The pharmacology and molecular pharmacology of various gastrointestinal (GI) peptides are being investigated. One peptide receptor family was investigated during the year: those for bombesin- (Bn) related peptides. Bn-related peptides (GRP, NMB) interact primarily with three distinct receptors (GRP-R, NMB-R) and the orphan receptor, BRS-3 to mediate a number of effects in the GI tract and central nervous system (CNS). In a recent study we examined the molecular basis for the unique ability of a synthetic ligand of bombesin to function as a universal, high affinity agonist at all human bombesin receptor subtypes. In this study using various criteria 74 amino acids were identified in human GRPR that might contribute to its high affinity for this ligand and 101 mutant receptors were made and expressed in CHOP cells to assess receptor affinities. 22 amino acids were identified that contributes to this high affinity of which 4 were especially important. The chemical properties of each of these was investigated in detail and the particular properties of each amino contributing to the high affinity was investigated. In another study the selectivity of NMB for the NMB-R and of various synthetic analogues of Bn for the orphan receptor, BRS-3. The identification of these molecular determinants will provide information that may allow development of even more selective ligands for these receptors in the future. In other studies we have addressed the ability of various described classes of agonists/antagonists of thee receptors to interact with human bombesin receptor members. We first synthesized 35 different synthetic putative peptide antagonists composed of 10 different chemical classes and examined their abilities to interact with human Bn receptors on both cell lines possessing a single subtype or Bn-R transfected cells containing receptor densities similar to seen on native cells. A number of classes of high affinity selective antagonists were identified which should be useful for in vivo studies. A similar study examined the ability of 24 natural occurring and synthetic Bn analogues to interact with these receptors as agonists and compared the results to the pharmacology in rat Bn expressing cells, which are commonly used in experimental studies. The results identified a number of selective high affinity agonists for the GRP and NMB receptor and showed that the rat GRP receptor pharmacophore differs substantially from that of the human and thus should not be used for extrapolating possible results to human receptors.
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