IDENTIFICATION OF A UNIQUE LIGAND WHICH HAS HIGH-AFFINITY FOR ALL 4 BOMBESIN RECEPTOR SUBTYPES

Four subtypes of bombesin receptors are identified (gastrin-releasing peptide receptor, neuromedin B receptor, the orphan receptor bombesin receptor subtype 3 (BB3 or BRS-3) and bombesin receptor subtype 4 (BB4 )), however, only the pharmacology of the gastrin-releasing peptide re ceptor has been well studied. This lack of data is due in part to the absence of a general ligand. Recently we have discovered a ligand, I-1 25-[D-Tyr(6),beta Ala(11),Phe(13),Nle(14)]bombesin-(6-14) that binds t o BRS-3 receptors. In this study we investigate its ability to interac t with all four bombesin receptor subtypes. In rat pancreatic acini co ntaining only gastrin-releasing peptide receptor and in BB4 transfecte d BALB cells, this ligand and I-125-[Tyr(4)]bombesin, the conventional gastrin-releasing peptide receptor ligand, gave similar results for r eceptor number, affinity for bombesin and affinity for the unlabeled l igand. In neuromedin B receptor transfected BALB cells, this ligand an d I-125-[D-Tyr(0)]neuromedin B, the generally used neuromedin B recept or ligand, gave similar results for receptor number, neuromedin B affi nity or the unlabeled ligand affinity. Lastly, in BRS-3 transfected BA LB cells, only this ligand had high affinity. For all four bombesin re ceptors this ligand had an affinity of 1-8 nM and was equal or greater in affinity than any other specific ligands for any receptor. The unl abeled ligand is specific for gastrin-releasing peptide receptors on r at pancreatic acini and did not inhibit binding of I-125-cholecystokin in octapeptide (I-125-CCK-8), I-125-vasoactive intestinal peptide (I-1 25-VIP) or I-125-endothelin to their receptors. The unlabeled ligand w as an agonist only at the gastrin-releasing peptide receptor in rat ac ini and did not interact with CCKA receptors or muscarinic M-3 acetylc holine receptors to increase [H-3]inositol phosphates. These results d emonstrate I-125-[D-Tyr(6),beta Ala(11),Phe(13),Nle(14)]bombesin-(6-14 ) is a unique ligand with high affinity for all subtypes of bombesin r eceptors. Because of the specificity for bombesin receptors, this liga nd will be a valuable addition for such pharmacological studies as scr eening for bombesin receptor agonists or antagonists and, in particula r, for investigating BRS-3 cell biology, a receptor for which no ligan d currently exists. Published by Elsevier Science B.V.