Comparative biochemical and pharmacological characterization of a novel, NOP receptor selective hexapeptide, Ac-RYYRIR-ol.
Nociceptin/orphanin FQ (N/OFQ) is an endogenous neuropeptide, which is widely distributed in central and peripheral nervous system. Some N/OFQ sequence unrelated hexapeptides can effectively bind to the N/OFQ peptide (NOP) receptor and they were used as template for structure-activity studies that lead to discovery of the new NOP selective ligands. In the present study, the pharmacological profile of the novel hexapeptide Ac-RYYRIR-ol was investigated using various in vitro assays including receptor binding and G-protein activation in rat brain membranes, mouse and rat vas deferens, guinea pig ileum, mouse colon and Ca(2+) mobilization assay in chinese hamster ovary (CHO) cells co-expressing the human recombinant NOP receptor and the C-terminally modified Galpha(qi5) protein. In rat brain membranes Ac-RYYRIR-ol displaced both [(3)H]nociceptin/OFQ and [(3)H]Ac-RYYRIK-ol with high affinity (pK(i) 9.35 and 8.81, respectively) and stimulated [(35)S]GTPgammaS binding showing however lower maximal effects than N/OFQ (alpha=0.28). The stimulatory effect of Ac-RYYRIR-ol was antagonized by the selective NOP receptor antagonist UFP-101. In the electrically stimulated mouse vas deferens Ac-RYYRIR-ol displayed negligible agonist activity while antagonizing in a competitive manner (pA(2) 7.99) the inhibitory effects of N/OFQ. Similar results were obtained in the rat vas deferens. In the mouse colon Ac-RYYRIR-ol produced concentration dependent contractile effects with similar potency and maximal effects as N/OFQ. Finally, in the Ca(2+) mobilization assay performed with CHO-hNOP-Galpha(qi5) cells Ac-RYYRIR-ol displayed lower potency and maximal effects (alpha=0.87) compared with N/OFQ. In conclusion, the novel NOP receptor selective hexapeptide Ac-RYYRIR-ol has been shown to have fine selectivity, high potency, furthermore agonist and antagonist effects toward the NOP receptors were measured in various assays; this is likely due to its partial agonist pharmacological activity.
Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, 6726 Szeged, Temesvari krt 62, Hungary.
Engin
E
Bojnik
Fruzsina
F
Babos
Carmela
C
Fischetti
Anna
A
Magyar
Valeria
V
Camarda
Anna
A
Borsodi
Sándor
S
Bajusz
Girolamo
G
Calo'
Sándor
S
Benyhe
D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D003106:Colon / Q000187:drug effects / Q000502:physiology; D006224:Cricetinae; D003412:Cricetulus; D004558:Electric Stimulation; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000235:genetics / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000737:chemistry / Q000378:metabolism; D006168:Guinea Pigs; D006801:Humans; D008297:Male; D051379:Mice; D009119:Muscle Contraction / Q000187:drug effects; D009842:Oligopeptides / Q000378:metabolism* / Q000494:pharmacology*; D018847:Opioid Peptides / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding; D051381:Rats; D017207:Rats, Sprague-Dawley; D017208:Rats, Wistar; D011957:Receptors, Opioid / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000094942:Nociceptin Receptor; D000097629:Nociceptin
D002118:Calcium; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D009842:Oligopeptides; D018847:Opioid Peptides; D011957:Receptors, Opioid; D011994:Recombinant Proteins; D000094942:Nociceptin Receptor; D000097629:Nociceptin; C512381:Ac-RYYRIK-ol; C549028:acetyl-arginyl-tyrosyl-tyrosyl-arginyl-isoleucyl-arginine-ol