REGULATION OF DNA IN CAMPTOTHECIN TREATED CELLS

描述：喜树碱(CPT)及其类似物为拓扑异构酶I(Topo 一)抑制剂，并显示出对实体肿瘤的疗效 历史上对大多数癌症化疗药物具有抗药性。这个 本应用的长期目标是阐明分子 CPT处理细胞中DNA复制抑制的机制。这个 喜树碱的细胞毒作用主要作用于S期细胞，并与 抑制DNA复制。这种对DNA复制的抑制 被认为是由顺式作用过程调节的，结果是 前进的复制叉与可切割的CPT-TOPO的碰撞 I-DNA复合体。这个实验室进行的研究表明，在 除了顺式作用的抑制过程，还有反式作用 在CPT处理的细胞中活跃的抑制过程。调查员 假设CPT产生的受损/修饰的DNA是一种中介 招募RPA进行修复并抑制其在DNA复制中的功能。它是 进一步假设Ku可能会与RPA竞争绑定到 损坏/修饰的DNA，或者拆解或重塑RPA-DNA复合体以 释放RPA。因此，允许RPA再次启动DNA复制，并且 因此，减少了抑制作用。关于分子的信息 这种DNA复制调控的决定因素可能提供新的方法 临床干预，实现CPT致敏。具体的 旨在重点研究RPA和KU作为负责的活动 CPT处理的细胞中DNA复制的调控 以上型号。我们的目标是：1.描述 与DNA复制抑制相关的RPA的特性 CPT处理细胞和2.研究Ku对DNA复制抑制的影响 在CPT处理的细胞中，以及它通过什么机制相互作用和调节 RPA的活动。猴病毒40(SV40)的体外DNA复制 化验将在拟议的实验中使用。拟议的研究是 基于初步结果的信息，并结合当前知识 用DNA复制和CPT细胞毒性表征其调控作用 在CPT处理的细胞中的DNA复制。

DESCRIPTION: Camptothecin (CPT) and its analogs are topoisomerase I (Topo I) inhibitors and show efficacy against solid tumors which have been historically resistant to most cancer chemotherapeutic agents. The long-term objective of the present application is to elucidate the molecular mechanism of DNA replication inhibition in CPT-treated cells. The cytotoxicity of CPT is mainly exerted on S-phase cells and is associated with an inhibition of DNA replication. This inhibition of DNA replication is thought to be mediated by cis-acting processes as a result of the collision of the advancing replication fork with the cleavable CPT-Topo I-DNA complex. Work carried out in this laboratory suggests that in addition to cis-acting inhibitory processes, there are also trans-acting inhibitory processes active in CPT-treated cells. The investigator hypothesizes that damaged/modified DNA produced by CPT is a mediator that recruits RPA for repair and inhibits its function in DNA replication. It is further hypothesized that Ku may either compete with RPA for binding to damaged/modified DNA, or disassemble or remodel the RPA-DNA complex to release RPA. Thus RPA is allowed to initiate DNA replication again, and therefore the inhibition is reduced. Information on the molecular determinants of this regulation of DNA replication may offer new ways of intervention in the clinic for achieving CPT sensitization. The Specific Aims focus on studying RPA and Ku as the activities responsible for the regulation of DNA replication in CPT-treated cells in the framework of the above model. The goals are: 1. To characterize modifications in the properties of RPA that are related to DNA replication inhibition in CPT-treated cells and 2. To study how Ku affects DNA replication inhibition in CPT-treated cells, and by what mechanism it interacts and modulates the activity of RPA. The Simian virus 40 (SV40) based in vitro DNA replication assay will be used in the proposed experiments. The proposed research is based on information from preliminary results and combines current knowledge on the DNA replication and CPT cytotoxicity to characterize the modulation of DNA replication in CPT-treated cells.