Agonist inverse agonist characterization at CB1 and CB2 cannabinoid receptors of L759633, L759656 and AM630

1 We have tested our prediction that AM630 is a CB2 cannabinoid receptor ligand and also investigated whether L759633 and L759656, are CB2 receptor agonists.

2 Binding assays with membranes from CHO cells stably transfected with human CB1 or CB2 receptors using [H-3]-CP55940, confirmed the CB2-selectivity of L759633 and L759656 (CB2/CB1 affinity ratios=163 and 414 respectively) and showed AM630 to have a K-1 at CB2 receptors of 31.2 nM and a CB2/CB1 affinity ratio of 165.

3 In CB2-transfected cells, L759633 and L759656 were potent inhibitors of forskolin-stimulated cyclic AMP production, with EC50 values of 8.1 and 3.1 nM respectively and CB1/CB2 EC50 ratios of >1000 and >3000 respectively.

4 AM630 inhibited [S-35]-GTP gamma S binding to CB2 receptor membranes (EC50 = 76.6 nM), enhanced forskolin-stimulated cyclic AMP production in CB2-transfected cells (5.2 fold by 1 mu M), and antagonized the inhibition of forskolin-stimulated cyclic AMP production in this cell line induced by CP55940.

5 In CB1-transfected cells, forskolin-stimulated cyclic AMP production was significantly inhibited by AM630 (22.6% at 1 mu M and 45.9% at 10 mu M) and by L759633 at 10 mu M (48%) but not 1 mu M. L759656 (10 mu M) was not inhibitory. AM630 also produced a slight decrease in the mean inhibitory effect of CP55940 on cyclic AMP production which was not statistically significant.

6 We conclude that AM630 is a CB2-selective ligand that behaves as an inverse agonist at CB2 receptors and as a weak partial agonist at CB1 receptors. L759633 and L759656 are both potent CB2-selective agonists.