Genomic Analysis of Tumor Context Vulnerabilities in Human Metastatic Melanoma

描述（由申请人提供）：尽管经过数十年的研究，转移性恶性黑色素瘤仍然是一种不治之症。只有少数诊断为转移性黑色素瘤的患者显示出对化疗的临床反应，而治愈的患者更少;播散性疾病患者的5年生存率为5- 18%。该项目的重点和基本目标是确定黑色素瘤中的特定遗传变化，通过功能基因组筛选和体内验证证实，这提供了可以在晚期黑色素瘤中利用的遗传脆弱性的背景。具体来说，我们建议进行高通量功能性RNA干扰（RNAi）筛选，以系统地识别介导黑色素瘤细胞敏感性的基因（具体目标1）。随后，我们将通过确认siRNA介导的基因敲除体外验证这些基因的作用（特异性目标2），使用组织微阵列（TMA）表征人黑色素瘤中的蛋白质表达，进行高含量的机制分析以阐明这些基因介导黑色素瘤细胞体外敏感性的机制，并评估体外抑制这些基因与选择剂组合的有效性。最后，我们将评估我们验证的遗传靶点是否可以添加到针对黑素瘤异种移植模型的体内分子信息组合疗法中（具体目标3）。我们假设，这种基于功能的基因组方法将确定与肿瘤进展相关的遗传扰动的背景，这些遗传扰动对于使黑色素瘤细胞对化疗更具抗性至关重要。我们推测，这些经过验证的基因可以促进这些肿瘤背景脆弱性的联合靶向，并可能与临床开发直接相关。相关性：诊断为晚期黑色素瘤的患者的长期预后很差，5年生存率为5- 18%。这项拟议的研究旨在确定转移性黑色素瘤细胞获得的遗传脆弱性，这些遗传脆弱性可能被用来提高对这种疾病的临床干预的有效性。

DESCRIPTION (provided by applicant): Despite decades of research, metastatic malignant melanoma remains an incurable disease. Only a minority of patients diagnosed with metastatic melanoma show a clinical response to chemotherapy, while still fewer are cured; the 5-year survival rate for patients with disseminated disease ranges from 5-18%. The focus and underlying goal of this project is to identify specific genetic changes in melanoma, confirmed through both functional genomic screening and in vivo validation, which provide a context of genetic vulnerability that can be exploited in advanced stage melanoma. Specifically, we are proposing to undertake a high-throughput functional RNA interference (RNAi) screen to systematically identify genes that mediate melanoma cell sensitivity (Specific Aim 1). We will subsequently validate the role of these genes (Specific Aim 2) by confirming siRNA-mediated gene knockdown in vitro, characterizing protein expression in human melanomas using tissue microarrays (TMAs), performing high-content mechanistic assays to elucidate the mechanism by which these genes mediate melanoma cell sensitivity in vitro, and evaluating the effectiveness of inhibiting these genes in vitro in combination with the selecting agent. Lastly, we will evaluate whether our validated genetic targets can add to molecularly-informed combination therapies in vivo against xenograft models of melanoma (Specific Aim 3). We hypothesize that this functional-based genomic approach will identify the context of genetic perturbations associated with tumor progression that are critical to rendering melanoma cells more resistant to chemotherapy. We speculate that these validated genes could facilitate the combined targeting of these tumor-context vulnerabilities and could be of direct relevance to clinical exploitation. Relevance: The long-term prognosis for patients diagnosed with advanced stage melanoma is dismal, with a five-year survival rate ranging from 5-18%. The proposed research aims to identify genetic vulnerabilities acquired by metastatic melanoma cells that may be exploited to improve the effectiveness of clinical intervention in this disease.