Rapid Kinase Profiling with Luminescent Reporters

描述（由申请人提供）：超过500种不同的蛋白激酶已在人类中被鉴定，占人类基因组的1.7%。蛋白激酶的作用是催化ATP的3-磷酸基转移到特定蛋白质底物上的羟基。激酶在疾病中的核心作用及其在人类生物学中的基本作用促使了有效和选择性抑制剂的发展，作为治疗剂和在信号转导途径背景下解剖激酶功能的新工具。由于许多人类激酶在遗传和结构上都是如此密切相关，许多抑制剂对单个靶点缺乏特异性，而作用于多个激酶，通常是在活性位点与ATP竞争。因此，需要全面的激酶面板和工具来评估药物的选择性。激酶筛选和分析市场在2008年估计为1.06亿美元，并且需求每年都在稳步上升。我们申请的目的是开发一种基于发光的、均匀的、非放射性的、敏感的、无细胞的检测方法，可以快速适应于高通量（HTP）筛选和分析小分子激酶拮抗剂。本实验对市售药物/候选药物的效用以及这些分子对激酶面板的选择性将被证明。

DESCRIPTION (provided by applicant): More than 500 different protein kinases have been identified in humans and represent 1.7% of the human genome. Protein kinases function by catalyzing the transfer of the 3-phosphoryl group of ATP to the hydroxyl group on a specific protein substrate. The central role of kinases in disease and their fundamental role in human biology have prompted the development of potent and selective inhibitors as therapeutic agents and as novel tools for dissecting kinase function in the context of signal transduction pathways. Because many human kinases are so closely related both genetically and structurally, many inhibitors lack specificity for a single target and act on more than one kinase, typically by out-competing ATP in the active site. Hence there is a need for comprehensive kinase panels and tools for selectivity assessment of drugs. The kinase screening and profiling market was estimated to be $106 MM in 2008 and the demand is steadily rising every year. The purpose of our application is to develop a luminescence-based, homogeneous, non- radioactive, sensitive, cell-free assay that can be rapidly adapted for high throughput (HTP) screening and profiling of small molecule antagonists of kinases. The utility of this assay against commercially available drugs/dug candidates and the selectivity of these molecules against a kinase panel will be demonstrated. PUBLIC HEALTH RELEVANCE: Kinases constitute 1.7% of the genome and are implicated in various diseases, from cancer to diabetes. Assessment of selectivity of drugs against kinases is critical to evaluate their physiological safety. The purpose of our application is to develop a method to identify selective drugs by profiling them against a kinase panel.