PT  - JOURNAL ARTICLE
AU  - Tamara A.M. Mocking
AU  - Eleonore W.E. Verweij
AU  - Henry F Vischer
AU  - Rob Leurs
TI  - Homogeneous, real-time NanoBRET binding assays for the histamine H<sub>3</sub> and H<sub>4</sub> receptor on living cells
AID  - 10.1124/mol.118.113373
DP  - 2018 Jan 01
TA  - Molecular Pharmacology
PG  - mol.118.113373
4099  - http://molpharm.aspetjournals.org/content/early/2018/09/24/mol.118.113373.short
4100  - http://molpharm.aspetjournals.org/content/early/2018/09/24/mol.118.113373.full
AB  - Receptor binding affinity and ligand-receptor residence time are key parameters for the selection of drug candidates, and are routinely determined using radioligand competition binding assays. Recently, a novel bioluminescence resonance energy transfer (BRET) method utilizing a NanoLuc-fused receptor was introduced to detect fluorescent ligand binding. Moreover, this NanoBRET method gives the opportunity to follow fluorescent ligand-binding on intact cells in real time, and results might therefore better reflect in vivo conditions as compared to the routinely used cell homogenates or purified membrane fractions. In this study, a real-time NanoBRET-based binding assay was established and validated to detect binding of unlabeled ligands to the histamine H3 and histamine H4 receptor on intact cells. Obtained residence times of clinically tested H3R antagonists was reflected by their duration of H3R antagonism in a functional receptor recovery assay.