Molecular Modeling of Pediatric Skeletal Muscle Tumors

描述（申请人提供）：横纹肌肉瘤（RMS）是一种表现出不同程度骨骼肌分化的异质性癌症集合。虽然RMS占儿童恶性实体瘤的8%，但它是14岁以下儿童最常见的软组织肉瘤。RMS的两种主要组织学亚型是胚胎型（eRMS）和腺泡型（aRMS）。高风险患者的5年生存率为30%，而表达PAX 3-FKHR融合基因的肿瘤儿童的结局非常差;当转移时，他们的5年生存率<8%。这种标志性遗传变化仅在aRMS中发现，并被认为是肿瘤特异性致癌基因，但没有分子靶向治疗。为了解决RMS知识的差距，我们已经为这种疾病创建了一个新的模型，该模型基于原代人类骨骼肌细胞向其致瘤对应物的转化，使用一组定义的遗传变化。使用这个模型，我们发现，人骨骼肌成肌细胞可以转化为细胞，产生肿瘤模仿RMS时，作为异种移植物在免疫缺陷小鼠。在确定骨骼肌来源的原代人类细胞可以引起RMS之后，我们研究了在其中表达PAX 3-FKHR的影响，并发现了可能是其致癌行为基础的两种表型。首先，当PAX 3-FKHR作为早期遗传变化稳定表达时，它能够绕过衰老检查点，并作为骨骼肌肿瘤发展的起始致癌打击。第二，当PAX 3-FKHR作为晚期遗传变化稳定表达时，它将体内肿瘤形成的潜伏期从11周缩短至2周，这可能是通过Ras途径的激活，因为在对照实验中，PAX 3-FKHR可以在功能上替代RAS癌基因。在这个提议中，我们希望了解PAX 3-FKHR如何能够绕过衰老检查点，以及它如何加速先前转化细胞的肿瘤发生。为了实现这一点，我们将（1）使用功能获得和功能丧失方法检查PAX 3-FKHR下游的候选蛋白在克服衰老检查点中的作用，以及（2）检查加速肿瘤细胞在生长信号传导、凋亡和/或血管生成中增强的自给自足，以及Ras途径在这种PAX 3-FKHR增强的肿瘤发生中的作用。这些目标的实现将为深入了解这种儿科恶性肿瘤的起源提供帮助，并为研究提供新的治疗靶点。此外，这种基因定义的模型将作为其他人类肉瘤系统研究的模板。 公共卫生相关性：这项研究独特地模拟了导致小儿横纹肌肉瘤的一系列致癌事件。它有望深入了解这种癌症的起源，为研究提供新的治疗靶点，并作为其他人类肉瘤系统分析的模板。

DESCRIPTION (provided by applicant): Rhabdomyosarcoma (RMS) is a heterogeneous collection of cancers demonstrating varying degrees of skeletal muscle differentiation. Although accounting for ~8% of pediatric malignant solid tumors, RMS is the most common soft tissue sarcoma in children younger than 14 years. The two major histologic subtypes of RMS are embryonal (eRMS) and alveolar (aRMS). High risk patients have a 5-year survival of 30%, and outcome is very poor for children whose tumors express the PAX3-FKHR fusion gene; when metastatic, their 5-year survival is <8%. This signature genetic change is found only in aRMS and considered a tumor-specific oncogene, but has no molecularly targeted treatment. To address gaps in knowledge of RMS, we have created a new model for this disease based on the conversion of primary human skeletal muscle cells to their tumorigenic counterpart, using a defined set of genetic changes. Using this model, we found that human skeletal muscle myoblasts may be converted to cells that generate tumors mimicking RMS when tested as xenografts in immunodeficient mice. Having established that primary human cells of skeletal muscle origin can give rise to RMS, we studied the repercussions of expressing PAX3-FKHR in them, and discovered two phenotypes that may underlie its oncogenic behavior. First, when PAX3-FKHR was stably expressed as an early genetic change, it enabled bypass of the senescence checkpoint and served as an initiating oncogenic hit for the development of skeletal muscle tumors. Second, when PAX3-FKHR was stably expressed as a late genetic change, it shortened the latency of in vivo tumor formation from 11 to 2 weeks, possibly through activation of the Ras pathway, since in control experiments PAX3-FKHR could functionally substitute for the RAS oncogene. In this proposal, we wish to understand how PAX3-FKHR enables bypass of the senescence checkpoint, and how it accelerates tumorigenesis in previously transformed cells. To accomplish this, we will (1) examine candidate proteins that are downstream of PAX3-FKHR for their role in overcoming the senescence checkpoint, using both gain-of-function and loss-of-function approaches, and (2) examine the accelerated tumor cells for enhanced self-sufficiency in growth signaling, apoptosis, and/or angiogenesis, and the role of the Ras pathway in this PAX3-FKHR-augmented tumorigenesis. The accomplishment of these aims will provide insight into the genesis of this pediatric malignancy, and provide new therapeutic targets for study. In addition, this genetically defined model will serve as a template for the systematic investigation of other human sarcomas. PUBLIC HEALTH RELEVANCE: This research uniquely models the series of oncogenic events causing the pediatric cancer rhabdomyosarcoma. It is expected to yield insight into the genesis of this cancer, provide new therapeutic targets for study, and serve as a template for the systematic analysis of other human sarcomas.