A novel functional genomic pipeline for target identification in sarcoma

描述(申请人提供)：这项工作的长期目标是改善尤因肉瘤的预后，尤因肉瘤是儿童中最致命的骨肿瘤。尤因肉瘤家族肿瘤(ESFT)是困扰儿童、青少年和年轻人的第二种最常见的骨和软组织癌症。ESFT的特征是染色体易位导致EWS与ETS转录因子家族的一个成员融合，最常见的是FLI1。ESFT接受强化化疗、放射治疗和手术治疗，但超过一半的患者在确诊后五年内死于疾病，而幸存者往往遭受治疗的长期有害影响。目前迫切需要开发有效的、降低毒性的靶向治疗方法。不幸的是，我们仍然有一个有限的 对ESFT肿瘤发生的分子机制的了解，这极大地阻碍了改进治疗方法的确定。我们的初步数据表明，尤文肉瘤中存在染色体获得和丢失的复发区，这表明这些复发区的协同基因的扩增或缺失促进了EWS-FLI1的细胞转化。识别和验证这些合作基因对于新疗法的开发至关重要，因为针对EWS-FLI1本身的策略到目前为止还没有成功。我们之前开发了一个斑马鱼ESFT模型，概括了人类疾病的关键特征。我们现在将使用FISH模型进行高通量功能基因组分析，以验证从我们对人类ESFT的高分辨率基因组分析中出现的有希望的EWS-FLI1效应基因。我们将进一步利用这一实验，进行小分子筛选，以确定作为EWS-FLI1介导的细胞转化抑制剂的先导化合物。为了实现这一目标，我们将1)通过对肿瘤的高分辨率基因组分析来确定关键的EWS-FLI1效应器；2)使用斑马鱼体内EWS-FLI1活性的模型来验证基因组拷贝数分析中确定的候选ESFT效应器；以及3)在斑马鱼模型中识别能够抑制EWS-FLI1功能的小分子。这一新的渠道将利用我们在拷贝数量分析和使用斑马鱼模型的比较肿瘤学方面的综合专业知识。这一新的渠道及其新的方法将导致对肉瘤的快速测试和引入新的治疗剂，肉瘤是儿童和成人最致命的癌症之一。

DESCRIPTION (provided by applicant): The long-term goal of this work is to improve outcomes in Ewing Sarcoma, the most lethal bone tumor of children. Ewing Sarcoma Family Tumors (ESFT) are the second most common bone and soft tissue cancer afflicting children, adolescents, and young adults. ESFT are characterized by a chromosomal translocation resulting in a fusion between EWS and a member of the ETS family of transcription factors, most commonly FLI1. ESFT are treated with intensive chemotherapy, radiation, and surgery, yet more than half of patients die of disease within five years of diagnosis, while survivors often suffer long-term deleterious effects of treatment. An immediate need exists for the development of effective and targeted therapies with decreased toxicity. Unfortunately, we still have a limited understanding of the molecular mechanisms of ESFT tumorigenesis, which has greatly impeded the identification of improved therapies. Our preliminary data indicate that recurrent areas of chromosomal gain and loss occur in Ewing Sarcoma, suggesting that amplification or deletion of cooperating genes in these recurring regions facilitate cell transformation by EWS-FLI1. Identification and validation of these cooperating genes is essential for the development of new therapies, because strategies to target EWS-FLI1 itself have not to date been successful. We previously developed a zebrafish model of ESFT that recapitulates key features of the human disease. We will now use the fish model for high-throughput functional genomic assays to validate promising candidate EWS-FLI1 effector genes emerging from our high- resolution genomic analyses of human ESFT. We will further capitalize on this assay by conducting small- molecule screens to identify lead compounds that act as inhibitors of EWS-FLI1-mediated cellular transformation. To achieve this goal we will 1) identify critical EWS-FLI1 effectors though high-resolution genomic analysis of tumors; 2) use zebrafish in vivo models of EWS-FLI1 activity to validate candidate ESFT effectors identified in genomic copy number analyses; and 3) identify small molecules capable of inhibiting EWS-FLI1 function in the zebrafish model At the completion of this study, we will have demonstrated the effectiveness of a novel genomic pipeline for discovery and functional analysis of cooperating genes in translocation positive sarcomas. This novel pipeline will take advantage of our combined expertise in copy number analysis and comparative oncology using zebrafish models. This new pipeline and its novel approach will lead to the rapid testing and introduction of new therapeutic agents for sarcomas, one of the most deadly types of cancer in children and adults.