Fluorescent Tools for Imaging Class A G-protein Coupled Receptors.

Abstract

G protein-coupled receptors (GPCRs) are pivotal in biological processes and represent a significant class of drug targets, with 516 approved drugs acting on 121 GPCRs. Many GPCRs, particularly orphan receptors, remain underexplored, emphasizing the need for innovative investigative tools. Fluorescent ligands provide a powerful means to characterize GPCRs including their functional mechanisms and spatial organization, bridging fundamental research and drug discovery. This review presents recent advances (2018-2024) in fluorescent probe development for Class A GPCRs, analyzing over 120 newly developed probes covering 60 GPCRs. We examine their distribution across receptor subclasses, comparing pre-2018 data with contemporary findings and identifying previously uncharted GPCRs that now have fluorescent ligands. Notably, novel probes have been developed for 12 new receptor subtypes and 6 orphan receptors such as GPR6, GPR52, GPR84, MAS1, MRGPRX₂, and MRGPRX₄. Advances in GPCR structural biology, driven by cryo-EM and AlphaFold technologies, have significantly enhanced probe development, facilitating the design of selective fluorescent ligands across aminergic, peptidergic, lipid, nucleotide, alicarboxylic, melatonin, protein, and orphan GPCRs. These innovations support a broad range of applications, from single-molecule imaging and in vivo bioimaging to diagnostics and fluorescence-guided surgery. By integrating fluorescence-based approaches with structural and pharmacological insights, this field continues to refine polypharmacology profiling, optimize drug-receptor interactions, and accelerate GPCR-targeted drug discovery.