Delineating the Mode of Action of Adenosine A 1 Receptor Allosteric Modulators □ S

ABSTRACT Despite the identification of 2-amino-3-benzoylthiophenes (2A3BTs) as the first example of small-molecule allosteric potentiators of agonist function at a G protein-coupled receptor (GPCR)-the adenosine A 1 receptor-their mechanism of action is still not fully understood. We now report the mechanistic basis for the complex behaviors noted for 2A3BTs at A 1 receptors. Using a combination of membranebased and intact-cell radioligand binding, multiple signaling assays, and a native tissue bioassay, we found that the allosteric interaction between 2A3BTs and the agonists 2-chloro- 3 H]8-cyclopentyl-1,3-dipropylxanthine is consistent with a ternary complex model involving recognition of a single extracellular allosteric site. However, when allowed access to the intracellular milieu, 2A3BTs have a secondary action as direct G protein inhibitors; this latter property is receptor-independent as it is observed in nontransfected cells and also after stimulation of another GPCR. In addition, we found that 2A3BTs can signal as allosteric agonists in their own right but show bias toward certain pathways relative to the orthosteric agonist, R-PIA. These results indicate that 2A3BTs have a dual mode of action when interacting with the A 1 receptor and that they can engender novel functional selectivity in A 1 signaling. These mechanisms need to be factored into allosteric ligand structure-activity studies

Assessment of the molecular mechanisms of action of novel 4-phenylpyridine-2-one and 6-phenylpyrimidin-4-one allosteric modulators at the M1 muscarinic acetylcholine receptors

Positive allosteric modulators (PAMs) that target the M1 muscarinic acetylcholine (ACh) receptor (M1 mAChR) are potential treatments for cognitive deficits in conditions such as Alzheimer's disease and schizophrenia. We recently reported novel 4-phenylpyridine-2-one and 6-phenylpyrimidin-4-one M1 mAChR PAMs with the potential to display different modes of positive allosteric modulation and/or agonism (Mistry et al., 2016), but their molecular mechanisms of action remain undetermined. The current study compared the pharmacology of three such novel PAMs with the prototypical first-generation PAM, BQCA, in a recombinant Chinese hamster ovary (CHO) cell line stably expressing the human M1 mAChR. Interactions between the orthosteric agonists and the novel PAMs or BQCA suggested their allosteric effects were solely governed by modulation of agonist affinity. The greatest degree of positive co-operativity was observed with higher efficacy agonists, whereas minimal potentiation was observed when the modulators were tested against the lower efficacy agonist, xanomeline. Each PAM was investigated for its effects on the endogenous agonist, ACh, on three different signalling pathways, (ERK1/2 phosphorylation, IP1 accumulation and β-arrestin-2 recruitment), revealing that the allosteric potentiation generally tracked with the efficiency of stimulus-response coupling and that there was little pathway bias in the allosteric effects. Thus, despite the identification of novel allosteric scaffolds targeting the M1 mAChR, the molecular mechanism of action of these compounds is largely consistent with a model of allostery previously described for BQCA, suggesting that this may be a more generalized mechanism for M1 mAChR PAM effects than previously appreciated

Acetylcholine receptors (muscarinic) (version 2019.4) in the IUPHAR/BPS Guide to Pharmacology Database

Muscarinic acetylcholine receptors (nomenclature as agreed by the NC-IUPHAR Subcommittee on Muscarinic Acetylcholine Receptors [45]) are GPCRs of the Class A, rhodopsin-like family where the endogenous agonist is acetylcholine. In addition to the agents listed in the table, AC-42, its structural analogues AC-260584 and 77-LH-28-1, N-desmethylclozapine, TBPB and LuAE51090 have been described as functionally selective agonists of the M1 receptor subtype via binding in a mode distinct from that utilized by non-selective agonists [243, 242, 253, 155, 154, 181, 137, 11, 230]. There are two pharmacologically characterised allosteric sites on muscarinic receptors, one defined by it binding gallamine, strychnine and brucine, and the other defined by the binding of KT 5720, WIN 62,577, WIN 51,708 and staurosporine [161, 162]

Development of novel 4‐arylpyridin‐2‐one and 6‐arylpyrimidin‐4‐one positive allosteric modulators of the M1 muscarinic acetylcholine receptor

This study investigated the structure‐activity relationships of 4‐phenylpyridin‐2‐one and 6‐phenylpyrimidin‐4‐one muscarinic M1 acetylcholine receptor (M1 mAChRs) positive allosteric modulators (PAMs). The presented series focuses on modifications to the core and top motif of the reported leads, MIPS1650 (1) and MIPS1780 (2). Profiling of our novel analogues showed that these modifications result in more nuanced effects on the allosteric properties compared to our previous compounds with alterations to the biaryl pendant. Further pharmacological characterisation of the selected compounds in radioligand binding, IP1 accumulation and β‐arrestin 2 recruitment assays demonstrated that despite primarily acting as affinity modulators, the PAMs displayed different pharmacological properties across the two cellular assays. The novel PAM 7f is a potential lead candidate for further development of peripherally‐restricted M1 PAMs, due to its lower blood‐brain‐barrier (BBB) permeability and improved exposure in the periphery compared to lead 2

Acetylcholine receptors (muscarinic) in GtoPdb v.2021.3

Muscarinic acetylcholine receptors (mAChRs) (nomenclature as agreed by the NC-IUPHAR Subcommittee on Muscarinic Acetylcholine Receptors [50]) are activated by the endogenous agonist acetylcholine. All five (M1-M5) mAChRs are ubiquitously expressed in the human body and are therefore attractive targets for many disorders. Functionally, M1, M3, and M5 mAChRs preferentially couple to Gq/11 proteins, whilst M2 and M4 mAChRs predominantly couple to Gi/o proteins. Both agonists and antagonists of mAChRs are clinically approved drugs, including pilocarpine for the treatment of elevated intra-ocular pressure and glaucoma, and atropine for the treatment of bradycardia and poisoning by muscarinic agents such as organophosphates

Acetylcholine receptors (muscarinic) in GtoPdb v.2023.1

Muscarinic acetylcholine receptors (mAChRs) (nomenclature as agreed by the NC-IUPHAR Subcommittee on Muscarinic Acetylcholine Receptors [53]) are activated by the endogenous agonist acetylcholine. All five (M1-M5) mAChRs are ubiquitously expressed in the human body and are therefore attractive targets for many disorders. Functionally, M1, M3, and M5 mAChRs preferentially couple to Gq/11 proteins, whilst M2 and M4 mAChRs predominantly couple to Gi/o proteins. Both agonists and antagonists of mAChRs are clinically approved drugs, including pilocarpine for the treatment of elevated intra-ocular pressure and glaucoma, and atropine for the treatment of bradycardia and poisoning by muscarinic agents such as organophosphates. Of note, it has been observed that mAChRs dimerise reversibly [134] and that dimerisation/oligomerisation can be affected by ligands [183, 196]

Synthesis and pharmacological profiling of analogues of benzyl quinolone carboxylic acid (BQCA) as allosteric modulators of the M1 muscarinic receptor

Established therapy in Alzheimer’s disease involves potentiation of the endogenous orthosteric ligand, acetylcholine, at the M1 muscarinic receptors found in higher concentrations in the cortex and hippocampus. Adverse effects, due to indiscriminate activation of other muscarinic receptor subtypes, are common. M1 muscarinic positive allosteric modulators/allosteric agonists such as BQCA offer an attractive solution, being exquisitely M1-selective over other muscarinic subtypes. A common difficulty with allosteric ligands is interpreting SAR, based on composite potency values derived in the presence of fixed concentration of agonist. In reality these values encompass multiple pharmacological parameters, each potentially and differentially sensitive to structural modification of the ligand. We report novel BQCA analogues which appear to augment ligand affinity for the receptor (pKB), intrinsic efficacy (τB), and both binding (α) and functional (β) cooperativity with acetylcholine. Ultimately, development of such enriched SAR surrounding allosteric modulators will provide insight into their mode of action

THE CONCISE GUIDE TO PHARMACOLOGY 2021/22: G protein-coupled receptors.

The Concise Guide to PHARMACOLOGY 2021/22 is the fifth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of nearly 1900 human drug targets with an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. Although the Concise Guide constitutes over 500 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/bph.15538. G protein-coupled receptors are one of the six major pharmacological targets into which the Guide is divided, with the others being: ion channels, nuclear hormone receptors, catalytic receptors, enzymes and transporters. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2021, and supersedes data presented in the 2019/20, 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate

The Concise Guide to PHARMACOLOGY 2023/24: G protein-coupled receptors.

peer reviewedThe Concise Guide to PHARMACOLOGY 2023/24 is the sixth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of approximately 1800 drug targets, and about 6000 interactions with about 3900 ligands. There is an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (https://www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. Although the Concise Guide constitutes almost 500 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/bph.16177. G protein-coupled receptors are one of the six major pharmacological targets into which the Guide is divided, with the others being: ion channels, nuclear hormone receptors, catalytic receptors, enzymes and transporters. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2023, and supersedes data presented in the 2021/22, 2019/20, 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate