Treatment of acute organophosphate poisoning by using a cocaine hydrolase engineered from human butyrylcholinesterase

Johnathan E. LeSaint; Shurong Hou; Nellore Bhanu Chandar; Annet Kyomuhangi; Huimei Wei; Fang Zheng; Chang Guo Zhan

doi:10.1016/j.cbi.2025.111552

Treatment of acute organophosphate poisoning by using a cocaine hydrolase engineered from human butyrylcholinesterase

Johnathan E. LeSaint
, Shurong Hou
, Nellore Bhanu Chandar
, Annet Kyomuhangi
, Huimei Wei
, Fang Zheng
, Chang Guo Zhan

Research output: Contribution to journal › Article › peer-review

5 Scopus citations

Abstract

Organophosphate (OP) chemical warfare nerve agents and pesticides are potent, irreversible inhibitors of acetylcholinesterase (AChE), and paraoxon is often used as a surrogate compound in the studies of OP poisoning. For a truly effective treatment of OP poisoning, it is desirable that a protein-based OP bioscavenger can react with OP significantly faster than AChE reacting with OP to protect AChE from further inhibition reaction with OP. In the present study, our in vitro reactivity assays revealed that CocH3-Fc(M3), a potent cocaine hydrolase engineered from human butyrylcholinesterase (BChE), has a ∼20-fold improved bimolecular rate constant for the reaction with paraoxon compared to wild-type BChE. Due to the improved in vitro reactivity with paraoxon, CocH3-Fc(M3) at a modest dose of 25 mg/kg was able to effectively rescue all mice that had been injected with a lethal dose of 0.66 mg/kg paraoxon and accelerate the recovery of the mice from paraoxon-induced toxicity symptoms. All the in silico, in vitro, and in vivo data consistently suggest that CocH3-Fc(M3) can be used to effectively detoxify paraoxon.

Original language	English
Article number	111552
Journal	Chemico-Biological Interactions
Volume	416
DOIs	https://doi.org/10.1016/j.cbi.2025.111552
State	Published - Aug 1 2025

Bibliographical note

Publisher Copyright:
© 2025 Elsevier B.V.

Funding

This work was supported in part by the National Institutes of Health (NIH grants UG3/UH3 NS134920, U01 DA051079, UH2/UH3 DA041115, R01 DA056646, U18 DA052319, R01 DA035552, R01 DA032910, and R01 DA013930) and the Molecular Modeling and Biopharmaceutical Center (MMBC).The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:Chang-Guo Zhan reports financial support was provided by National Institutes of Health. Fang Zheng reports a relationship with National Institutes of Health that includes: funding grants. Chang-Guo Zhan has patent #Cocaine Hydrolase-Fc Fusion Proteins for Cocaine and Methods for Utilizing the Same (U.S. Patent No. 10, 772, 940); Increased polypeptide production yields of butyrylcholinesterase polypeptides for therapeutic use (U.S. Patent No. 11, 473, 070). Issued to Not yet licensed. The patents are owned by the University of Kentucky. Fang Zheng has patent #Cocaine Hydrolase-Fc Fusion Proteins for Cocaine and Methods for Utilizing the Same (U.S. Patent No. 10, 772, 940); Increased polypeptide production yields of butyrylcholinesterase polypeptides for therapeutic use (U.S. Patent No. 11, 473, 070). Issued to Not yet licensed. The patents are owned by the University of Kentucky. None. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. This work was supported in part by the National Institutes of Health (NIH grants UG3/UH3 NS134920 , U01 DA051079 , UH2/UH3 DA041115 , R01 DA056646 , U18 DA052319 , R01 DA035552 , R01 DA032910 , and R01 DA013930 ) and the Molecular Modeling and Biopharmaceutical Center (MMBC) .

Funders	Funder number
University of Kentucky
National Institutes of Health (NIH)	R01 DA035552, UG3/UH3 NS134920, R01 DA013930, U01 DA051079, UH2/UH3 DA041115, R01 DA056646, R01 DA032910, U18 DA052319
Molecular Modeling and Biopharmaceutical Center	772, 940, 070

Keywords

Cocaine hydrolase
Esterase
Organophosphate
Organophosphate poisoning
Phosphorylation

ASJC Scopus subject areas

Toxicology

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10.1016/j.cbi.2025.111552

Cite this

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title = "Treatment of acute organophosphate poisoning by using a cocaine hydrolase engineered from human butyrylcholinesterase",

abstract = "Organophosphate (OP) chemical warfare nerve agents and pesticides are potent, irreversible inhibitors of acetylcholinesterase (AChE), and paraoxon is often used as a surrogate compound in the studies of OP poisoning. For a truly effective treatment of OP poisoning, it is desirable that a protein-based OP bioscavenger can react with OP significantly faster than AChE reacting with OP to protect AChE from further inhibition reaction with OP. In the present study, our in vitro reactivity assays revealed that CocH3-Fc(M3), a potent cocaine hydrolase engineered from human butyrylcholinesterase (BChE), has a ∼20-fold improved bimolecular rate constant for the reaction with paraoxon compared to wild-type BChE. Due to the improved in vitro reactivity with paraoxon, CocH3-Fc(M3) at a modest dose of 25 mg/kg was able to effectively rescue all mice that had been injected with a lethal dose of 0.66 mg/kg paraoxon and accelerate the recovery of the mice from paraoxon-induced toxicity symptoms. All the in silico, in vitro, and in vivo data consistently suggest that CocH3-Fc(M3) can be used to effectively detoxify paraoxon.",

keywords = "Cocaine hydrolase, Esterase, Organophosphate, Organophosphate poisoning, Phosphorylation",

author = "LeSaint, \{Johnathan E.\} and Shurong Hou and Chandar, \{Nellore Bhanu\} and Annet Kyomuhangi and Huimei Wei and Fang Zheng and Zhan, \{Chang Guo\}",

note = "Publisher Copyright: {\textcopyright} 2025 Elsevier B.V.",

year = "2025",

month = aug,

day = "1",

doi = "10.1016/j.cbi.2025.111552",

language = "English",

volume = "416",

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T1 - Treatment of acute organophosphate poisoning by using a cocaine hydrolase engineered from human butyrylcholinesterase

AU - LeSaint, Johnathan E.

AU - Hou, Shurong

AU - Chandar, Nellore Bhanu

AU - Kyomuhangi, Annet

AU - Wei, Huimei

AU - Zheng, Fang

AU - Zhan, Chang Guo

PY - 2025/8/1

Y1 - 2025/8/1

N2 - Organophosphate (OP) chemical warfare nerve agents and pesticides are potent, irreversible inhibitors of acetylcholinesterase (AChE), and paraoxon is often used as a surrogate compound in the studies of OP poisoning. For a truly effective treatment of OP poisoning, it is desirable that a protein-based OP bioscavenger can react with OP significantly faster than AChE reacting with OP to protect AChE from further inhibition reaction with OP. In the present study, our in vitro reactivity assays revealed that CocH3-Fc(M3), a potent cocaine hydrolase engineered from human butyrylcholinesterase (BChE), has a ∼20-fold improved bimolecular rate constant for the reaction with paraoxon compared to wild-type BChE. Due to the improved in vitro reactivity with paraoxon, CocH3-Fc(M3) at a modest dose of 25 mg/kg was able to effectively rescue all mice that had been injected with a lethal dose of 0.66 mg/kg paraoxon and accelerate the recovery of the mice from paraoxon-induced toxicity symptoms. All the in silico, in vitro, and in vivo data consistently suggest that CocH3-Fc(M3) can be used to effectively detoxify paraoxon.

AB - Organophosphate (OP) chemical warfare nerve agents and pesticides are potent, irreversible inhibitors of acetylcholinesterase (AChE), and paraoxon is often used as a surrogate compound in the studies of OP poisoning. For a truly effective treatment of OP poisoning, it is desirable that a protein-based OP bioscavenger can react with OP significantly faster than AChE reacting with OP to protect AChE from further inhibition reaction with OP. In the present study, our in vitro reactivity assays revealed that CocH3-Fc(M3), a potent cocaine hydrolase engineered from human butyrylcholinesterase (BChE), has a ∼20-fold improved bimolecular rate constant for the reaction with paraoxon compared to wild-type BChE. Due to the improved in vitro reactivity with paraoxon, CocH3-Fc(M3) at a modest dose of 25 mg/kg was able to effectively rescue all mice that had been injected with a lethal dose of 0.66 mg/kg paraoxon and accelerate the recovery of the mice from paraoxon-induced toxicity symptoms. All the in silico, in vitro, and in vivo data consistently suggest that CocH3-Fc(M3) can be used to effectively detoxify paraoxon.

KW - Cocaine hydrolase

KW - Esterase

KW - Organophosphate

KW - Organophosphate poisoning

KW - Phosphorylation

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Treatment of acute organophosphate poisoning by using a cocaine hydrolase engineered from human butyrylcholinesterase

Abstract

Bibliographical note

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Keywords

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