Nociceptin/orphanin FQ (N/OFQ) is the endogenous 17 amino acid peptide ligand for the Gi-protein-coupled N/OFQ receptor (NOP). In an attempt to improve the metabolic stability of N/OFQ, we have produced a truncated cyclic analogue with cysteine residues at positions 7 and 10, c[Cys7,10]N/OFQ(1-13)NH2 (c[Cys7,10]). c[Cys7,10], the template N/OFQ(1-13)NH2 and N/OFQ displaced the binding of [3H]N/OFQ to Chinese hamster ovary cells expressing recombinant human NOP (CHOhNOP) with pK i values of 9.98, 9.83 and 9.18, respectively. In addition, c[Cys7,10], N/OFQ(1-13)NH2 and N/OFQ stimulated the binding of guanosine triphosphate gamma [35S] to CHOhNOP cells with pEC50/E max (stimulation factor) of 9.16/5.5, 9.11/4.9 and 8.35/5.5, respectively. c[Cys7,10], N/OFQ(1-13)NH2 and N/OFQ inhibited forskolin-stimulated cyclic adenosine monophosphate (cAMP) formation with pEC50 values of 10.08, 10.11 and 9.78, respectively. All ligands produced complete inhibition of cAMP formation. In both functional assays, c[Cys7,10] was a full agonist. In a series of metabolism experiments, incubation of 1 nM c[Cys7,10], N/OFQ(1-13)NH2 and N/OFQ with a rat brain homogenate produced a time-dependent loss of peptide that was greatest for the native peptide N/OFQ. Amidation in N/OFQ(1-13)NH2 produced some metabolic protection, but this was not significantly improved by further inclusion of c[Cys7,10]. In summary, c[Cys7,10] is a high-affinity, high-potency full agonist of the NOP receptor. However, we were unable to demonstrate clear metabolic protection.
In vitro pharmacological characterisation of a novel cyclic nociceptin/orphanin FQ analogue c[Cys(7,10)]N/OFQ(1-13)NH2
CALO', Girolamo;GUERRINI, Remo;
2007
Abstract
Nociceptin/orphanin FQ (N/OFQ) is the endogenous 17 amino acid peptide ligand for the Gi-protein-coupled N/OFQ receptor (NOP). In an attempt to improve the metabolic stability of N/OFQ, we have produced a truncated cyclic analogue with cysteine residues at positions 7 and 10, c[Cys7,10]N/OFQ(1-13)NH2 (c[Cys7,10]). c[Cys7,10], the template N/OFQ(1-13)NH2 and N/OFQ displaced the binding of [3H]N/OFQ to Chinese hamster ovary cells expressing recombinant human NOP (CHOhNOP) with pK i values of 9.98, 9.83 and 9.18, respectively. In addition, c[Cys7,10], N/OFQ(1-13)NH2 and N/OFQ stimulated the binding of guanosine triphosphate gamma [35S] to CHOhNOP cells with pEC50/E max (stimulation factor) of 9.16/5.5, 9.11/4.9 and 8.35/5.5, respectively. c[Cys7,10], N/OFQ(1-13)NH2 and N/OFQ inhibited forskolin-stimulated cyclic adenosine monophosphate (cAMP) formation with pEC50 values of 10.08, 10.11 and 9.78, respectively. All ligands produced complete inhibition of cAMP formation. In both functional assays, c[Cys7,10] was a full agonist. In a series of metabolism experiments, incubation of 1 nM c[Cys7,10], N/OFQ(1-13)NH2 and N/OFQ with a rat brain homogenate produced a time-dependent loss of peptide that was greatest for the native peptide N/OFQ. Amidation in N/OFQ(1-13)NH2 produced some metabolic protection, but this was not significantly improved by further inclusion of c[Cys7,10]. In summary, c[Cys7,10] is a high-affinity, high-potency full agonist of the NOP receptor. However, we were unable to demonstrate clear metabolic protection.I documenti in SFERA sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
