Lack of selectivity of URB602 for 2‐oleoylglycerol compared to anandamide hydrolysis in vitro

Abstract

Background and purpose: Two compounds, URB602 and URB754, have been reported in the literature to be selective inhibitors of monoacylglycerol lipase, although a recent study has questioned their ability to prevent 2-arachidonoyl hydrolysis by brain homogenates and cerebellar membranes. In the present study, the ability of these compounds to inhibit monoacylglycerol lipase and fatty acid amide hydrolase has been reinvestigated. Experimental approach: Homogenates and cell lines were incubated with test compounds and, thereafter, with either3 H]-anandamide. Labelled reaction products were separated from substrate using chloroform: methanol extraction. Key results: In cytosolic fractions from rat brain, URB602 and URB754 inhibited the hydrolysis of 2-oleoylglycerol with IC 50 values of 25 and 48 mM, respectively. Anandamide hydrolysis by brain membranes was not sensitive to URB754, but was inhibited by URB602 (IC 50 value 17 mM). Hydrolysis of 2-oleoylglycerol by human recombinant monoacylglycerol lipase was sensitive to URB602, but not URB754. The lack of selectivity of URB602 for 2-oleoylglycerol compared to anandamide hydrolysis was also observed for intact RBL2H3 basophilic leukaemia cells. C6 glioma expressed mRNA for monoacylglycerol lipase, and hydrolyzed 2-oleoylglycerol in a manner sensitive to inhibition by methyl arachidonoyl fluorophosphonate but not URB754 or URB597. MC3T3-E1 mouse osteoblastic cells, which did not express mRNA for monoacylglycerol lipase, hydrolyzed 2-oleoylglycerol in the presence of URB597, but the hydrolysis was less sensitive to methyl arachidonoyl fluorophosphonate than for C6 cells. Conclusions and implications:The data demonstrate that the compounds URB602 and URB754 do not behave as selective and/or potent inhibitors of monoacylglycerol lipase.

The synthesis and biological evaluation of para-substituted phenolic N-alkyl carbamates as endocannabinoid hydrolyzing enzyme inhibitors

“…Moreover, inhibition of FAAH and MGL by reference compounds, FAAH inhibitor URB597 (4) and previously reported selective MGL inhibitor URB602 (5), were evaluated. The results of reference compounds showed good correlation with the ones reported in the literature [29,42,43].…”

“…URB602 has also been reported to inhibit AEA hydrolysis by rat brain membranes (IC 50 ; 17 mM) [43]. In the present study URB602 inhibited FAAH with an IC 50 value of 108 mM.…”

“…Additionally, the N-biphenyl-3-cyclohexylcarbamic acid cyclohexyl ester (5, URB602) ( Fig. 2) was originally suggested to be a selective MGL inhibitor [41], although its selectivity data has recently been questioned [42,43]. Sulfhydryl-reactive compounds such as p-chloromercuribenzoic acid, N-ethylmaleimide (NEM) [9,44,45] and disulfiram [46] inhibit MGL activity, indicating the presence of an essential sulfhydryl group at the active site of the enzyme.…”

Microglia produce and hydrolyze palmitoylethanolamide

“…Accordingly, recombinant FAAH is capable of using both AEA and PEA as substrates (Boger et al 2000;Wei et al 2006). Note that one study showed that recombinant FAAH activity measured in homogenates is inhibited by micromolar concentrations of URB602 ( (Vandevoorde et al 2007). Second, it is possible that an unknown, URB602-sensitive, enzymatic activity is expressed by BV-2 cells and is located where PEA is synthesized in intact cells.…”

Therapeutic Potential of Fatty Acid Amide Hydrolase, Monoacylglycerol Lipase, and N-Acylethanolamine Acid Amidase Inhibitors

“…156 The finding that 150 elevates 2-AG levels without affecting AEA levels at a 100 μM concentration in rat brain slice cultures 155 is probably due to the fact that more than 80% inhibition of FAAH is required for a detectable increase of AEA level. 157 Compound 150 was determined to covalently but partially reversibly bind to MAGL by dialysis experiments.…”