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Novel M₂ -selective, G_i -biased agonists of muscarinic acetylcholine receptors.

ABSTRACT:

Background and purpose

More than 30% of currently marketed medications act via GPCRs. Thus, GPCRs represent one of the most important pharmacotherapeutic targets. In contrast to traditional agonists activating multiple signalling pathways, agonists activating a single signalling pathway represent a new generation of drugs with increased specificity and fewer adverse effects.

Experimental approach

We have synthesized novel agonists of muscarinic ACh receptors and tested their binding and function (on levels of cAMP and inositol phosphates) in CHO cells expressing individual subtypes of muscarinic receptors, primary cultures of rat aortic smooth muscle cells and suspensions of digested native tissues from rats. Binding of the novel compounds to M₂ receptors was modelled in silico.

Key results

Two of the tested new compounds (1-(thiophen-2-ylmethyl)-3,6-dihydro-2H-pyridinium and 1-methyl-1-(thiophen-2-ylmethyl)-3,6-dihydro-2H-pyridinium) only inhibited cAMP synthesis in CHO cells, primary cultures, and native tissues, with selectivity for M₂ muscarinic receptors and displaying bias towards the G_i signalling pathway at all subtypes of muscarinic receptors. Molecular modelling revealed interactions with the orthosteric binding site in a way specific for a given agonist followed by agonist-specific changes in the conformation of the receptor.

Conclusions and implications

The identified compounds may serve as lead structures in the search for novel non-steroidal and non-opioid analgesics acting via M₂ and M₄ muscarinic receptors with reduced side effects associated with activation of the phospholipase C signalling pathway.

SUBMITTER: Randakova A

PROVIDER: S-EPMC7161557 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Efficacy describes the property of a ligand that enables the receptor to change its behavior towards the host cell, while biased agonism defines the ability of a ligand to differentially activate some of the vectorial pathways over others mediated through the receptor. However, little is known about the molecular basis defining the efficacy of ligands at G protein-coupled receptors. Here we characterize the biased agonism and cell phenotypic efficacy of seven agonists at the endogenous muscarinic M3 receptors in six different cell lines including HT-29, PC-3, HeLa, SF268, CCRF-CEM and HCT-15 cells.Quantitative real-time PCR and multiple label-free whole cell dynamic mass redistribution (DMR) assays were used to determine the functional muscarinic receptors in each cell line. DMR pathway deconvolution assay was used to determine the pathway biased activity of the muscarinic agonists. Operational agonism model was used to quantify the pathway bias, while macro-kinetic data reported in literature was used to analyze the biochemical mechanism of action of these agonists.Quantitative real-time PCR and ligand pharmacology studies showed that all the native cell lines endogenously express functional M3 receptors. Furthermore, different agonists triggered distinct DMR signals in a specific cell line as well as in different cell lines. DMR pathway deconvolution using known G protein modulators revealed that the M3 receptor in all the six cell lines signals through multiple G protein-mediated pathways, and certain agonists display biased agonism in a cell line-dependent manner. The whole cell efficacy and potency of these agonists were found to be sensitive to the assay time as well as the cell background. Correlation analysis suggested that the whole cell efficacy of agonists is correlated well with their macro-dissociation rate constants.This study implicates that the endogenous M3 receptors are coupled to multiple pathways, and the muscarinic agonists can display distinct biased agonism and whole cell phenotypic efficacy.

Dataset Information

Novel M₂ -selective, G_i -biased agonists of muscarinic acetylcholine receptors.

Background and purpose

Experimental approach

Key results

Conclusions and implications

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OmicsDI is part of the ELIXIR infrastructure

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Novel M2 -selective, Gi -biased agonists of muscarinic acetylcholine receptors.

Background and purpose

Experimental approach

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Conclusions and implications

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OmicsDI is part of the ELIXIR infrastructure

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Novel M₂ -selective, G_i -biased agonists of muscarinic acetylcholine receptors.