Exploiting Biased Agonism at the Ghrelin Receptor (GHSR1a) for Opioid Addiction

摘要 在美国，诊断为阿片类药物使用障碍(OUD)的人数已超过200万。尽管现有的音频 药物(如美沙酮、丁丙诺啡、纳曲酮)可改善症状和复发率。 并不是没有问题的副作用，停药仍然是一个问题。因此，NIDA 已经优先开发新的药物来预防和/或治疗OUD。NIDA评选出的十大最 乌德的药理靶标是生长激素释放素受体，生长激素促分泌素受体1a。 (GHSR1a)。GHSR1a在中脑边缘(“奖赏”)系统的多巴胺(DA)神经元中密集表达， 它的激活导致DA释放增加，并增强了获得奖励/强化刺激的动机 (例如，滥用的食物、药物)。与其他GPCR一样，GHSR1a的生理效应取决于G蛋白是否- 或b-arrestin(Barr)依赖的信号通路优先参与，这一过程称为功能性 选择性(或有偏见的激动主义)。相当多的先前证据支持利用GPCR功能选择性 可提供一种提高药物治疗效果和/或减少有害副作用的方法- 效果。Caron研究小组此前已经证明，GHSR1a拮抗剂可以阻断可卡因的多动- 通过DA神经元的Barr依赖机制致敏小鼠。此外，Caron集团还 证明了GHSR1a诱导的肌动蛋白应激纤维的形成--这是一个完整的分子过程 强化物质所产生的神经适应性变化--是巴尔介导的。在大规模的药物筛查中 与药物化学相结合的新型GHSR1a配体，Caron小组与NCATS合作，已经 确定了一种先导化合物8279，它作为GHSR1a选择性G蛋白偏向激动剂，因此， 减弱GHSR1a介导的Barr信号。初步数据显示，8279减少了基础多动 多巴胺转运蛋白基因敲除(DAT KO)小鼠(一种高DAergia的动物模型)，但不影响享乐性 喂食。这项应用的目的是描绘G蛋白偏向配体的机制 GHSR1a，并在阿片类药物滥用的小鼠模型中询问其治疗潜力。中心假说 偏向G蛋白的GHSR1a激动剂是否通过抑制阿片类药物诱导的神经可塑性和自我给药 在GHSR1a激活后，Barr招募和/或信号转导减少。这一假设将通过以下方式检验 使用8279或其衍生物作为我们的主要工具化合物来追求三个特定目标：1)阐明 GHSR1a偏向信号的生化机制，2)确定偏向GHSR1a的影响 信号对神经可塑性的影响，以及3)评估GHSR1a偏向的信号对阿片诱导的行为的影响。 这三个目标将使用已建立的GPCR交易、蛋白质-蛋白质相互作用和信号传递来测试 测试，基于病毒载体的脑组织神经可塑性测量，以及小鼠自我给药。这些 研究是有意义的和新颖的，因为它们将提供对潜在的临床前研究。 利用GHSR1a功能选择性预防和/或治疗尿毒症的治疗作用。

Abstract In the U.S., the number of Opioid Use Disorder (OUD) diagnoses has exceeded 2 million. Although existing OUD medications (e.g., methadone, buprenorphine, naltrexone) ameliorate OUD symptoms and relapse rates, they are not without problematic side-effects and medication discontinuation remains an issue. Consequently, NIDA has prioritized the development of novel medications to prevent and/or treat OUD. Among NIDA's "10 most wanted" pharmacologic targets for OUD is the ghrelin receptor, growth hormone secretagogue receptor 1a (GHSR1a). GHSR1a is densely expressed within dopamine (DA) neurons of mesolimbic ("reward") system and its activation leads to increased DA release and enhanced motivation to acquire rewarding/reinforcing stimuli (e.g., food, drugs of abuse). Like other GPCRs, the physiological effects of GHSR1a depend whether G protein- or b-arrestin (barr)-dependent signaling pathways are preferentially engaged, a process known as functional selectivity (or biased agonism). Considerable prior evidence supports that leveraging GPCR functional selectivity may provide a means by which to enhance pharmacotherapeutic efficacy and/or diminish deleterious side- effects. The Caron group has previously shown that GHSR1a antagonism blocks hyperlocomotion in cocaine- sensitized mice via a barr-dependent mechanism in DA neurons. Additionally, the Caron group has also demonstrated that GHSR1a-induced actin stress fiber formation -- a molecular process integral to the neuroadaptive changes produced by reinforcing substances -- is barr-mediated. In a large-scale drug screen for novel GHSR1a ligands coupled with medicinal chemistry, the Caron group in collaboration with NCATS, has identified a lead compound, 8279, which acts as a GHSR1a-selective G protein-biased agonist and thus, attenuates GHSR1a-mediated barr signaling. Initial data shows that 8279 reduces basal hyperlocomotion in dopamine transporter knockout (DAT KO) mice (an animal model of hyperDAergia), but does not affect hedonic feeding. The objectives of this application are to delineate the mechanisms of G protein-biased ligands at GHSR1a and interrogate their therapeutic potential in mouse models of opioid abuse. The central hypothesis is that G protein-biased GHSR1a agonism attenuates opioid-induced neuroplasticity and self-administration via reduced barr recruitment and/or signaling following GHSR1a activation. This hypothesis will be tested by pursuing three specific aims using 8279, or its derivatives, as our primary tool compound: 1) Elucidate the biochemical mechanisms underlying GHSR1a biased signaling, 2) Identify the effects of GHSR1a biased signaling on neuroplasticity, and 3) Evaluate the effects of GHSR1a biased signaling on opioid-induced behavior. These three aims will be tested using established GPCR trafficking, protein-protein interaction, and signaling assays, viral vector-based measures of neuroplasticity in brain tissue, and mouse self-administration. These studies are significant and novel in that they will provide a thorough preclinical investigation into the potential therapeutic utility of exploiting GHSR1a functional selectivity for the prevention and/or treatment of OUD.