Mechanistic Insights into Allosteric Structure-Function Relationships at the M1 Muscarinic Acetylcholine Receptor

Abdul-Ridha, Alaa; Lane, J. Robert; Mistry, Shailesh N.; Lopez, Laura; Sexton, Patrick M.; Scammells, Peter J.; Christopoulos, Arthur; Canals, Meritxell

Abstract

Benzylquinolone carboxylic acid (BQCA) is the first highly selective positive allosteric modulator (PAM) for the M1 muscarinic acetylcholine receptor (mAChR), but it possesses low affinity for the allosteric site on the receptor. More recent drug discovery efforts identified 3-((1S,2S)-2-hydroxycyclohexyl)-6-((6-(1-methyl-1H-pyrazol-4-yl)pyridin-3-yl)methyl)benzo[h]-quinazolin-4(3H)-one (referred to herein as benzoquinazolinone 12) as a more potent M1 mAChR PAM with a structural ancestry originating from BQCA and related compounds. In the current study, we optimized the synthesis of and fully characterized the pharmacology of benzoquinazolinone 12, finding that its improved potency derived from a 50-fold increase in allosteric site affinity as compared with BQCA, while retaining a similar level of positive cooperativity with acetylcholine. We then utilized site-directed mutagenesis and molecular modeling to validate the allosteric binding pocket we previously described for BQCA as a shared site for benzoquinazolinone 12 and provide a molecular basis for its improved activity at the M1 mAChR. This includes a key role for hydrophobic and polar interactions with residues Tyr-179, in the second extracellular loop (ECL2) and Trp-4007.35 in transmembrane domain (TM) 7. Collectively, this study highlights how the properties of affinity and cooperativity can be differentially modified on a common structural scaffold and identifies molecular features that can be exploited to tailor the development of M1 mAChR-targeting PAMs. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Citation

Abdul-Ridha, A., Lane, J. R., Mistry, S. N., Lopez, L., Sexton, P. M., Scammells, P. J., …Canals, M. (2014). Mechanistic Insights into Allosteric Structure-Function Relationships at the M1 Muscarinic Acetylcholine Receptor. Journal of Biological Chemistry, 289(48), 33701-33711. https://doi.org/10.1074/jbc.m114.604967

• Journal Article Type: Article
• Acceptance Date: Oct 6, 2014
• Online Publication Date: Oct 17, 2014
• Publication Date: Nov 28, 2014
• Deposit Date: Oct 9, 2015
• Publicly Available Date: Oct 9, 2015
• Journal: Journal of Biological Chemistry
• Electronic ISSN: 0021-9258
• Publisher: American Society for Biochemistry and Molecular Biology
• Peer Reviewed: Peer Reviewed
• Volume: 289
• Issue: 48
• Pages: 33701-33711
• DOI: https://doi.org/10.1074/jbc.m114.604967
• Public URL: https://nottingham-repository.worktribe.com/output/737982
• Publisher URL: http://www.jbc.org/content/289/48/33701
• Additional Information: This research was originally published in Journal of Biological Chemistry. Alaa Abdul-Ridha, J. Robert Lane, Shailesh N. Mistry, Laura López, Patrick M. Sexton, Peter J. Scammells, Arthur Christopoulos and Meritxell Canals. Mechanistic insights into allosteric structure-function relationships at the M1 muscarinic acetylcholine receptor. Journal of Biological Chemistry. 2015. 289:33701-33711. © the American Society for Biochemistry and Molecular Biology.