According to the implication of

According to the implication of the endocannabinoid system in IBD, we decided in our laboratory to develop selective CB agonists and FAAH inhibitors to treat these diseases. Recently, we described 3-carboxamido-5-aryl-isoxazoles as selective CB agonists. This series of isoxazoles possesses a 2-substituted phenyl group at position 5 and a bulky aliphatic group on the carboxamide function. Our best selective CB agonist is orexins (ALIAE809, ) with a of 9.0nM on CB receptors. Then, we decided to further explore our 3-carboxamido-5-aryl-isoxazoles selective CB agonists for their capacity to inhibit FAAH. Indeed, because CB agonists and FAAH inhibitors are very potent modulators of the endocannabinoid system, it would be very interesting to combine both activities in one molecule in order to obtain a synergistic activity of the compound and enhance its therapeutic properties. Thus, 18 of our previously described isoxazoles were tested for their FAAH inhibition potential. A comparison of the CB affinity and FAAH inhibition capacity of these compounds was carried out and structure–activity relationships were established. As previously described, the 3-carboxamido-5-aryl-isoxazoles – were obtained in four steps from various ketones. Their synthesis consists in a Claisen condensation of the corresponding ketones, followed by a cyclization into ethyl isoxazole-3-carboxylates. The target compounds – were finally obtained by saponification of the ethyl ester function followed by amidation. The 3-carboxamido-5-aryl-isoxazoles – were evaluated for their ability to inhibit the hydrolysis of [H]-AEA by a recombinant human FAAH preparation (expressed in )., , All compounds were first screened at a concentration of 10μM for their inhibitory activity toward the AEA-degradation enzyme. The half maximal inhibitory concentrations (IC) were determined for compounds exhibiting a specific inhibition superior to 50% for FAAH. In , all biological results towards CB and FAAH were summarized. As previously described, the best selective CB agonists among these 3-carboxamido-5-aryl-isoxazoles are compounds bearing an -alkyl chain at the position of the phenyl group at position 5 (R) and a bulky aliphatic group on the carboxamide function at position 3 (R) (). The optimal length for the -alkyl chain is 5–6 carbons. As shown in and 7 isoxazoles possess an IC on FAAH less than 10μM. Among these 7molecules, compounds and , which bear an adamantyl group on the carboxamide function and an -pentyl chain at the and position on the phenyl group at position 5 respectively, showed the best FAAH inhibitory activities with values of 63nM and 8nM. On the other hand, compound with the -pentyl chain at the position induced no FAAH inhibition. In contrast with the results obtained with the in vitro CB affinity assay, an -pentyl chain at the and position but not at the position of the phenyl group is favorable for FAAH inhibitory activity. Furthermore, compounds and , which possessed no affinity for the CB receptor, present a weak FAAH inhibitory activity. We observed that compound , with a 1-adamantyl group on the carboxamide function and a non-substituted phenyl group at position 5, present a 10-fold better FAAH inhibitory activity compared to compound with a 2-adamantyl group on the carboxamide function. When the 1-adamantyl group is replaced by a 1-adamantylmethyl group (compound ), the FAAH inhibitory activity is lost. Interestingly, 3 of our selective CB ligands (, and ) inhibit FAAH in moderate concentrations (: IC=0.912μM; : IC=5.43μM; : IC=2.33μM). Compound , with an adamantyl group on the carboxamide function at position 5 and an -methyl moiety on the phenyl group at the position, possessed a FAAH inhibitory activity with an IC value of 0.912μM and a good CB affinity with a value of 70.1±5.4 nM. Herein, we can consider compound as a dual drug in spite of a moderate FAAH activity.