Guanidine Derivatives: How Simple Structural Modification of Histamine H₃R Antagonists Has Led to the Discovery of Potent Muscarinic M₂R/M₄R Antagonists

Abstract

This article describes the discovery of novel potent muscarinic receptor antagonists identified during a search for more active histamine H 3 receptor (H 3 R) ligands. The idea was to replace the flexible seven methylene linker with a semirigid 1,4-cyclohexylene or p -phenylene substituted group of the previously described histamine H 3 R antagonists ADS1017 and ADS1020 . These simple structural modifications of the histamine H 3 R antagonist led to the emergence of additional pharmacological effects, some of which unexpectedly showed strong antagonist potency at muscarinic receptors. This paper reports the routes of synthesis and pharmacological characterization of guanidine derivatives, a novel chemotype of muscarinic receptor antagonists binding to the human muscarinic M 2 and M 4 receptors (hM 2 R and hM 4 R, respectively) in nanomolar concentration ranges. The affinities of the newly synthesized ADS10227 (1-{4-{4-{[4-(phenoxymethyl)cyclohexyl]methyl}piperazin-1-yl}but-1-yl}-1-(benzyl)guanidine) at hM 2 R and hM 4 R were 2.8 nM and 5.1 nM, respectively.

A Versatile Sub-Nanomolar Fluorescent Ligand Enables NanoBRET Binding Studies and Single-Molecule Microscopy at the Histamine H3 Receptor

“…Using 200 nM of 12 , we observed little off-target effects with only the muscarinic hM 2 R and hM 4 R displaying mild but significant response (13.2 ± 4.5 and 6.7 ± 5.3% of hH 3 R response, respectively; Figure ). Some ligand binding at the hM 2 R and hM 4 R was unsurprising as the binding sites of these receptors share features with the hH 3 R, and several characteristics of known H 3 R ligands have been proven to be useful in developing muscarinic antagonists. , It should be noted that BRET signals from binding experiments to different NLuc-GPCR constructs can only be compared with caution given the different N-terminal lengths and localizations of orthosteric binding pockets. Therefore, the low BRET signals observed at hM 2 R and hM 4 R (only ∼10% of the BRET signal detected with NLuc-hH 3 R) do not provide a quantitative value but rather an estimate for the binding of 12 to the receptors.…”

“…56,57 It should be noted that BRET signals from binding experiments to different NLuc-GPCR constructs can only be compared with caution given the different N-terminal lengths and localizations of orthosteric binding pockets. Therefore, the low BRET signals observed at hM 2 R and hM 4 R (only ∼10% of the BRET signal detected with NLuc-hH 3 R) do not provide a quantitative value but rather an estimate for the binding of 12 to the receptors.…”

Multitargeting nature of muscarinic orthosteric agonists and antagonists

“…An interesting aspect of interneurotransmitter effects has recently been shown—the simple replacement of a flexible alkyl chain with a semirigid aryl or cycloalkyl ring leads to increased mAChR affinity of specific ligands, detrimentally affecting histamine receptor binding ( Staszewski et al, 2021 ). However, these newly synthesized ligands continued to bind to histamine receptors, although with lower affinity.…”

Modular access to substituted cyclohexanes with kinetic stereocontrol

“…Cyclohexanes bearing one substituent in the equatorial position and another in the axial position, namely the 1,2- cis , 1,3- trans , and 1,4- cis substitution patterns (Fig. 1A), represent a key scaffold in a wide range of molecules of pharmaceutical interest, such as linrodostat ( 7 ), an anticancer phase 3 drug; cathepsin S inhibitor ( 8 ); oxysterols receptor LXR-beta ( 9 ); and histamine H 3 R ADS10227 ( 10 ) (Fig. 1B).…”

“…1B). Furthermore, they frequently exhibit better bioactivity than their thermodynamically favored isomers (10,11). Although studies toward the development of efficient methods for the construction of cyclohexane skeletons have not stopped since the discovery of the Diels-Alder reaction (12)(13)(14), modular and variable approaches for the synthesis of thermodynamically disfavored substituted cyclohexanes are still lacking.…”