋亡。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PCSC是否在体内诱导CRPC生长发育，以及在该模型系统中Stat5抑制剂是否阻断PCSC诱导的CRPC生长发育。除了为CRPC的发展提供机制方面的见解外，拟议的研究还探索了Stat5抑制作为CRPC的新一线/辅助治疗。这些研究具有重要的意义和创新性，因为他们提出了史无前例的新颖的Stat5抑制概念，作为一种双重策略，能够同时针对AR依赖和AR非依赖的生存途径，消除CRPC的生长，而无论AR状态如何。对于对将基础科学发现转化为治疗进步以改善患者结果感兴趣的医学博士/博士学生来说，这项建议代表了一种杰出的前列腺癌研究培训经验。

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.