title: Muscarinic receptors as model targets and antitargets for structure-based ligand discovery.
abstract: G protein-coupled receptors (GPCRs) regulate virtually all aspects of human physiology and represent an important class of therapeutic drug targets. Many GPCR-targeted drugs resemble endogenous agonists, often resulting in poor selectivity among receptor subtypes and restricted pharmacologic profiles. The muscarinic acetylcholine receptor family exemplifies these problems; thousands of ligands are known, but few are receptor subtype-selective and nearly all are cationic in nature. Using structure-based docking against the M2 and M3 muscarinic receptors, we screened 3.1 million molecules for ligands with new physical properties, chemotypes, and receptor subtype selectivities. Of 19 docking-prioritized molecules tested against the M2 subtype, 11 had substantial activity and 8 represented new chemotypes. Intriguingly, two were uncharged ligands with low micromolar to high nanomolar Ki values, an observation with few precedents among aminergic GPCRs. To exploit a single amino-acid substitution among the binding pockets between the M2 and M3 receptors, we selected molecules predicted by docking to bind to the M3 and but not the M2 receptor. Of 16 molecules tested, 8 bound to the M3 receptor. Whereas selectivity remained modest for most of these, one was a partial agonist at the M3 receptor without measurable M2 agonism. Consistent with this activity, this compound stimulated insulin release from a mouse β-cell line. These results support the ability of structure-based discovery to identify new ligands with unexplored chemotypes and physical properties, leading to new biologic functions, even in an area as heavily explored as muscarinic pharmacology.
authors.affiliation: Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California (A.C.K., B.K.K.); Department of Pharmaceutical Chemistry, University of California, San Francisco, California (D.R.W., B.K.S.); Faculty of Pharmacy, University of Toronto, Toronto, Ontario, Canada (B.K.S.); Molecular Signaling Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, Maryland (M.R., J.H., K.H., J.W.); and Department of Chemistry and Pharmacy, Friedrich Alexander University, Erlangen, Germany (K.E., P.G.).
authors.forename: Andrew C
authors.initials: AC
authors.lastname: Kruse
authors.forename: Dahlia R
authors.initials: DR
authors.lastname: Weiss
authors.forename: Mario
authors.initials: M
authors.lastname: Rossi
authors.forename: Jianxin
authors.initials: J
authors.lastname: Hu
authors.forename: Kelly
authors.initials: K
authors.lastname: Hu
authors.forename: Katrin
authors.initials: K
authors.lastname: Eitel
authors.forename: Peter
authors.initials: P
authors.lastname: Gmeiner
authors.forename: Jürgen
authors.initials: J
authors.lastname: Wess
authors.forename: Brian K
authors.initials: BK
authors.lastname: Kobilka
authors.forename: Brian K
authors.initials: BK
authors.lastname: Shoichet
mesh_terms: D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D018678:Cholinergic Agents / Q000008:administration & dosage / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D016503:Drug Delivery Systems / Q000639:trends*; D055808:Drug Discovery / Q000639:trends*; D006801:Humans; D007313:Insecta; D008024:Ligands; D051379:Mice; D011485:Protein Binding / Q000502:physiology; D051381:Rats; D011976:Receptors, Muscarinic / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D018678:Cholinergic Agents; D008024:Ligands; D011976:Receptors, Muscarinic