Science 356, 1084–1087 (2017)

Inhibitors of fatty acid amide hydrolase (FAAH), which degrades amidated lipids, including anandamide, have been developed to treat pain and nervous system disorders. However, in a 2016 phase 1 trial of the FAAH inhibitor BIA 10-2474, one participant died and four others were hospitalized with neurotoxicity after receiving multiple doses of the drug. Previous clinical trials of other FAAH inhibitors reported no serious adverse events. van Esbroeck et al. tested whether off-target effects of BIA 10-2474—mediated by its covalent bonding to serine hydrolase active sites through its electrophilic imidazole urea—might account for the observed neurotoxicity. To do so, the authors incubated cells with BIA 10-2474 or a different FAAH inhibitor and treated lysates with a biotinylated fluorophosphonate probe specific for serine hydrolase active sites. Streptavidin enrichment for biotin-labeled proteins by affinity purification enabled the identification of probe-bound enzymes by MS. Putative inhibitor targets failed to bind the probe. This activity-based profiling of serine hydrolases revealed that BIA 10-2474 and its major metabolite BIA 10-2639 inhibited not only FAAH, but also lipid hydrolases and drug-metabolizing enzymes at a range of concentrations. As many of these enzymes are expressed in human brain, it's possible that the neurotoxicity observed in the BIA 10-2474 trial could be attributed to off-target inhibition resulting in altered brain lipid metabolism.