Pharmacological profiles of selective non-peptidic delta opioid receptor ligands

Several non-peptidic opioids have been synthesized recently as part of a pr ogram to develop selective delta receptor agonists. In this study, the affi nities of a set of compounds for cloned delta and mu opioid receptors expre ssed in HEK 293 cell lines were determined by competition analysis of [H-3] bremazocine binding to membrane preparations. All compounds studied exhibit ed high affinity and selectivity, with apparent dissociation constants in t he range of 0.6-1.7 nM for the delta opioid receptor and 140-1165 nhl for t he mu opioid receptor. We next sought to determine which domain of the delt a receptor was critical for mediating the highly selective binding by analy sis of ligand affinities for mu/delta receptor chimeras. Receptor binding p rofiles suggested that a critical site of receptor/ligand interaction was l ocated between transmembrane domain 5 (TM5) and TM7 of the delta receptor. Substitution of tryptophan 284, located at the extracellular surface of TM6 , with lysine, which is: found at the equivalent position in the CL opioid receptor, led to a spectrum, of effects on affinities, depending on the lig and tested. Affinities of SE 219825 and SE 222941 were particularly sensiti ve to the substitution, displaying a 50-fold and 70-fold decrease in affini ty, respectively. Activities of the delta receptor-selective agonists were tested in two functional assays. Brief exposure of HEK 293 cells expressing delta opioid receptors with selective ligands induced phosphorylation of M AP kinase, although the non-peptidic ligands were less efficacious than the enkephalin derivative DADL (Tyr-D-Ala-Gly-Phe-D-Leu). Similarly, chronic e xposure of HEK; 293 cells expressing delta opioid receptors with selective, non-peptidic ligands, with the exception of SE 206848, caused receptor dow nregulation, however, the SE compounds were less efficacious than DADL. (C) 2000 Elsevier Science B.V. All rights reserved.