New Horizons on Molecular Pharmacology Applied to Drug Discovery: When Resonance Overcomes Radioligand Binding

Author(s): Larissa Pernomian, Mayara Santos Gomes, Josimar Dornelas Moreira, Carlos Henrique Tomich de Paula da Silva, Joaquin Maria Campos Rosa, Cristina Ribeiro de Barros Cardoso

Journal Name: Current Radiopharmaceuticals

Volume 10 , Issue 1 , 2017

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Graphical Abstract:


One of the cornerstones of rational drug development is the measurement of molecular parameters derived from ligand-receptor interaction, which guides therapeutic windows definition. Over the last decades, radioligand binding has provided valuable contributions in this field as key method for such purposes. However, its limitations spurred the development of more exquisite techniques for determining such parameters. For instance, safety risks related to radioactivity waste, expensive and controlled disposal of radioisotopes, radiotracer separation-dependence for affinity analysis, and one-site mathematical models-based fitting of data make radioligand binding a suboptimal approach in providing measures of actual affinity conformations from ligands and G proteincoupled receptors (GPCR). Current advances on high-throughput screening (HTS) assays have markedly extended the options of sparing sensitive ways for monitoring ligand affinity. The advent of the novel bioluminescent donor NanoLuc luciferase (Nluc), engineered from Oplophorus gracilirostris luciferase, allowed fitting bioluminescence resonance energy transfer (BRET) for monitoring ligand binding. Such novel approach named Nluc-based BRET (NanoBRET) binding assay consists of a real-time homogeneous proximity assay that overcomes radioligand binding limitations but ensures the quality in affinity measurements. Here, we cover the main advantages of NanoBRET protocol and the undesirable drawbacks of radioligand binding as molecular methods that span pharmacological toolbox applied to Drug Discovery. Also, we provide a novel perspective for the application of NanoBRET technology in affinity assays for multiple-state binding mechanisms involving oligomerization and/or functional biased selectivity. This new angle was proposed based on specific biophysical criteria required for the real-time homogeneity assigned to the proximity NanoBRET protocol.

Keywords: G protein-coupled receptors (GPCR), ligand binding affinity, drug discovery, radioligand binding assay, bioluminescence resonance energy transfer (BRET), NanoLuc luciferase (Nluc), Nluc-based BRET (NanoBRET), real-time proximity assays.

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Article Details

Year: 2017
Published on: 07 March, 2017
Page: [16 - 20]
Pages: 5
DOI: 10.2174/1874471010666170208152420
Price: $65

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PDF: 43