Next generation OP-bioscavengers: a circulatory long-lived 4-PEG hypolysine mutant of F338A-HuAChE with optimal pharmacokinetics and pseudo-catalytic characteristics.
We have shown previously that conjugation of polyethylene glycol (PEG) chains to recombinant human acetylcholinesterase (rHuAChE) results in the extension of its residence time in the circulation of mice and monkeys [1,2]. By profiling the pharmacokinetic behavior of an array of well-defined hypolysine human mutant AChE molecules following PEGylation, we now determine that the duration of these enzyme forms in the circulation of rhesus macaques correlates with their number of appended PEG moieties, and is influenced by the actual location of the PEG chains at the molecule surface, as well. These findings, which concur with those we have previously established in mice, indicate that a common set of rules dictates the circulatory fate of PEGylated HuAChEs in rodents and non-human primates. In addition to its effect on circulatory residence, PEGylation reduces the ability of the rHuAChE bioscavenger to elicit an immune response in the heterologous mouse animal system. Thus, an inverse relationship between anti-AChE antibody production and PEG loading was observed following repeated administration of the different PEGylated hypolysine human AChEs to mice. We note however, that in rhesus macaques, the essentially homologous (human) AChE does not induce specific anti-AChE antibodies after repeated administration of high doses of the enzyme in its PEGylated form, and even in its non-PEGylated form. Taken together, these findings indicate that PEG acts by veiling enzyme-related epitopes, which would otherwise interact with host circulatory elimination pathways and immune system. The barring of such interactions by obstructive PEGs, confers the enzyme molecule with both extended circulatory residence and mitigated immunogenic properties. Further modulation by incorporation of the F338A mutation into the PEGylated hypolysine rHuAChE enzyme mold, resulted in the generation of an OP-bioscavenger that displayed reduced aging rates and could effectively protect mice against repeated exposure to CW agents. This selected 4-PEG F338A-AChE can serve as a paradigm for new generation OP-bioscavengers, specifically tailored for prophylactic treatment against toxic OP-agents.
Kronman
Chanoch
C
Department of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, 24 Rueven St., P.O. Box 19, Ness-Ziona 74100, Israel.
Cohen
Ofer
O
Mazor
Ohad
O
Ordentlich
Arie
A
Raveh
Lily
L
Velan
Baruch
B
Shafferman
Avigdor
A
D000110:Acetylcholinesterase / Q000097:blood / Q000235:genetics / Q000378:metabolism* / Q000493:pharmacokinetics*; D000818:Animals; D000931:Antidotes / Q000378:metabolism / Q000493:pharmacokinetics; D055162:Biocatalysis; D002460:Cell Line; D000939:Epitopes / Q000276:immunology; D005260:Female; D006801:Humans; D008239:Lysine; D008253:Macaca mulatta; D008297:Male; D051379:Mice; D009154:Mutation; D009943:Organophosphorus Compounds / Q000378:metabolism*; D011092:Polyethylene Glycols / Q000737:chemistry*; D011994:Recombinant Proteins / Q000097:blood / Q000235:genetics / Q000378:metabolism / Q000493:pharmacokinetics
D000931:Antidotes; D000939:Epitopes; D009943:Organophosphorus Compounds; D011994:Recombinant Proteins; D011092:Polyethylene Glycols; D000110:Acetylcholinesterase; D008239:Lysine