Isolation and small molecule targeting of Ewing's Sarcoma stem cells

描述（由申请人提供）：“癌症干细胞假说”假定每个肿瘤内存在对化疗具有抗性的稀缺干细胞群体，并负责产生和维持大量肿瘤群体。尤文肉瘤家族肿瘤（ESFT）代表了由癌症干细胞驱动的肿瘤对治疗的预期反应。目前的治疗方法很可能是针对干细胞的后代（肿瘤的大部分），而不是真正的癌症干细胞。因此，虽然大多数患者获得缓解，但复发是常见的。ESFT含有一个特征性易位t（11：22），它编码致癌转录， 因子EWS-FLI 1。EWS-FLI 1被认为是驱动肿瘤发生的因素，因此抑制EWS-FLI 1应该是一种有效的治疗策略。由于EWS-FLI 1缺乏内在的酶活性，我们的药理学抑制方法是通过靶向蛋白质-蛋白质相互作用。RNA解旋酶A是EWSFLI 1的关键结合伴侣。我们已经确定了一种先导化合物YK-4-279，它可以阻断EWS-FLI 1与RHA的相互作用。此外，我们的初步数据表明，ESFT细胞系中具有干细胞样特性的亚群仍然表达EWS-FLI 1。因此，EWS-FLI 1有望成为ESFT干细胞的关键分子靶点。我们假设阻断RHA与EWS-FLI 1相互作用的药物将靶向化疗耐药的ESFT干细胞。这些研究将提供必要的临床前数据，以证明专门针对ESFT干细胞设计的化合物的临床试验的合理性。本研究的主要目的是开发尤文肉瘤干细胞（ESSC）的靶向治疗。我们正在提出一个综合的，合作的研究计划，旨在识别和表征ESSC，这将允许开发一种有效的治疗策略，以靶向和消除患者中的这些细胞。本研究的第一个目的是识别和表征ESSC。我们将采用各种已提出的检测方法来鉴定癌症干细胞，包括醛脱氢酶的表达、染料外排、在非粘附条件下形成球形聚集体的能力以及特征基因的表达。基因表达谱将是假设驱动的，并将专门进行测试的假设，尤文肉瘤是来自间充质干细胞的肿瘤转化。然后，我们将证明EWS-FLI 1/RHA相互作用的抑制对化疗耐药的ESSC是致命的。该研究的最终目的是确定其他适合小分子靶向的关键信号通路。这将解决单独抑制EWS-FLI 1活性可能不足以消除ESSC的可能性。同时靶向多种信号通路可能是一种更有效的治疗策略。我们预计，在该项目结束时，ESSC将被分离并充分表征，以允许开发可快速转化为临床试验的靶向治疗，并显着改善尤文肉瘤家族肿瘤患者的预后。

DESCRIPTION (provided by the applicant): The "Cancer Stem Cell Hypothesis" posits the existence of a scarce stem cell population within each tumor that is resistant to chemotherapy and is responsible for generating and maintaining the bulk tumor population. Ewing sarcoma family tumors (ESFT) typify the response to treatment expected of a tumor driven by cancer stem cells. Current therapies most likely target the stem cells' progeny (the bulk of the tumor) and spare the true cancer stem cells. Consequently, although the majority of patients achieve a remission, relapse is common. ESFT contains a characteristic translocation, t(11:22), which encodes the oncogenic transcription factor EWS-FLI1. EWS-FLI1 is thought to drive tumorigenesis, so inhibition of EWS-FLI1 should be an effective treatment strategy. Because EWS-FLI1 lacks intrinsic enzymatic activity, our approach to pharmacologic inhibition is through targeting protein-protein interactions. RNA Helicase A is a critical binding partner of EWSFLI1. We have identified a lead compound, YK-4-279, that blocks EWS-FLI1 interaction with RHA. Furthermore, our preliminary data suggest that a subpopulation within ESFT cell lines with stem-like properties still expresses EWS-FLI1. Therefore, EWS-FLI1 is expected to be a key molecular target in ESFT stem cells. We hypothesize that a drug that blocks the interaction of RHA with EWS-FLI1 will target chemotherapyresistant ESFT stem cells. These studies will provide the necessary preclinical data to justify clinical trials with a compound designed specifically to target ESFT stem cells. The primary objective of this study is to develop a targeted therapy for Ewing's sarcoma stem cells (ESSC). We are proposing an integrative, collaborative research program aimed at the identification and characterization of ESSC which will allow the development of an effective therapeutic strategy to target and eliminate these cells in patients. The first aim of the study is to identify and characterize ESSC. We will employ a variety of assays that have been proposed to identify cancer stem cells, including expression of aldehyde dehydrogenase, dye efflux, the ability to form spherical aggregates under nonadherent conditions, and the expression of characteristic genes. Gene expression profiling will be hypothesis-driven, and will be performed specifically to test the hypothesis that Ewing's sarcoma is derived from the neoplastic transformation of mesenchymal stem cells. We will then demonstrate that inhibition of the EWS-FLI1/RHA interaction is lethal to chemotherapy-resistant ESSC. The final aim of the study is to identify other key signaling pathways that are amenable to small molecule targeting. This will address the possibility that inhibition of EWS-FLI1 activity alone might be insufficient to eliminate ESSC. Simultaneous targeting of multiple signaling pathways is likely to be a substantially more effective therapeutic strategy. We anticipate that at the conclusion of the project, ESSC will have been isolated and sufficiently characterized to allow the development of targeted therapies that can be rapidly translated into clinical trials and dramatically improve the prognosis of patients with Ewing's sarcoma family tumors.