Role of ABHD6 in 2-AG Signaling

描述（由申请人提供）：内源性大麻素花生四烯酸和2-花生四烯酸甘油（2-AG）通过大麻素CB1和CB2受体起作用，并通过两步过程灭活：两者都被转运到细胞中，然后花生四烯酸被脂肪酸酰胺水解酶（FAAH）水解，2-AG被单酰基甘油脂肪酶（MGL）水解。FAAH或MGL的选择性抑制分别导致花生四烯酸酰胺或2-AG的积累，并导致非重叠的治疗效果。 在一项由NIDA-CEBRA资助的研究中，我们发现小胶质细胞系BV-2不表达MGL，但能有效水解2-AG，这表明存在一种新的2-AG水解酶，可以靶向治疗与神经炎症相关的病理。在与Ben Cravatt的合作中，我们使用功能蛋白质组学方法并将ABHD6鉴定为BV-2细胞中2-AG水解的候选酶。最近，我们使用shRNA敲除和新开发的ABHD 6抑制剂，并表明该酶在BV-2细胞和神经元中的2-AG水解中起主要作用。由于亨廷顿病（HD）与CB1受体的早期下调和慢性神经炎症反应有关，因此抑制ABHD6可能是治疗这种退行性疾病的有效治疗方法。 在这项资助提案中，我们概述了旨在：1）绘制健康和HD小鼠大脑中ABHD6表达和活性的实验，2）确定ABHD6的药理学特征并开发新型抑制剂3）测试ABHD6是否控制小胶质细胞和神经元中的2-AG信号传导，并构成治疗HD小鼠的有效靶标。 这三个目标的完成将为全面了解ABHD6在健康和退化大脑中的小胶质细胞和神经元中的表达、活性和作用提供基础。我们的长期目标是产生选择性抑制内源性大麻素失活的药理学和遗传学工具，作为开发缺乏滥用倾向和大麻素激动剂产生的不良反应的治疗方法的手段。 公共卫生相关性：在这项资助提案中，我们概述了旨在1）绘制新型内源性大麻素水解酶ABHD 6在健康小鼠大脑和两种亨廷顿病小鼠模型中的表达和活性的实验，2）确定药理学特征并开发这种新型酶的抑制剂和3）测试其在控制内源性大麻素调节小胶质细胞和神经元细胞功能的能力中的作用，以及其抑制是否影响亨廷顿病小鼠模型的疾病进展。

DESCRIPTION (provided by applicant): The endocannabinoids anandamide and 2-arachidonoylglycerol (2-AG) act through cannabinoid CB1 and CB2 receptors, and are inactivated by a two-step process: Both are transported into cells, and then anandamide is hydrolyzed by fatty acid amide hydrolase (FAAH) and 2-AG by monoacylglycerol lipase (MGL). Selective inhibition of either FAAH or MGL results in the accumulation of either anandamide or 2-AG, respectively, and in non-overlapping therapeutic effects. In a study funded by a NIDA-CEBRA grant, we found that the microglial cell line, BV-2, does not express MGL and yet efficiently hydrolyzes 2-AG, suggesting the existence of a new 2-AG-hydrolyzing enzyme that could be targeted for the treatment of pathologies associated with neuroinflammation. In collaboration with Ben Cravatt, we used a functional proteomics approach and identified ABHD6 as a candidate enzyme for 2-AG hydrolysis in BV-2 cells. More recently, we used shRNA knockdown and a newly developed ABHD6 inhibitor and showed that this enzyme plays a major role in 2-AG hydrolysis in both BV-2 cells and neurons. Because Hungtington's disease (HD) is associated with an early down-regulation of CB1 receptors and a chronic neuroinflammatory response, inhibiting ABHD6 might represent a valid therapeutic approach to treat this degenerative disease. In this grant proposal, we outline experiments designed to: 1) Map ABHD6 expression and activity in healthy and HD mice brains, 2) Determine the pharmacological profile of ABHD6 and develop novel inhibitors 3) Test if ABHD6 controls 2-AG signaling in microglia and neurons, and constitutes a valid target for treating HD mice. The completion of these three aims will provide a comprehensive understanding of the expression, activity and role of ABHD6 in microglia and neurons in healthy and degenerating brain. Our long-term goal is to generate pharmacological and genetic tools that selectively inhibit endocannabinoid inactivation as means to develop therapeutics that lack the abuse liability and adverse effects produced by cannabinoid agonists. PUBLIC HEALTH RELEVANCE: In this grant proposal, we outline experiments designed to 1) Map the expression and activity of the novel endocannabinoid-hydrolyzing enzyme, ABHD6, in the brains of healthy mice, and of two mice models of Huntington's disease, 2) Determine the pharmacological profile and develop inhibitors of this novel enzyme and 3) Test its role in controlling endocannabinoid's ability to regulate microglial and neuronal cell functions, and whether its inhibition affects disease progression in a mouse model of Huntington's disease.