title: Probe-Dependent Negative Allosteric Modulators of the Long-Chain Free Fatty Acid Receptor FFA4.
abstract: High-affinity and selective antagonists that are able to block the actions of both endogenous and synthetic agonists of G protein-coupled receptors are integral to analysis of receptor function and to support suggestions of therapeutic potential. Although there is great interest in the potential of free fatty acid receptor 4 (FFA4) as a novel therapeutic target for the treatment of type II diabetes, the broad distribution pattern of this receptor suggests it may play a range of roles beyond glucose homeostasis in different cells and tissues. To date, a single molecule, 4-methyl-N-9H-xanthen-9-yl-benzenesulfonamide (AH-7614), has been described as an FFA4 antagonist; however, its mechanism of antagonism remains unknown. We synthesized AH-7614 and a chemical derivative and demonstrated these to be negative allosteric modulators (NAMs) of FFA4. Although these NAMs did inhibit FFA4 signaling induced by a range of endogenous and synthetic agonists, clear agonist probe dependence in the nature of allosteric modulation was apparent. Although AH-7614 did not antagonize the second long-chain free fatty acid receptor, free fatty acid receptor 1, the simple chemical structure of AH-7614 containing features found in many anticancer drugs suggests that a novel close chemical analog of AH-7614 devoid of FFA4 activity, 4-methyl-N-(9H-xanthen-9-yl)benzamide (TUG-1387), will also provide a useful control compound for future studies assessing FFA4 function. Using TUG-1387 alongside AH-7614, we show that endogenous activation of FFA4 expressed by murine C3H10T1/2 mesenchymal stem cells is required for induced differentiation of these cells toward a more mature, adipocyte-like phenotype.
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Kenneth R
authors.initials: KR
authors.lastname: Watterson
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Steffen V F
authors.initials: SVF
authors.lastname: Hansen
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Brian D
authors.initials: BD
authors.lastname: Hudson
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Elisa
authors.initials: E
authors.lastname: Alvarez-Curto
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Sheikh Zahir
authors.initials: SZ
authors.lastname: Raihan
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Carlos M G
authors.initials: CMG
authors.lastname: Azevedo
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Gabriel
authors.initials: G
authors.lastname: Martin
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Julia
authors.initials: J
authors.lastname: Dunlop
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Yarwood
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.).
authors.forename: Trond
authors.initials: T
authors.lastname: Ulven
authors.affiliation: Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom (K.R.W., B.D.H., E.A.-C., S.Z.R., J.D., S.J.Y., G.M.); Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark (S.V.F.H., C.M.G.A., G.M., T.U.); and Institute of Biological Chemistry, Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh, United Kingdom (S.J.Y.) Graeme.Milligan@glasgow.ac.uk.
authors.forename: Graeme
authors.initials: G
authors.lastname: Milligan
mesh_terms: D000494:Allosteric Regulation / Q000187:drug effects / Q000502:physiology; D000818:Animals; D001713:Biphenyl Compounds / Q000494:pharmacology*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D057809:HEK293 Cells; D006801:Humans; D008809:Mice, Inbred C3H; D010666:Phenylpropionates / Q000494:pharmacology*; D043562:Receptors, G-Protein-Coupled / Q000819:agonists* / Q000037:antagonists & inhibitors / Q000502:physiology*
chemical_list: C585065:3-(4-((4-fluoro-4'-methyl-(1,1'-biphenyl)-2-yl)methoxy)phenyl)propanoic acid; D001713:Biphenyl Compounds; C519853:FFAR4 protein, human; D010666:Phenylpropionates; D043562:Receptors, G-Protein-Coupled