Radiolabeled Dibenzodiazepinone-Type Antagonists Give Evidence of Dualsteric Binding at the M 2 Muscarinic Acetylcholine Receptor

J Med Chem. 2017 Apr 27;60(8):3314-3334. doi: 10.1021/acs.jmedchem.6b01892. Epub 2017 Apr 17.


The dualsteric ligand approach, aiming at ligands with improved subtype selectivity, has been increasingly applied to muscarinic receptors (MRs). In this article, we present the synthesis and characterization of a M2R subtype-preferring radiolabeled dibenzodiazepinone-type antagonist ([3H]UNSW-MK259, [3H]19) and its homodimeric analogue [3H]UR-AP060 ([3H]33). Saturation binding studies at the M2R, using the orthosteric antagonist atropine to determine unspecific binding, proved that the monomeric and the dimeric compound bind to the orthosteric binding site (apparent Kd: 0.87 and 0.31 nM, respectively). Various binding studies with [3H]19 and [3H]33 at the M2R, for instance, saturation binding experiments in the presence of the allosteric MR modulators W84 (8) or LY2119620 (9) (Schild-like analysis) suggested a competitive mechanism between the allosteric modulator and the dibenzodiazepinone derivatives, and thus a dualsteric binding mode of both 19 and 33. This was consistent with the results of M2R MD simulations (≥2 μs) performed with 19 and 33.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Benzodiazepinones / metabolism*
  • Binding Sites
  • Molecular Dynamics Simulation
  • Muscarinic Antagonists / pharmacology*
  • Radioisotopes / chemistry*
  • Receptor, Muscarinic M2 / antagonists & inhibitors*
  • Receptor, Muscarinic M2 / metabolism


  • Benzodiazepinones
  • Muscarinic Antagonists
  • Radioisotopes
  • Receptor, Muscarinic M2