title: Comparison of [Dmt1]DALDA and DAMGO in binding and G protein activation at mu, delta, and kappa opioid receptors.
abstract: [Dmt1]DALDA (H-Dmt-d-Arg-Phe-Lys-NH2; Dmt = 2',6'-dimethyltyrosine) binds with high affinity and selectivity to the mu opioid receptor and is a surprisingly potent and long-acting analgesic, especially after intrathecal administration. In an attempt to better understand the unique pharmacological profile of [Dmt1]DALDA, we have prepared [3H][Dmt1]DALDA and compared its binding properties with that of [3H]DAMGO ([d-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin). Kinetic studies revealed rapid association of [3H][Dmt1]DALDA when incubated with mouse brain membranes (K+1 = 0.155 nM(-1) min(-1)). Dissociation of [3H][Dmt1]DALDA was also rapid (K(-1) = 0.032 min(-1)) and indicated binding to a single site. [3H][Dmt1]DALDA binds with very high affinity to human mu opioid receptor (hMOR) (Kd = 0.199 nM), and Kd and Bmax were reduced by sodium but not Gpp(NH)p [guanosine 5'-(beta,gamma-imido)triphosphate]. Similar Kd values were obtained in brain and spinal cord tissues and SH-SY5Y cells. The hMOR:hDOR (human delta opioid receptor) selectivity of [Dmt1]DALDA ( approximately 10,000) is 8-fold higher than DAMGO. However, [Dmt1]DALDA is less selective than DAMGO against hKOR (human kappa opioid receptor) (26-versus 180-fold). The Ki values for a number of opioid ligands were generally higher when determined by competitive displacement binding against [3H][Dmt1]DALDA compared with [3H]DAMGO, with the exception of Dmt1-substituted peptide analogs. All Dmt1 analogs showed much higher affinity for the mu receptor than corresponding Tyr1 analogs. [35S]GTPgammaS (guanosine 5'-O -(3-[35S]thio)triphosphate) binding showed that [Dmt1]DALDA and DAMGO are full agonists at hMOR and hDOR but are only partial agonists at hKOR. The very high affinity and selectivity of [3H][Dmt1]DALDA for the mu receptor, together with its very low nonspecific binding (10-15%) and metabolic stability, make [3H][Dmt1]DALDA an ideal radioligand for labeling mu receptors.
authors.affiliation: Department of Pharmacology, Weill Medical College of Cornell University, 1300 York Ave., New York, NY 10021, USA.
authors.forename: Guo-Min
authors.initials: GM
authors.lastname: Zhao
authors.forename: Xuanxuan
authors.initials: X
authors.lastname: Qian
authors.forename: Peter W
authors.initials: PW
authors.lastname: Schiller
authors.forename: Hazel H
authors.initials: HH
authors.lastname: Szeto
mesh_terms: D000701:Analgesics, Opioid / Q000378:metabolism*; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D001711:Biotransformation / Q000502:physiology; D001921:Brain / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003001:Cloning, Molecular; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)- / Q000378:metabolism* / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008297:Male; D051379:Mice; D009842:Oligopeptides / Q000494:pharmacology*; D011869:Radioligand Assay; D051381:Rats; D017207:Rats, Sprague-Dawley; D017465:Receptors, Opioid, delta / Q000378:metabolism*; D017473:Receptors, Opioid, kappa / Q000378:metabolism*; D017450:Receptors, Opioid, mu / Q000378:metabolism*; D012964:Sodium / Q000494:pharmacology; D013116:Spinal Cord / Q000378:metabolism
chemical_list: C424424:2',6'-dimethyltyrosyl-arginyl-phenylalanyl-lysinamide; D000701:Analgesics, Opioid; D009842:Oligopeptides; D017465:Receptors, Opioid, delta; D017473:Receptors, Opioid, kappa; D017450:Receptors, Opioid, mu; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D006165:Guanylyl Imidodiphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D012964:Sodium; D019204:GTP-Binding Proteins