Toward a Better Understanding of Fatty Acid Amide Hydrolase Inhibition by β-Lactams

Fatty acid amide hydrolase (FAAH) inhibition holds therapeutic promise by enhancing endocannabinoid signaling. We previously identified β-lactam compounds as reversible and selective hFAAH inhibitors with nanomolar potency. Here, we describe the synthesis of new β-lactam derivatives to evaluate the effect of imide conformational constraints on activity and to eliminate potential metabolic soft spots. Bicyclic derivatives were designed to lock the imide in a syn configuration, but showed reduced potency compared with non-cyclic analogs. Docking studies revealed that this weaker inhibition arises from an altered binding mode within the FAAH active site. In parallel, removal of ester and allyl groups did not affect inhibitory potency. Importantly, the optimized inhibitor enhanced N-acylethanolamine levels in J774 cells, supporting target engagement and suggesting improved metabolic stability. These results provide insights into the mode of action of β-lactam FAAH inhibitors and guide the development of more potent, stable derivatives.