Characterization And Pharmacology Of Receptors For Gastrointestinal Peptides

胃肠肽受体的表征和药理学

• 批准号: 7593650
• 负责人: Robert Jensen
• 金额: $ 35.97万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7593650
• 关键词: Affinity; Agonist; Amino Acid Substitution; Amino Acids; BRS3 gene; Binding; Bombesin; Bombesin Receptor; Camptothecin; Cytotoxic agent; Family; GRP gene; Gastrin releasing peptide; Gastrointestinal Hormone Receptors; Gastrointestinal Hormones; Gastrointestinal tract structure; Goals; Human; Ligands; Mediating; Molecular; Neuraxis; Numbers; Orphan; PACAP27; Pathologic Processes; Peptide Receptor; Peptides; Pharmacology; Physiological; Protocols documentation; Role; Secretin; Site-Directed Mutagenesis; Somatostatin; Structure; Therapeutic; Topoisomerase Inhibitors; Vasoactive Intestinal Peptide; Vasoactive Intestinal Peptide Receptors; analog; bombesin receptor subtype 3; cancer cell; gastrointestinal; member; molecular modeling; neoplastic cell; neuromedin B; receptor; somatostatin analog; vasoactive intestinal peptide receptor 1

The pharmacology and molecular pharmacology of various gastrointestinal (GI) peptides are being investigated. Three peptide receptor families investigated during the year are those for bombesin- (Bn) related peptides, somatatostatin (SS) family and for VIP-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). Using information from structure-function studies by us and others we have synthesized high affinity Bn, VIP and somatostatin analogues that were metabolically stable and that be could to cytotoxic agents such as the topoisomerase inhibitor, camptothecin (CPT), which had potent antitumor activity against various human tumor cells. We demonstrated using inactive chemically identical analogues that this tumoricidal effect was mediated by the peptide receptors expressed on the cancer cells. To identify key amino acids responsible for selective high affinity binding of various agonists for different members of the bombesin receptor family we used an analysis of the structural homologies of different receptors of this family and site-directed mutagenesis. Between the GRP receptor and BRS-3 receptor which different by >1000 in their affinity for bombesin (BN) we found 14 important amino acid differences which were then substituted in each receptor. Molecular modeling and pharmacology studies showed 7 of these were important for high affinity for Bn and contributed to the selectivity of these receptors demonstrating this is a useful approach to identify amino acid important for selectivity and affinity. A second aspect of our studies is to develop selective receptor ligands for Bn or VIP receptor subtypes that could be metabolically stable. To accomplish this for the human BRS-3 receptor, which has no selective ligands, we made N-Methyl substitutions, D-amino acid substitutions, truncations and substitutions of various amino acids in a nonselective ligand that we had discovered that interacted with this receptor with high affinity. One analog DTyr6, Apa-4 Cl, Phe13, Nle14-Bn (6-14) was found to have a marked enhanced selectivity for the human BRS3 receptor over the other receptors of this family. This analogue was found to have greater affinity and selectivity than other recently described BRS-3 selective ligands and thus should be useful to investigate its role in physiological and pathological processes. Furthermore, using our previous ligand structure function studies we were able to construct simplified analogs of the 28 amino acid peptide VIP that were metabolically stable and had high affinity and selectivity for VPAC1 receptors.

正在研究各种胃肠道（GI）肽的药理学和分子药理学。在这一年研究的三个肽受体家族是孟买（BN）相关的肽，生物抑素（SS）家族和VIP相关肽的肽受体家族。与BN相关的肽（GRP，NMB）主要与三种不同的受体（GRP-R，NMB-R）和孤儿受体BRS-3相互作用，以介导GI道和中枢神经系统（CNS）中的许多影响。使用我们和其他人的结构功能研究中的信息，我们已经合成了高亲和力BN，VIP和生长抑素类似物，这些类似物在代谢上稳定，可以与细胞毒素剂，例如topoisomerase抑制剂，凸轮皮塞克蛋白（CPT），这些毒素具有强有力的抗肿瘤活性，抗体细胞对各种人类肿瘤细胞。我们证明了使用非活性化学相同的类似物表明，这种肿瘤作用是由在癌细胞上表达的肽受体介导的。 为了鉴定负责选择性高亲和力的关键氨基酸对孟买受体家族的不同成员的选择性高亲和力结合，我们使用了该家族不同受体的结构同源性和定向诱变的分析。 在GRP受体和BRS-3受体之间，它们对孟买（BN）的亲和力> 1000不同，我们发现了14种重要的氨基酸差异，然后在每个受体中取代。 分子建模和药理学研究表明，其中7个对于对BN的高亲和力很重要，并有助于这些受体的选择性，证明这是鉴定氨基酸对选择性和亲和力很重要的有用方法。我们研究的第二个方面是开发用于代谢稳定的BN或V​​IP受体亚型的选择性受体配体。为了为没有选择性配体的人类BRS-3受体来实现这一目标，我们在非选择性配体中的各种氨基酸的d-氨基酸取代，D-氨基酸取代，截断和取代，我们发现我们发现与该受体相互作用具有高亲和力。发现一种模拟DTYR6，APA-4 CL，PHE13，NLE14-BN（6-14）对人类BRS3受体的选择性明显增强，而不是该家族的其他受体。发现该类似物比最近描述的BRS-3选择性配体具有更大的亲和力和选择性，因此对于研究其在生理和病理过程中的作用应该很有用。 此外，使用我们以前的配体结构功能研究，我们能够构建28个氨基酸肽VIP的简化类似物，这些氨基酸肽VIP具有代谢稳定，并且对VPAC1受体具有高亲和力和选择性。