Molecular determinants of PARP inhibitor sensitivity in 11q-deleted malignancy

11q 缺失恶性肿瘤中 PARP 抑制剂敏感性的分子决定因素

• 批准号: 8881505
• 负责人: Sonia Franco
• 金额: $ 18.45万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2017-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8881505
• 关键词: 11q; 11q22; 11q23; Aftercare; Alleles; Antineoplastic Agents; B-Cell Lymphomas; B-Lymphocytes; BRCA1 gene; Biochemical; Biological Assay; Biological Markers; Cell Cycle Progression; Cell Line; Cell Survival; Cells; Chromosomal Stability; Clinical; Clinical Trials; DNA Damage; DNA Double Strand Break; DNA Repair; Data; Defect; Development; Dose; Double Strand Break Repair; Embryo; Fibroblasts; Future; Gene Dosage; Genes; Genetic; Genetic Epistasis; Genetic Recombination; Genome; Genomic Instability; Human; Ionizing radiation; Kinetics; Knowledge; Lesion; Malignant Neoplasms; Mediating; Modeling; Molecular; Molecular Cytogenetics; Mus; Mutation; Natural History; Neuroblastoma; Outcome; PARP inhibition; Pathway interactions; Phenotype; Phosphotransferases; Post-Translational Protein Processing; Pre-Clinical Model; Radiation Tolerance; Radiation induced double strand break; Relative (related person); Reporting; Role; Series; Signal Transduction; Testing; Work; ataxia telangiectasia mutated protein; base; cancer cell; cell transformation; chemotherapeutic agent; chemotherapy; genetic approach; homologous recombination; human H2AX protein; inhibitor/antagonist; insight; knock-down; neoplastic cell; neuroblastoma cell; novel; public health relevance; repaired; research clinical testing; research study; response; tumor

 DESCRIPTION (provided by applicant): Deletions at 11q are common in human malignancies and associate with poor clinical outcomes. In this context, deletions at 11q22, containing the Ataxia-Telangiectasia Mutated (ATM) locus, are frequently observed in B cell lymphomas (BCL) and, combined with mutation of the second allele, render these tumors ATM null. In contrast, deletions at 11q23, containing the histone H2AX locus, result in monoallelic H2AX expression in a subset of neuroblastomas (NB). Given the prominent roles for the ATM kinase and its substrate histone H2AX in the signaling and repair of DNA double-strand breaks (DSB), alterations in their copy number may result in differential responses to DNA damaging agents in cancer cells. In this context, PARP inhibitors (PARPi) are a novel class of DNA damaging chemotherapeutic agents that preferentially eliminate cells with defects in replication-associated DSBs. Based on these previous observations, we propose here to test the hypothesis that monoallelic expression of H2AX in 11q-deleted malignancies sensitizes them to PARPi, by conferring a defect in Homologous Recombination (HR)-mediated repair of PARPi-induced DSBs in human cancer cells. In support of our hypothesis, we provide strong genetic evidence that H2AX deficiency is synthetic lethal with PARP inhibition. First, we find that combined loss of H2AX and either of the two main PARPi targets, PARP1 or PARP2, results in embryonic lethality in the mouse. Secondly, we demonstrate a defect in the repair of replication-associated DSBs after treatment with PARPi, in an H2AX gene dose- dependent manner. Lastly, we demonstrate that H2AX is also limiting for DSB repair in NB cells with 11q23 deletion and monoallelic H2AX expression. To test our hypotheses, we will employ a combination of biochemical, molecular and cytogenetic assays on mouse primary and transformed cells and in human NB cells. Specifically, experiments in Aim 1 will employ murine cells to test the hypothesis that H2AX becomes limiting for HR-mediated repair of PARPi-induced lesions, characterize the underlying genetic pathway and define the relative contribution of PARP1 and PARP2 to these phenotypes. In Aim 2, we will assess PARPi sensitivity in 11q23-deleted NB cells as a function of H2AX gene dose and define roles for PARP1 and PARP2 in DSB repair in human cancer cells with reduced H2AX expression. In Aim 3, we will employ a novel murine model to examine nonoverlapping functions for ATM and H2AX in the repair of PARPi-induced lesions, modeling the subset of human cancers that co-delete the two factors. In the longer term, knowledge gained from these exploratory studies will increase our understanding of how PARPi interfere with the mechanisms that normally protect the genome during replication and facilitate the development of future clinical trials for biomarker development in a variety of human malignancies with 11q abnormalities.

 描述（由申请人提供）：11q 缺失在人类恶性肿瘤中很常见，并且与不良临床结果相关。在这种情况下，包含共济失调毛细血管扩张突变 (ATM) 位点的 11q22 缺失经常在 B 细胞淋巴瘤 (BCL) 中观察到。 ）并且结合第二个等位基因的突变，使这些肿瘤 ATM 无效。相反，包含组蛋白的 11q23 处的缺失。鉴于 ATM 激酶及其底物组蛋白 H2AX 在 DNA 双链断裂 (DSB) 的信号传导和修复中的重要作用，以及其拷贝数的改变，H2AX 基因座导致神经母细胞瘤 (NB) 亚群中单等位基因 H2AX 的表达。可能会导致癌细胞对 DNA 损伤剂产生不同的反应。在这种情况下，PARP 抑制剂 (PARPi) 是一类新型 DNA 损伤化疗剂，可优先消除复制相关缺陷的细胞。基于这些先前的观察，我们在此建议通过赋予同源重组（HR）介导的人类 PARPi 诱导的 DSB 修复缺陷来检验以下假设：11q 缺失的恶性肿瘤中 H2AX 的单等位基因表达使它们对 PARPi 敏感。为了支持我们的假设，我们提供了强有力的遗传证据，证明 H2AX 缺陷与 PARP 抑制联合导致死亡。 H2AX 和两个主要 PARPi 靶标 PARP1 或 PARP2 中的任何一个都会导致小鼠胚胎致死。其次，我们证明了用 PARPi 治疗后复制相关 DSB 的修复存在缺陷，且呈 H2AX 基因剂量依赖性。 ，我们证明 H2AX 也限制了 11q23 缺失和单等位基因 H2AX 表达的 NB 细胞中的 DSB 修复。为了测试我们的最终假设，我们将采用生化、具体来说，目标 1 中的实验将使用小鼠细胞来测试 H2AX 限制 HR 介导的 PARPi 诱导损伤修复的假设，从而表征潜在的遗传途径。并定义 PARP1 和 PARP2 对这些表型的相对贡献 在目标 2 中，我们将评估 11q23 缺失的 NB 细胞中 PARPi 敏感性作为 H2AX 基因剂量的函数，并定义其作用。 H2AX 表达减少的人类癌细胞 DSB 修复中的 PARP1 和 PARP2 在目标 3 中，我们将采用一种新型小鼠模型来检查 ATM 和 H2AX 在 PARPi 诱导的损伤修复中的非重叠功能，对人类癌症子集进行建模。从长远来看，从这些探索性研究中获得的知识将增加我们对 PARPi 如何干扰复制过程中正常保护基因组的机制的理解，并促进未来临床试验的发展。具有 11q 异常的多种人类恶性肿瘤的生物标志物开发。