Abstract
Positive allosteric modulation of metabotropic glutamate subtype 5 (mGlu5) receptor has emerged as a potential new therapeutic strategy for the treatment of schizophrenia and cognitive impairments. However, positive allosteric modulator (PAM) agonist activity has been associated with adverse side effects, and neurotoxicity has also been observed for pure PAMs. The structural and pharmacological basis of therapeutic versus adverse mGlu5 PAM in vivo effects remains unknown. Thus, gaining insights into the signaling fingerprints, as well as the binding kinetics of structurally diverse mGlu5 PAMs, may help in the rational design of compounds with desired properties. We assessed the binding and signaling profiles of N-methyl-5-(phenylethynyl)pyrimidin-2-amine (MPPA), 3-cyano-N-(2,5-diphenylpyrazol-3-yl)benzamide (CDPPB), and 1-[4-(4-chloro-2-fluoro-phenyl)piperazin-1-yl]-2-(4-pyridylmethoxy)ethenone [compound 2c, a close analog of 1-(4-(2-chloro-4-fluorophenyl)piperazin-1-yl)-2-(pyridin-4-ylmethoxy)ethanone] in human embryonic kidney 293A cells stably expressing mGlu5 using Ca2+ mobilization, inositol monophosphate (IP1) accumulation, extracellular signal–regulated kinase 1/2 (ERK1/2) phosphorylation, and receptor internalization assays. Of the three allosteric ligands, only CDPPB had intrinsic agonist efficacy, and it also had the longest receptor residence time and highest affinity. MPPA was a biased PAM, showing higher positive cooperativity with orthosteric agonists in ERK1/2 phosphorylation and Ca2+ mobilization over IP1 accumulation and receptor internalization. In primary cortical neurons, all three PAMs showed stronger positive cooperativity with (S)-3,5-dihydroxyphenylglycine (DHPG) in Ca2+ mobilization over IP1 accumulation. Our characterization of three structurally diverse mGlu5 PAMs provides further molecular pharmacological insights and presents the first assessment of PAM-mediated mGlu5 internalization.
SIGNIFICANCE STATEMENT Enhancing metabotropic glutamate receptor subtype 5 (mGlu5) activity is a promising strategy to treat cognitive and positive symptoms in schizophrenia. It is increasingly evident that positive allosteric modulators (PAMs) of mGlu5 are not all equal in preclinical models; there remains a need to better understand the molecular pharmacological properties of mGlu5 PAMs. This study reports detailed characterization of the binding and functional pharmacological properties of mGlu5 PAMs and is the first study of the effects of mGlu5 PAMs on receptor internalization.
Footnotes
- Received October 20, 2020.
- Accepted January 27, 2021.
↵1 Current affiliation: QIMR Berghofer Medical Research Institute, Brisbane, QLD, Australia.
↵2 A.A. and T.C.M. contributed equally to this work.
↵3 H.B.-O. and K.J.G. contributed equally to this work.
A.A. acknowledges financial support from the University of Copenhagen, Oticon Foundation, and Torben and Alice Frimodts Foundation. H.B.-O. acknowledges financial support from the Augustinus Foundation, the Lundbeck Foundation, and the Independent Research Fund Denmark. This project received funding from the European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement (797497) (T.C.M.). S.R.F. acknowledges financial support from the Lundbeck Foundation and the Independent Research Fund Denmark. This work was supported by the National Health and Medical Research Council of Australia (NHMRC): Project Grants APP1084775 (K.J.G.) and APP1127322 (K.J.G.). K.J.G. is supported by an Australian Research Council Future Fellowship: FT170100392.
J.L.H. was an employee and shareholder of Novo Nordisk A/S at the time of the study.
A prior version of the paper was included in the following PhD thesis: Arsova A (2018) Biased signaling and allosteric modulation of metabotropic glutamate receptor 5. Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
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This article has supplemental material available at molpharm.aspetjournals.org.
- Copyright © 2021 by The American Society for Pharmacology and Experimental Therapeutics
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