Parallel Multimodal High-throughput screening to identify activators of the orexin receptors

项目摘要 配体调节的受体的GPCR超家族已被证明是靶点的丰富来源， 发展出治疗无数人类疾病的方法。食欲素1和食欲素2受体是A类 GPCR在中枢神经系统中的表达不同增食欲素1受体是最丰富的 表达于蓝斑，被认为控制情绪、奖励和自主神经方面 神经系统，而食欲素2受体表达在控制觉醒的区域，如 乳头状结节核是调节睡眠/觉醒的重要部位。在他们离开后近20年， 最初的发现，第一个有效的双重食欲素1 /食欲素2受体拮抗剂治疗已被带到 用于治疗失眠症（Belsomera ®，suvorexant，Merck）。尽管人们付出了巨大的努力来识别这种病毒的对手， 食欲素受体，在原始或专利文献中几乎没有小分子激动剂的报道。优雅 使用食欲素敲除动物的遗传消除实验和脑室内给药的实验 食欲素A或食欲素B肽的作用表明食欲素受体激动剂或增强剂对许多潜在的 适应症包括：1）抑郁症; 2）学习和记忆（认知）; 3）注意缺陷多动障碍 （ADHD）; 4）结肠癌的治疗;和5）睡眠障碍，包括嗜睡症。虽然食欲肽是 作为两种食欲素受体的有效激动剂，它们不是选择性的，也不能很好地穿过血脑屏障， 使它们成为体内药理学研究的不良探针底物。这为识别小型 分子，食欲素受体的脑渗透激活剂，以询问受体在以下情况下的功能： 疾病状态。我们已经优化了基于细胞的高通量筛选兼容的初级测定， 特异性测量食欲素1和食欲素2的功能活性。一个初步的10 k试点屏幕是 导致鉴定出这些受体的许多小分子激动剂， 该测定是稳健的和可重现的。斯克里普斯机构药物的全面HTS筛选活动 发现库（~ 640 k化合物）将导致识别多种类别的食欲素受体激动剂 以及进一步发展的增效剂。在迭代过程中，将对这些经验证的命中进行表征 通过一系列基于细胞的体外试验来确定效力、选择性和作用机制。 初步药物化学铅优化将确定早期结构活性关系和体外 将评估药物代谢以确认早期导联的可追踪性。这些努力将提供一流的 化学工具，用于进一步探测动物疾病模型中的食欲素受体功能。

PROJECT SUMMARY The GPCR superfamily of ligand regulated receptors has proven to be a rich source of targets for development of therapeutics for a myriad of human diseases. The orexin 1 and orexin 2 receptors are class A GPCR’s differentially expressed in the central nervous system. The orexin 1 receptor is most abundantly expressed in the locus coeruleus and is thought to control aspects of emotion, reward, and the autonomic nervous system, whereas the orexin 2 receptor is expressed in regions controlling arousal such as the tubermammillary nucleus, an important site for the regulation of sleep/wakefulness. Almost 20 years after their initial discovery, the first potent dual orexin 1 / orexin 2 receptor antagonist therapeutic has been brought to market for insomnia (Belsomera ®, suvorexant, Merck). Despite a massive effort to identify antagonists of the the orexin receptor, almost no reports of small molecule agonists appear in the primary or patent literature. Elegant genetic ablation experiments using orexin knock-out animals and experiments with intracerebroventricular dosing of orexin A or orexin B peptides suggests a role for orexin receptor agonists or potentiators for a number of potential indications including: 1) depression; 2) learning and memory (cognition); 3) attention deficit hyperactivity disorder (ADHD); 4) treatment for colon cancer; and 5) sleep disorders including narcolepsy. While the orexin peptides are potent agonists of both orexin receptors, they are not selective nor do they cross the blood brain barrier well, making them poor probe substrates for in vivo pharmacology studies. This provides the impetus to identify small molecule, brain penetrant activators of the orexin receptors to interrogate the receptors function in the context of disease states. We have optimized a cell-based high-throughput screening compatible primary assay that specifically measures the functional activity of orexin 1 and orexin 2. A preliminary 10k pilot screen was performed leading to the identification of a number of small molecule agonists of these receptors demonstrating this assay to be robust and reproducible. A full HTS screening campaign of the Scripps Institutional Drug Discovery Library (~640k compounds) will lead to the identification of multiple classes of orexin receptor agonists and potentiators for further development. In an iterative process, these validated hits will be characterized through a cascade of in vitro cell-based assays to determine potency, selectivity and mechanism of action. Preliminary medicinal chemistry lead optimization will identify early structure activity relationships and in vitro drug metabolism will be assessed to confirm tractibility of early leads. These efforts will provide first in-class chemical tools to be used to further probe orexin receptor function in animal models of disease.