Novel pyrazole cannabinoids: insights into CB(1) receptor recognition and activation.

Synthesis of an antagonist, SR141716A, that selectively binds to brain cannabinoid (CB(1)) receptors without producing cannabimimetic activity in vivo, suggests that recognition and activation of cannabinoid receptors are separable events. In the present study, a series of SR141716A analogs were synthesized and were tested for CB(1) binding affinity and in a battery of in vivo tests, including hypomobility, antinociception, and hypothermia in mice. These analogs retained the central pyrazole structure of SR141716A with replacement of the 1-, 3-, 4-, and/or 5-substituents by alkyl side chains or other substituents known to impart potent agonist activity in traditional tricyclic cannabinoid compounds. Although none of the analogs alone produced the profile of cannabimimetic effects seen with full agonists, several of the 3-substituent analogs with higher binding affinities showed partial agonism for one or more measures. Cannabimimetic activity was most noted when the 3-substituent of SR141716A was replaced with an alkyl amide or ketone group. None of the 3-substituted analogs produced antagonist effects when tested in combination with 3 mg/kg Delta(9)-tetrahydrocannabinol (Delta(9)-THC). In contrast, antagonism of Delta(9)-THC's effects without accompanying agonist or partial agonist effects was observed with substitutions at positions 1, 4, and 5. These results suggest that the structural properties of 1- and 5-substituents are primarily responsible for the antagonist activity of SR141716A.

Highly selective CB(1) cannabinoid receptor ligands and novel CB(1)/VR(1) vanilloid receptor "hybrid" ligands.

Anandamide and the metabolically stabler analogs, (R)-1'-methyl-2'-hydroxy-ethyl-arachidonamide (Met-AEA) and N-(3-methoxy-4-hydroxy-benzyl)-arachidonamide (arvanil), are CB(1) cannabinoid and VR(1) vanilloid receptors agonists. We synthesized 1',1'-dimethylheptyl-arvanil (O-1839) and six other AEA analogs obtained by addition of either a hydroxy, cyano, or bromo group on the C-20 atom of 1,1'-dimethylpentyl-Met-AEA (O-1811, O-1812 and O-1860, respectively) or 1,1'-dimethylpentyl-arvanil (O-1856, O-1895 and O-1861, respectively). The compounds were tested for their (i) affinity for CB(1) and CB(2) receptors, (ii) capability to activate VR1 receptors, (iii) inhibitory effect on the anandamide hydrolysis and on the anandamide membrane transporter, and (iv) cannabimimetic activity in the mouse 'tetrad' of in vivo assays. O-1812 is the first ligand ever proven to be highly (500- to 1000-fold) selective for CB(1) vs both VR(1) and CB(2) receptors, while O-1861 is the first true "hybrid" agonist of CB(1)/VR(1) receptors and a compound with potential therapeutic importance. The activities of the seven compounds in vivo did not correlate with their activities at either CB(1) or VR(1) receptors, thus suggesting the existence of other brain sites of action mediating some of their neurobehavioral actions in mice.

Influence of phenylmethylsulfonyl fluoride on anandamide brain levels and pharmacological effects.

The endogenous cannabinoid anandamide produces cannabimimetic effects similar to those produced by delta9-tetrahydrocannabinol (delta9-THC), but has a much shorter duration of action due to its rapid metabolism to arachidonic acid and polar metabolites via action of fatty acid amide hydrolase (FAAH). Our earlier observations that anandamide's effects persisted after brain levels of anandamide itself had substantially dropped prompted us to examine the influence of the irreversible amidase inhibitor, phenylmethyl sulfonyl fluoride (PMSF), on the brain levels and pharmacological effects of anandamide. As shown previously, pretreatment with PMSF resulted in a leftward shift of the anandamide dose effect curves for antinociception and hypothermia in male mice. Brain and plasma levels of anandamide, arachidonic acid and polar metabolites peaked at 1 min after i.v. injection with 3H-anandamide and remained high at 5 min post-injection, with levels falling sharply thereafter. Pretreatment with PMSF (30 mg/kg, i.p.) prior to an injection of 1 or 10 mg/kg 3H-anandamide resulted 5 min later in enhanced brain levels of anandamide compared to those obtained with 3H-anandamide plus vehicle injection. Levels of arachidonic acid and polar metabolites in brain were not significantly increased. The clear correspondence between brain levels of anandamide following pretreatment with PMSF and pharmacological activity suggests that this parent compound is responsible for the antinociception and hypothermia that occurred 5 min after injection. These results further suggest that metabolite contribution to anandamide's effects, if any, would occur primarily at later times.