Development of drugs that modify CNS innate immunity for the treatment of Multiple Sclerosis

项目摘要 由中枢神经系统（CNS）中的常驻细胞（如星形胶质细胞和小胶质细胞）驱动的病理学 在多发性硬化症（MS）中发挥重要作用，特别是对于最终达到的进展阶段， 大多数MS患者然而，目前尚无药物可用于调节星形胶质细胞和小胶质细胞的致病性 活动我们开发了一种体外试验来鉴定调节星形胶质细胞致病活性的化合物。 这些研究鉴定了化合物A-38，其是产生红细胞生成素的肝细胞癌的抑制剂。 癌受体B3（EphB 3），作为星形胶质细胞致病活性的抑制剂，因此，作为 治疗进展性MS（PMS）先导化合物。此外，我们的数据表明，小胶质细胞驱动的 星形胶质细胞中的EphB 3信号传导促进CNS炎症和神经变性。肝配蛋白受体是 已知参与CNS发育，然而，我们的发现确定了EphB 3在CNS中的新作用， 炎症并将其鉴定为治疗干预潜在靶点。我们假设EphB 3 激酶抑制将为PMS和潜在其它疾病提供有效的治疗方法 神经系统疾病因此，我们建议开发新的A-38类似物来治疗PMS。我们的具体目标是： 特定目的1：EPHB 3激酶抑制剂的优化和体外评价。我们将 设计、合成和评估多达150种A-38类似物的性质。这些研究的目的是 优化A-38新类似物的效力和激酶选择性，同时保持良好的药物样性质 以及口服给药后的脑暴露。在本项目的R61阶段，我们将集成迭代 构效关系（SAR）研究和基于结构的药物设计，以优化A-38的效力， 激酶选择性，同时保持良好的药物样性质。我们将首先评估EphB 3的A-38类似物 EphB 2、EphB 4和EphA 4激酶测定中的活性和选择性。有希望的化合物将发展到 表型测定以鉴定调节星形胶质细胞致病活性的那些分子。化合物 然后将在药物样性质测定中表征感兴趣化合物，以选择用于PO PK研究的先导化合物。 在该项目的R61阶段结束时，我们期望鉴定出一种主要的EphB 3抑制剂和1-2种备用的EphB 3抑制剂。 适合于推进到R33阶段的体内功效测试。 特定目的2：在PMS临床前模型中评价铅化合物。期间 在该项目的R33阶段，我们将评估NOD EAE中的主要EphB 3抑制剂和1-2种备用药物， 铜腙诱导的小鼠临床前模型，其概括了PMS的几个方面。我们将评估 所述化合物对疾病发展、CNS炎症、轴突损失和脱髓鞘的作用， 通过组织病理学、流式细胞术和基因表达分析确定。 总之，该项目将确定一种新的EphB 3抑制剂，具有药物样特性，适合于开发 作为PMS和其他神经系统疾病的候选治疗药物，得到NIH和/或行业支持。

PROJECT SUMMARY Pathology driven by resident cells in the central nervous system (CNS) such as astrocytes and microglia plays an important role in multiple sclerosis (MS), particularly for its progressive stage which is eventually achieved by most MS patients. However, no drugs are available for the modulation astrocyte and microglia pathogenic activities. We developed an in vitro assay to identify compounds that modulate astrocyte pathogenic activities. These studies identified compound A-38, an inhibitor of the Erythropoietin-producing hepatocellular carcinoma receptor B3 (EphB3), as a suppressor of astrocyte pathogenic activities and, consequently, as a lead therapeutic compound for progressive MS (PMS). Moreover, our data suggest that microglia-driven EphB3 signaling in astrocytes promotes CNS inflammation and neurodegeneration. Ephrin receptors are known to participate in CNS development, however our findings define a novel role for EphB3 in CNS inflammation and identify it as potential target for therapeutic intervention. We hypothesize that EphB3 kinase inhibition will provide an efficacious therapeutic approach for PMS, and potentially other neurologic diseases. Thus, we propose to develop new A-38 analogs to treat PMS. Our Specific Aims are: SPECIFIC AIM 1: OPTIMIZATION AND IN VITRO EVALUATION OF EPHB3 KINASE INHIBITORS. We will design, synthesize and evaluate the properties of up to 150 analogs of A-38. The goal of these studies is to optimize the potency and kinase selectivity of new analogs of A-38, while maintaining good drug-like properties and brain exposure following oral delivery. During the R61 phase of this project, we will integrate iterative structure-activity relationship (SAR) studies and structure based drug design to optimize A-38 potency and kinase selectivity, while maintaining good drug-like properties. We will first evaluate A-38 analogs for EphB3 activity and for selectivity in EphB2 EphB4, and EphA4 kinase assays. Promising compounds will advance to phenotypic assays to identify those molecules that modulate astrocyte pathogenic activities. Compounds of interest will then be characterized in drug-like property assays to select leading compounds for PO PK studies. At the end of the R61 phase of this project we expect to identify a lead EphB3 inhibitor and 1-2 back-ups suitable for advancement into in vivo efficacy testing in the R33 phase. SPECIFIC AIM 2: EVALUATION OF LEAD COMPOUND(S) IN PMS PRE-CLINICAL MODELS. During the R33 phase of this project, we will evaluate the lead EphB3 inhibitor and 1-2 back-ups in the NOD EAE and cuprizone-induced murine pre-clinical models which recapitulate several aspects of PMS. We will evaluate the effects of the compounds on disease development, CNS inflammation, axonal loss and demyelination, as determined by histopathology, flow cytometry and gene expression analyses. In summary, this project will identify a novel EphB3 inhibitor with drug-like properties suitable for development as a candidate therapeutic for PMS and other neurologic diseases with NIH and/or industry support.