Inhibition of Recombination DNA Repair in Pancreatic Cancer Cells

描述（由申请人提供）：胰腺癌仍然是美国癌症死亡的第四大原因。这些肿瘤似乎对标准可用疗法具有相对抗性。一种能够使胰腺癌细胞对肿瘤治疗敏感的药物可能会改善这些患者的预后。该项目的一个中心假设是，同源重组（HR）DNA修复水平的升高可导致人类胰腺肿瘤对某些化疗和放疗产生耐药性，并且HR的特异性抑制可能有助于克服这种对治疗的耐药性。拟议的研究计划将开发一个多步骤的筛选，以确定人类RAD 51的小分子抑制剂，这是参与启动HR的中心蛋白。该项目的初始提交包括一个目标，提出开发一个高通量（HT）测定和筛选一个幼稚的小分子库，以寻找可以抑制单链DNA上RAD 51细丝形成的化合物。初步工作的一部分已经完成，现总结在初步研究/进展报告部分。10 k文库的筛选鉴定了72种抑制RAD 51丝形成≥ 50%的化合物。这项工作成功地验证了细丝形成试验作为HT筛选，它提供了重要的信息，包括改进试验技术和数据分析。使用这些信息，现在将使用更大的起始文库（130 k化合物）和机器人液体处理来重复筛选，这可能会产生具有更大活性和特异性的化合物（目标1）。为了确定哪些具有特定活性，将在二级和三级测定中进一步测试从两种筛选中得到的化合物。目的2将采用一组体外二级测定，旨在鉴定那些可以特异性阻断纯化的RAD 51蛋白的特定功能的化合物。在第三个目标中，将用基于三级细胞的测定进一步表征化合物，以鉴定能够通过HR特异性机制使胰腺癌细胞对DNA损伤疗法的致死作用敏感的亚组。最终目标是确定至少一种适合在临床试验中进一步开发为肿瘤药物的最佳化合物。胰腺癌仍然是美国癌症死亡的第四大原因。这些肿瘤似乎对目前可用的疗法具有相对抗性。能够克服细胞对放疗和/或化疗药物的耐药性的药物可以改善这些患者的结果。该提案的目标是开发一种能够使胰腺癌细胞对这些常见肿瘤治疗敏感的药物。

DESCRIPTION (provided by applicant): Pancreatic cancer remains the 4th leading cause of cancer death in the United States. These tumors appear to be relatively resistant to standard available therapies. An agent capable of sensitizing pancreatic cancer cells to oncology therapies may improve the outcomes of these patients. A central hypothesis of this project is that elevated levels of homologous recombinational (HR) DNA repair can cause human pancreatic tumors to be resistant to some chemotherapies and radiotherapy, and that specific inhibition of HR may help overcome this resistance to therapy. The proposed research plan will develop a multi-step screen for identifying small molecule inhibitors of human RAD51, which is the central protein involved in initiating HR. The initial submission of this project included an aim proposing to develop a high-throughput (HT) assay and to screen a na¿ve library of small molecules in search of compounds that can inhibit the formation of RAD51 filaments on single-stranded DNA. A portion of initial work has been completed and is now summarized in the Preliminary Studies / Progress Report section. The screen of a 10k library identified 72 compounds that inhibit RAD51 filament formation by = 50%. This work successfully validated the filament formation assay as an HT screen, and it provided important information including refinement of assay techniques and data analysis. Using this information, the screen will be now be repeated using a larger starting library (130k compounds) and robotic liquid handling, which will likely yield compounds with even greater activities and specificities (Aim 1). To determine which have specific activities, the compounds resulting from both screens will be tested further in secondary and tertiary assays. Aim 2 will employ a set of in-vitro secondary assays aimed at identifying those compounds that can specifically block particular functions of purified RAD51 protein. In the third aim, compounds will be further characterized with tertiary cell-based assays, to identify the subset capable of sensitizing pancreatic cancer cells to the lethal effects of DNA damaging therapies via HR-specific mechanisms. The ultimate goal is to identify at least one optimal compound suitable for further development in clinical trials as an oncology drug. Pancreatic cancer remains the 4th leading cause of cancer death in the United States. These tumors appear to be relatively resistant to presently available therapies. A drug capable of overcoming the cellular resistance to radiotherapy and/or chemotherapeutic drugs may improve on the outcomes of these patients. The goal of this proposal is to develop an agent capable of sensitizing pancreatic cancer cells to these common oncology therapies.