Pathogenesis of Malignant Mesothelioma by the Human Polycomb Complex BAP1-ASXL

项目摘要/摘要： 恶性间皮瘤(MM)是一种与石棉接触有关的侵袭性、抗药性癌症。这个 多发性骨髓瘤的遗传学基础历来集中在CDKN2A和NF2的体细胞突变作为关键变化 影响启动和进展的。最近，BAP1泛素羧基末端水解酶，第一 被分离为一株CH.3p21肿瘤抑制因子在1998年(Rauscher Lab)，已被强烈牵连为主要 基于遗传分析的MM球员。在两个高发的家系中发现了BAP1基因突变 MM和其他癌症的发生率，以及MMS中体细胞BAP1的改变，与双等位基因一致 肿瘤抑制因子的失活(Testa Lab)。此外，体细胞BAP1突变在散发性疾病中很常见。 散发性葡萄膜黑色素瘤和家族性葡萄膜黑色素瘤(UM)。遗传和生化机制的研究 哪些BAP1突变易患MM和UM以及BAP1与CDKN2A和NF2如何在基因上相互作用 影响多发性骨髓瘤的病理、预后和治疗反应在很大程度上是未知的。然而，事实是 BAP1编码一种核定位的去泛素酶，它与ASXL1/2结合，ASXL1/2是专有的多梳家族伴侣 以组蛋白为靶点的蛋白质强烈暗示在基因转录的表观遗传调控中发挥作用。此外， 由于BAP1与ASXL1/2的结合是酶活性和肿瘤抑制所必需的，因此替代 失活BAP1抑癌活性可能是通过突变或沉默ASXL1/2基因来实现的。要同时定义 多发性骨髓瘤BAP1、ASXL1/2、CDKN2A和NF2的生化机制及遗传互作 Rauscher和Testa实验室联手追求以下具体目标：1)在体内直接使用 用遗传学方法确定杂合子BAP1突变(+/mut)小鼠是否易患 各种自发性肿瘤，包括MM和加速石棉诱导的MM。2)使用条件基因敲除 小鼠确定BAP1在间皮细胞中的躯体丢失是否能够驱动间皮细胞 如果体细胞BAP1失活结合NF2和CDKN2a的丢失导致 更快的发展和/或更具侵袭性的疾病表型，具有潜在的预后和新奇 治疗方面的影响。3)确定人类多发性骨髓瘤标本/细胞中BAP1和ASXL1/2突变的谱 并确定这些突变如何影响与ASXL1/2和ASXL1/2的复合体中BAP1的结合和功能 其他PR-DUB复合蛋白，并定义转录组和细胞表型，它们被改变为 MM中BAP1和其他PR-DUB成分的变化(通过击倒和重建)的后果 细胞。4)确定含有野生-DUB的天然PR-DUB大分子蛋白质复合体的组成 间皮细胞和BAP1和/或ASXL1/2缺陷型MM细胞中BAP1和ASXL1/2的比较 正常和肿瘤特异性亚单位结构以及这些复合体在表观遗传失调中的作用。这个 由两个具有互补专业知识的私人投资机构进行的拟议多管齐下的研究代表了一种全面的方法 为多发性骨髓瘤的发病机制提供新的见解，同时为治疗和预防提供新的靶点。

PROJECT SUMMARY/ABSTRACT: Malignant mesothelioma (MM) is an aggressive, treatment-resistant cancer linked to asbestos exposure. The genetic basis for MM has historically focused on somatic mutations of CDKN2A and NF2 as key alterations influencing initiation and progression. Very recently, the BAP1 ubiquitin carboxy-terminal hydrolase, first isolated as a Ch. 3p21 tumor suppressor in 1998 (Rauscher Lab), has been strongly implicated as a major player in MM based on genetic analyses. Germline BAP1 mutations were found in two families with a high incidence of MM and other cancers, and somatic BAP1 alterations occurred in MMs, consistent with biallelic inactivation of a tumor suppressor (Testa Lab). Moreover, somatic BAP1 mutations are common in sporadic MMs and in both sporadic and familial uveal melanoma (UM). The genetic and biochemical mechanisms by which BAP1 mutations predispose to MM and UM and how BAP1 interacts genetically with CDKN2A and NF2 to influence MM pathology, prognosis, and therapeutic response are largely unknown. However, the fact that BAP1 encodes a nuclear-localized de-ubiquitinase that binds to ASXL1/2, an obligate polycomb family partner protein that targets histones, strongly suggests a role in epigenetic regulation of gene transcription. Moreover, since binding of BAP1 to ASXL1/2 is required for enzyme activity and tumor suppression, an alternative way to inactivate BAP1 tumor suppressor activity may be to mutate or silence ASXL1/2 genes. To define both the biochemical mechanisms and the genetic interactions among BAP1, ASXL1/2, CDKN2A and NF2 in MM, the Rauscher and Testa Labs have joined forces to pursue the following Specific Aims: 1) Use a direct in vivo genetic approach to determine if heterozygous Bap1-mutant (+/mut) mice are predisposed to the development of various spontaneous tumors, including MM, and accelerate asbestos-induced MM. 2) Use conditional knockout mice to determine if somatic loss of Bap1 alone in the mesothelium is capable of driving mesothelial hyperplasia and/or frank MM, and if somatic Bap1 inactivation combined with loss of Nf2 and Cdkn2a results in more rapid development and/or a more aggressive disease phenotype, with potential prognostic and novel therapeutic implications. 3) Define the spectrum of BAP1 and ASXL1/2 mutations in human MM specimens/cell lines and determine how these mutations affect binding and function of BAP1 in complex with ASXL1/2 and other PR-DUB complex proteins, and define the transcriptome and cellular phenotype which are altered as a consequence of changes (via knockdown and reconstitution) in BAP1 and other PR-DUB components in MM cells. 4) Determine the composition of the native PR-DUB macromolecular protein complexes that contain wild- type BAP1 and ASXL1/2 in mesothelial cells and in BAP1- and/or ASXL1/2-deficient MM cells to compare normal- and tumor-specific subunit structure and the role of these complexes in epigenetic dysregulation. The proposed multipronged studies by two PIs with complementary expertise represent a comprehensive approach to yield novel insights into MM pathogenesis while providing new targets for therapy and prevention.