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A Kinetic Fluorescence-based Ca2+ Mobilization Assay to Identify G Protein-coupled Receptor Agonists, Antagonists, and Allosteric Modulators
Published on: February 20, 2018
Molecular Basis of Action of a Small-Molecule Positive Allosteric Modulator Agonist at the Type 1 Cholecystokinin
Aditya J Desai1, Ingrid Mechin1, Karthigeyan Nagarajan1
1Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic, Scottsdale, Arizona (A.J.D., L.J.M.); In Silico Drug Discovery Department, Icagen Tucson Innovation Center, Oro Valley, Arizona (I.M., K.N., A.N.); Drug Discovery Biology Theme, Monash Institute of Pharmaceutical Sciences and Department of Pharmacology, Monash University, Parkville, Australia (C.V., D.W., P.M.S., A.C., L.J.M.); Molsoft, La Jolla, California (P.C.H.L., A.O., R.A.); Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California (R.A.); and School of Pharmacy, Fudan University, Shanghai, China (D.W., P.M.S.).
Abstract:
Allosteric modulation of receptors provides mechanistic safety while effectively achieving biologic endpoints otherwise difficult or impossible to obtain by other means. The theoretical case has been made for the development of a positive allosteric modulator (PAM) of the type 1 cholecystokinin receptor (CCK1R) having minimal intrinsic agonist activity to enhance meal-induced satiety for the treatment of obesity, while reducing the risk of side effects and/or toxicity. Unfortunately, such a drug does not currently exist. In this work, we have identified a PAM agonist of the CCK1R, SR146131, and determined its putative binding mode and receptor activation mechanism by combining molecular modeling, chimeric CCK1R/CCK2R constructs, and site-directed mutagenesis. We probed the structure-activity relationship of analogs of SR146131 for impact on agonism versus cooperativity of the analogs. This identified structural features that might be responsible for binding affinity and potency while retaining PAM activity. SR146131 and several of its analogs were docked into the receptor structure, which had the natural endogenous peptide agonist, cholecystokinin, already in the bound state (by docking), providing a refined structural model of the intact CCK1R holoreceptor. Both SR146131 and its analogs exhibited unique probe-dependent cooperativity with orthosteric peptide agonists and were simultaneously accommodated in this model, consistent with the derived structure-activity relationships. This provides improved understanding of the molecular basis for CCK1R-directed drug development.
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