Desmethionine alkylamide bombesin analogs: a new class of bombesin receptor antagonists with potent antisecretory activity in pancreatic acini and antimitotic activity in Swiss 3T3 cells

Abstract

Bombesin-related peptides have a large number of physiological functions as well as having an autocrine growth mechanism for the regulation of small cell lung cancer cells. In the present study we have synthesized 21 des-Met amide or alkylamide analogues of bombesin and compared their abilities to function as bombesin receptor antagonists in guinea pig pancreatic acini and Swiss 3T3 cells with those of the previously most potent antagonist described, [Leu13 psi(CH2NH)Leu14]bombesin (analogue I). All des-Met analogues functioned as antagonists. Bn(1-13)NH2 was approximately equipotent to I (Ki = 60-80 nM) whereas Bn(6-13)NH2 was 30-fold less potent (Ki = 1800 nM). Formation of an ethylamide, Bn(6-13)ethylamide, increased the potency 30-fold such that this octapeptide was equipotent to I. The addition of a D-Phe6 moiety to I did not change potency but caused a 30-fold increase in potency of Bn(6-13)NH2 and a 8-fold increase in the potency of Bn(6-13)ethylamide (Ki = 16 nM). Additional studies of both NH2- and COOH-terminal alterations in Bn(6-13)NH2 demonstrated that the most potent antagonist was [D-Phe6]Bn(6-13)propylamide (PA), having IC50's of 1.6 nM and 0.8 nM for bombesin-stimulated amylase release and Swiss 3T3 cell growth, respectively. Detailed studies of the most potent amide analogue, [D-Phe6]Bn(6-13)NH2, and alkylamide analogue, [D-Phe6]Bn(6-13)PA, demonstrated that these analogues functioned as competitive antagonists and that their action was selective for the bombesin receptor. These results demonstrate that, as with CCK- and gastrin-related peptides, the C-terminal amino acid is important for initiating a biologic response but not essential for determining receptor affinity.(ABSTRACT TRUNCATED AT 250 WORDS)

Discovery of a High Affinity Radioligand for the Human Orphan Receptor, Bombesin Receptor Subtype 3, Which Demonstrates That It Has a Unique Pharmacology Compared with Other Mammalian Bombesin Receptors

“…At present, the explanation for the differences from the study on [Ca 2ϩ ] i (4) is unclear. The differences are not due to inactivity of the peptide in our study, because it had a high affinity for the GRP-R, as reported previously (13,29). It remains possible that, in contrast to the GRP-R and NMB-R, there could be large receptor spareness in hBRS-3 receptors such that minimal changes in receptor occupation cause marked changes in [Ca 2ϩ ] i , and therefore the biologic activity dose-response curve for agonist-induced changes in [Ca 2ϩ ] i is far to the left of the receptor occupation curve.…”

International Union of Pharmacology. LXVIII. Mammalian Bombesin Receptors: Nomenclature, Distribution, Pharmacology, Signaling, and Functions in Normal and Disease States

“…Structure-function studies of all natural occurring bombesin-related peptides for BB 2 and BB 1 receptors suggested that primarily the presence of His for Leu and to a lesser extent the presence of Leu for Phe were the most important differences in GRP from NMB determining high affinity and selectivity for the BB 1 receptor (Lin et al, 1996). Correlating biological activity with binding affinity, especially of antagonists, demonstrated that the presence of a COOH-terminal amino acid in position 14 of bombesin is not essential for high affinity for the BB 2 receptor, but it is essential for biologic activity Wang et al, 1990aWang et al, , 1992.…”

“…From studies correlating binding results with biological activity, especially for COOH-terminal pseudopeptides, a model was proposed for the biologically active conformation of GRP/Bn at the BB 2 receptor (Coy et al, , 1991bWang et al, 1990a). In a study of the effects on the affinity and potency of bombesin for the BB 2 receptor of substitution of a bond (i.e., CH 2 NH 2 instead of CONH) between each amino acid pair at the COOH terminus, it was found only 13-14 and 9 -10 substitutions resulted in peptides that retained affinity for the BB 2 receptor but did not activate it and thus functioned as antagonists.…”

“…In this model the hydrogen bonding between Leu 13 -Leu 14 CO groups and Ala 9 -Val 10 -CO groups is important, and their destruction by a pseudopeptide bond would lead to a conformational shift and loss of efficacy. Support for this conformation has come from studies of both agonists and antagonists Wang et al, 1990a;Coy et al, 1991a). Only the agonist results will be discussed here with the antagonist result in the next section.…”

“…BB 2 Receptor Antagonist Binding. Numerous structure-function studies of primarily peptide antagonists demonstrated that the COOH-terminal amino acid of GRP or bombesin was not required for high-affinity interaction with the BB 2 receptor; however it was required to activate the receptor Heimbrook et al, 1989;Wang et al, 1990aWang et al, , 1992. A number of results from these studies and molecular modeling studies supported the model proposed by Coy et al (1988) in which the COOH terminus of GRP existed in a folded conformation, stabilized by hydrogen bonding, with the rest of the amino acid chains arranged as an antiparallel ␤-pleated sheet, 1988).…”