Pharmacology and selectivity of various natural and synthetic bombesin related peptide agonists for human and rat bombesin receptors differs

Abstract

The mammalian bombesin (Bn)-receptor family[gastrin-releasing peptide-receptor(GRPR-receptor), neuromedin B-receptor(NMB-receptor)], their natural ligands,GRP/NMB, as well as the related orphan-receptor,BRS-3, are widely-distributed, and frequently overexpressed by tumors. There is increased interest in agonists for this receptor family to explore their roles in physiological/pathophysiological processes, and for receptor-imaging/cytotoxicity in tumors. However, there is minimal data on human pharmacology of Bn-receptor agonists and most results are based on nonhuman receptor studies, particular rodent-receptors, which with other receptors frequently differ from human-receptors. To address this issue we compared hNMB/GRP-receptor affinities and potencies/efficacies of cell-activation(assessing phospholipase C activity) for 24 putative Bn-agonists(12-natural,12-synthetic) in four different cells with these receptors, containing native-receptors or receptors expressed at physiological densities, and compared the results to native rat-GRP-receptor-containing cells-(AR42J–cells) or rat-NMB-receptor cells(C6-glioblastoma cells). There were close correlations(r=0.92–99,p<0.0001) between their affinities/potencies for the two hGRP- or hNMB-receptor cells. Twelve analogues had high affinities(≤ 1 nM) for hGRP-receptor with 15 selective for it(greatest=GRP,NMC), 8 had high affinity/potencies for hNMB-receptors and 4 were selective for it. Only synthetic Bn-analogues containing β−alanine11 had high affinity for hBRS-3, but t also had high affinities/potencies for all GRP-/hNMB-receptor cells. There was no correlation between affinities for human-GRP-receptors and rat-GRP-receptors(r=0.131,p=0.54), but hNMB-receptor results correlated with rat-NMB-receptor(r=0.71, p<0.0001). These results elucidate the human- and rat-GRP-receptor pharmacophore for agonists differ markedly,whereas they do not for NMB-receptors, therefore potential GRP-receptor agonists for human studies(such as Bn-receptor-imaging/cytotoxicity) must be assessed on human-Bn-receptors. The current study provides affinities/potencies on a large number of potential agonists that might be useful for human studies.

Comparative Pharmacology of Bombesin Receptor Subtype-3, Nonpeptide Agonist MK-5046, a Universal Peptide Agonist, and Peptide Antagonist Bantag-1 for Human Bombesin Receptors

“…Balb 3T3 cells stably expressing human GRP receptor (hGRP-R), human NMB receptor (hNMB-R), and human BRS-3-receptor (hBRS-3) were prepared and used as described previously elsewhere (Benya et al, 1995;Mantey et al, 1997;Ryan et al, 1998a). In addition, NCI-H1299 were transfected to overexpress human NMB receptor (Gonzalez et al, 2009;Uehara et al, 2011). Balb 3T3 cells hGRP-R, hNMB-R, and hBRS-3 were grown in DMEM with 300 mg/l G418 sulfate.…”

“…Balb 3T3 cells hGRP-R, hNMB-R, and hBRS-3 were grown in DMEM with 300 mg/l G418 sulfate. The HuTu-80 cells, which contain native hGRP-receptor (Gonzalez et al, 2009;Uehara et al, 2011), were grown in DMEM. The hNMB-R transfected NCI-H1299 cells were grown in RPMI 1640 medium with 300 mg/l G418 sulfate and NCI-N417, which contains native human BRS-3-receptors (Ryan et al, 1998b;Sancho et al, 2010) in RPMI 1640 medium.…”

“…The total [ 3 H]IP was isolated by anion exchange chromatography as described previously elsewhere (Rowley et al, 1990;Benya et al, 1995;Ryan et al, 1998a;Uehara et al, 2011 Western Blot Analysis. Western blot analysis of the various cells was performed as previously described elsewhere (Berna et al, 2007;Sancho et al, 2010).…”

“…Until recently, the only known high-affinity ligand was the synthetic Bn-related agonist peptide #1 ,bAla 11 ,Phe 13 , Nle 14 ]Bn-(6-14)]), but peptide #1 also has high affinity for GRP-R and NBM-R from all species as well as human bombesin receptor subtype-3 (hBRS-3) (Mantey et al, 1997;Pradhan et al, 1998;Sancho et al, 2010;Uehara et al, 2011). Recent studies have reported that the nonpeptide MK-5046 [(2S)-1,1,1-trifluoro-2-[4-(1H-pyrazol-1-yl)phenyl]-3-(4-{[1-(trifluoromethyl)cyclopropyl]methyl}-1H-imidazol-2-yl)propan-2-ol] is an orally active, potent, and selective BRS-3 agonist (Sebhat et al, 2011;Guan et al, 2011;Reitman et al, 2012), with activity in mice, rats, dogs , and humans (Reitman et al, 2012).…”

“…There are limited data on the pharmacologic characteristics of these compounds (Guan et al, 2010Feng et al, 2011;Sebhat et al, 2011). In addition, for the Bn-receptor family, similar to other gastrointestinal hormone/neurotransmitter G protein-coupled receptors, nonpeptide-agonists are uncommon (Freidinger, 1993;Jensen et al, 2008;Uehara et al, 2011); in fact, MK-5046 is the first high-affinity, widely used nonpeptide agonist for any member of the Bn family of receptors. Therefore, for most of these receptors there are limited data on the comparison of the pharmacological properties of receptor interaction/ activation by nonpeptide agonists with peptide-receptor agonists (Freidinger, 1993).…”