Quantitative cell-based high-content screening for vasopressin receptor agonists using transfluor technology

Area of Science:

• Pharmacology
• Cell Biology
• Biochemistry

Background:

• G-protein-coupled receptors (GPCRs) are crucial drug targets.
• Developing robust assays for GPCR agonist screening is essential for drug discovery.
• Vasopressin V2 receptor (V2R) plays a role in regulating water balance.

Purpose of the Study:

• To demonstrate a universal assay for screening GPCR agonists.
• To identify novel agonists for the vasopressin V2 receptor (V2R).
• To validate the utility of high-content screening (HCS) for GPCR drug discovery.

Main Methods:

• Utilized cells with stable expression of V2R and a green fluorescent protein (GFP)-labeled beta-arrestin fusion protein.
• Employed a high-content screening (HCS) assay to analyze a peptide library for V2R agonists.
• Automated imaging and quantitative analysis of intracellular fluorescent spots indicative of receptor activation.

Main Results:

• Successfully screened a peptide library for V2R agonists using the developed GPCR assay.
• Identified V2R agonists characterized by V2R internalization and associated GFP-beta-arrestin in endosomes.
• Hits were further evaluated for potency, demonstrating the assay's effectiveness in lead identification.

Conclusions:

• The developed GPCR assay is a simple and universal method for screening agonists.
• Automated HCS combined with beta-arrestin recruitment provides a powerful platform for GPCR drug discovery.
• This approach facilitates the identification and validation of potent GPCR-targeting drug leads.