Family of Structurally Related Bioconjugates Yields Antibodies with Differential Selectivity against Ketamine and 6-Hydroxynorketamine
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High Resolution Image Download MS PowerPoint Slide The dissociative-hypnotic compound ketamine is being used in an increasingly wide range of therapeutic contexts, including anesthesia, adjunctive analgesia, treatment-resistant depression, but it also continues to be a notable substance of abuse. No specific antidotes exist for ketamine intoxication or overdose. Immunopharmacotherapy has demonstrated the ability to offer overdose protection through production of highly specific antibodies that prevent psychoactive drug penetration across the blood–brain barrier, although antiketamine antibodies have not yet been assessed or optimized for use in this approach. Moreover, generation of specific antibodies also provides an opportunity to address the role of 6-hydroxynorketamine metabolites in ketamine’s rapid-acting antidepressant effect through selective restriction of metabolite access to the central nervous system. Hapten design is a critical element for tuning immune recognition of small molecules, as it affects the presentation of the target antigen and thus the quality and selectivity of the response. Here, we report the synthesis and optimization of carrier protein and conjugation conditions for an initial hapten, norketamine-N-COOH (NK-N-COOH), to optimize vaccination conditions and assess the functional consequences of such vaccination on ketamine-induced behavioral alterations occurring at dissociative-like (50 mg/kg) doses. Iterating from this initial approach, two additional haptens, ketamine-N-COOH (KET-N-COOH) and 6-hydroxynorketamine-N-COOH (HNK-N-COOH), were synthesized to target either ketamine or 6-hydroxynorketamine with greater selectivity. The ability of these haptens to generate antiketamine, antinorketamine, and anti-6-hydroxynorketamine immune responses in mice was then assessed using enzyme-linked immunosorbent assay (ELISA) and competitive surface plasmon resonance (SPR) methods. All three haptens provoked immune responses in vivo, although the KET-N-COOH and 6-HNK-N-COOH haptens yielded antibodies with 5- to 10-fold improvements in affinity for ketamine and/or 6-hydroxynorketamine, as compared to NK-N-COOH. Regarding selectivity, vaccines bearing a KET-N-COOH hapten yielded an antibody response with approximately equivalent K d values against ketamine (86.4 ± 3.2 nM) and 6-hydroxynorketamine (74.1 ± 7.8 nM) and a 90-fold weaker K d against norketamine. Contrastingly, 6-HNK-N-COOH generated the highest affinity and most selective antibody profile, with a 38.3 ± 4.7 nM IC 50 against 6-hydroxynorketamine; K d values for ketamine and norketamine were 33- to 105-fold weaker, at 1290 ± 281.5 and 3971 ± 2175 nM, respectively. Overall, these findings support the use of rational hapten design to generate antibodies capable of distinguishing between structurally related, yet mechanistically distinct, compounds arising from the same precursor molecule. As applied to the production of the first-reported anti-6-hydroxynorketamine antibodies to date, this approach demonstrates a promising path forward for identifying the individual and combinatorial roles of ketamine and its metabolites in supporting rewarding effects and/or rapid-acting antidepressant activity.
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Production of 6-HNKs is important for the full antidepressant-like behavioral effects of ketamine.
“Production of the 6-HNKs is important for the full antidepressant-like behavioral effects of ketamine (Zanos, Highland, Liu, et al., 2019; Zanos et al., 2016; Zheng et al., 2021).”
From Mechanisms of ketamine and its metabolites as antidepressants (2022) · cites this paper as Zheng et al., 2021
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Cites
NMDAR inhibition-independent antidepressant actions of ketamine metabolites
Zanos, P., Moaddel, R., Morris, P. J., Georgiou, P., Fischell, J., Elmer, G. I., Alkondon, M., Yuan, P., Pribut, H. J., Singh, N. S., et al. (2016) · Nature · doi:10.1038/nature17998 · Cited by 1,609 · see citing works
Ketamine and Ketamine Metabolite Pharmacology: Insights into Therapeutic Mechanisms
Zanos, P., Moaddel, R., Morris, P. J., Riggs, L. M., Highland, J. N., Georgiou, P., Pereira, E. F. R., Albuquerque, E. X., Thomas, C. J., Zarate, C. A., et al. (2018) · Pharmacological Reviews · doi:10.1124/pr.117.015198 · Cited by 1,260 · see citing works
Ketamine: A Review of Clinical Pharmacokinetics and Pharmacodynamics in Anesthesia and Pain Therapy
Peltoniemi, M. A., Hagelberg, N. M., Olkkola, K. T., & Saari, T. I. (2016) · Clinical Pharmacokinetics · doi:10.1007/s40262-016-0383-6 · Cited by 672 · see citing works
Harper, L., Powell, J., & Pijl, E. M. (2017) · Harm Reduction Journal · doi:10.1186/s12954-017-0179-5 · Cited by 339 · see citing works
Deep posteromedial cortical rhythm in dissociation
Vesuna, S., Kauvar, I. V., Richman, E., Gore, F., Oskotsky, T., Sava-Segal, C., Luo, L., Malenka, R. C., Henderson, J. M., Nuyujukian, P., et al. (2020) · Nature · doi:10.1038/s41586-020-2731-9 · Cited by 281 · see citing works
Convergent Mechanisms Underlying Rapid Antidepressant Action
Zanos, P., Thompson, S. M., Duman, R. S., Zarate, C. A., & Gould, T. D. (2018) · CNS Drugs · doi:10.1007/s40263-018-0492-x · Cited by 171 · see citing works
Yang, C., Qu, Y., Abe, M., Nozawa, D., Chaki, S., & Hashimoto, K. (2016) · Biological Psychiatry · doi:10.1016/j.biopsych.2016.12.020 · Cited by 162 · see citing works
Lumsden, E. W., Troppoli, T. A., Myers, S. J., Zanos, P., Aracava, Y., Kehr, J., Lovett, J., Kim, S., Wang, F.-H., Schmidt, S., et al. (2019) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1816071116 · Cited by 158 · see citing works
Zanos, P., Highland, J. N., Stewart, B. W., Georgiou, P., Jenne, C. E., Lovett, J., Morris, P. J., Thomas, C. J., Moaddel, R., Zarate, C. A., et al. (2019) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1819540116 · Cited by 152 · see citing works
Bremer, P. T., Kimishima, A., Schlosburg, J. E., Zhou, B., Collins, K. C., & Janda, K. D. (2016) · Angewandte Chemie International Edition · doi:10.1002/anie.201511654 · Cited by 138 · see citing works
Parkin, M. C., Turfus, S. C., Smith, N. W., Halket, J. M., Braithwaite, R. A., Elliott, S. P., Osselton, M. D., Cowan, D., & Kicman, A. T. (2008) · Journal of Chromatography B · doi:10.1016/j.jchromb.2008.09.036 · Cited by 109 · see citing works
Ketamine Use for Cancer and Chronic Pain Management
Culp, C. J., Kim, H. K., & Abdi, S. (2021) · Frontiers in Pharmacology · doi:10.3389/fphar.2020.599721 · Cited by 98 · see citing works
Impact of Distinct Chemical Structures for the Development of a Methamphetamine Vaccine
Moreno, A. Y., Mayorov, A. V., & Janda, K. D. (2011) · Journal of the American Chemical Society · doi:10.1021/ja108807j · Cited by 80 · see citing works
Orson, F. M., Kinsey, B. M., Singh, R. A. K., Wu, Y., Gardner, T., & Kosten, T. R. (2008) · Annals of the New York Academy of Sciences · doi:10.1196/annals.1441.027 · Cited by 79 · see citing works
Monoclonal Antibodies for Combating Synthetic Opioid Intoxication
Smith, L. C., Bremer, P. T., Hwang, C. S., Zhou, B., Ellis, B. A., Hixon, M. S., & Janda, K. D. (2019) · Journal of the American Chemical Society · doi:10.1021/jacs.9b04872 · Cited by 70 · see citing works
Rudra, J. S., Ding, Y., Neelakantan, H., Ding, C., Appavu, R., Stutz, S., Snook, J. D., Chen, H., Cunningham, K. A., & Zhou, J. (2016) · ACS Chemical Neuroscience · doi:10.1021/acschemneuro.5b00345 · Cited by 58 · see citing works
Baruffaldi, F., Kelcher, A. M. H., Laudenbach, M., Gradinati, V., Limkar, A. R., Roslawski, M. J., Birnbaum, A. K., Lees, A., Haßler, C., Runyon, S. P., et al. (2018) · Molecular Pharmaceutics · doi:10.1021/acs.molpharmaceut.8b00592 · Cited by 56 · see citing works
Development of active and passive human vaccines to treat methamphetamine addiction
Gentry, W. B., Rüedi‐Bettschen, D., & Owens, S. M. (2009) · Human Vaccines · doi:10.4161/hv.5.4.7456 · Cited by 54 · see citing works
Hwang, C. S., Bremer, P. T., Wenthur, C. J., Ho, S. O., Chiang, S., Ellis, B. A., Zhou, B., Fujii, G., & Janda, K. D. (2018) · Molecular Pharmaceutics · doi:10.1021/acs.molpharmaceut.7b00933 · Cited by 50 · see citing works
Self-citationMatyas, G. R., Rice, K. C., Cheng, K., Li, F., Antoline, J. F. G., Iyer, M. R., Jacobson, A. E., Mayorov, A. V., Beck, Z., Torres, O. B., et al. (2014) · Vaccine · doi:10.1016/j.vaccine.2014.01.028 · Cited by 48 · see citing works
Corkery, J., Hung, W.-C., Claridge, H., Goodair, C., Copeland, C. S., & Schifano, F. (2021) · Journal of Psychopharmacology · doi:10.1177/02698811211021588 · Cited by 48 · see citing works
Strategies to Prolong Ketamine’s Efficacy in Adults with Treatment-Resistant Depression
McMullen, E. P., Lee, Y. L., Lipsitz, O., Lui, L. M. W., Vinberg, M., Ho, R., Rodrigues, N. B., Rosenblat, J. D., Cao, B., Gill, H., et al. (2021) · Advances in Therapy · doi:10.1007/s12325-021-01732-8 · Cited by 44 · see citing works
Antidepressant Actions of Ketamine Versus Hydroxynorketamine
Collingridge, G. L., Lee, Y., Bortolotto, Z. A., Kang, H., & Lodge, D. (2016) · Biological Psychiatry · doi:10.1016/j.biopsych.2016.06.029 · Cited by 41 · see citing works
A Conjugate Vaccine Using Enantiopure Hapten Imparts Superior Nicotine-Binding Capacity
Lockner, J. W., Lively, J. M., Collins, K. C., Vendruscolo, J. C. M., Azar, M. R., & Janda, K. D. (2014) · Journal of Medicinal Chemistry · doi:10.1021/jm501625j · Cited by 37 · see citing works
Ketamine Metabolite (2<i>R</i>,6<i>R</i>)-Hydroxynorketamine Interacts with μ and κ Opioid Receptors
Joseph, T. T., Bu, W., Lin, W., Zoubak, L., Yeliseev, A., Liu, R., Eckenhoff, R. G., & Brannigan, G. (2021) · ACS Chemical Neuroscience · doi:10.1021/acschemneuro.0c00741 · Cited by 28 · see citing works
Rapid and sensitive detection of ketamine in blood using novel fluorescence genosensor
Ding, Y., Li, X.-M., Guo, Y.-D., Yan, J., Ling, J., Li, W., Lan, L.-M., Chang, Y.-F., Cai, J.-F., & Zha, L. (2017) · Analytical and Bioanalytical Chemistry · doi:10.1007/s00216-017-0650-x · Cited by 27 · see citing works
Negrusz, A., Adamowicz, P., Saini, B. K., Webster, D. E., Juhascik, M. P., Moore, C. M., & Schlemmer, R. F. (2005) · Journal of Analytical Toxicology · doi:10.1093/jat/29.3.163 · Cited by 24 · see citing works
Is Metabolism of (R)-Ketamine Essential for the Antidepressant Effects?
Chaki, S. (2017) · The International Journal of Neuropsychopharmacology · doi:10.1093/ijnp/pyx120 · Cited by 17 · see citing works
Broadly Neutralizing Synthetic Cannabinoid Vaccines
Lin, M., Lee, J. C., Blake, S., Ellis, B. A., Eubanks, L. M., & Janda, K. D. (2020) · JACS Au · doi:10.1021/jacsau.0c00057 · Cited by 16 · see citing works
Torres, O. B., Duval, A. J., Sulima, A., Antoline, J. F. G., Jacobson, A. E., Rice, K. C., Alving, C. R., & Matyas, G. R. (2018) · Analytical and Bioanalytical Chemistry · doi:10.1007/s00216-018-1060-4 · Cited by 15 · see citing works
Vaccine-driven pharmacodynamic dissection and mitigation of fenethylline psychoactivity
Wenthur, C. J., Zhou, B., & Janda, K. D. (2017) · Nature · doi:10.1038/nature23464 · Cited by 14 · see citing works
Self-citationLanger, N., Steinicke, F., Lindigkeit, R., Ernst, L., & Beuerle, T. (2017) · Forensic Science International · doi:10.1016/j.forsciint.2017.09.011 · Cited by 14 · see citing works
A Vision for Vaccines: Combating the Opioid Epidemic
Hwang, C. S., & Janda, K. D. (2017) · Biochemistry · doi:10.1021/acs.biochem.7b00948 · Cited by 9 · see citing works
Ketamine Metabolism: Identification and Synthesis of a Deaminated Product
Lai, I., Hong, S.-C., & Davisson, J. N. (1985) · Journal of Pharmaceutical Sciences · doi:10.1002/jps.2600740425 · Cited by 2 · see citing works
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1 of the 35 references couldn’t be matched to a known paper.
Research works printed but not linked: 1
- [9]Orhurhu VJ, Vashisht R, Claus LE, Ketamine Toxicity. [Updated 2020 Nov 15]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2021 Jan-. Available from: https://www.ncbi.nlm.nih.gov/books/NBK541087/. 31082131
Cited by
Sorted by citations
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Hess, E. M., Riggs, L. M., Michaelides, M., & Gould, T. D. (2022) · Biochemical Pharmacology · doi:10.1016/j.bcp.2021.114892 · Cited by 169 · see citing works
Hanson, E. K., & Whelan, R. J. (2023) · Sensors · doi:10.3390/s23104831 · Cited by 15 · see citing works
Zheng, Z., Ma, M., Jia, Y., Cui, Y., Zhao, R., Li, S., Wenthur, C. J., Li, L., & Li, G. (2023) · Analytical Chemistry · doi:10.1021/acs.analchem.3c00223 · Cited by 11 · see citing works
Self-citationCelik, M. A., & Fuehrlein, B. (2022) · ImmunoTargets and Therapy · doi:10.2147/itt.s370435 · Cited by 10 · see citing works
Wang, K., Luo, J., Wang, H., & Wang, X. (2025) · European Journal of Medicinal Chemistry · doi:10.1016/j.ejmech.2025.117274 · Cited by 2 · see citing works
Zhang, Y., Guo, Y., Qi, Y., & Chen, L. (2023) · Indian Journal of Pharmaceutical Sciences · doi:10.36468/pharmaceutical-sciences.1222 · Cited by 1 · see citing works
Ketamine in Prehospital Emergency Medicine
Schober, P. R., & Schwarte, L. A. (2025) · Neuromethods · doi:10.1007/978-1-0716-4603-8_22
Generation of enantiospecific monoclonal antibodies against (2R,6R)-hydroxynorketamine.
Enriquez, U. M., Velázquez, N. M. G., Hossain, M. M., Kyzer, J. L., Guzei, I. A., & Wenthur, C. J. (2026) · Bioorganic & Medicinal Chemistry · doi:10.1016/j.bmc.2026.118573
Self-citation
- Authors
- Zheng, Z., Kyzer, J. L., Worob, A., & Wenthur, C. J.
- Affiliations
- University of Wisconsin–Madison (US)
- Corresponding author
- Wenthur, Cody J.
- Venue
- ACS Chemical Neuroscience
- Publisher
- American Chemical Society (ACS)
- Published
- 2021-10-15
- Volume
- 12
- Issue
- 21
- Pages
- 4113-4122
- ISSN
- 1948-7193 (ISSN-L)
- PMC
- PMC9358770
- OpenAlex
- W3205594036
- Cited by
- 8
- References
- 35
- Citations by year
- 2026: 1 · 2025: 2 · 2023: 3 · 2022: 1 · 2021: 1
- FWCI
- 0.66
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- ACS Chemical Neuroscience (publishedVersion, cc-by-nc-nd)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- Funders
- National Institute of Mental Health (R01MH122742)
- Topics
- Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Monoclonal and Polyclonal Antibodies Research
- Health Sciences › Medicine › Pharmacology › Treatment of Major Depression
- Physical Sciences › Chemistry › Organic Chemistry › Synthesis and pharmacology of benzodiazepine derivatives
- Keywords
- ketamine, norketamine, hapten design, bioconjugates, immunopharmacotherapy, antibody selectivity, antibody affinity, vaccine immunization, dissociative states, blood-brain barrier
- MeSH terms
- Animals, Antidepressive Agents, Ketamine, Mice
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