Inhibition of endocannabinoid hydrolases MAGL, FAAH and ABHD6 by AKU-005 reduces ex vivo cortical spreading depression.

backgroundMigraine is a common neurovascular disorder that remains currently untreated in half of the patients. One third of migraine patients experience aura, which is associated with the development of cortical spreading depolarization (CSD), a wave of depolarization involving neurons and glial cells. Cannabinoids have proven to be a promising class of compounds for the treatment of migraine pain. In this study, we are proposing a new strategy to counteract development of CSD and downstream events via multicomponent enhancement of the endocannabinoid system (ECS) by using a AKU-005, to simultaneously target several key endocannabinoids hydrolases. To this end, we profiled the activity of selective endocannabinoid hydrolases and their inhibition by AKU-005 and analyzed the effect of AKU-005 on the development of CSD in an ex vivo cortical slice model.

methodsThe inhibitory profile of AKU-005 was evaluated by a glycerol assay of lysates from HEK293 cells expressing mouse and human MAGL and ABHD6. After ex vivo treatment of cortex slices of Wistar rats and C57 BL/6 J-OlaHsd mice, endocannabinoids were quantified by mass spectrometry (LC-MS/MS), and activity of the hydrolases MAGL, FAAH, and ABHD6 were measured by activity-based protein profiling (ABPP). The effect of AKU-005 on ex vivo CSD wave in cortical slices was studied by live calcium imaging.

resultsEx vivo, AKU-005 inhibited MAGL, FAAH, and ABHD6, increasing 2-arachidonoylglycerol (2-AG) and anandamide (AEA) levels in rat cortex under both basal and CSD conditions. In mice, AKU-005 showed a milder effect, inhibiting MAGL only under CSD conditions and increasing 2-AG levels in both basal and CSD states. In vitro analyses confirmed the ex vivo findings for rats and revealed basal MAGL inhibition in mice cortex. AKU-005, previously reported as a double MAGL/FAAH-inhibitor, also inhibited overexpressed mouse and human ABHD6, a little studied 2-AG-hydrolyzing enzyme in brain. In line with these results, AKU-005 reduced CSD events in cortical slices from both rodent species, with higher efficacy in rats.

conclusionsGiven the distinct profile of endocannabinoids hydrolases activities between rats and mice in the brain areas associated with migraine, AKU-005 may target multiple endocannabinoid hydrolases to serve as an efficient treatment option for migraine with aura.