title: An in vitro and in vivo evaluation of the efficacy of recombinant human liver prolidase as a catalytic bioscavenger of chemical warfare nerve agents.
abstract: In this study, we determined the ability of recombinant human liver prolidase to hydrolyze nerve agents in vitro and its ability to afford protection in vivo in mice. Using adenovirus containing the human liver prolidase gene, the enzyme was over expressed by 200- to 300-fold in mouse liver and purified to homogeneity by affinity and gel filtration chromatography. The purified enzyme hydrolyzed sarin, cyclosarin and soman with varying rates of hydrolysis. The most efficient hydrolysis was with sarin, followed by soman and by cyclosarin {apparent kcat/Km [(1.9 ± 0.3), (1.7 ± 0.2), and (0.45 ± 0.04)] × 10(5 )M(-1 )min(-1), respectively}; VX and tabun were not hydrolyzed by the recombinant enzyme. The enzyme hydrolyzed P (+) isomers faster than the P (-) isomers. The ability of recombinant human liver prolidase to afford 24 hour survival against a cumulative dose of 2 × LD50 of each nerve agent was investigated in mice. Compared to mice injected with a control virus, mice injected with the prolidase expressing virus contained (29 ± 7)-fold higher levels of the enzyme in their blood on day 5. Challenging these mice with two consecutive 1 × LD50 doses of sarin, cyclosarin, and soman resulted in the death of all animals within 5 to 8 min from nerve agent toxicity. In contrast, mice injected with the adenovirus expressing mouse butyrylcholinesterase, an enzyme which is known to afford protection in vivo, survived multiple 1 × LD50 challenges of these nerve agents and displayed no signs of toxicity. These results suggest that, while prolidase can hydrolyze certain G-type nerve agents in vitro, the enzyme does not offer 24 hour protection against a cumulative dose of 2 × LD50 of G-agents in mice in vivo.
authors.affiliation: Physiology & Immunology Branch, Research Division, US Army Medical Research Institute of Chemical Defense , Aberdeen Proving Ground , MD, USA.
authors.forename: Peter E
authors.initials: PE
authors.lastname: Rezk
authors.forename: Pierre
authors.initials: P
authors.lastname: Zdenka
authors.forename: Praveena
authors.initials: P
authors.lastname: Sabnekar
authors.forename: Takwen
authors.initials: T
authors.lastname: Kajih
authors.forename: David G
authors.initials: DG
authors.lastname: Mata
authors.forename: Chester
authors.initials: C
authors.lastname: Wrobel
authors.forename: Douglas M
authors.initials: DM
authors.lastname: Cerasoli
authors.forename: Nageswararao
authors.initials: N
authors.lastname: Chilukuri
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D055162:Biocatalysis; D002091:Butyrylcholinesterase / Q000235:genetics / Q000494:pharmacology; D002619:Chemical Warfare Agents / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity*; D004150:Dipeptidases / Q000097:blood / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology*; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D006868:Hydrolysis; D066298:In Vitro Techniques; D007928:Lethal Dose 50; D008099:Liver / Q000201:enzymology*; D008297:Male; D051379:Mice; D011994:Recombinant Proteins / Q000097:blood / Q000235:genetics / Q000494:pharmacology
chemical_list: D002619:Chemical Warfare Agents; D011994:Recombinant Proteins; D002091:Butyrylcholinesterase; D004150:Dipeptidases; C022449:proline dipeptidase
keywords: Adenovirus; butyrylcholinesterase; catalytic bioscavenger; chemical warfare nerve agents; human liver prolidase; in vivo delivery