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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：
10334506
负责人：
ELENA B PASQUALE
金额：
$ 75.76万
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/10334506
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 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 Western Blotting 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 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破坏性疾病，涉及慢性、进行性神经退行性过程和衰退最终导致死亡的大脑认知功能。 β-淀粉样蛋白和淀粉样蛋白聚集体的积累大脑中过度磷酸化的 tau 蛋白会导致突触功能障碍和神经元损失，尽管该疾病的确切机制仍不清楚。可以治疗的药物很少阿尔茨海默病，它们只能提供有限的益处。事实上，阿尔茨海默病造成的经济损失仅在美国，每年因疾病造成的损失就达到数千亿美元。许多候选药物已被过去十年进行了临床试验评估，但尚未获得批准。因此，识别新的基于新靶点的药物。 EphA4受体酪氨酸激酶最近成为一种新的有前途的药物对抗阿尔茨海默病和其他疾病的神经退行性变和认知缺陷的目标神经退行性疾病。 EphA4 受体信号传导异常诱导时可促进神经毒性淀粉样蛋白-β 寡聚物和肝配蛋白配体。 EphA4 的激酶结构域，负责神经毒性作用，是一个可药物靶标。然而，针对 EphA4 的小分子激酶抑制剂选择性和高亲和力仍有待确定。这里我们建议进行两次高通量筛选 EphA4 的活动。一个屏幕将部署蛋白质热位移 (PTS) 作为测定格式，另一个屏幕将部署蛋白质热位移 (PTS) 作为检测格式 screen 将部署激酶活性测定格式。这些屏幕的目标是确定广泛的面板 EphA4 激酶依赖性功能的调节剂可能。使用两种格式应该最大限度地提高我们的能够识别具有不同作用模式的大量调节剂，并最终开发出对EphA4具有高选择性的抑制化合物。我们预计 PTS 筛选将识别化合物不仅通过与激酶结合来稳定 EphA4 胞内区域的非活性构象域，还包括该域之外的监管区域。第二屏将部署一部体外小说我们开发了激酶测定配置，其中非活性 EphA4 胞内区域作为具有催化能力的 EphA4 细胞内区域的生理底物。我们预计该检测不仅会识别出针对 EphA4 ATP 结合袋的化合物，还会识别出具有以下特性的变构抑制剂：新颖的作用机制。两个屏幕中确定的热门点击及其衍生产品将是通过多轮二级和三级生化和细胞检测进行表征和改进已在 PI 及其合作者的实验室中建立，并以结构指导为补充诸如计算机对接和 X 射线晶体学等方法。这些测定将提供正式的命中验证，神经元作用和活动机制的选择性分析和初步表征。我们预计该项目将鉴定出几种新型 EphA4 选择性抑制剂，这些抑制剂适用于阿尔茨海默病和其他神经退行性疾病动物模型的未来研究取得进展。