Opiate aromatic pharmacophore structure-activity relationships in CTAP analogues determined by topographical bias, two-dimensional NMR, and biological activity assays

Topographically constrained analogues of the highly mu-opioid-receptor-sele ctive antagonist CTAP (H-D-Phe-c[Cys-Tyr-D-Trp-Arg-Thr-Pen]-Thr-NH2, 1) wer e prepared by solid-phase peptide synthesis. Replacement of the D-Phe resid ue with conformationally biased beta-methyl derivatives of phenylalanine or tryptophan (2R,3R; 2R,3S; 2S,3R; 2S,3S) yielded peptides that displayed wi dely varying types of biological activities. In an effort to correlate the observed biological activities of these analogues with their structures, tw o-dimensional H-1 NMR and molecular modeling was performed. Unlike the pare nt (1), which is essentially a pure mu antagonist with weak delta agonist a ctivities in the MVD bioassay, the diastereomeric beta-MePhe(1)-containing peptides exhibited simultaneous delta agonism and mu antagonism by the (2R, 3R)-containing isomer 2; mu antagonism by the (2R,3S)-containing isomer 3; weak mu agonism by the (2S,3R)-containing isomer 4; and delta agonism by th e (2S,3S)-containing isomer 5. Incorporation of beta-MeTrp isomers into pos ition 1 led to peptides that were mu antagonists (2R,3R), 8; (2R,3S), 9, or essentially inactive (<10%) in the MVD and GPI assays (2S,3R), 10; (2S,3S) , 11. Interestingly, in vivo antinociceptive activity was predicted by neit her MVD nor GPI bioactivity. When D-Trp was incorporated in position 1, the result (7) is a partial, yet relatively potent mu agonist which also displ ayed weak delta agonist activity. Molecular modeling based on 2D NMR reveal ed that low energy conformers of peptides with similar biological activitie s had similar aromatic pharmacophore orientations and interaromatic distanc es. Peptides that exhibit mu antagonism have interaromatic distances of 7.0 -7.9 Angstrom and have their amino terminal aromatic moiety pointing in a d irection opposite to the direction that the amino terminus points. Peptides with delta opioid activity displayed an interaromatic distance of <7 Angst rom and had their amino terminal aromatic moiety pointing in the same direc tion as the amino terminus.