Selective Fluorescent Nonpeptidic Antagonists For Vasopressin V2 GPCR: Application To Ligand Screening and Oligomerization Assays.

A series of fluorescent benzazepine ligands for the arginine–vasopressin
V2 receptor (AVP V2R) was synthesized using
“Click” chemistry. Their in vitro pharmacological profile
at AVP V2R, V1aR, V1bR, and oxytocin
receptor was measured by binding assay and functional studies. Compound 9p, labeled with Lissamine Rhodamine B using novel solid-phase
organic tagging (SPOrT) resin, exhibited a high affinity for V2R (4.0 nM), an excellent selectivity toward V2R
and antagonist properties. By changing the nature of the dye, DY647
and Lumi4-Tb probes 44 and 47 still display
a high affinity for V2R (5.6 and 5.8 nM, respectively).
These antagonists constitute the first high-affinity selective nonpeptidic
fluorescent ligands for V2R. They enabled the development
of V2R time-resolved FRET-based assay readily amenable
to high-throughput screening. Taking advantage of their selectivity,
these compounds were also successfully involved in the study of V1aR–V2R dimerization on cell surface.

利用“点击”化学技术合成了一系列针对精氨酸加压素V2受体（AVP V2R）的荧光苯并噁唑啉配体。通过结合试验和功能研究，测定了这些配体在AVP V2R、V1aR、V1bR和催产素受体上的体外药理学特性。化合物9p，采用新型固相有机标记（SPOrT）树脂标记的异硫氰酸荧光素（Lissamine Rhodamine B），对V2R表现出极高的亲和力（4.0 nM），对V2R具有卓越的选择性，并展现出拮抗活性。通过改变染料的性质，DY647和Lumi4-Tb探针44和47仍对V2R保持高亲和力（分别为5.6 nM和5.8 nM）。这些拮抗剂构成了针对V2R的第一个高亲和力选择性非肽荧光配体。它们使得V2R时间分辨荧光共振能量转移（FRET）分析的快速发展成为可能，这种分析方法易于进行高通量筛选。利用其选择性，这些化合物还成功应用于研究细胞表面V1aR-V2R二聚化过程。