MOLECULAR PHARM. AND BIOCHEM. OF NOVEL DUAL SPECIIFICITY PHOSPHATASE INHIBITORS

该项目是一个多学科的努力，利用化学多样性和智能分析来发现抗肿瘤药物。本项目的总体目标是检查从化学库中获得并在其他子项目中合成的小分子，以了解它们破坏双特异性磷酸酶(DSPase)活性的能力，以及它们对人类实体瘤的抗增殖活性。DSPase已成为未来针对癌症的治疗药物的重要分子靶点。CDC25 DSPase是控制细胞周期检查点的细胞周期蛋白依赖性蛋白激酶(CDK)磷酸化状态的主要决定因素。最近，在这项计划项目拨款的支持下，我们确定了最有效和 已知的CDC25磷酸酶的选择性抑制物。尽管这些化合物有相当大的希望成为工具药物，但我们建议定义更有效、更有选择性和更像药物一样的CDC25抑制剂。因此，我们的工作假设仍然是致癌DSPase的磷酸酶活性与CDC25的致癌特性有关，并且这种催化活性的选择性抑制剂可能产生医学上有用的和独特的化合物。本项目中的生化分析已明确设计用于评估其他子项目中概述的新的、固相和溶液相组合化学策略。我们的具体目标是(1)表征所有新的文库化合物的体外抗磷酸酶活性，(2)评估优先考虑的化合物的抗磷酸酶、抗增殖和细胞周期的作用 (3)用新的类似物来确定CDc25B的配体相互作用和三维结构，以及(4)建立一种可靠的智能分析方法来评估新的针对CDc25B的配体的动态化学库。我们将继续使用人类乳腺、前列腺和卵巢肿瘤细胞系。我们建议使用非恶性细胞以及具有温度敏感CDC25底物的小鼠细胞和缺乏功能性CDC25B或CDC25C基因的小鼠成纤维细胞来检测这些化合物的细胞活性。

This Project is a multidisciplinary effort to employ chemical diversity and smart assays to antitumor drug discovery. The overall goal of this Project is to examine small molecules obtained from chemical libraries and synthesized in other subprojects for their ability to disrupt dual specificity phosphatase (DSPase) activity and for their antiproliferative activity against human solid tumors. DSPases have emerged as important molecular targets for future therapeutic agents directed against cancer. The Cdc25 DSPases are major determinants of the phosphorylation state of cyclin-dependent kinases (Cdks), which control cell cycle checkpoints. Recently, with the support of this Program Project Grant, we identified the most potent and selective inihibitors of the Cdc25 phosphatases known. Although these compounds hold considerable promises as tool drugs, we propose to define more potent, selective and drug-like Cdc25 inhibitors. Thus, our working hypothesis continues to be that the phosphatase activity of oncogenic DSPases is responsible for the oncogenic properties of Cdc25 and that selective inhibitors of this catalytic activity are likely to generate medicinally useful and unique compounds. The biochemical assays in this Project have been explicitly designed to evaluate the novel, solid phase and solution-phase combinatorial chemistry strategies outlined in the other subprojects. Our Specific Aims are to (1) characterize the in vitro antiphosphatase activity of all novel library compounds, (2) assess the antiphosphatase, antiproliferative and cell cycle effects of prioritized compounds, (3) determine the ligand interactions and 3-dimensional structure of Cdc25B with novel analogs and, (4) develop a robust smart assay to evaluate new dynamic chemical libraries for ligands against Cdc25B. We will continue to use human breast, prostate and ovarian tumor cell lines. We propose to use nonmalignant cells as well as murine cells with a temperature sensitive Cdc25 substrate and fibroblasts from mice that lack functional Cdc25B or Cdc25C genes to examine the cellular activity of these compounds.