Structure-activity studies on nociceptin analogues: ORL1 receptor binding and biological activity of cyclic disulfide-containing analogues of nociceptin peptides

Nociceptin/Orphanin FQ is an endogenous peptide ligand for the opioid recep tor-like 1 (ORL1) receptor. To investigate the structural and conformationa l requirements of the nociceptin (NC)receptor interaction, six cyclic analo gues containing Cys disulfide linkages were designed and synthesized. Analo gues cyclized at the N-terminal part, cyclo[Cys(0), Cys(7)]NC(1-13)-NH2 (2) and cyclo[Cys(0), Cys(11)]NC(1-13)-NH2 (4), and their corresponding linear peptides had very low activities in both the receptor binding and the GTP gammaS functional assays using human ORL1 transfected cell membranes. On th e contrary, analogues cyclized at the C-terminal parts by the disulfide lin kages at positions 6-10, 7-11, 7-14, and 10-14 sustained relatively high po tencies in both assays. Notably, cyclo[Cys(10), Cys(14)]NC(1-14)-NH2 (12) w as found to be a potent NC agonist nearly as active as the parent peptide o r NC. The maximum efficacy (Emax) of the C-terminally cyclized analogues an d their linear counterparts in the GTP gammaS functional assay showed more than 94% (vs NC as 100%), suggesting that these analogues are full agonists . Analogue 12 is the first conformationally constrained NC analogue with al most full activity, and thus may serve to analyze the bioactive conformatio ns of NC at the receptor site as well as serving as a template for more pot ent NC agonists.