Cyclic EphA4 peptide antagonists for neuroprotection in ALS

 描述(由申请人提供)：受体酪氨酸激酶EphA4与几个神经退行性变过程有关。值得注意的是，最近的一项研究表明，EphA4的低表达和功能丧失突变与肌萎缩侧索硬化症(ALS)的较晚发病和延长生存期有关。ALS是一种毁灭性的神经退行性疾病，目前尚无治愈方法，FDA批准的唯一疗法只能将生存期延长几个月。即使部分EphA4基因失活也在ALS动物模型中显示出有益的效果，使抑制EphA4功能成为对抗神经退行性变的一种有吸引力的策略。因此，我们之前发现的一种选择性EphA4拮抗剂的多肽在经典的大鼠SOD1 G93A ALS模型中显示出了希望，在该模型中，它显著抑制了ALS的发病机制，证明了EphA4拮抗剂的治疗潜力。尽管有这些令人兴奋的结果，我们的第一代EphA4拮抗剂在生物系统中缺乏所需的效力和稳定性，无法使它们适合作为药物开发的先导。因此，在这项申请中，我们建议开发和评估新的有效的EphA4拮抗剂，其药理特性适合于使它们成为未来ALS治疗的真正治疗线索。在我们的初步工作中，我们开发了一种环状十二肽的衍生物，它能特异性地抑制EphA4-ePhrin的结合，IC50值为~25 nM。这一原型是一种惊人的80倍于任何先前已知的EphA4拮抗剂的效力，并具有优化成为所需的治疗先导的潜力。为了开发相关的环状EphA4拮抗剂以实现这一目标，我们提出了一种高度集成的迭代策略，利用多肽药物化学、结构生物学和我们在EphA4神经生物学方面的长期专业知识。合理设计的新型多肽拮抗剂的活性将通过生化分析和神经细胞培养模型进行评估。作为对这些研究的补充，我们最有希望的新型EphA4拮抗剂将根据它们在血浆和脑脊液中的特性以及它们穿越血脑屏障的倾向以及它们在血液循环中的半衰期进行分析。最后，与著名的肌萎缩侧索硬化症专家Wim Robberecht博士合作进行的活体研究将在肌萎缩侧索硬化症小鼠模型中确定最好的拮抗剂的特征，以延缓疾病的发病并提高存活率。我们预计，拟议的研究将产生一种有效的EphA4抑制剂，其药理特性适合立即开发为单独或与其他治疗方法联合使用的治疗剂。它们还将为EphA4抑制神经退行性变的益处的潜在机制提供有价值的见解。

 DESCRIPTION (provided by applicant): The receptor tyrosine kinase EphA4 has been implicated in several neurodegenerative processes. Strikingly, a recent study has shown that low EphA4 expression and loss-of-function mutations are linked to late onset and prolonged survival in amyotrophic lateral sclerosis (ALS), a devastating neurodegenerative illness for which there is no cure and the only FDA-approved therapy can increase survival by only several months. Even partial EphA4 gene inactivation has shown beneficial effects in animal models of ALS, making inhibition of EphA4 function an attractive strategy to counteract neurodegeneration. Accordingly, a peptide that we previously discovered as a selective EphA4 antagonist has shown promise in the classic rat SOD1 G93A ALS model, where it significantly dampened ALS pathogenesis demonstrating the therapeutic potential of EphA4 antagonistic agents. Despite these exciting results, our first generation EphA4 antagonists lack the required potency and stability in biological systems to make them suitable as leads for drug development. Thus, in this application we propose to develop and evaluate new potent EphA4 antagonists with pharmacological properties suitable to make them bona fide therapeutic leads for future treatment of ALS. In our preliminary work, we have developed a derivative of a cyclic dodecapeptide that specifically inhibits EphA4-ephrin binding with an IC50 value of ~25 nM. This prototype is a striking 80 fold more potent than any previously known EphA4 antagonist and has the potential for optimization to become a desired therapeutic lead. To develop related cyclic EphA4 antagonists towards this goal, we propose a highly integrated iterative strategy using peptide medicinal chemistry, structural biology and our long-standing expertise in EphA4 neurobiology. The activities of rationally designed new peptide antagonists will be evaluated using biochemical assays and neuronal cell culture models. Complementing these studies, our most promising new EphA4 antagonists will be profiled for their properties in plasma and cerebrospinal fluid and their propensity to cross the blood-brain barrier, as well as their half-lie in the blood circulation. Finally, in vivo studies in collaboration with renowned ALS expert Dr. Wim Robberecht will characterize the best antagonists in delaying disease onset and promoting survival in an ALS mouse model. We anticipate that the proposed studies will result in a potent EphA4 inhibitor with a pharmacologic profile suited for immediate development as a therapeutic agent alone or in combination with other treatments. They will also provide valuable insight into the mechanism underlying the benefits of EphA4 inhibition against neurodegeneration.