Development and Characterization of a Fluorescent Tracer for the Free Fatty Acid Receptor 2 (FFA2/GPR43)

Anders Højgaard Hansen; Eugenia Sergeev; Sunil K. Pandey; Brian D Hudson; Elisabeth Rexen Ulven; Graeme Milligan; Trond Ulven

doi:10.1021/acs.jmedchem.7b00338

Development and Characterization of a Fluorescent Tracer for the Free Fatty Acid Receptor 2 (FFA2/GPR43)

Anders Højgaard Hansen, Eugenia Sergeev, Sunil K. Pandey, Brian D Hudson, Elisabeth Rexen Ulven, Graeme Milligan, Trond Ulven

22 Citations (Scopus)

Abstract

The free fatty acid receptor 2 (FFA2/GPR43) is considered a potential target for treatment of metabolic and inflammatory diseases. Here we describe the development of the first fluorescent tracer for FFA2 intended as a tool for assessment of thermodynamic and kinetic binding parameters of unlabeled ligands. Starting with a known azetidine FFA2 antagonist, we used a carboxylic acid moiety known not to be critical for receptor interaction as attachment point for a nitrobenzoxadiazole (NBD) fluorophore. This led to the development of 4 (TUG-1609), a fluorescent tracer for FFA2 with favorable spectroscopic properties and high affinity, as determined by bioluminescence resonance energy transfer (BRET)-based saturation and kinetic binding experiments, as well as a high specific to nonspecific BRET binding signal. A BRET-based competition binding assay with 4 was also established and used to determine binding constants and kinetics of unlabeled ligands.

Original language	English
Journal	Journal of Medicinal Chemistry
Volume	60
Issue number	13
Pages (from-to)	5638-5645
Number of pages	8
ISSN	0022-2623
DOIs	https://doi.org/10.1021/acs.jmedchem.7b00338
Publication status	Published - 13 Jul 2017
Externally published	Yes

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title = "Development and Characterization of a Fluorescent Tracer for the Free Fatty Acid Receptor 2 (FFA2/GPR43)",

abstract = "The free fatty acid receptor 2 (FFA2/GPR43) is considered a potential target for treatment of metabolic and inflammatory diseases. Here we describe the development of the first fluorescent tracer for FFA2 intended as a tool for assessment of thermodynamic and kinetic binding parameters of unlabeled ligands. Starting with a known azetidine FFA2 antagonist, we used a carboxylic acid moiety known not to be critical for receptor interaction as attachment point for a nitrobenzoxadiazole (NBD) fluorophore. This led to the development of 4 (TUG-1609), a fluorescent tracer for FFA2 with favorable spectroscopic properties and high affinity, as determined by bioluminescence resonance energy transfer (BRET)-based saturation and kinetic binding experiments, as well as a high specific to nonspecific BRET binding signal. A BRET-based competition binding assay with 4 was also established and used to determine binding constants and kinetics of unlabeled ligands.",

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T1 - Development and Characterization of a Fluorescent Tracer for the Free Fatty Acid Receptor 2 (FFA2/GPR43)

AU - Hansen, Anders Højgaard

AU - Sergeev, Eugenia

AU - Pandey, Sunil K.

AU - Hudson, Brian D

AU - Rexen Ulven, Elisabeth

AU - Milligan, Graeme

AU - Ulven, Trond

PY - 2017/7/13

Y1 - 2017/7/13

N2 - The free fatty acid receptor 2 (FFA2/GPR43) is considered a potential target for treatment of metabolic and inflammatory diseases. Here we describe the development of the first fluorescent tracer for FFA2 intended as a tool for assessment of thermodynamic and kinetic binding parameters of unlabeled ligands. Starting with a known azetidine FFA2 antagonist, we used a carboxylic acid moiety known not to be critical for receptor interaction as attachment point for a nitrobenzoxadiazole (NBD) fluorophore. This led to the development of 4 (TUG-1609), a fluorescent tracer for FFA2 with favorable spectroscopic properties and high affinity, as determined by bioluminescence resonance energy transfer (BRET)-based saturation and kinetic binding experiments, as well as a high specific to nonspecific BRET binding signal. A BRET-based competition binding assay with 4 was also established and used to determine binding constants and kinetics of unlabeled ligands.

AB - The free fatty acid receptor 2 (FFA2/GPR43) is considered a potential target for treatment of metabolic and inflammatory diseases. Here we describe the development of the first fluorescent tracer for FFA2 intended as a tool for assessment of thermodynamic and kinetic binding parameters of unlabeled ligands. Starting with a known azetidine FFA2 antagonist, we used a carboxylic acid moiety known not to be critical for receptor interaction as attachment point for a nitrobenzoxadiazole (NBD) fluorophore. This led to the development of 4 (TUG-1609), a fluorescent tracer for FFA2 with favorable spectroscopic properties and high affinity, as determined by bioluminescence resonance energy transfer (BRET)-based saturation and kinetic binding experiments, as well as a high specific to nonspecific BRET binding signal. A BRET-based competition binding assay with 4 was also established and used to determine binding constants and kinetics of unlabeled ligands.

U2 - 10.1021/acs.jmedchem.7b00338

DO - 10.1021/acs.jmedchem.7b00338

M3 - Journal article

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JO - Journal of Medicinal Chemistry

JF - Journal of Medicinal Chemistry

IS - 13

ER -

Development and Characterization of a Fluorescent Tracer for the Free Fatty Acid Receptor 2 (FFA2/GPR43)

Abstract

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