Benzisothiazolinone Derivatives as Potent Allosteric Monoacylglycerol Lipase Inhibitors That Functionally Mimic Sulfenylation of Regulatory Cysteines

We describe a set of benzisothiazolinone (BTZ) derivatives
that
are potent inhibitors of monoacylglycerol lipase (MGL), the primary
degrading enzyme for the endocannabinoid 2-arachidonoyl-sn-glycerol (2-AG). Structure–activity relationship studies
evaluated various substitutions on the nitrogen atom and the benzene
ring of the BTZ nucleus. Optimized derivatives with nanomolar potency
allowed us to investigate the mechanism of MGL inhibition. Site-directed
mutagenesis and mass spectrometry experiments showed that BTZs interact
in a covalent reversible manner with regulatory cysteines, Cys201
and Cys208, causing a reversible sulfenylation known to modulate MGL
activity. Metadynamics simulations revealed that BTZ adducts favor
a closed conformation of MGL that occludes substrate recruitment.
The BTZ derivative 13 protected neuronal cells from oxidative
stimuli and increased 2-AG levels in the mouse brain. The results
identify Cys201 and Cys208 as key regulators of MGL function and point
to the BTZ scaffold as a useful starting point for the discovery of
allosteric MGL inhibitors.