Nomad Biosensors: A New Multiplexed Technology for the Screening of GPCR Ligands

Abstract

Nomad Technology (Innoprot [Innovative Technologies in Biological Systems], Derio, Spain), a novel tool for multiplexing high-throughput cell-based G protein-coupled receptor (GPCR) assays, is described in this work. This new technology comprises a family of fluorescent biosensors called Nomad Biosensors that allow for the measurement of responses mediated by G proteins through their interactions with second-messenger transduction proteins. GPCRs are one of the largest protein families of receptors in eukaryotes, and their signaling mediates important physiological processes within cells. Thus, GPCRs are associated with a wide variety of diseases, and considered major targets in therapeutic research. Nomad constitutes a novel tool for unraveling the mechanism of GPCR signal transduction by simultaneously tracing different pathways. GPCR activation changes the structural folding of the biosensor and promotes its vesicularization, as well as an increase in the fluorescence intensity. Based on this technology, the _MPXNomad cellular model was developed to discriminate between the Ca²⁺-mediated pathway and the cyclic adenosine monophosphate (cAMP)-mediated pathway. To validate this model, endothelin receptor B (ET_BR) was coexpressed into the _MPXNomad cell line and assessed with a specific agonist, an antagonist, and a chemical library of compounds. Nomad Technology optimizes the identification of novel GPCR ligands and enables the testing of large numbers of compounds.

Keywords: GPCR; HTS; fluorescence; high-throughput screening; multiplex and cell-based assay.