In vitro and in vivo pharmacological characterization of J-113397, a potent and selective non-peptidyl ORL1 receptor antagonist

1-[(3 R,4 R)-1-cyclooctylmethyl-3-hydroxymethyl-4-piperidyl]-3-ethyl-1,3-dihydro-2 H-benzimidazol-2-one (J-113397) was found to be the first potent nonpeptidyl ORL1 receptor antagonist ( K i: cloned human ORL1=1.8 nM) with high selectivity over other opioid receptors ( K i: 1000 nM for human μ-opioid receptor, >10,000 nM for human δ-opioid receptor, and 640 nM for human κ-opioid receptor). In vitro, J-113397 inhibited nociceptin/orphanin FQ-stimulated [ 35S]guanosine 5′- O-(γ-thio)triphosphate (GTPγS) binding to Chinese Hamster Ovary (CHO) cells expressing ORL1 (CHO-ORL1) with an IC 50 value of 5.3 nM but had no effect on [ 35S]GTPγS binding by itself. Schild plot analysis of the [ 35S]GTPγS binding assay and cAMP assay using CHO-ORL1 indicated competitive antagonism of J-113397 on the ORL1 receptor. In CHO cells expressing μ-, δ- or κ-opioid receptors, J-113397 had no effects on [ 35S]GTPγS binding up to a concentration of 100 nM, indicating selective antagonism of the compoud on the ORL1 receptor. In vivo, J-113397, when administered subcutaneously (s.c.), dose-dependently inhibited hyperalgesia elicited by intracerebroventricular (i.c.v.) administration of nociceptin/orphanin FQ in a tail-flick test with mice. An in vitro binding study using mouse brains indicated that J-113397 possesses high affinity for the mouse ORL1 receptor ( K i: 1.1 nM) as well as the human receptor. In summary, J-113397 is the first potent, selective ORL1 receptor antagonist that may be useful in elucidating the physiological roles of nociceptin/orphanin FQ.