Small Molecules to Promote Regeneration and Recovery Following Spinal Cord Injury

小分子促进脊髓损伤后的再生和恢复

• 批准号: 10016825
• 负责人: Samy Meroueh
• 金额: --
• 依托单位: RLR VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-10-01 至 2024-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10016825
• 关键词: Affinity Chromatography; Animal Model; Animals; Axon; Binding; Biological Assay; Biology; Blood; Blood - brain barrier anatomy; Brain; Cell Culture Techniques; Clinical Trials; Contusions; Data; Development; Docking; Dorsal; Dose; Drug Industry; Drug Kinetics; Event; Exhibits; Fiber; Forelimb; Goals; Grant; Growth; Guanosine Triphosphate Phosphohydrolases; In Vitro; Injury; Investigational Drugs; Laboratories; Lead; Length; Lesion; Libraries; Lower Extremity; Mass Spectrum Analysis; Maximum Tolerated Dose; Mediating; Methods; Modeling; Monomeric GTP-Binding Proteins; Motor; Mus; Natural regeneration; Neurites; Neurons; Outcome; Paraplegia; Patients; Permeability; Pharmaceutical Chemistry; Positioning Attribute; Property; Proteins; Recovery; Recovery of Function; Sensory; Signal Transduction; Site; Small Molecule Chemical Library; Spinal Cord Contusions; Spinal cord injury; Spinal cord injury patients; Structural Protein; Structure; Testing; Therapeutic Agents; United States; aggrecan; axon regeneration; base; blood-brain barrier crossing; chemical standard; clinically relevant; design; druggable target; economic cost; effective therapy; in vitro Assay; in vivo; in vivo Model; in vivo regeneration; innovation; insight; neuroprotection; protein structure; repaired; screening; small molecule; small molecule libraries; small molecule therapeutics; success; transcriptome sequencing

A. ABSTRACT A significant impediment towards improvement of outcome after spinal cord injury (SCI) is the limited axonal regeneration due to insufficient regrowth of the axonal fibers at the lesion site. Signaling mediated by RhoA GTPases presents a significant impediment to growth. To date, there exists no small-molecule RhoA antagonists that have shown in vivo efficacy. This is attributed to the fact that small GTPases like RhoA do not have druggable pockets, making the development of small molecules that directly bind to them extremely difficult. However, over the years, several proteins with druggable pockets have been discovered to interact with RhoA. We stipulated that small molecules that bind to these druggable targets will promote axonal regeneration and recovery to a similar or greater extent than direct inhibition of RhoA activity. To identify such compounds we pursued a unique approach that consisted of structure-based computational docking of large chemical libraries to druggable targets that interact with RhoA. We tested the top 50-100 compounds for their effect on neurite outgrowth and axonal regeneration. One compound, RLA-47, inhibited in vitro neurite outgrowth and axonal regeneration. The compound also showed substantial axon crossings in an animal model of spinal cord injury. Our central hypothesis is that RLA-47 and its derivatives will promote axonal regeneration and recovery following spinal cord injury. Our preliminary data puts us in a strong position to test our hypothesis. In our first aim, we propose to design and synthesize derivatives of RLA-47 that will be tested in our neurite outgrowth and axonal regeneration in vitro assays. The most promising candidates will be explored in our in vivo hemisection model of spinal cord injury. In our second aim, we propose to develop RLA-47 derivatives with suitable pharmacokinetics and blood- brain permeability that promote axonal regeneration in vivo. We also test the most promising compounds using a more clinically relevant contusion model of spinal cord injury. We will use a standard chemical biology strategy of affinity purification and mass spectrometry to uncover targets of RLA-47 and its derivatives. Along with RNA sequencing, these studies will provide deeper insight into the mechanism of action of compounds. We expect 1-2 derivatives of RLA-47 to exhibit superior pharmacokinetics, brain permeability and efficacy. These compounds will lead to investigational new drug filing that we expect will eventually lead to candidates that can be explored in clinical trials either alone, or in combination with other therapies to promote regeneration and recovery in spinal cord injury patients.

A. 摘要 脊髓损伤 (SCI) 后改善预后的一个重大障碍是轴突受限 由于病变部位轴突纤维再生不足而导致的再生。 RhoA 介导的信号传导 GTPases 对生长构成重大障碍。迄今为止，尚无小分子 RhoA 拮抗剂 已显示出体内功效。这是因为像 RhoA 这样的小 GTP 酶不具有药物作用 口袋，使得开发直接与其结合的小分子变得极其困难。然而，超过 多年来，人们发现了几种具有可药物口袋的蛋白质与 RhoA 相互作用。我们规定 与这些可药物靶标结合的小分子将促进轴突再生和恢复 与直接抑制 RhoA 活性相似或更大程度。为了识别此类化合物，我们寻求了一种独特的方法 方法包括大型化学库与可药物靶标的基于结构的计算对接 与 RhoA 相互作用。我们测试了前 50-100 种化合物对神经突生长和轴突的影响 再生。一种化合物 RLA-47 可抑制体外神经突生长和轴突再生。这 化合物还在脊髓损伤的动物模型中显示出大量的轴突交叉。我们的中央 假设 RLA-47 及其衍生物将促进脊髓损伤后的轴突再生和恢复 受伤。我们的初步数据使我们有能力检验我们的假设。在我们的第一个目标中，我们建议 设计和合成 RLA-47 衍生物，将在我们的神经突生长和轴突再生中进行测试 体外测定。最有希望的候选者将在我们的脊髓半切模型中进行探索 受伤。我们的第二个目标是开发具有合适的药代动力学和血液动力学的 RLA-47 衍生物。 促进体内轴突再生的脑通透性。我们还使用以下方法测试最有前途的化合物 更具临床相关性的脊髓损伤挫伤模型。我们将使用标准的化学生物学策略 亲和纯化和质谱分析以揭示 RLA-47 及其衍生物的靶标。与RNA一起 测序，这些研究将为化合物的作用机制提供更深入的见解。我们期望 RLA-47 的 1-2 衍生物表现出优异的药代动力学、脑通透性和功效。这些 化合物将导致研究性新药备案，我们预计最终将产生可以的候选药物 在临床试验中单独探索，或与其他疗法结合使用，以促进再生和 脊髓损伤患者的康复。