PROTEIN-ASSOCIATED DNA BREAKS AS INDICATOR OF TOPOISOMERASE INHIBITION

蛋白质相关 DNA 断裂作为拓扑异构酶抑制的指标

• 批准号: 3853153
• 负责人: Y POMMIER
• 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/3853153
• 关键词: DNA binding protein; DNA gyrase; DNA repair; DNA topoisomerases; active sites; anthracyclines; biological signal transduction; calcium; camptothecin; cell death; clone cells; drug metabolism; drug resistance; enzyme complex; enzyme inhibitors; enzyme mechanism; etoposide; molecular oncology; neoplasm /cancer chemotherapy; neoplastic cell culture for noncancer research; oncogenes; protein kinase C

DNA topoisomerases are major targets for cancer chemotherapy. Topoisomerase I is concentrated in nucleoli and topoisomerase II is a major component of both the nuclear scaffold in interphase nuclei and the chromosome scaffold during mitosis. Topoisomerase II activity is required for chromosome segregation during mitosis and both enzymes are involved in DNA replication and transcription functions. Camptothecin inhibits topoisomerase I, and DNA intercalators (amsacrine, anthracyclines) and demethylepipodophyllotoxins (VP-16 & VM-26) inhibit topoisomerase II. The drugs act by stabilizing enzymelinked DNA strand breaks, and these may be the initial cytotoxic lesions produced by the drugs. However, the breaks reverse quickly upon drug removal, indicating that other processes are involved in the sequence of events leading to cell death. This project aims to elucidate these events. We have obtained evidence that the topoisomerase I-DNA complexes induced by camptothecin kill rapidly proliferating Chinese hamster DC3F and slowly proliferating human colon cancer cells by interacting with DNA replication complexes. We have also demonstrated that topoisomerase-mediated DNA breaks are not toxic in cells which have been depleted of calcium and that protein kinases can modulate cellular sensitivity to topoisomerase inhibitors. Further, it now appears possible that topoisomerase I is regulated by signal transduction pathways including protein kinase C. We have started mapping cleavage sites in the c-myc oncogene and find that m-AMSA selectively inhibits topoisomerase II by stabilizing cleavage complexes in the P2 promoter, a site most often used for c-myc transcription in tumor cells. A new project has been the development of camptothecin-resistant cells. We have now obtained a highly resistant cell line and are studying its topoisomerases.

DNA拓扑异构酶是癌症化疗的主要靶点。 拓扑异构酶I集中在核仁中，而拓扑异构酶II是 中间相核和相间核中的核支架的主要成分 有丝分裂过程中的染色体支架。拓扑异构酶II的活性为 有丝分裂过程中染色体分离所必需的，两种酶都是 参与DNA复制和转录功能。喜树碱 抑制拓扑异构酶I和DNA嵌入物(阿马西林， 去甲基鬼臼毒素(VP-16和Vm-26)抑制 拓扑异构酶II.药物通过稳定酶连接的DNA链起作用 断裂，这些可能是由 毒品。然而，在药物清除后，断裂很快就会逆转， 指示事件序列中涉及其他进程 导致细胞死亡。该项目旨在阐明这些事件。我们 已经获得证据表明，由DNA诱导的拓扑异构酶I-DNA复合体 喜树碱杀死快速增殖的中国仓鼠DC3F和缓慢 DNA相互作用对人结肠癌细胞增殖的影响 复制复合体。我们也证明了 拓扑异构酶介导的DNA断裂对已经被 耗尽了钙，蛋白激酶可以调节细胞 对拓扑异构酶抑制剂的敏感性。此外，现在看来 拓扑异构酶I可能受信号转导调控 包括蛋白激酶C在内的通路我们已经开始绘制裂解图谱 C-myc癌基因上的位点，发现m-AMSA选择性抑制 稳定P2启动子a中切割复合体的拓扑异构酶II 在肿瘤细胞中最常用于c-myc转录的位点。一种新的 该项目一直在开发耐喜树碱细胞。我们 现在已经获得了一个高度抗性的细胞系，并正在研究它的 拓扑异构酶。