Discovery of Selective Inhibitors for the EphA4 Kinase

EphA4 激酶选择性抑制剂的发现

基本信息

批准号：
9888253
负责人：
ELENA B PASQUALE
金额：
$ 75.18万
依托单位：
SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-03-15 至 2023-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9888253
关键词：
Affect Affinity Age Alzheimer&apos s Disease Alzheimer&apos s disease model Amyloid beta-Protein Animal Model Automation Binding Biochemical Biological Assay Biological Models Biophysics Blood - brain barrier anatomy Brain Cells Cessation of life Characteristics Chemicals Chronic Clinical Trials Cognitive deficits Complement Computer Models Crystallization Cultured Cells Disease Docking Dose Drug Targeting EGF-Like Domain Economics Eph Family Receptors EphA4 Receptor Ephrins Evaluation Functional disorder Future Goals Immunoblotting In Vitro Laboratories Lead Ligand Binding Ligand Binding Domain Ligands Measures Memory Loss Molecular Conformation Mus Nerve Degeneration Neurodegenerative Disorders Neurons Nucleic Acid Regulatory Sequences Pathogenesis Penetrance Peptides Pharmaceutical Preparations Pharmacology Phosphorylation Phosphotransferases Physiological Powder dose form Procedures Process Production Proteins Protocols documentation Receptor Protein-Tyrosine Kinases Receptor Signaling SAM Domain Safety Site Societies Specificity Structure Sushi Domain Synapses Tertiary Protein Structure Testing Therapeutic Therapeutic Effect Thinking Validation X-Ray Crystallography abeta oligomer analog base cheminformatics cognitive function data modeling design drug candidate drug discovery druggable target high throughput screening hyperphosphorylated tau improved in silico inhibitor/antagonist kinase inhibitor mouse model nerve injury neuron loss neurotoxic neurotoxicity neurotransmission novel novel strategies novel therapeutics pre-clinical relating to nervous system repaired response scaffold scale up screening small molecule small molecule libraries tau Proteins therapeutic development tool

项目摘要

Alzheimer’s is a devastating disease involving chronic, progressive neurodegenerative processes and decline in brain cognitive function that ultimately lead to death. Accumulation of aggregates of amyloid-beta and hyperphosphorylated tau protein in the brain causes synaptic dysfunction and loss of neurons, although the precise mechanisms underlying the disease remain unknown. There are few drugs available to treat Alzheimer’s disease, and they provide only limited benefits. In fact, the economic damages from Alzheimer’s disease reach hundreds of billion dollars every year in the US alone. Many candidate drugs have been evaluated in clinical trials in the last decade, but none has been approved. Thus, it is important to identify new drugs based on novel targets. The EphA4 receptor tyrosine kinase has recently emerged as a novel promising target for counteracting neurodegeneration and cognitive deficits in Alzheimer’s disease and other neurodegenerative diseases. EphA4 receptor signaling can promote neurotoxicity when aberrantly induced by amyloid-beta oligomers and ephrin ligands. The kinase domain of EphA4, which is responsible for the neurotoxic effects, is a druggable target. However, small molecule kinase inhibitors targeting EphA4 with selectivity and high affinity remain to be identified. Here we propose to perform two high-throughput screening campaigns of EphA4. One screen will deploy Protein Thermal Shift (PTS) as the assay format and the other screen will deploy a kinase activity assay format. The goal of these screens is to identify as broad a panel of modulators of the kinase-dependent functions of EphA4 as possible. Use of two formats should maximize our ability to identify a large repertoire of modulators with diverse modes of action and ultimately develop an inhibitory compound with high selectivity for EphA4. We anticipate that the PTS screen will identify compounds that stabilize the inactive conformation of the EphA4 intracellular region by binding not only to the kinase domain but also to regulatory regions outside this domain. The second screen will deploy a novel in vitro kinase assay configuration we have developed, where the inactive EphA4 intracellular region serves as the physiological substrate for the catalytically competent EphA4 intracellular region. We anticipate that this assay will identify not only compounds that target the EphA4 ATP binding pocket, but also allosteric inhibitors with novel mechanisms of action. The top hits identified in the two screens and their derivatives will be characterized and improved through rounds of secondary and tertiary biochemical and cell-based assays already established in the laboratories of the PIs and their collaborators, complemented by structure-guided approaches such as in silico docking and X-ray crystallography. These assays will provide formal hit validation, selectivity profiling and initial characterization of mechanisms of action and activities in neurons. We anticipate that this project will result in the identification of several novel EphA4 selective inhibitors that are suitable for advancement to future studies in animal models of Alzheimer’s and other neurodegenerative diseases.

阿尔茨海默氏症是一种毁灭性的疾病，涉及慢性，进行性神经退行性过程和下降在最终导致死亡的大脑认知功能中。淀粉样蛋白β骨料的积累和大脑中的高磷酸化tau蛋白会引起突触功能障碍和神经元的丧失，尽管该疾病的确切机制仍然未知。几乎没有药物可以治疗阿尔茨海默氏病，它们仅提供有限的好处。实际上，阿尔茨海默氏症的经济损害仅在美国，疾病每年就达到数十亿美元。许多候选药物已经在过去十年中在临床试验中进行了评估，但没有批准。那很重要基于新靶标的药物。 Epha4受体酪氨酸激酶最近成为一种新颖的有希望的在阿尔茨海默氏病和其他方面抵消神经变性和认知缺陷的目标神经退行性疾病。当被异常诱导的EPHA4受体信号传导可以促进神经毒性淀粉样蛋白-beta低聚物和Ephrin配体。 EPHA4的激酶域，该领域负责神经毒性作用是可吸毒的靶标。然而，针对以EPHA4为目标的小分子激酶抑制剂选择性和高亲和力仍有待确定。在这里，我们建议进行两个高通量筛查 Epha4的运动。一个屏幕将部署蛋白质热移（PTS）作为测定格式，另一种是屏幕将部署激酶活动分析格式。这些屏幕的目的是确定一个广泛的面板 EPHA4的激酶依赖性功能的调节剂。使用两种格式应最大化我们的能够识别具有潜水动作模式的大量调制器的能力，并最终发展抑制性化合物对EPHA4的选择性高。我们预计PTS屏幕将识别化合物通过与激酶结合，稳定Epha4细胞内区域的无活性构象领域，也要用于该领域之外的监管区域。第二个屏幕将在体外部署小说激酶测定构型我们已经开发了，其中无活性EPHA4细胞内区域作为催化能力的EPHA4细胞内区域的生理底物。我们预计这个测定法将不仅鉴定针对EPHA4 ATP结合袋的化合物，还可以识别具有的化合物新型作用机理。两个屏幕中确定的最高命中及其导数将是通过二级和第三纪生化和基于细胞的测定的圆圈进行表征和改进已经在PIS及其合作者的实验室中建立，由结构引导完成诸如计算机对接和X射线晶体学之类的方法。这些测定将提供正式的命中验证，神经元中作用和活动机制的选择性分析和初始表征。我们期待该项目将导致鉴定几种适合的EPHA4选择性抑制剂在阿尔茨海默氏症和其他神经退行性疾病的动物模型中未来研究的进步。