Development of the first potent and specific inhibitors of endocannabinoid biosynthesis

详细信息    查看全文

• 作者：Tiziana Bisogno ; Maria Grazia Cascio ; Bijali Saha ; Anu Mahadevan ; Paolo Urbani ; Alberto Minassi ; Giovanni Appendino ; Carmela Saturnino ; Billy Martin ; Raj Razdan and Vincenzo Di Marzo
• 关键词：Inhibitor ; Diacylglycerol ; Cannabinoid ; 2-Arachidonoylglycerol ; Lipase
• 刊名：Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids
• 出版年：2006
• 出版时间：February 2006
• 年：2006
• 卷：1761
• 期：2
• 页码：205-212
• 全文大小：241 K

文摘

Enzymes for the biosynthesis and degradation of the endocannabinoid 2-arachidonoyl glycerol (2-AG) have been cloned and are the sn-1-selective-diacylglycerol lipases α and β (DAGLα and β) and the monoacylglycerol lipase (MAGL), respectively. Here, we used membranes from COS cells over-expressing recombinant human DAGLα to screen new synthetic substances as DAGLα inhibitors, and cytosolic fractions from wild-type COS cells to look for MAGL inhibitors. DAGLα and MAGL activities were assessed by using sn-1-[¹⁴C]-oleoyl-2-arachidonoyl-glycerol and 2-[³H]-arachidonoylglycerol as substrates, respectively. We screened known compounds as well as new phosphonate derivatives of oleic acid and fluoro-phosphinoyl esters of different length. Apart from the general lipase inhibitor tetrahydrolipstatin (orlistat®) (IC₅₀ 60 nM), the most potent inhibitors of DAGLα were O-3640 [octadec-9-enoic acid-1-(fluoro-methyl-phosphoryloxymethyl)-propylester] (IC₅₀ = 500 nM), and O-3841 [octadec-9-enoic acid 1-methoxymethyl-2-(fluoro-methyl-phosphinoyloxy)-ethyl ester] (IC₅₀ = 160 nM). Apart from being almost inactive on MAGL, these two compounds showed high selectivity over rat liver triacylglycerol lipase, rat N-acylphosphatidyl-ethanolamine-selective phospholipase D (involved in anandamide biosynthesis), rat fatty acid amide hydrolase and human recombinant cannabinoid CB₁ and CB₂ receptors. Methylarachidonoyl-fluorophosphonate and the novel compound UP-101 [O-ethyl-O-p-nitro-phenyl oleylphosphonate] inhibited both DAGLα and MAGL with similar potencies (IC₅₀ = 0.8–0.1 and 3.7–3.2 μM, respectively). Thus, we report the first potent and specific inhibitors of the biosynthesis of 2-AG that may be used as pharmacological tools to investigate the biological role of this endocannabinoid.