Mechanistic Studies of Type II Topoisomerases and Topoisomerase-Targeted Agents

II 型拓扑异构酶和拓扑异构酶靶向药物的机理研究

基本信息

批准号：
10533336
负责人：
NEIL OSHEROFF
金额：
$ 35.58万
依托单位：
VANDERBILT UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-02-01 至 2025-11-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10533336
关键词：
Acute Promyelocytic Leukemia Affect Antineoplastic Agents Apoptosis Bacillus anthracis Biological CDC2 gene Catalysis Catalytic DNA Catenated DNA Cell Death Cell Proliferation Cell Survival Cells Chemotherapy-Oncologic Procedure Chromatids Chromosomal translocation Chromosome Segregation Chromosome Structures Complex Cultured Cells Cyclin B DNA DNA Damage DNA Double Strand Break DNA Topoisomerase IV DNA biosynthesis DNA topoisomerase II alpha Daughter Development Drug Prescriptions Drug Targeting Enzyme Inhibition Enzymes Escherichia coli Etoposide Gene Mutation Generations Genetic Materials Genetic Recombination Genetic Transcription Geometry Handedness Herb Human Human Activities Impairment In Vitro Learning MLL gene Magnetism Malignant Neoplasms Mechanical Stress Mediating Medicine Mitosis Mitoxantrone Multiple Sclerosis Mycobacterium tuberculosis Naphthoquinones Natural Products Neurofibrillary Tangles PML gene Pathway interactions Patients Pharmaceutical Preparations Physiological Play Poison Property Protein Isoforms Reaction Regulation Relaxation Research Research Design Role Specificity Study models Techniques Time Topoisomerase Topoisomerase II Topoisomerase Inhibitors Toxin Type I DNA Topoisomerases Xenopus laevis anti-cancer cell growth cell killing drug action drug development egg environmental chemical enzyme activity experimental study genome integrity in vitro activity inhibitor innovation leukemia neuron development novel novel therapeutics pollutant single molecule targeted agent

项目摘要

Type II topoisomerases are ubiquitous enzymes that are required for proper chromosome structure and segregation and play important roles in DNA replication, transcription, and recombination. These enzymes relax DNA and remove knots and tangles from the genetic material by passing an intact double helix (transport segment) through a transient double-stranded break that they generate in a separate DNA segment (gate segment). Humans encode two closely related isoforms of the type II enzyme, topoisomerase IIα and topoisomerase IIβ. Topoisomerase IIα is essential for the survival of proliferating cells and topoisomerase IIβ plays critical roles during development. However, because these enzymes generate requisite double-stranded DNA breaks during their crucial catalytic functions, they assume a dual persona. Although essential to cell survival, they also pose an intrinsic threat to genomic integrity every time they act. Beyond their critical physiological functions, topoisomerase IIα and IIβ are the primary targets for some of the most active and widely prescribed drugs currently used for the treatment of human cancers. These agents kill cells by stabilizing covalent topoisomerase II-cleaved DNA complexes (cleavage complexes) that are normal, but fleeting, intermediates in the catalytic DNA strand passage reaction. When the resulting enzyme- associated DNA breaks are present in sufficient concentrations, they can trigger cell death pathways. Anticancer drugs that target type II enzymes are referred to as topoisomerase II poisons because they convert these indispensable enzymes to potent physiological toxins that generate DNA damage in treated cells. Although topoisomerase IIα and IIβ are important targets for cancer chemotherapy, they also have the potential to trigger specific leukemias. For example, a small percentage of patients with cancer or multiple sclerosis who are treated with the topoisomerase II-targeted drug mitoxantrone go on to develop acute promyelocytic leukemias (APLs) with 15:17 translocations. Despite the importance of type II topoisomerases in cell growth and cancer, we still have much to learn about how the human enzymes function and interact with DNA and anticancer drugs. Thus, this proposal will further define the catalytic mechanism of type II topoisomerases and examine how enzyme activity is regulated in the cell. It also will define mechanisms by which established and novel topoisomerase II-targeted agents, environ- mental chemicals, and natural products increase levels of enzyme-mediated DNA breaks or inhibit enzyme activity and determine the cellular consequences of topoisomerase II poisons. The primary research models for this study will be human topoisomerase IIα and IIβ, cultured human cells, and Xenopus laevis egg extracts. Gyrase and topoisomerase IV from Escherichia coli and Bacillus anthracis will be used as counterpoints to mechanistic experiments and Mycobacterium tuberculosis gyrase will be used to assess relationships between the mechanisms of action of drugs targeted to prokaryotic and eukaryotic type II enzymes.

II 型拓扑异构酶是普遍存在的酶，是正确的染色体结构和分离并在 DNA 复制、转录和重组中发挥重要作用。通过完整的双螺旋（运输片段）通过短暂的双链断裂，它们在单独的 DNA 片段（门人类编码两种密切相关的 II 型酶亚型：拓扑异构酶 IIα 和拓扑异构酶 IIβ 对于增殖细胞和拓扑异构酶 IIβ 的生存至关重要。然而，由于这些酶产生必需的双链，因此在发育过程中起着至关重要的作用。 DNA 在其关键的催化功能过程中断裂，但它们具有双重角色，尽管它们对细胞至关重要。为了生存，它们每次行动都会对基因组完整性构成内在威胁。除了其关键的生理功能外，拓扑异构酶 IIα 和 IIβ 也是某些药物的主要靶标。目前用于治疗人类癌症的最活跃和最广泛使用的药物。通过稳定共价拓扑异构酶 II 切割的 DNA 复合物（切割复合物）来杀死细胞正常但短暂的中间体，在催化 DNA 链传递反应中产生酶-。相关的 DNA 断裂以足够的浓度存在，它们可以触发细胞死亡途径。针对 II 型酶的抗癌药物被称为拓扑异构酶 II 毒物，因为它们会转化这些酶是强效生理毒素不可或缺的酶，可在受治疗的细胞中产生 DNA 损伤。虽然拓扑异构酶 IIα 和 IIβ 是癌症化疗的重要靶点，但它们也具有可能引发特定的白血病，例如一小部分患有癌症或多发性白血病的患者。使用拓扑异构酶 II 靶向药物米托蒽醌治疗的硬化症会继续发展为急性具有 15:17 易位的早幼粒细胞白血病 (APL)。尽管 II 型拓扑异构酶在细胞生长和癌症中很重要，但我们仍有很多东西需要了解因此，这一提议将进一步研究人类酶如何发挥作用并与 DNA 和抗癌药物相互作用。定义 II 型拓扑异构酶的催化机制并研究酶活性如何在它还将定义已建立的和新型拓扑异构酶 II 靶向药物、环境药物的机制。精神化学物质和天然产物会增加酶介导的 DNA 断裂水平或抑制酶活性并确定拓扑异构酶 II 毒物的细胞后果。这项研究将使用人类拓扑异构酶 IIα 和 IIβ、培养的人类细胞以及非洲爪蟾卵提取物。来自大肠杆菌和炭疽芽孢杆菌的旋转酶和拓扑异构酶 IV 将用作机械实验和结核分枝杆菌促旋酶将用于评估之间的关系针对原核和真核 II 型酶的药物的作用机制。