Bacterial cell killing by topoisomerase I mediated DNA lesion

项目描述（由申请人提供）：本项目以细菌IA型拓扑异构酶为目标，发现新型抗菌药物，作为多重耐药细菌病原体（包括革兰氏阴性）的对策。通过捕获由IB型和IIA型拓扑异构酶形成的共价切割复合物而启动细胞杀伤的药物被广泛应用于当前的抗癌和抗菌治疗中。IA型拓扑异构酶切割复合物的积累可通过细胞氧化死亡途径引发细菌细胞的快速死亡，但IA型细菌拓扑异构酶I的特异性抑制剂仍有待发现，可以开发成新的抗菌药物。下一个资助期的拟研究活动将开发和利用基于快速细胞的适用于HTS检测格式，以鉴定由于与细菌拓扑异构酶I特异性相互作用而具有抗菌活性的小分子化合物。从筛选中鉴定出的细菌拓扑异构酶I抑制剂将进行生物化学分析，以确定其作用机制和结构活性关系。将优先考虑对革兰氏阴性大肠杆菌和鼠疫耶尔森菌有活性的低毒化合物，以便确定可用于开发针对革兰氏阴性病原体的治疗药物的线索。处理被捕获的拓扑异构酶切割复合物的细菌蛋白将通过遗传和生化实验进行鉴定。所涉及的修复蛋白可能是针对IA型和IIA型拓扑异构酶的抗生素联合治疗的有用的新靶点。这些拟议实验的成功将为开发新型抗菌药物提供工具和线索，以解决细菌病原体对所有现有抗生素具有耐药性的紧迫公共卫生问题。

DESCRIPTION (provided by applicant): This project targets bacterial type IA topoisomerase for discovery of novel antibacterial drugs as countermeasure for multi-drug resistant bacterial pathogens, including gram negatives. Drugs that initiate cell killing by trapping the covalent cleavage complex formed by type IB and type IIA topoisomerases are widely used in current anti-cancer and anti-bacterial therapy. Accumulation of type IA topoisomerase cleavage complex can trigger rapid bacterial cell death via the oxidative cell death pathway but specific inhibitors of type IA bacterial topoisomerase I that can be developed into new antibacterial drugs remain to be discovered. The proposed research activities for the next funding period would develop and utilize rapid cell based applicable for HTS assay format to identify small molecule compounds that have antibacterial activity due to specific interaction with bacterial topoisomerase I. Inhibitors of bacterial topoisomerase I identified from screening would be analyzed biochemically for mechanism of action and structure activity relationship. Priority would be given to compounds with low toxicity that are active against gram negative Escherichia coli and Yersinia pestis in order to identify leads that can be used in development of therapeutic agents against gram negative pathogens. Bacterial proteins that process trapped topoisomerase cleavage complexes will be identified with genetic and biochemical experiments. The repair proteins involved may be useful new targets for combination therapy with antibiotics targeting both type IA and type IIA topoisomerases. Success in these proposed experiments would provide tools and leads for development of novel antibacterial drugs for the urgent public health problem of bacterial pathogens resistant to all current antibiotics. PUBLIC HEALTH RELEVANCE: This project targets bacterial topoisomerase I enzyme for discovery of specific inhibitors. The proposed work would greatly aid the development of new antibacterial compounds against a novel target as part of the much needed arsenal to combat the global health problem of multi-drug drug resistant bacterial pathogens, including gram negatives.