CELL CYCLE CHECKPOINTS AND CHEMOSENSITIVITY OF HUMAN CANCER CELLS

人类癌细胞的细胞周期检查点和化学敏感性

• 批准号: 5201364
• 负责人: P M O'CONNOR
• 金额: --
• 依托单位: DIVISION OF CANCER TREATMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/5201364
• 关键词: DNA damage; DNA repair; antimitotics; antineoplastics; cell cycle; cell cycle proteins; chemical information system; cyclins; drug screening /evaluation; enzyme activity; gene mutation; microtubules; neoplasm /cancer genetics; neoplastic cell; protein kinase; protein structure function; tissue /cell culture; transfection; tumor suppressor genes

This research project focuses on cell cycle checkpoints and the roles these systems play in determining chemosensitivity. We also aim to utilize our emerging knowledge of these checkpoint systems to design new therapeutic stratagems for cancer treatment. We are elucidating the checkpoints that respond to DNA damage to arrest cells in G1 and G2 phases. We are also defining defects in these systems in cancer cells. We aim to trace these systems from the point where DNA damage or unreplicated DNA is sensed by the cell to the response elements that interact with the cyclin-dependent kinases to arrest cell cycle progression or in some cases induce apoptosis. Our results suggest that p53 mutations i lymphoma cell lines confer decreased sensitivity to DNA damaging agents. but not antimitotic agents. Decreased sensitivity is related to an evasion of p53-mediated apoptosis, although other factors can also contribute to the outcome. We have been exploring domains in the p53 regulated gene product, Waf1/Cip1, that interact with cyclin E/Cdk2 and PCNA. Our results suggest two regions in the amino terminus of Waf1/Cip1 are involved in cyclin E/Cdk2 interaction and/or inhibition of this kinase while the carboxy-terminal region of Waf1/Cip1 is involved in PCNA interaction. The G2 checkpoint appears to protect cells from DNA damage by extending the time available for DNA repair. We have found that pentoxifylline preferentially abrogates the G2 checkpoint in cells with disrupted p53 and such cells can be preferentially killed by a combination of a DNA damaging agent and pentoxifylline. We are presently exploring the mechanisms underlying these activities with a special focus on the formation and activation of the G2 cyclin-dependent kinases. We are also investigating several G2 checkpoint abrogators that might be clinically useful in combination protocols to preferentially kill mutant p53 tumors. In conjunction with several other laboratories we have found that the majority of cell lines in the NCI cell screen have disrupted p53 function. We have also found these p53 disrupted cells tend to be less sensitive to a wide variety of currently used chemotherapeutic agents. A notable exception in this analysis was the microtubule agents, whose activity appeared independent of p53 status. Searching the 45,000 compound data base revealed a number of agents which might be preferentially active in cells with disrupted p53. These agents will undergo detailed analysis in isogeneic cell lines in which p53 function has been disrupted by transfection with either the HPV-E6 gene or a dominant negative mutant p53 gene.

该研究项目重点关注细胞周期检查点及其作用 这些系统在确定化学敏感性中发挥作用。 我们还旨在 利用我们对这些检查点系统的新兴知识来设计新的 癌症治疗的治疗策略。 我们正在阐明 对 DNA 损伤作出反应的检查点可将细胞阻滞在 G1 和 G2 期 阶段。我们还定义了癌细胞中这些系统的缺陷。 我们 旨在从 DNA 损伤或未复制的点开始追踪这些系统 DNA 被细胞感知到与 DNA 相互作用的反应元件 细胞周期蛋白依赖性激酶可阻止细胞周期进展或在某些情况下 诱导细胞凋亡。 我们的结果表明淋巴瘤细胞中的 p53 突变 线导致对 DNA 损伤剂的敏感性降低。但不是 抗有丝分裂剂。 敏感性降低与逃避有关 p53 介导的细胞凋亡，尽管其他因素也可能导致 结果。 我们一直在探索 p53 调控基因的域 产品 Waf1/Cip1 与细胞周期蛋白 E/Cdk2 和 PCNA 相互作用。 我们的 结果表明 Waf1/Cip1 氨基末端的两个区域是 参与细胞周期蛋白 E/Cdk2 相互作用和/或抑制该激酶 而 Waf1/Cip1 的羧基末端区域则参与 PCNA 相互作用。 G2 检查点似乎可以保护细胞免受 DNA 损伤 通过延长 DNA 修复的时间。 我们发现 己酮可可碱优先废除细胞中的 G2 检查点 破坏的 p53 和此类细胞可以通过组合优先杀死 DNA损伤剂和己酮可可碱。 我们目前正在探索 这些活动的基础机制，特别关注 G2 细胞周期蛋白依赖性激酶的形成和激活。 我们也是 调查几个可能在临床上起作用的 G2 检查点废除者 可用于优先杀死突变 p53 肿瘤的组合方案。 与其他几个实验室合作，我们发现 NCI 细胞筛选中的大多数细胞系都破坏了 p53 功能。 我们还发现这些 p53 破坏的细胞往往对 目前使用的化疗药物种类繁多。 一个值得注意的 该分析中的例外是微管剂，其活性 似乎与 p53 状态无关。 搜索 45,000 个化合物数据 基地揭示了许多可能优先活跃于 p53 被破坏的细胞。 这些代理将接受详细分析 p53 功能已被破坏的同基因细胞系 用 HPV-E6 基因或显性失活突变体 p53 转染 基因。