NOVEL DOUBLE STRAND BREAK PRODUCING COMPOUNDS

新型双链断裂产生化合物

• 批准号: 6173608
• 负责人: JOHN R LAMB
• 金额: $ 25.72万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-07-01 至 2002-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6173608
• 关键词: DNA damage; DNA gyrase; DNA repair; DNA topoisomerases; antineoplastics; cytotoxicity; double stranded RNA; enzyme activity; enzyme structure; gene expression; genetic strain; mutant; neoplasm /cancer pharmacology; tissue /cell culture; yeasts

We propose to identify novel double strand break (DSB) producing compounds which interact with the human tumor cellular context in a manner which is distinct from existing agents. We hypothesize that by using a cell based approach we have identified an entirely new class of DSB producing compounds. For simplicity, these compounds will be referred to throughout this proposal as 'putative DSB producing' compounds. A subset of the new compounds are structurally novel and do not behave like topo I or II poisons. They may thus represent entirely new classes of DSB inducing agents. Topoisomerase poisons are successfully used in the clinic against different cancers. Unfortunately these successes are plagued by severe dose-limiting toxicities and ovelapping mechanisms of resistance. The compounds we propose to study were isolated in a genetic defect-based rather than a narrowly defined target-based screen. The selection criteria eliminate the majority of agents that directly damage DNA. This approach makes no selection based on chemistry or mechanism of action, and may have identified compounds that interact with new classes of proteins to damage DNA. We propose to carry out indicative toxicity tests in yeast and mammalian cells and on purified topoisomerases to clearly categorize the compounds into topoisomerase I poisons, topoisomerase II poisons, and mechanistically novel putative double strand break-causing agents. We will then confirm the production of double strand breaks in vivo. Finally we will identify the protein targets and other cellular factors which influence the cytotoxicity of the compounds. This will be achieved by profiling the toxicity of compounds in a panel of mammalian matched pairs of cell lines +/- defects associated with cancer, and identifying cellular factors in yeast, by overexpression and deletion studies. The yeast experiments will involve both classical genetic approaches and genome-wide analyses based on DNA microarray technology.

我们建议以与现有药物不同的方式鉴定出与人类肿瘤细胞上下文相互作用的新型双链破裂（DSB）。 我们假设通过使用基于细胞的方法，我们已经确定了一类全新的DSB产生化合物。 为简单起见，这些化合物将在整个建议中称为“推定的DSB生产”化合物。 新化合物的一个子集在结构上是新颖的，并且不像托普I或II毒药那样行事。 因此，它们可能代表全新的DSB诱导剂。 拓扑异构酶毒物成功用于诊所，以针对不同的癌症。 不幸的是，这些成功受到严重的剂量限制性毒性和抗药性机制的困扰。 我们建议研究的化合物是在基于遗传缺陷的基于遗传缺陷的基于遗传缺陷的基于靶标的筛查中分离出来的。 选择标准消除了直接损害DNA的大多数药物。 这种方法没有基于化学或作用机理的选择，并且可能已经确定了与新类别蛋白质相互作用的化合物损害DNA。 我们建议在酵母和哺乳动物细胞和纯化的拓扑异构酶中进行指示性毒性测试，以将化合物清楚地分类为拓扑异构酶I毒素，拓扑异构酶II毒药，以及机械学的新型假定假定的双链断裂剂。 然后，我们将确认体内双链断裂的产生。 最后，我们将确定影响化合物细胞毒性的蛋白质靶标和其他细胞因子。 这将通过在哺乳动物匹配的细胞系+/-缺陷对中介绍与癌症相关的细胞系+/-缺陷以及通过过表达和缺失研究鉴定酵母中细胞因子的毒性来实现这一目标。 酵母实验将涉及基于DNA微阵列技术的经典遗传方法和全基因组分析。