Large-scale production of human serum butyrylcholinesterase as a bioscavenger

Abstract

Disclosed herein are methods for the large-scale preparation of human butyrylcholinesterase (HuBChE) preparations from Cohn Fraction IV-4. As disclosed, the methods provide HuBChE preparations that are about 99% or more pure with recovery yields of about 60%. Also disclosed are the pharmacokinetics, safety and toxicity, stability and efficacy of the HuBChE preparations.

Description

CROSS REFERENCE TO RELATED APPLICATIONS [0001] This application claims the benefit of U.S. Provisional Patent Application No. 60 / 618,064, filed Oct. 9, 2004, pending, and is herein incorporated by reference in its entirety.ACKNOWLEDGEMENT OF GOVERNMENT SUPPORT [0002] Employees of the United States Army made this invention. The government has rights in the invention.BACKGROUND OF THE INVENTION [0003] 1. Field of the Invention [0004] The present invention generally relates to a method for the large-scale production of human serum butyrylcholinesterase (HuBChE) from Cohn Fraction IV-4 paste. [0005] 2. Description of the Related Art [0006] Exposure to organophosphorus compounds (OPs) in the form of nerve agents and pesticides poses an ever increasing threat to military and civilian populations. The serious medical challenges posed by chemical warfare agents to both the military and civilian health care systems became evident in the Iran-Iraq conflict and the 1995 Tokyo subway incident, ...

AI Technical Summary

Benefits of technology

[0015] In some embodiments, the present invention provides a method of treating, preventing, or inhibiting toxicity to an organophosphorus compound in a subject which comprises administering to the subject the human butyrylcholinesterase preparation made by subjecting about 2 or more kilograms, preferably about 10 to 500 kilograms, more preferably about 100 to 500 kilograms, most preferably about 300 to 500 kilograms, of Cohn Fraction IV-4 paste to affinity chromatography followed by anion exchange chromatography. In some embodiments, the amount of Cohn Fraction IV-4 is about 80 kilograms or more, preferably about 80 kilograms. In some embodiments, the method comprises diluting the Cohn Fraction IV-4 paste by about ten-fold with water to obtain a suspension and then adjusting the suspension to a pH of about 4.5 to about 5.5, preferably about 4.7 to about 5.2, more preferably about 4.8 to about 5.0, most preferably about 4.9. In some embodiments, the method comprises centrifuging the suspension using a continuous flow centrifuge at about 7400 to about 7900 rpm, preferably about 7500 to about 7800 rpm, more preferably about 7600 to about 7700 rpm, most preferably about 7663 rpm at a flow rate of about 2 to about 6 kilograms per minute, preferably about 3 to about 5 kilograms per minute, more preferably about 4 kilograms per minute to obtain a supernatant. In some embodiments, the method comprises adjusting the supernatant to a pH of about 7.0 to about 9.0, preferably about 7.5 to about 8.5, more preferably about 8.0. In some embodiments, the method comprises filtering the supernatant with a 0.65 μm filter cartridge

Problems solved by technology

Exposure to organophosphorus compounds (OPs) in the form of nerve agents and pesticides poses an ever increasing threat to military and civilian populations.

The serious medical challenges posed by chemical warfare agents to both the military and civilian health care systems became evident in the Iran-Iraq conflict and the 1995 Tokyo subway incident, respectively.

The resultant increase in acetylcholine concentration manifests at the cholinergic synapses of both the peripheral and central nervous systems, which precipitates a cholinergic crisis characterized by miosis, increased tracheobronchial and salivary secretions, bronchoconstriction, bradycardia, fasciculations, behavioral inc

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