Mechanisms of tumorigenesis in Brg1 mutant lung cancer

总结/摘要 肺癌是全球和美国癌症相关死亡的主要原因。也是 非小细胞肺癌（NSCLC）是遗传学上最复杂的疾病之一， 肺鳞状细胞癌和肺腺癌的突变数分别排在第2位和第3位。 在21种不同肿瘤类型的研究中，将这些突变定位到基因区域的努力 已经证明，BRG 1相关因子（BAF）复合物（也称为 哺乳动物的SWI/SNF复合物是常见的。BRG 1中的失活突变，其编码ATP酶， BAF复合物的解旋酶存在于8-20%的NSCLC中。BRG 1（也称为SMARCA 4）通常与 突变的KRAS，并且没有针对这种遗传亚型的肺癌的靶向疗法。生物曝气滤池 复合物长期以来一直被认为是含EZH 2的Polycomb抑制蛋白的表观遗传拮抗剂。 复合物2（PRC 2），但这些复合物在癌症背景下如何相互作用还不清楚。 最近，我们证明了EZH 2抑制剂与拓扑异构酶II（TopoII）抑制剂，如 与依托泊苷在BRG 1突变型NSCLC中的作用相同。我们的数据表明BRG 1在肺癌细胞中的关键作用 包括促进TopoII复杂功能。此外，我们的数据表明，EZH 2抑制的组合 TopoII抑制揭示了BRG 1突变肿瘤细胞对DNA修复缺陷的脆弱性。为了 了解EZH 2，BRG 1和TopoII之间相互作用的机制基础，我们建议使用 一组人类和小鼠同基因模型，其中唯一的差异是存在或不存在BRG 1。 在目的1中，我们将确定BRG 1缺陷在同基因小鼠中发生的DNA修复缺陷的机制 和人肺癌细胞系通过评估双链断裂修复机制的标志物，非- 同源末端连接（NHEJ）和同源重组（HR）。我们还将确定DNA缺陷 修复导致BRG 1突变型肺肿瘤对EZH 2 i/依托泊苷双重治疗的敏感性。在目标2中， 使用小鼠模型来检查EZH 2抑制和依托泊苷的组合对 BRG 1-null肺肿瘤。将使用基因工程小鼠和患者来源的异种移植物（PDX）模型 确定EZH 2和TopoII抑制的组合是否上级现有的标准治疗 BRG 1突变型肺癌在目标3中，我们将描述BRG 1缺陷型小鼠的免疫微环境。 并且我们将评估用依托泊苷治疗的EZH 2抑制对肿瘤免疫细胞的影响。的 这些研究的潜在影响是巨大的，因为BRG 1是最常见的突变基因之一， NSCLC，目前还没有针对该基因亚型的靶向治疗。此外，EZH 2抑制剂具有 转移到其他肿瘤类型的I期试验，以及EZH 2抑制和口服生物可利用的组合 如果这些临床前研究表明有效，依托泊苷（VP-16）可以迅速进入试验阶段。

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.