NOVEL DNA DOUBLE STRAND BREAK REPAIR TARGETING THERAPEUTICS FOR CANCER TREATMENT

新型 DNA 双链断裂修复靶向治疗癌症

• 批准号: 9187077
• 负责人: LARRY A. SKLAR
• 金额: $ 14.56万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2017-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9187077
• 关键词: NOVEL; DNA; DOUBLE; STRAND; BREAK

DESCRIPTION: Cancer cells frequently inactivate DNA double strand break (DSB) repair proteins causing genomic instability and the genetic evolution of tumors. Inactivation of these proteins usually does not cause overt defects in DNA DSB repair due to the activity of compensatory repair pathways. We reason that identifying compounds that cripple these compensatory pathways will be the first step in developing therapeutic agents that can sensitize tumor cells, but not normal tissues to genotoxic cancer therapies. The DNA damage response histone protein H2AX is a dosage-dependent tumor suppressor that exhibits reduced expression in many cancers. H2AX deficiency leads to genomic instability, but not to overt defects in DNA DSB repair by non-homologous end joining (NHEJ). Deficiency in the NHEJ factor XLF also does not lead to demonstrable defects in NHEJ. However, the combined deficiency of H2AX and XLF leads to a severe block in DSB repair by NHEJ. These findings demonstrate that H2AX and XLF are in compensatory DSB repair pathways and that inhibition of one pathway in cells deficient in the other will lead to a profound block in NHEJ. In conjunction with the University of New Mexico Center for Molecular Discover (UNMCMD), we have developed a flow cytometric high throughput screen for chemical compounds that inhibit NHEJ. This screen has been optimized and used to screen a small library of compounds (1,500) achieving Z' scores of >0.6. Here, we propose to use this approach to screen a larger, more complex chemical library to identify compounds that inhibit NHEJ specifically in H2AX-deficient, but not wild type cells. We refer to these compounds as H2AX-specific Synthetic Inhibitors of NHEJ (SINs). A linear, multistep approach to validate the function of these compounds will also be developed. We believe that developing agents that selectively inhibit NHEJ in cancer cells will ultimately enable us to exploit DNA repair pathway defects to treat cancers. Moreover, this approach has significant advantages over current efforts to selectively inhibit the other major pathway of DSB repair, homologous recombination (HR) in cancer cells. Whereas HR only functions in DSB repair in dividing cells, NHEJ is required for DSB repair in all cells in the G0-G1 phases of the cell cycle. Thus, agents identified by our screen, which inhibit NHEJ will be effective in treating all cancer cells including the vast majority that are not actively dividing. Importantly, cancer stem cells, which are thought to be the basis for relapse of many cancers after treatment, exist primarily in G1 and would also be sensitized by SINs. In addition to identifying H2AX-specific SINs, completion of these studies will establish a linear pipeline for th future identification of SINs for treating cancers defective in a broad range of DNA DSB repair proteins. Moreover, they will also identify novel DSB repair pathways used by normal cells and cancer cells.

描述：癌细胞经常使 DNA 双链断裂 (DSB) 修复蛋白失活，导致基因组不稳定和肿瘤的遗传进化。由于补偿性修复途径的活性，这些蛋白质的失活通常不会导致 DNA DSB 修复的明显缺陷。我们推断，识别削弱这些补偿途径的化合物将是开发可以使肿瘤细胞（而不是正常组织）对基因毒性癌症疗法敏感的治疗药物的第一步。 DNA 损伤反应组蛋白 H2AX 是一种剂量依赖性肿瘤抑制因子，在许多癌症中表现出表达降低。 H2AX 缺陷会导致基因组不稳定，但不会导致非同源末端连接 (NHEJ) 造成的 DNA DSB 修复中的明显缺陷。 NHEJ 因子 XLF 的缺乏也不会导致 NHEJ 明显的缺陷。然而，H2AX 和 XLF 的联合缺陷导致 NHEJ 的 DSB 修复严重受阻。这些发现表明，H2AX 和 XLF 处于补偿性 DSB 修复途径中，并且在缺乏另一种途径的细胞中抑制一种途径将导致 NHEJ 的严重阻断。我们与新墨西哥大学分子发现中心 (UNMCMD) 合作，开发了一种流式细胞术高通量筛选方法，用于筛选抑制 NHEJ 的化合物。该筛选已经过优化，可用于筛选一个小型化合物库 (1,500)，实现 Z' 分数 >0.6。在这里，我们建议使用这种方法来筛选更大、更复杂的化学库，以鉴定特异性抑制 H2AX 缺陷细胞而非野生型细胞中 NHEJ 的化合物。我们将这些化合物称为 H2AX 特异性 NHEJ 合成抑制剂 (SIN)。还将开发一种线性、多步骤的方法来验证这些化合物的功能。我们相信，开发选择性抑制癌细胞中 NHEJ 的药物最终将使我们能够利用 DNA 修复途径缺陷来治疗癌症。此外，与目前选择性抑制 DSB 修复的另一个主要途径——癌细胞中的同源重组 (HR) 的方法相比，这种方法具有显着的优势。 HR 仅在分裂细胞中的 DSB 修复中发挥作用，而 NHEJ 是细胞周期 G0-G1 期所有细胞中 DSB 修复所必需的。因此，我们筛选出的抑制 NHEJ 的药物将有效治疗所有癌细胞，包括绝大多数不活跃分裂的癌细胞。重要的是，癌症干细胞被认为是许多癌症治疗后复发的基础，主要存在于 G1 期，并且也会被 SIN 致敏。除了鉴定 H2AX 特异性 SIN 之外，这些研究的完成还将为未来鉴定 SIN 建立线性管道，用于治疗多种 DNA DSB 修复蛋白缺陷的癌症。此外，他们还将确定正常细胞和癌细胞使用的新 DSB 修复途径。