Mechanisms of tumorigenesis in Brg1 mutant lung cancer

摘要/摘要 肺癌是全球和美国与癌症相关的死亡的主要原因。它也是 基因最复杂的疾病之一--非小细胞肺癌(NSCLC)的两大亚型， 其中肺鳞癌和肺腺癌的突变最多，分别排在第二和第三位 在一项对21种不同肿瘤类型的研究中发现了大量的DNA。将这些突变映射到基因区域的努力 已经证明BRG1相关因子(BAF)复合体的突变，也被称为 哺乳动物的SWI/SNF复合体是常见的。编码ATPase和ATPase的BRG1基因突变失活 BAF复合体的解旋酶存在于8%-20%的非小细胞肺癌中。BRG1(也称为SMARCA4)通常是联合 与KRAS突变，目前还没有针对这种基因亚型肺癌的靶向治疗。《BAF》 长期以来，人们一直知道复合体是含有EZH2的Polycomb抑制的表观遗传拮抗剂 复合体2(PRC2)，但这些复合体在癌症环境中究竟如何相互作用尚不清楚。 最近，我们证明了EZH2抑制剂与拓扑异构酶II(Topo II)抑制剂有很好的协同作用，如 在BRG1突变型NSCLC中为依托泊苷。我们的数据表明BRG1在肺癌细胞中的关键作用 涉及促进TopoII的复杂功能。此外，我们的数据表明，EZH2抑制的组合 Topo II的抑制揭示了BRG1突变的肿瘤细胞对DNA修复缺陷的脆弱性。为了 为了了解EZH2、BRG1和TopO II之间相互作用的机制基础，我们建议使用一种 人类和小鼠等基因模型的面板，其中唯一的区别是BRG1的存在或不存在。 在目标1中，我们将确定等基因小鼠BRG1缺陷所导致的DNA修复缺陷的机制 和人类肺癌细胞系通过评估双链断裂修复机制的标记，非 同源末端连接(NHEJ)和同源重组(HR)。我们还将确定DNA中的缺陷 修复导致BRG1突变肺肿瘤对双重EZH2/依托泊苷治疗的敏感性。在《目标2》中我们将 用小鼠模型检测EZH2抑制和依托泊苷联合应用的临床前效应 BRG1阴性的肺部肿瘤。将使用基因工程小鼠和患者来源的异种移植(PDX)模型 确定EZH2和TopOII抑制的组合是否优于现有的标准护理治疗 针对BRG1基因突变的肺癌。在目标3中，我们将描述BRG1缺陷的免疫微环境 我们将评估用依托泊苷治疗抑制EZH2对肿瘤免疫细胞的影响。这个 这些研究的潜在影响是巨大的，因为BRG1是人类最常见的突变基因之一 NSCLC，目前还没有针对这种基因亚型的靶向治疗。此外，EZH2抑制剂有 转移到其他肿瘤类型的I期试验，以及EZH2抑制和口服生物利用度的组合 如果这些临床前研究表明有效，依托泊苷(VP-16)可能会迅速进入试验阶段。

Summary/Abstract Lung cancer is the leading cause of cancer-related deaths both worldwide and in the United States. It is also one of the most genetically complex diseases – the two major subtypes of non-small cell lung cancer (NSCLC), which are lung squamous cell carcinoma and lung adenocarcinoma rank 2nd and 3rd for the most mutations per megabase of DNA in a study of 21 different tumor types. Efforts to map these mutations to genetic regions have demonstrated that mutations in the BRG1-Associated Factor (BAF) complex, also known as the mammalian SWI/SNF complex, are common. Inactivating mutations in BRG1, which encodes the ATPase and helicase of the BAF complex, are present in 8-20% of NSCLCs. BRG1 (also known as SMARCA4) is often co- mutated with KRAS, and there are no targeted therapies for this genetic subtype of lung cancer. The BAF complex has long been known to be the epigenetic antagonist of the EZH2-containing Polycomb Repressive Complex 2 (PRC2), but exactly how these complexes interact in the cancer setting is not well understood. Recently, we demonstrated that EZH2 inhibitors synergize well with Topoisomerase II (TopoII) inhibitors such as etoposide in BRG1 mutant NSCLCs. Our data suggest that a key role for BRG1 in lung cancer cells involves facilitating TopoII complex function. Furthermore, our data suggest that combination of EZH2 inhibition and TopoII inhibition uncovers the vulnerabilities of BRG1 mutant tumor cells to DNA repair defects. In order to understand the mechanistic basis of the interactions between EZH2, BRG1 and TopoII, we propose to use a panel of human and mouse isogenic models in which the only difference is the presence or absence of BRG1. In Aim 1 we will identify mechanisms of DNA repair defects that occur with BRG1 deficiency in isogenic murine and human lung cancer cell lines by assessing markers of double strand break repair mechanisms, non- homologous end joining (NHEJ) and homologous recombination (HR). We will also determine if defects in DNA repair cause the sensitivity of BRG1 mutant lung tumors to dual EZH2i/etoposide treatment. In Aim 2 we will use mouse models to examine the preclinical effects of combination of EZH2 inhibition and etoposide against BRG1-null lung tumors. Genetically engineered mice and patient derived xenograft (PDX) models will be used to determine if the combination of EZH2 and TopoII inhibition is superior to standard of care therapies in place for BRG1-mutant lung cancers. In Aim 3 we will characterize the immune microenvironment of BRG1-deficient tumors and we will assess the impact of EZH2 inhibition with etoposide treatment on tumor immune cells. The potential impact of these studies is large because BRG1 is one of the most commonly mutated genes in NSCLC and there is currently no targeted therapy for this genetic subtype. In addition, EZH2 inhibitors have moved to Phase I trials for other tumor types, and the combination of EZH2 inhibition and orally bioavailable etoposide (VP-16) could rapidly move to trials if these preclinical studies suggest efficacy.