Abstract
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.
Original language | English |
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Pages (from-to) | 205-212 |
Number of pages | 8 |
Journal | Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids |
Volume | 1761 |
Issue number | 2 |
Early online date | 23 Jan 2006 |
DOIs | |
Publication status | Published - Feb 2006 |
Bibliographical note
Funding Information:The authors acknowledge the National Institute of Drug Abuse (Grants DA-09789, DA-08904 and subaward 520479/PO P642467) for partly funding this work. We are grateful to Prof. N. Ueda, Kagawa University, Japan, for the generous gift of NAPE-PLD cDNA construct.
Keywords
- 2-Arachidonoylglycerol
- Cannabinoid
- Diacylglycerol
- Inhibitor
- Lipase