Visualizing viral transduction of a cocaine-hydrolyzing, human butyrylcholinesterase in rats.
Human plasma butyrylcholinesterase (BChE) is essential for cocaine detoxification even though its catalytic efficiency for that substrate is relatively poor. Site-directed mutagenesis of this protein has recently been used to obtain much-improved cocaine esterases, one of which we designate CocE. We previously showed that adenoviral transduction of such esterases caused up to 50,000-fold increases in circulating cocaine hydrolase activity, led to drastically shortened cocaine half-life, and blunted the cardiovascular responses to cocaine in rats. In those experiments, gene transduction of cocaine esterase was sustained at high levels for up to 1 week but then declined steeply. Our eventual goal is to use long-term esterase expression as a means of reducing drug reward and extinguishing intake in models of cocaine-addiction. Therefore, we investigated the site of enzyme transduction for clues to the local reactions that may limit the duration of CocE expression. Histological and immunohistochemical observations demonstrated that hepatocytes were the primary focus for transduction of modified human BChE. Rats were administered 2.2 x 10(10) plaque forming units of a replication-incompetent, type-5 adenoviral vector incorporating CocE cDNA. Within days the livers showed intense thiocholine staining for BChE activity. Selective immunohistochemistry for human BChE proved that this activity represented CocE transgene. By 5 days, however, pockets of mononuclear cells had invaded the hepatic parenchyma, and a meshwork of IgM-like immunoreactivity had lined the hepatic sinusoids. These phenomena probably represent early responses of the immune system, either to the transduced CocE or to the hepatocytes producing this protein.
Gao
Yang
Y
Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic, 200 First St. SW, Rochester MN 55905, USA.
Brimijoin
Stephen
S
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D002091:Butyrylcholinesterase / Q000235:genetics* / Q000378:metabolism*; D002265:Carboxylic Ester Hydrolases / Q000378:metabolism; D003042:Cocaine / Q000378:metabolism*; D005822:Genetic Vectors / Q000235:genetics; D022781:Hepatocytes / Q000378:metabolism / Q000821:virology; D006801:Humans; D006868:Hydrolysis; D007150:Immunohistochemistry; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000378:metabolism; D014161:Transduction, Genetic
D011994:Recombinant Proteins; D002265:Carboxylic Ester Hydrolases; C498183:CocE protein; D002091:Butyrylcholinesterase; D003042:Cocaine