Disulfiram is an Inhibitor of Human Purified Monoacylglycerol Lipase, the Enzyme Regulating 2‐Arachidonoylglycerol Signaling

Abstract

Monoacylglycerol lipase (MAGL) is a key enzyme responsible for the termination of endocannabinoid signaling. Its crucial role in 2-arachidonoylglycerol (2-AG) metabolism, together with the numerous pharmacological properties mediated by this endocannabinoid, emphasize the interest in MAGL as therapeutic target, along with the need to design potent and selective inhibitors. Meanwhile, the complexity of 2-AG degradation pathways underscores the need to use a purified source of enzyme in evaluation studies of new inhibitors. We report here the first heterologous expression and purification of human MAGL. A highly pure protein was obtained and allowed us to measure the affinity of several MAGL inhibitors for the human enzyme. Importantly, disulfiram (tetraethylthiuram disulfide), a compound used to treat alcoholism, and other disulfide-containing compounds were shown to inhibit MAGL with good potency, likely through an interaction with cysteine residues.

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

“…In a series of sulfhydryl-specific maleimide derivatives, N-arachidonylmaleimide (NAM, 6) ( Fig. 2) was found to be a potent inhibitor of MGL, inhibiting 2-AG hydrolysis in rat cerebellar membranes with an IC 50 value of 140 nM [45], a result confirmed using human recombinant MGL (hrMGL), and 2-OG as substrate [46]. Despite containing a highly reactive maleimide group, NAM has been shown to exhibit rather high selectivity for MGL relative to other brain serine hydrolases [37].…”

“…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. In a series of sulfhydryl-specific maleimide derivatives, N-arachidonylmaleimide (NAM, 6) ( Fig.…”

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

“…148 Additionally, two disulfide derivatives, compounds 137 (IC50 = 1.6 µM) and 138 (IC50 = 0.13 µM), were shown to inactivate MAGL-mediated activity in the micromolar range ( Figure 49). 148 These disulfide derivatives were hypothesized to interact with the cysteine residues of MAGL to form a mixed adduct or a disulfide bond. 148 The exploration of numerous analogs of 136 with different substitutions led to discovering a series of selective MAGL inhibitors.…”

“…148 These disulfide derivatives were hypothesized to interact with the cysteine residues of MAGL to form a mixed adduct or a disulfide bond. 148 The exploration of numerous analogs of 136 with different substitutions led to discovering a series of selective MAGL inhibitors. Typically, compound 139 ( Figure 49) exerted optimal inhibitory capacity against hMAGL with an IC50 of 0.11 µM and more than 1000-fold selectivity over hFAAH.…”

Crystal structure of a soluble form of human monoglyceride lipase in complex with an inhibitor at 1.35 Å resolution

“…Contrary to our experiences, two recent independent reports published after this work was initiated demonstrated successful production of a soluble wildtype human MGL that did not require detergent during purification and was not aggregated. 24,25,31 The protein produced was the 313 aminoacids MGL isoform (GenBank AAH06230) versus the 303 aminoacid isoform (GenBank CAC43316) described in this manuscript. Production of the soluble wild-type long isoform of MGL was achieved in Escherichia coli BL21 and Rosetta strain while expression in the baculovirus expression system produced only insoluble, aggregated protein as described above.…”