THE GENETICS OF OCULAR MELANOMA

描述（由申请人提供）： 翻译后摘要：转移是恶性肿瘤的一个定义特征，是癌症相关死亡的最常见原因。然而，转移的遗传学知之甚少。葡萄膜黑色素瘤（UM）是眼睛最常见的原发性癌症，也是黑色素瘤的第二常见形式。UM具有高度特征性的肝转移模式，对常规化疗具有抗性，通常是致命的。UM具有非常小的基因组不稳定性，很少的细胞遗传学改变和罕见的遗传突变。因此，当在这些肿瘤中发现突变时，它们很可能是驱动突变而不是乘客突变。基于经验证的基因表达特征，可以根据转移性死亡的风险将UM分为1类（低风险）和2类（高风险）。研究的一个主要重点是确定特定的遗传变化，赋予转移能力的UM。2类特征通常伴随着3号染色体的一个拷贝的丢失（3号单体），这导致了广泛的预期，即在UM细胞中3号染色体的一个拷贝的丢失揭示了3号染色体的剩余拷贝上的一个基因（或多个基因）中的隐性失活突变，其赋予转移能力。其他变化包括染色体8 q的增加和染色体8 p的丢失。在1类肿瘤中，染色体6p的获得是常见的。这项建议的研究人员是UM分子遗传学分析的先驱。我们是第一组应用最近描述的外显子组捕获技术，然后进行大规模平行测序，以确定BAP 1作为UM 3号染色体的转移抑制基因。在目前的研究中，我们将利用我们最近的成功与这些技术，以确定额外的肿瘤抑制基因和癌基因突变的UM。在目标1中，我们将生成1类和2类肿瘤的额外外显子组序列，将它们与其匹配的生殖系DNA进行比较。将使用桑格测序确认携带有害突变的潜在肿瘤抑制基因和携带驱动UM发展的潜在激活突变的癌基因，并在另外10-30例1类和2类肿瘤和匹配的生殖系DNA中进行评价。在目标2中，我们将使用靶向捕获来重新测序额外样品（每种肿瘤类型>100个）中这些基因内新鉴定的额外突变，以确定它们对UM的贡献。在目标3中，我们将在细胞系中对5个新鉴定的基因进行有限的功能研究。我们将进行结合试验，以评估错义和框内编码变化对与已知和新型伴侣相互作用的影响，并过度表达活化癌基因和敲低肿瘤抑制因子，以确定其对细胞形态和基因表达的影响。然后将肿瘤分子改变的信息与临床信息结合，开始发展UM的预后分类。这是一个来自眼科和遗传学部门的多学科合作小组的合作提案，该小组拥有经过验证的专业知识来完成本提案的目标。) 公共卫生相关性： 个人叙述。葡萄膜（眼黑色素瘤）是眼睛最常见的原发性癌症，也是黑色素瘤的第二常见形式。它可以根据其因转移而死亡的风险进行分类。与某些癌症不同，UM相对简单，并且可能有许多易于处理的遗传改变推动其发展。我们最近使用外显子组捕获和NexGen测序的方法来鉴定UM的转移抑制因子。当前应用的目标是识别剩余的重要癌症基因。这具有诊断和预后意义，并可能导致这种毁灭性癌症的新形式治疗的发展。

DESCRIPTION (provided by applicant): Abstract: Metastasis is a defining feature of malignant tumors and is the most common cause of cancer-related death. However, the genetics of metastasis are poorly understood. Uveal melanoma (UM) is the most common primary cancer of the eye and the second most common form of melanoma. UMs have a highly characteristic pattern of metastasis to the liver that is resistant to conventional chemotherapy and is usually fatal. UMs have remarkably little genomic instability, few cytogenetic alterations, and rare genetic mutations. Thus, when mutations are found in these tumors, they are highly likely to be driver rather than passenger mutations. UMs can be grouped according to risk of metastatic death into class 1 (low risk) and class 2 (high risk) based on a validated gene expression signature. A major focus of research has been to identify the specific genetic changes that confer metastatic competency in UM. The class 2 signature is usually accompanied by loss of one copy of chromosome 3 (monosomy 3), which has led to the widespread expectation that loss of one copy of chromosome 3 in UM cells unmasks recessive inactivating mutations in a gene (or genes) on the remaining copy of chromosome 3 that confer metastatic capacity. Other changes include gain of chromosome 8q and loss of chromosome 8p. In the class of class 1 tumors, gain of chromosome 6p is common. The)investigators of this proposal are pioneers in the analysis of the molecular genetics of UM. We were the first group to apply the recently described technique of exome capture followed by massively parallel sequencing to identify BAP1 as the metastasis suppressor mapping to chromosome 3 in UM. In the current study we will capitalize on our recent success with these technologies to identify additional tumor suppressor genes and oncogenes mutated in UM. In Aim 1 we will generate additional exome sequences of both class 1 and class 2 tumors, comparing them with their matched germline DNA. Potential tumor suppressor genes harboring deleterious mutations, and oncogenes harboring potential activating mutations driving the development of UM will be confirmed with Sanger sequencing and evaluated in an additional 10-30 class 1 and class 2 tumors and matched germline DNA. In Aim 2 we will use targeted capture to re-sequence newly identified additional mutations within these genes in additional samples (>100 of each tumor type) to determine their contribution to UM. In Aim 3 we will perform limited functional studies of 5 newly identified genes in cell lines. We will perform binding assays to evaluate the effects of mis-sense and in-frame coding changes on interactions with known and novel partners, and over-express activating oncogenes and knockdown tumor suppressors to determine their effect on cell morphology and gene expression. Information on molecular alterations in tumors will then be incorporated with clinical information to begin to develop a prognostic classification of UM. This is a collaborative proposal from a multidisciplinary collaborative group from the departments of Ophthalmology and Genetics that has the proven expertise to complete the aims of this proposal. ) PUBLIC HEALTH RELEVANCE: Personal Narrative. Uveal (ocular melanoma) is the most common primary cancer of the eye and the second most common form of melanoma. It can be classified according to their risk of death due to metastasis. Unlike some cancers, UM is relatively simple, and there are likely to be a tractable number of genetic alterations driving their development. We recently used the approach of exome capture and NexGen sequencing to identify a metastasis suppressor for UM. The goal of the current application is to identify the remaining important cancer genes. This has both diagnostic and prognostic implications, and may lead to the development of novel forms of treatment for this devastating cancer.