DNA LESIONS AS ENDOGENOUS TOPOISOMERASE POISONS

DNA 损伤作为内源性拓扑异构酶毒物

• 批准号: 6131038
• 负责人: NEIL OSHEROFF
• 金额: $ 25.77万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-05-01 至 2004-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6131038
• 关键词: DNA binding protein; DNA damage; DNA repair; DNA replication; DNA topoisomerases; Drosophilidae; Saccharomyces cerevisiae; adduct; antineoplastics; cell death; chemical binding; chemical cleavage; cytotoxicity; enzyme activity; enzyme induction /repression; enzyme mechanism; genetic recombination; mutant; pharmacokinetics; tissue /cell culture

DESCRIPTION (As Adapted From the Investigator's Abstract): Topoisomerase II is an essential enzyme that is required for proper chromosome structure and segregation and plays important roles in DNA replication and recombination. Beyond its critical cellular functions, topoisomerase II is the primary target for some of the most active and widely prescribed drugs used for the treatment of human cancers. These agents elicit their cytotoxic effects by a mechanism that is markedly different than that of other drugs. Rather than inhibiting the catalytic activity of topoisomerase II, anticancer drugs targeted to the enzyme dramatically increase levels of covalent topoisomerase II-cleaved DNA complexes that are normal, but fleeting, catalytic intermediates. When the resulting topoisomerase II-associated double-stranded DNA breaks are present in high concentrations, they generate mutations, chromosomal translocations, and trigger cell death pathways. Because topoisomerase II-targeted anticancer drugs convert this dispensable enzyme into a potent physiological toxin, they are referred to as topoisomerase II poisons. Although topoisomerase II is one of the most important targets for cancer chemotherapy, there is compelling circumstantial evidence that the enzyme also has the potential to trigger the disease. Together with the unique mechanism of action of topoisomerase II poisons, this suggests that topoisomerase II-targeted drugs may represent exogenous counterparts of cellular components that induce DNA recombination, mutagenesis, or cell death pathways. Previous results form this laboratory indicated that abasic sites, the most commonly formed lesions in DNA, stimulate topoisomerase II-mediated double-stranded DNA cleavage with a potency that is greater than 1000-fold higher than that of etoposide, one of the most widely prescribed anticancer drugs in clinical use. Therefore, the ultimate goals of the proposal are to further define interactions between topoisomerase II and DNA damage and to determine whether DNA lesions function in vivo as endogenous topoisomerase II poisons. The specific aims of this proposal are to: 1) further define the spectrum of DNA damage that alters the catalytic function of type II topoisomerases; 2) define the mechanism by which DNA lesions enhance topoisomerase II-mediated DNA cleavage; 3) determine whether abasic intermediates generated by base excision repair can trigger the formation of permanent topoisomerase II-mediated double-stranded DNA breaks; and 4) determine whether DNA lesions act as topoisomerase II poisons in the cell. The primary enzymological model for this study will be human topoisomerase II alpha and beta. Physiological studies will employ human cell lines and yeast (Saccharomyces cerevisiae). Cellular DNA damage will be induced by a combination of chemical and genetic approaches.

描述（改编自研究者摘要）：拓扑异构酶II是 一种必需的酶，是正确的染色体结构所必需的， 分离，并在DNA复制和重组中起重要作用。 除了其关键的细胞功能，拓扑异构酶II是主要的目标 对于一些最活跃和最广泛的处方药用于治疗 人类的癌症。这些药物通过一种机制引起其细胞毒性作用， 这与其他药物有很大不同。而不是抑制 拓扑异构酶II的催化活性，靶向该酶的抗癌药物 显著增加共价拓扑异构酶II切割的DNA复合物的水平 它们是正常的，但转瞬即逝的催化中间体。当所得 拓扑异构酶II相关的双链DNA断裂存在于高 浓度，它们产生突变，染色体易位， 触发细胞死亡途径。因为拓扑异构酶II靶向抗癌药物 将这种酶转化为一种强有力的生理毒素， 称为拓扑异构酶II毒物。虽然拓扑异构酶II是一种 癌症化疗最重要的目标，有令人信服的 间接证据表明，这种酶也有可能引发 疾病加上拓扑异构酶II的独特作用机制 这表明拓扑异构酶II靶向药物可能代表 诱导DNA重组的细胞组分的外源对应物， 诱变或细胞死亡途径。本实验室先前的结果 表明脱碱基位点，DNA中最常见的损伤，刺激 拓扑异构酶II介导的双链DNA切割， 比依托泊苷高1000倍以上，依托泊苷是最广泛的 临床使用的抗癌药物。因此， 这些建议是进一步定义拓扑异构酶II和 DNA损伤，并确定DNA损伤是否在体内作为内源性功能 拓扑异构酶II毒素。该提案的具体目标是：（1）进一步 定义改变II型胶原催化功能的DNA损伤谱 拓扑异构酶; 2）定义DNA损伤增强的机制 拓扑异构酶II介导的DNA切割; 3）确定是否脱碱基 由碱基切除修复产生的中间体可以触发 永久性拓扑异构酶II介导的双链DNA断裂;和4） 确定DNA损伤是否在细胞中作为拓扑异构酶II毒物。的 本研究的主要酶学模型是人拓扑异构酶II α 和beta。生理学研究将使用人类细胞系和酵母 （酿酒酵母）。细胞DNA损伤将由一种 化学和遗传方法的结合。