Structural elucidation and development of agonists for the human orexin receptors

本提案的目的是获得对人食欲素受体激动剂的生物物理学理解 结合和激活，并利用这一知识开发小分子食欲素受体激动剂， 神经药理学工具和用于发作性睡病/紧张症和其它疾病的潜在治疗剂。 嗜睡症是一种终身衰弱性疾病，影响大约20万美国人， 以无法保持清醒、睡眠发作、肌肉功能突然丧失和睡眠为特征的 瘫痪目前的嗜睡症治疗（如精神兴奋剂药物）不治疗潜在的 神经化学缺陷并表现出不希望的副作用。人类的动物模型和临床研究 患者表明，发作性睡病是由食欲素（下丘脑分泌素）神经肽缺乏引起的， 神经元的外侧下丘脑，并认为外源性替代食欲素活性可能治愈疾病。 然而，食欲素不能用作治疗剂，因为它们是肽，其不能穿透胃肠道。 在口服给药后，由于代谢分解，血脑屏障显示出较差的活性。 我们建议使用GPCR结构生物学的新技术进展来确定 食欲素受体在食欲素结合和小分子激动剂结合状态，揭示了详细的非- 稳定这些复合物的共价相互作用以及受体构象的变化， 是激动剂结合的结果。在第二个目标中，我们将开发构象特异性纳米抗体 结合并稳定食欲素受体的活性状态，并解决纳米抗体共晶体结构，以了解 跨膜传播的结构变化连接细胞外神经肽结合位点 和细胞内G蛋白偶联位点。在第三个目标中，我们将把结构性见解与 计算对接/模拟和药物化学，以提高小分子的亲和力和效力， 分子食欲素受体激动剂，其先前在高通量筛选中鉴定。我们的组合 在GPCR结构生物学、合成和药物化学以及计算化学方面的优势 使我们处于一个独特的位置，设计具有药物样特性的小分子食欲素模拟物， 进一步发展成为治疗发作性睡病和其他神经障碍的治疗剂。

The goal of this proposal is to gain a biophysical understanding of human orexin receptor agonist binding and activation, and to use this knowledge to develop small-molecule orexin receptor agonists as neuropharmacological tools and potential therapeutics for narcolepsy/cataplexy and other diseases. Narcolepsy is a life-long debilitating disorder affecting approximately 200,000 Americans, which is characterized by an inability to maintain wakefulness, sleep attacks, sudden loss of muscle function, and sleep paralysis. Current treatments for narcolepsy (such as psychostimulant drugs) do not treat the underlying neurochemical deficits and exhibit undesirable side-effects. Animal models and clinical investigations of human patients show that narcolepsy is caused by deficiency of the orexin (hypocretin) neuropeptides produced by neurons of the lateral hypothalamus, and that exogenous replacement of orexin activity may cure the disease. However, orexins cannot be used as therapeutic agents because they are peptides, which do not penetrate the blood-brain barrier and show poor activity after oral administration due to metabolic decomposition. We propose to use new technical advances in GPCR structural biology to determine X-ray structures of the orexin receptors in orexin-bound and small-molecule agonist-bound states, revealing the detailed non- covalent interactions that stabilize these complexes as well as changes in conformation of the receptors that are a consequence of agonist binding. In the second Aim, we will develop conformation-specific nanobodies that bind and stabilize the orexin receptor active state, and solve nanobody co-crystal structures to understand the propagated structural changes across the membrane that link the extracellular neuropeptide binding site and the intracellular G protein coupling site. In the third Aim, we will integrate structural insights with computational docking/simulation and medicinal chemistry to improve the affinity and potency of small- molecule orexin receptor agonists that were previously identified in a high-throughput screen. Our combination of strengths in GPCR structural biology, synthetic and medicinal chemistry, and computational chemistry places us in a unique position to design small-molecule orexin mimics with drug-like properties that can be further developed into therapeutics for the treatment of narcolepsy and other neurological disorders.