Development of a potent inhibitor of 2-arachidonoylglycerol hydrolysis with antinociceptive activity in vivo.

Although inhibitors of the enzymatic hydrolysis of the endocannabinoid 2-arachidonoylglycerol are available, they are either rather weak in vitro (IC50>30 µM) or their selectivity towards other proteins of the endocannabinoid system has not been tested. Here we describe the synthesis and activity in vitro and in vivo of a tetrahydrolipstatin analogue, OMDM169, as a potent inhibitor of 2-AG hydrolysis, capable of enhancing 2-AG levels and of exerting analgesic activity via indirect activation of cannabinoid receptors. OMDM169 exhibited 0.13 µM10 µM) at human CB1 and CB2 receptors. However, OMDM169 shared with tetrahydrolipstatin the capability of inhibiting the human pancreatic lipase (IC50=0.6 µM). OMDM169 inhibited fatty acid amide hydrolase and diacylglycerol lipase only at higher concentrations (IC50=3.0 and 2.8 µM, respectively), and, accordingly, it increased by ~1.6-fold the levels of 2-AG, but not anandamide, in intact ionomycin-stimulated N18TG2 neuroblastoma cells. Acute intraperitoneal (i.p.) administration of OMDM169 to mice inhibited the second phase of the formalininduced nocifensive response with an IC50 of ~2.5 mg/kg, and concomitantly elevated 2-AG, but not anandamide, levels in the ipsilateral paw of formalin-treated mice. The antinociceptive effect of OMDM169 was antagonized by antagonists of CB1 and CB2 receptors, AM251 and AM630, respectively (1 mg/kg, i.p.). OMDM69 might represent a template for the development of selective and even more potent inhibitors of 2- AG hydrolysis.