Characterization of the fatty acid amide hydrolase inhibitor cyclohexyl carbamic acid 3'- carbamoyl-biphenyl-3-yl ester (URB597): effects on anandamide and oleoylethanolamide

Fatty acid amide hydrolase (FAAH) is an intracellular serine enzyme that catalyzes the hydrolysis of bioactive fatty acid ethanolamides such as anandamide and oleoylethanolamide (OEA). Genetic deletion of the faah gene in mice elevates brain anandamide levels and amplifies the effects of this endogenous cannabinoid agonist. Here, we show that systemic administration of the selective FAAH inhibitor URB597 (cyclohexyl carbamic acid 3 -carbamoyl-biphenyl-3-yl ester; 0.3 mg/kg i.p.) increases anandamide levels in the brain of rats and wild-type mice but has no such effect in FAAH-null mutants. Moreover, URB597 enhances the hypothermic actions of anandamide (5 mg/kg i.p.) in wild-type mice but not in FAAH-null mice. In contrast, the FAAH inhibitor does not affect anandamide or OEA levels in the rat duodenum at doses that completely inhibit FAAH activity. In addition, URB597 does not alter the hypophagic response elicited by OEA (5 and 10 mg/kg i.p.), which is mediated by activation of peroxisome proliferator-activated receptor type. Finally, exogenously administered OEA (5 mg/kg i.p.) was eliminated at comparable rates in wild-type and FAAH / mice. Our results indicate that URB597 increases brain anandamide levels and magnifies anandamide responses by inhibiting intracellular FAAH activity. The results also suggest that an enzyme distinct from FAAH catalyzes OEA hydrolysis in the duodenum, where this lipid substance acts as a local satiety factor. The endogenous cannabinoid anandamide (Devane et al., 1992; Di Marzo et al., 1994), the satiety factor oleoylethanolamide (OEA) (Rodrı́guez de Fonseca et al., 2001; Fu et al., 2003), and the analgesic and anti-inflammatory factor palmitoylethanolamide (PEA) (Kuehl et al., 1957; Calignano et al., 1998; Jaggar et al., 1998) are all members of the fatty acid ethanolamide (FAE) family of lipid mediators. FAEs are found in most mammalian tissues, where they are thought to be stored as the phospholipid precursor N-acylphosphatidylethanolamine (Schmid et al., 1996; Piomelli, 2003) and to be produced in a stimulus-dependent manner by activation of an N-acylphosphatidylethanolamine-specific phospholipase D (Okamoto et al., 2004). After release from cells, polyunsaturated FAEs such as anandamide may be eliminated via a two-step process consisting of high-affinity transport into cells (Di Marzo et al., 1994; Beltramo et al., 1997; Hillard et al., 1997; Fegley et al., 2004; Ortega-Gutierrez et al., 2004) followed by intracellular degradation, catalyzed by fatty acid amide hydrolase (FAAH) (Cravatt and Lichtman, 2002). On the other hand, monounsaturated and saturated FAEs such as OEA and PEA are poor substrates for anandamide transport, and their inactivation may be primarily mediated by intracellular hydrolysis catalyzed by FAAH and/or by a distinct but as-yet-uncharacterized FAE amidase that operates at acid pH values (Ueda et al., 1999, 2001). Mutant mice lacking the gene encoding for FAAH (faah) have reduced FAE hydrolysis and elevated brain levels of This work was supported by National Institutes of Health Grants DA12413, DA12447, and DA12653 and by a University of California Discovery grant. Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. doi:10.1124/jpet.104.078980. ABBREVIATIONS: OEA, oleoylethanolamide; PEA, palmitoylethanolamide; FAE, fatty acid ethanolamide; FAAH, fatty acid amide hydrolase; URB597, cyclohexyl carbamic acid 3 -carbamoyl-biphenyl-3-yl ester; PPAR, peroxisome proliferator-activated receptor type; HPLC/MS, high-performance liquid chromatography/mass spectrometry; 2-AG, 2-arachidoloylglycerol. 0022-3565/05/3131-352–358$20.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 313, No. 1 Copyright © 2005 by The American Society for Pharmacology and Experimental Therapeutics 78980/1194978 JPET 313:352–358, 2005 Printed in U.S.A.