Highly Selective, Reversible Inhibitor Identified by Comparative Chemoproteomics Modulates Diacylglycerol Lipase Activity in Neurons

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• 作者：Marc P. Baggelaar ; Pascal J. P. Chameau ; Vasudev Kantae ; Jessica Hummel ; Ku-Lung Hsu ; Freek Janssen ; Tom van der Wel ; Marjolein Soethoudt ; Hui Deng ; Hans den Dulk ; Marco Allar脿 ; Bogdan I. Florea ; Vincenzo Di Marzo ; Wytse J. Wadman ; Chris G. Kruse ; Herman S. Overkleeft ; Thomas Hankemeier ; Taco R. Werkman ; Benjamin F. Cravatt ; Mario van der Stelt
• 刊名：Journal of the American Chemical Society
• 出版年：2015
• 出版时间：July 15, 2015
• 年：2015
• 卷：137
• 期：27
• 页码：8851-8857
• 全文大小：453K
• ISSN：1520-5126

文摘

Diacylglycerol lipase (DAGL)-伪 and -尾 are enzymes responsible for the biosynthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG). Selective and reversible inhibitors are required to study the function of DAGLs in neuronal cells in an acute and temporal fashion, but they are currently lacking. Here, we describe the identification of a highly selective DAGL inhibitor using structure-guided and a chemoproteomics strategy to characterize the selectivity of the inhibitor in complex proteomes. Key to the success of this approach is the use of comparative and competitive activity-based proteome profiling (ABPP), in which broad-spectrum and tailor-made activity-based probes are combined to report on the inhibition of a protein family in its native environment. Competitive ABPP with broad-spectrum fluorophosphonate-based probes and specific 尾-lactone-based probes led to the discovery of 伪-ketoheterocycle LEI105 as a potent, highly selective, and reversible dual DAGL-伪/DAGL-尾 inhibitor. LEI105 did not affect other enzymes involved in endocannabinoid metabolism including abhydrolase domain-containing protein 6, abhydrolase domain-containing protein 12, monoacylglycerol lipase, and fatty acid amide hydrolase and did not display affinity for the cannabinoid CB₁ receptor. Targeted lipidomics revealed that LEI105 concentration-dependently reduced 2-AG levels, but not anandamide levels, in Neuro2A cells. We show that cannabinoid CB₁-receptor-mediated short-term synaptic plasticity in a mouse hippocampal slice model can be reduced by LEI105. Thus, we have developed a highly selective DAGL inhibitor and provide new pharmacological evidence to support the hypothesis that 鈥渙n demand biosynthesis鈥?of 2-AG is responsible for retrograde signaling.