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Probe-Dependent Negative Allosteric Modulators of the Long-Chain Free Fatty Acid Receptor FFA4
Kenneth R Watterson1, Steffen V F Hansen1, Brian D Hudson1
1Centre 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.).
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.