Exploration of the DTrp-NMeLys Motif in the Search for potent somatostatin antagonists Original Research Article

Previous studies from this laboratory demonstrated that N-methylation at Lys5 residue in somatostatin octapeptide antagonist analogues increased the GH release inhibition potency by as much as 300%. We have now further investigated N-methylation of this Lys5 residue in conjunction with a number of N- and C-terminal modifications previously found to give highly potent somatostatin receptor antagonists. Synthetic analogues were tested in a functional assay for their ability to inhibit somatostatin-inhibited GH release from rat pituitary cells in culture and to displace 125I-labeled somatostatin from CHO cells transfected with the five known human somatostatin receptors. Several interesting observations resulted from the study. Replacement of liphophilic Nal8 at the C-terminus with a hydrophilic His8 resulted in the increased affinity and selectivity for type 2 receptor to give the most potent antagonist analogue yet discovered (Ki, 1.5 nM), although in the rat pituitary cells inhibitory activity on somatostatin inhibited GH release decreased somewhat. A His3 substitution within the cyclic portion of the analogues retained pituitary cell potency and affinity for type 2 receptor as did substitution with Bip8 and Fpa1. Replacement of Cpa1 with Iph1 did not effect the affinity for type 2 receptor significantly, but did decrease the effects on rat cell GH release. Iph3 within-ring substitution increased the selectivity for sst2 appreciably although the affinity for that receptor was considerably decreased. Substitution of Npa3 resulted in good selectivity for sst2 receptor. Replacement of Nal8 with D-Trp8 also increased the selectivity for type 2 receptor. Use of a ‘bivalent ligand’ approach in which two peptides were joined by 4,4′-biphenyldicarbonyl as a spacer destroyed the affinity for all the subtypes, however, the bivalent ligand formed with the Ahp spacer displayed significant affinity and high selectivity for the type 2 receptor.