Beta-Arrestin-Biased Agonism at the D1 Receptor as a Novel Approach to Levodopa-Induced Dyskinesias in Advanced Parkinson's Disease

项目摘要 帕金森病（PD）是世界上第二大最常见的神经退行性疾病， 在60岁以上的人群中，患病率估计约为1%，使其成为一个越来越重要的 我们老龄化人口的医疗问题。左旋多巴是目前治疗PD的标准药物， 在多巴胺耗尽的黑质纹状体通路中集中增加多巴胺水平。然而，左旋多巴是 不是一种长期治疗用途的可行药物，因为其具有不良副作用，特别是称为 常报告左旋多巴诱导的运动障碍（LID）。据估计，LID发生在超过50%的 PD患者在治疗5至10年后，不成比例地影响老年PD患者， 通过实质上限制该人群亚组的治疗选择来控制晚期疾病阶段。 最近的研究已经阐明，除了G蛋白信号传导，多巴胺受体也可以 通过不同β-抑制蛋白2（B-ARR 2）依赖性途径进行信号传导。这条通路在调节 在多巴胺1受体（D1 R）的下游反应，并在转换多巴胺 信号进入运动功能。以前的研究表明，在D1 R基因上的b-arr 2信号的遗传调节， 啮齿动物和非人灵长类PD模型改善了运动功能，同时预防了LID。差动 通过配体激活这些不同的下游信号传导途径被称为“功能选择性”或 迄今为止，D1 R处的ARR-ARR 2信号通路尚未被靶向。𝛽 以功能选择性的方式潜在地减少与左旋多巴相关的副作用。 为了实现这一点，目标1提出了新的D1 R配体的合成和表征， 系统地修饰先前鉴定的D1 R-选择性、非儿茶酚先导化合物。配体 将使用三种检测其在激活G 蛋白和b-arr 2信号转导。将使用以下方法评估每种配体的血脑屏障通透性： 经验证的人工膜测定。在目标2中，在计算机对接研究中，基于分子动力学的自由基- 能量计算和模型验证实验，利用先前合成的类似物， 将进行不同的功能选择性概况，以鉴定D1 R结构元件， 对a-arr2偏差很重要。这些结果将提供关键的结构信息，以帮助推断结构 在D1 R处的B-arr 2募集机制和结构-功能选择性关系（SFSR）， 通知进一步支架优化成有效的、有β-arr 2偏向的化合物。 该项目的发现将通过提供化学工具， 这将使人们能够研究D1 R如何发生偏置信号，以及下游的分子通路如何发生偏置信号。 受体参与PD和LID病理生理学。从这些研究中获得的见解将提供 重要的线索，指导新的治疗方法，这一具有挑战性的，未解决的医学问题。 !

PROJECT SUMMARY Parkinson’s Disease (PD) is the second most common neurodegenerative disease in the world with a prevalence estimated to be approximately 1% in people over age 60, making it an increasingly important medical problem in our aging population. Levodopa is the current standard of care for PD and functions by increasing levels of dopamine centrally in the dopamine-depleted nigrostriatal pathway. Levodopa, however, is not a viable agent for long-term therapeutic use as undesirable side effects, notably motor fluctuations termed Levodopa-induced dyskinesias (LIDs), are commonly reported. LIDs are estimated to occur in over 50% of PD patients after 5 to 10 years of treatment and disproportionately impact older PD patients at more advanced disease stages by substantially limiting the therapeutic options for this population subset. Recent research has elucidated that, in addition to G protein signaling, dopamine receptors can also signal through a distinct b-arrestin2 (b-arr2)-dependent pathway. This pathway is important in regulating downstream responses at the Dopamine 1 Receptor (D1R) and plays a significant role in converting dopamine signaling into motor function. Previous studies showed that genetic modulation of b-arr2 signaling at D1R in rodent and non-human primate PD models improved motor functioning, while preventing LIDs. The differential activation of these distinct downstream signaling pathways by a ligand is termed “functional selectivity” or “biased agonism”, and to date, the 𝛽-arr2 signaling pathway at D1R has not been pharmacologically targeted in a functionally selective manner to potentially reduce the adverse effects associated with Levodopa. To accomplish this, Aim 1 proposes the synthesis and characterization of new D1R ligands by systematically modifying a previously identified D1R-selective, non-catechol lead compound. Ligands will be profiled for functional selectivity using three assays that detect their relative potencies at activating G protein and b-arr2 signaling at D1R. The blood-brain barrier permeability of each ligand will be assessed using a validated artificial membrane assay. In Aim 2, in silico docking studies, molecular dynamics-based free- energy calculations, and model validation experiments utilizing previously synthesized analogues with diverse functional selectivity profiles will be performed to identify D1R structural elements that are important for 𝜷-arr2 bias. These results will provide critical structural information to help deduce a structural mechanism for 𝛽-arr2 recruitment at D1R and a structure-functional selectivity relationship (SFSR) that will inform further scaffold optimization into a potent, 𝛽-arr2-biased compound. Findings from this project will greatly advance knowledge in the field by providing chemical tools that will enable the study of how biased signaling occurs at D1R and how molecular pathways downstream of this receptor contribute to PD and LID pathophysiologies. The insights gained from such studies will provide important clues guiding novel therapeutic approaches to this challenging, unsolved medical problem. !