Generic Fluorescent HTS Assay for Protein Kinases

描述(申请人提供)：人类基因组中有400多种不同的蛋白激酶编码；阐明它们在疾病中的作用并识别选择性抑制剂是一项主要的制药计划。激酶功能障碍与所有最重要的治疗领域有关，包括癌症、心血管疾病、炎症、神经退行性疾病和代谢紊乱。此外，最近在Herceptin和Gleevec的病例中显示了将激酶作为药物靶点的临床验证，这两种药物分别抑制导致乳腺癌和白血病的异常酪氨酸激酶。高通量筛选(HTS)-数千种化合物与药物靶点相互作用的平行测试-已成为药物发现的主导模式。2003年HTS检测试剂的总市场超过5亿美元，大约20%的筛查是针对蛋白激酶进行的。尽管人们对此有很高的兴趣，但目前检测方法的缺陷阻碍了将新的激酶纳入HTS计划和比较不同激酶的抑制剂谱的努力。为了克服这些技术障碍，我们建议开发一种基于BellBrook实验室专有的Transcreener(Tm)平台的通用激酶分析方法。该检测依赖于一种基于竞争荧光偏振的免疫分析来检测腺二磷酸(ADP)，这是所有激酶反应的产物。Transcreener(Tm)激酶分析将通过提供一种可与任何激酶和任何受体底物一起使用的通用催化分析，加快确定激酶底物特异性和识别新型抑制剂的工作。

DESCRIPTION (provided by applicant): There are more than 400 distinct protein kinases encoded in the human genome; elucidating their role in disease and identifying selective inhibitors is a major pharma initiative. Kinase malfunction has been linked to all of the most important therapeutic areas, including cancer, cardiovascular diseases, inflammation, neurodegenerative diseases, and metabolic disorders. Moreover, clinical validation of kinases as drug targets has recently been shown in the cases of Herceptin and Gleevec, which inhibit aberrant tyrosine kinases that contribute to breast cancer and leukemia, respectively. High throughput screening (HTS) - the parallel testing of many thousands of compounds for interaction with a drug target - has become the dominant mode of drug discovery. The total market for HTS assay reagents in 2003 was over $500M, and approximately 20% of screening was done on protein kinases. Despite the high level of interest, shortcomings with the current assay methods are hampering efforts to incorporate new kinases into HTS programs and to compare inhibitor profiles for different kinases. To overcome these technical hurdles we propose to develop a universal kinase assay method based on BellBrook Labs' proprietary Transcreener(tm) platform. The assay relies on a competitive fluorescence polarization-based immunoassay for detection of adenosine diphosphate (ADP), a product of all kinase reactions. The Transcreener(tm) kinase assay will accelerate efforts to define kinase substrate specificity and to identify novel inhibitors by providing a universal catalytic assay that can be used with any kinase and any acceptor substrate.