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Target deconvolution studies of (2R,6R)-hydroxynorketamine: an elusive search

Molecular Psychiatry volume 27, pages 4144–4156 (2022)Cite this article

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Abstract

The off-label use of racemic ketamine and the FDA approval of (S)-ketamine are promising developments for the treatment of depression. Nevertheless, racemic ketamine and (S)-ketamine are controlled substances with known abuse potential and their use is associated with undesirable side effects. For these reasons, research efforts have focused on identifying alternatives. One candidate is (2R,6R)-hydroxynorketamine ((2R,6R)-HNK), a ketamine metabolite that in preclinical models lacks the dissociative and abuse properties of ketamine while retaining its antidepressant-like behavioral efficacy. (2R,6R)-HNK’s mechanism of action however is unclear. The main goals of this study were to perform an in-depth pharmacological characterization of (2R,6R)-HNK at known ketamine targets, to use target deconvolution approaches to discover novel proteins that bind to (2R,6R)-HNK, and to characterize the biodistribution and behavioral effects of (2R,6R)-HNK across several procedures related to substance use disorder liability. We found that unlike (S)- or (R)-ketamine, (2R,6R)-HNK did not directly bind to any known or proposed ketamine targets. Extensive screening and target deconvolution experiments at thousands of human proteins did not identify any other direct (2R,6R)-HNK-protein interactions. Biodistribution studies using radiolabeled (2R,6R)-HNK revealed non-selective brain regional enrichment, and no specific binding in any organ other than the liver. (2R,6R)-HNK was inactive in conditioned place preference, open-field locomotor activity, and intravenous self-administration procedures. Despite these negative findings, (2R,6R)-HNK produced a reduction in immobility time in the forced swim test and a small but significant increase in metabolic activity across a network of brain regions, and this metabolic signature differed from the brain metabolic profile induced by ketamine enantiomers. In sum, our results indicate that (2R,6R)-HNK does not share pharmacological or behavioral profile similarities with ketamine or its enantiomers. However, it could still be possible that both ketamine and (2R,6R)-HNK exert antidepressant-like efficacy through a common and previously unidentified mechanism. Given its pharmacological profile, we predict that (2R,6R)-HNK will exhibit a favorable safety profile in clinical trials, and we must wait for clinical studies to determine its antidepressant efficacy.

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Fig. 1: (2R,6R)-HNK does not bind nor activate known ketamine binding sites.
Fig. 2: Target deconvolution assays and other suggested targets.
Fig. 3: Fast clearance and no brain regional specificity of (2R,6R)-HNK uptake.
Fig. 4: Changes in metabolic activity induced by (2R,6R)-HNK differ from those produced by (S)-ketamine.
Fig. 5: Lack of effects of (2R,6R)-HNK on locomotor activity, CPP and self-administration.

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Acknowledgements

Some Ki determinations, and agonist and/or antagonist functional data were generously provided by the National Institute of Mental Health’s Psychoactive Drug Screening Program, Contract # HHSN-271-2018-00023-C (NIMH PDSP). The NIMH PDSP is Directed by Bryan L. Roth MD, PhD at the University of North Carolina at Chapel Hill and Project Officer Jamie Driscoll at NIMH, Bethesda MD, USA. This work was supported by the NIDA Intramural Research Program (ZIA000069), the NIA, NIMH and NCATS Intramural Research Programs and by Grants RYC-2019-027371-I (JB) and PID2020-117989RA-I00 (JB) funded by MCIN/AEI /10.13039/501100011033 and by “ESF Investing in your future”. TDG was supported by NIH R01-MH107615 and RAI145211A, and VA Merit Awards 1I01BX004062 and 101BX003631-01A1.

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Authors and Affiliations

  1. Biobehavioral Imaging and Molecular Neuropsychopharmacology Unit, National Institute on Drug Abuse Intramural Research Program, Baltimore, 21224, MD, USA

    Jordi Bonaventura, Juan L. Gomez, Meghan L. Carlton, Sherry Lam & Michael Michaelides

  2. Departament de Patologia i Terapèutica Experimental, Institut de Neurociències, Universitat de Barcelona, L’Hospitalet de Llobregat, 08907, Catalonia, Spain

    Jordi Bonaventura

  3. Molecular Neuropharmacology Section, National Institute of Neurological Disorders and Stroke Intramural Research Program, Bethesda, 20892, MD, USA

    Marta Sanchez-Soto & David R. Sibley

  4. Division of Preclinical Innovation, National Center for Advancing Translational Sciences, Rockville, 20850, MD, USA

    Patrick J. Morris & Craig J. Thomas

  5. Biomedical Research Center, National Institute on Aging, National Institutes of Health, Baltimore, 21224, MD, USA

    Ruin Moaddel

  6. National Institute of Mental Health Psychoactive Drug Screening Program, Department of Pharmacology, University of North Carolina Chapel Hill Medical School, Chapel Hill, 27599, NC, USA

    Hye Jin Kang

  7. Department of Psychology, University of Cyprus, Nicosia, 2109, Cyprus

    Panos Zanos

  8. Veterans Affairs Maryland Health Care System, Baltimore, MD, 21201, USA

    Todd D. Gould

  9. Departments of Psychiatry, Pharmacology, and Anatomy and Neurobiology, University of Maryland School of Medicine, Baltimore, 21201, MD, USA

    Todd D. Gould

  10. Experimental Therapeutics and Pathophysiology Branch, National Institute of Mental Health, Intramural Research Program, Bethesda, 20892, MD, USA

    Carlos A. Zarate Jr.

  11. Department of Psychiatry & Behavioral Sciences, Johns Hopkins University School of Medicine, Baltimore, 21205, MD, USA

    Michael Michaelides

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  1. Jordi Bonaventura
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Contributions

JB, JLG, MLC, SL, MSS, RM, HJK and PZ performed the experiments. JB, JLG, MLC, SL, MSS, PJM, RM and HYJ analyzed the data. JB, PJM, RM, TDG, CJT, CJZ and MM supervised the experiments. PJM, CJT, DRS, CAZ, MM provided access to resources and support. JB and MM designed the study and wrote the manuscript with input from all coauthors. All authors critically reviewed the content and approved the final version before submission. All coauthors reviewed the manuscript and provided comments.

Corresponding author

Correspondence to Michael Michaelides.

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Competing interests

CZ is listed as a co-inventor on a patent for the use of ketamine in major depression and suicidal ideation. CZ and RM are co-inventors on a patent for the use of (2R,6R)-HNK, (S)-dehydronorketamine, and other stereoisomeric dehydroxylated and hydroxylated metabolites of (R,S)-ketamine metabolites in the treatment of depression and neuropathic pain. PZ, RM, PM, CJT, CAZ and TDG are co-inventors on a patent application for the use of (2R,6R)-HNK and (2S,6S)-HNK in the treatment of depression, anxiety, anhedonia, suicidal ideation, and post-traumatic stress disorders, and on a patent on the crystal forms and methods of synthesis of (2R,6R)-HNK and (2S,6S)-HNK. PM and CJT are co-inventors on a patent application for the salts of (2R,6R)-HNK, their crystal forms, and methods of making the same and the process for synthesis and purification of (2R,6R)-HNK. RM, PM, CAZ, and CT have assigned their patent rights to the U.S. government but will share a percentage of any royalties that may be received by the government. PZ and TDG have assigned their patent rights to the University of Maryland Baltimore but will share a percentage of any royalties that may be received by the University of Maryland Baltimore. MM has received research funding from AstraZeneca, Redpin Therapeutics and Attune Neurosciences. All other authors declare no conflicts of interests.

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Bonaventura, J., Gomez, J.L., Carlton, M.L. et al. Target deconvolution studies of (2R,6R)-hydroxynorketamine: an elusive search. Mol Psychiatry 27, 4144–4156 (2022). https://doi.org/10.1038/s41380-022-01673-w

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