title: Interaction of p-fluorofentanyl on cloned human opioid receptors and exploration of the role of Trp-318 and His-319 in mu-opioid receptor selectivity.
abstract: In this study, we investigated the interactions of p-fluorofentanyl, an opioid designer drug, fentanyl, sufentanyl, and morphine on cloned human mu-, kappa-, and delta-opioid receptors coexpressed with heteromultimeric G protein-coupled inwardly rectifying K(+) channels (GIRK1/GIRK2) and a regulator of G protein signaling (RGS4) in Xenopus oocytes. We demonstrate that p-fluorofentanyl more potently activates GIRK1/GIRK2 channels through opioid receptors than fentanyl and that the p-fluoro substitution also changes the potency profile from mu > kappa > delta (fentanyl) to mu > delta > or = kappa (p-fluorofentanyl). A comparison of ligand efficacy revealed that morphine, fentanyl, and its analogs less efficiently activate GIRK1/GIRK2 channels through human mu-opioid receptor than [D-Ala(2),N-Me-Phe(4),Gly(5)-ol]-enkephalin. Using site-directed mutagenesis, we investigated whether mutating residues Trp-318 and His-319 to their corresponding residues in kappa- and delta-opioid receptors provides the molecular basis for mu/delta selectivity and mu/kappa selectivity. Changes in EC(50) values for the W318L and W318Y/H319Y mu-opioid receptors show a partial contribution of these residues to the decreased GIRK1/GIRK2 channel activation by fentanyl analogs through kappa- and delta-opioid receptors. The most pronounced effect was observed for p-fluorofentanyl, suggesting that an interaction between the 4-fluorophenylpropanamide moiety of the drug and residues Trp-318 and His-319 is important for the resulting enhanced GIRK1/GIRK2 channel activation through the mu-opioid receptor. Finally, we demonstrate that mutation of W318L confers delta-like potency for morphine on the mutant mu-opioid receptor.
authors.affiliation: Laboratory of Toxicology, Faculty of Pharmaceutical Sciences, University of Leuven, Belgium.
authors.forename: C
authors.initials: C
authors.lastname: Ulens
authors.forename: M
authors.initials: M
authors.lastname: Van Boven
authors.forename: P
authors.initials: P
authors.lastname: Daenens
authors.forename: J
authors.initials: J
authors.lastname: Tytgat
mesh_terms: D000818:Animals; D003001:Cloning, Molecular; D015198:Designer Drugs; D005283:Fentanyl / Q000494:pharmacology; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D006639:Histidine / Q000235:genetics*; D006801:Humans; D008024:Ligands; D009020:Morphine / Q000494:pharmacology; D009154:Mutation; D009294:Narcotics / Q000494:pharmacology; D009865:Oocytes; D018408:Patch-Clamp Techniques; D010880:Piperidines / Q000494:pharmacology*; D015221:Potassium Channels / Q000819:agonists; D024661:Potassium Channels, Inwardly Rectifying; D017465:Receptors, Opioid, delta / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D017473:Receptors, Opioid, kappa / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D017450:Receptors, Opioid, mu / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D017409:Sufentanil / Q000494:pharmacology; D014364:Tryptophan / Q000235:genetics*; D014982:Xenopus laevis
chemical_list: D015198:Designer Drugs; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D008024:Ligands; D009294:Narcotics; D010880:Piperidines; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D017465:Receptors, Opioid, delta; D017473:Receptors, Opioid, kappa; D017450:Receptors, Opioid, mu; D006639:Histidine; D009020:Morphine; D014364:Tryptophan; C412892:4-fluorofentanyl; D017409:Sufentanil; D005283:Fentanyl