MECHANISM BASED INHIBITION OF CYCLIN DEPENDENT KINASES

描述（逐字摘自申请人摘要）：细胞周期蛋白依赖性激酶 细胞周期蛋白激酶（CDKs）在细胞周期调控中起着重要作用 事件特异于CDK的抑制剂可以潜在地用作 用于阐明信号转导途径的药理学探针， 作为控制和管理细胞周期相关疾病的治疗剂 疾病然而，目前大多数CDK的化学抑制剂作为一种抗肿瘤药物， 竞争性抑制剂的共同底物ATP，往往缺乏所需的 特异性和效力。这促使我们开发特异性CDK抑制剂 具有新颖的作用机制和普遍适用的方法， 蛋白丝氨酸/苏氨酸激酶抑制剂的设计。基于机制 使用特定底物序列设计的抑制剂作为模板， 提供保留模板特异性的最佳可能性， 从而损害了必要的抑制效力。为了验证这一假设，一些 通过掺入二肽， Gly-Ser置换成已知是CDK的良好底物的序列。的 将探索在靶向丝氨酸周围使用修饰的肽基团 在将磷酸化的肽类似物转化为更高活性的 物种，其可以与靶向激酶反应以产生共价结合的蛋白质。 失活酶（具体目标1）。更换产品的目的是覆盖广泛的 为了发现必要的基本化学过程， 用于制造CDK的“自杀”抑制剂。将对这些抑制剂进行检测， 它们对重组人细胞周期蛋白B/CDC 2的抑制类型和性质 激酶（特异性目的2）。将评价抑制剂的特异性， 与原始模板对细胞周期蛋白的底物特异性相比， B/CDC 2，相对于cAMP依赖性蛋白激酶和蛋白激酶C （具体目标3）。未来的工作将集中在确定活跃的网站 残余物改性和改善抑制剂的渗透性 细胞膜及其对蛋白水解的稳定性。这种方法 应普遍适用于快速发展的机制为基础的 对任何蛋白质丝氨酸/苏氨酸激酶具有特异性的抑制剂， 特定底物序列。

DESCRIPTION (verbatim from the applicant's abstract): Cyclin-dependent kinases (CDKs) play an important role in the control and regulation of cell cycle events. Inhibitors specific for CDKs could potentially be used as pharmacological probes for the elucidation of signal transduction pathways and as therapeutic agents in the control and management of cell cycle-related diseases. However, most current chemical inhibitors of CDKs act as a competitive inhibitor of the common substrate ATP and often lack the desired specificity and potency. This prompted us to develop specific CDK inhibitors with a novel mechanism of action and a generally applicable approach to the design of protein serine/threonine kinase inhibitors. Mechanism-based inhibitors designed using a specific substrate sequence, as a template should offer the best probability of retaining specificity of the template without compromising the necessary inhibitory potency. To test this hypothesis, several potential mechanism-based inhibitors are designed by incorporation of dipeptide Gly-Ser replacements into a sequence known to be a good substrate of CDKs. The use of modified peptide groups surrounding the targeted serine will be explored in converting the phosphorylated peptide analog to a more highly reactive species, which could react with the targeted kinase to produce a covalently inactivated enzyme (specific aim 1). Replacements are designed to cover a wide range of activation in order to discover the fundamental chemistry necessary for making "suicide" inhibitors of CDKs. These inhibitors will be tested for their type and nature of inhibition against a recombinant human cyclin B/CDC2 kinase (specific aim 2). The specificity of inhibitors will be evaluated and compared with the substrate specificity of the original template towards cyclin B/CDC2, relative to cAMP-dependent protein kinase and protein kinase C (specific aim 3). Future efforts will focus on identifying the active site residue(s) modified and on improving the inhibitors' permeability across cellular membranes and their stability towards proteolysis. This approach should be generally applicable to the quick development of mechanism-based inhibitors specific for any protein serine/threonine kinase with a known specific substrate sequence.