In vitro properties of GSK554418, a potent, selective and orally available CB2 cannabinoid receptor agonist

Roy Gray1, Chas Bountra1, Andrew Brown1, Iain Chessell1, Andrew Eatherton1, Gerrard Giblin1, David Hall2, Carl Haslam1, Bill Mitchell1, Alan Naylor1, Celestine O'Shaughnessy1, Sara Pritchard1, Julian Reeves1, Alex Wilson1, Anton Michel1. 1GSK, Harlow, Essex, CM19 5AW, United Kingdom, 2GSK, Stevenage, Hertfordshire, SG1 2NY, United Kingdom.

Cannabinoid receptor agonists exhibit analgesic properties in pre-clinical animal models and in the clinic but their therapeutic potential is limited by their psychotropic actions. Selective CB2 receptor agonists are also effective analgesic agents in pre-clinical pain models without psychotropic effects and represent more developable analgesic agents (Fox & Bevan, 2005). Although several selective CB2 receptor agonists have been studied in pre-clinical pain models, information is limited (Fox & Bevan, 2005). In this study we describe the in vitro properties of a novel CB2 agonist, GSK554418, N-(3-chlorophenyl)-1-methyl-7-(4-morpholinylcarbonyl)-1H-pyrrolo[3,2-c]pyridin-4-amine.

The potency of GSK554418 at human recombinant CB1 and CB2 receptors expressed in HEK293 cells was determined using cAMP-sensitive reporter gene assays (Michel et al., 2005). Effects at CB1 receptors in rat and human cortex membranes were examined using a GTPγS binding assay (Leggett et al., 2004). The effect of GSK554418 at a putative CB2 receptor in human eosinophils was examinee by studying its ability to change cell shape (Sabroe et al., 1999). Data are the mean+S.E.M of 4-6 experiments. The intrinsic activity (IA) was expressed relative to HU210. Equipotent molar ratio (EMR) was calculated as potency GSK554418 / potency HU210 in order to correct for inter-assay potency variation.

In cells expressing human CB2 receptors, GSK554418 produced a rightward shift in the forskolin concentration-effect curve similar to that seen with HU210 (Michel et al., 2005). The pIC50 of GSK554418 to block forskolin (1μM) responses was 7.7+0.01 (IA=98+3%, EMR=5.8). At human recombinant CB1 receptors, GSK554418 had a pIC50 of 5.8+0.03 (EMR>1515; IA 100%) but was less active at native CB1 receptors in rat cortex (IA 46+4% at 10μM) or human cortex (no effect at 10μM). HU210 (pEC50=9.0) and GSK554418 (pEC50=8.8+0.16, EMR=1.7, IA=100%) caused a shape change in human eosinophils. The CB2 antagonist, SR144528, blocked the effects of HU210 (pKB 9.1, n=2).

This study has identified GSK554418 as a potent and selective CB2 receptor agonist at native and recombinant receptors. The compound activated recombinant CB1 receptors at high concentrations but possessed little activity at rat or human native CB1 receptors. In contrast, GSK554418 was potent at causing shape change in human eosinophils, a response which appears to be CB2 mediated. This compound has good oral bioavailability and is active in pre-clinical pain models (Clayton et al., Wilson et al., this meeting).

Fox, A., and Bevan, A. (2005). Expert Opin. Investig. Drugs, 14, 695-703

Leggett, J.D., et al., (2004). Br. J. Pharmacol., 141, 253-262

Michel, A.D. et al., (2005). From Winter 2005 Meeting:Proceedings of the British Pharmacological Society at http://www.pa2online.org/abstracts/Vol3Issue4abst182P.pdf

Sabroe, I., et al., (1999). J. Immunol., 162, 2946-2955.