松弛素家族多肽受体4（RXFP4）属于A类GPCR，其天然配体为胰岛素样肽5（INSL5）。前期研究表明二者均与糖代谢、摄食以及胰岛功能等密切相关，故推测INSL5/RXFP4系统在胰岛β细胞功能维持及胰岛素分泌中发挥作用。申请人的最新研究成果显示RXFP4在胰岛β细胞上高表达，INSL5可促进胰岛细胞释放胰岛素并诱导肠L细胞分泌GLP-1，动物水平研究也证实INSL5可以显著改善小鼠的糖耐量，这表明RXFP4是糖尿病治疗的潜在靶点。然而，INSL5制备极其困难，为解决RXFP4配体匮乏的问题，我们建立了基于cAMP通路的高通量筛选方法并筛选发现了其小分子配体。本项目拟对小分子配体进行结构优化改造，研究受体配体的结构与功能相关性及其分子机制，并选取效果最优的活性化合物开展系统的体内外抗糖尿病药效评价。项目的成功实施可望阐释RXFP4信号转导机制，发现1-2个先导化合物用于抗糖尿病药用开发。

Relaxin/insulin like family peptide receptor 4 (RXFP4) belongs to class A G protein-coupled receptor (GPCR) family with its cognate ligand named insulin-like peptide 5 (INSL5) which is a member of insulin superfamily. Various studies suggest that INSL5/RXFP4 is involved in glucose metabolism, appetite control and pancreatic islet function. Our recent results demonstrated that RXFP4 is highly expressed in islet β cells, and INSL5 is capable of promoting insulin release from β cells and enhancing glucagon-like peptide-1 (GLP-1) secretion from intestinal L cells. Subsequent in vivo investigations established its effects on glucose-stimulated insulin secretion (GSIS) and blood glucose regulation suggesting that the INSL5/RXFP4 system may be a potential therapeutic target for metabolic disorders such as type 2 diabetes. Further studies, however, are hampered by extremely limited access to native INSL5 (very difficult to produce an adequate amount) and the lack of small molecule ligands of RXFP4. To solve this problem, we developed a high-throughput screening (HTS) assay based on measurement of cAMP content and performed HTS campaigns which identified a few small molecule agonist-like compounds for RXFP4. In this project, we plan to conduct structural modification and optimization of these confirmed hits, study their structure-activity relationship (SAR) and molecular mechanism of action, and evaluate anti-diabetic their property in vitro and in vivo. The outcome of this project will help us better understand the signal transduction pathways of RXFP4 and discover one to two lead compounds for anti-diabetic drug development.