Novel selective and metabolically stable inhibitors of anandamide cellular uptake

Novel aromatic analogues of N-oleoylethanolamine and N-arachidonoylethanolamine (anandamide, AEA) were synthesized and, based on the capability of similar compounds to interact with proteins of the endocannabinoid and endovanilloid signaling systems, were tested on: (i) cannabinoid CB1 and CB2 receptors; (ii) vanilloid VR1 receptors; (iii) anandamide cellular uptake (ACU); and (iv) the fatty acid amide hydrolase (FAAH). The (R)- and, particularly, the (S)-1'-(4-hydroxybenzyl) derivatives of N-oleoylethanolamine and AEA (OMDM-1, OMDM-2, OMDM-3, and OMDM-4) inhibited to a varied extent ACU in RBL-2H3 cells (K-i ranging between 2.4 and 17.7 muM), the oleoyl analogues (OMDM-1 and OMDM-2, K-i 2.4 and 3.0 muM, respectively) being 6- to 7-fold more potent than the arachidonoyl analogues (OMDM-3 and OMDM-4). These four compounds exhibited: (i) poor affinity for either CB1 (K(i)greater than or equal tomuM) or CB2 (K-i>10 muM) receptors in rat brain and spleen membranes, respectively; (ii) almost no activity at vanilloid receptors in the intracellular calcium assay carried out with intact cells over-expressing the human VR1 (EC(50)greater than or equal to10 muM); and (iii) no activity as inhibitors of FAAH in N18TG2 cell membranes (K-i>50 muM). The oleoyl- and arachidonoyl-N'-(4-hydroxy-3-methoxybenzyl)hydrazines (OMDM-5 and OMDM-6), inhibited ACU (K-i 4.8 and 7.0 muM, respectively), and were more potent as VR1 agonists (EC50 75 and 50 nM, respectively), weakly active as CB1 receptor ligands (K-i 4.9 and 3.2 muM, respectively), and inactive as CB2 ligands (K-i>5 muM) as well as on FAAH (K(i)greater than or equal to40 muM). In conclusion, we report two novel potent and selective inhibitors of ACU (OMDM-1 and OMDM-2) and one "hybrid" agonist of CB1 and VR1 receptors (OMDM-6). Unlike other compounds of the same type, OMDM-1, OMDM-2, and OMDM-6 were very stable to enzymatic hydrolysis by rat brain homogenates. (C) 2003 Elsevier Science Inc. All rights reserved.