Pharmacology of neuroendocrine peptides

促肾上腺皮质激素释放因子（CRF）和尿皮质素（Ucn）的肽类拮抗剂在了解不同应激刺激下的内分泌、免疫、心血管和胃肠道功能方面具有重要意义。非肽类CRF拮抗剂被开发用于治疗抑郁症。此外，由于CRF受体选择性激动剂和拮抗剂的可用性，现在更好地理解了两种不同CRF受体（CRFR1和CRFR2）及其亚型的实际功能、分布和特异性。然而，多个CRF天然配体（CRF和CCR1）之间的显著氨基酸序列差异是不确定的。 Ucn 1、2和3）以及CRFR1和CRFR2的广泛分布及其伴随的多种功能调节仍有待整合和理解。本项目有两个互补的终点：a）结构和B）药理学/生物学。两个终点都严重依赖于肽（SPPS）和蛋白质（片段化学连接）的合成。a）我们建议与Riek博士的项目合作，设计和合成CRF/Ucn类似物和B1类GPCR的胞外结构域1（ECD1），用于使用NMR进行功能和结构研究。初步结果已经产生了CRFR2的ECD 1和CRF拮抗剂astressin B之间相互作用的模型。更多的研究 必须识别配体和负责识别和结合的初始步骤的相应ECD1之间的实际接触点。这将通过突变的ECD1和所选配体的互补衍生类似物的结合测定来实现。从这些研究中，我们将推导出CRFR1选择性和CRFR2选择性配体和各自受体的药效团的共有生物活性/结合构象。B）我们将在模型中使用这些药效团，用于合理设计新的肽和非肽CRFR1和CRFR2配体。特别是，我们建议使用几种与用于CRFR1拮抗剂的方法不同的方法来设计肽CRFR1选择性拮抗剂。 本发明涉及对一种有效的CRFR1选择性激动剂和一种有效的长效CRFR2选择性拮抗剂astressin2-B的应激表征。我们将使用这些选择性和有效的试剂，通过本计划项目内的合作，并与外部研究人员，确定可能治疗的病理生理状态的病因。

Peptidic antagonists of corticotropin releasing factor (CRF) and the urocortins (Ucn) have been critical in the understanding of endocrine, immune, cardiovascular and gastrointestinal functions in response to different stress stimuli. Nonpeptide CRF antagonists are developed for the treatment of depression. Further, the actual function, distribution and specificity of the two different CRF receptors (CRFR1 and CRFR2) and their subtypes are now better understood due to the availability of CRF-receptor-selective agonists and antagonists. However, significant amino acid sequence differences between the multiple CRF native ligands (CRF and Ucn 1, 2 and 3 in mammals) and the wide distribution of CRFR1 and CRFR2 with their concomitant modulation of multiple functions remain to be integrated and understood. There are two complementary end points to this Project: a) structural and b) pharmacological/biological. Both end points depend heavily on the synthesis of peptides (SPPS) and proteins (fragment chemical ligation). a) We propose to design and synthesize CRF/Ucn analogs and extra cellular domains 1 (ECD1s) of class B1 GPCR for functional and structural investigations using NMR in collaboration with Dr. Riek's project. Preliminary results have yielded a model of the interaction between the ECD1 of CRFR2 and the CRF antagonist, astressin B. Additional studies are mandatory to identify the actual contact points between ligands and the respective ECDs1 responsible for the initial steps of recognition and binding. This will be achieved with binding assays of mutated ECD1s and complementarily derivatized analogs of selected ligands. From these studies, we will derive consensus bioactive/binding conformations of the CRFR1 -selective and CRFR2-selective ligands and the pharmacophores of the respective receptors, b) We will use these pharmacophores in models for the rational design of novel peptide and nonpeptide CRFR1 and CRFR2 ligands. In particular, we propose to pursue the design of peptide CRFR1-selective antagonists using several approaches distinct from that used for the characterization of stressing a potent and CRFR1-selective agonist and astressin2-B, a potent and long acting CRFR2-selective antagonist. We will use these selective and potent reagents to identify through collaborations within this Program Project, and with outside investigators, the etiology of pathophysiological states for possible treatment.