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Quantitative evaluation of liver-specific promoters from retroviral vectors
after in vivo transduction of hepatocytes
DG Hafenrichter, X Wu, SD Rettinger, SC Kennedy, MW Flye and KP Ponder
Department of Surgery, Washington University School of Medicine, St Louis,
MO.
Hepatic gene therapy could be used to treat a number of inherited blood
diseases such as hemophilia or thrombophilia. Although liver-directed
retroviral transduction can result in long-term gene expression in vivo,
the low level of protein production has limited its clinical application.
We reasoned that the insertion of liver-specific promoters into retroviral
vectors would increase gene expression in vivo. The 347- bp human alpha
1-antitrypsin (hAAT), the 810-bp murine albumin (mAIb), the 490-bp rat
phosphoenolpyruvate carboxykinase (rPECK), and the 596- bp rat liver fatty
acid binding protein promoters were inserted into a Moloney murine leukemia
retroviral backbone containing the hAAT reporter gene. Vectors that
produced appropriately sized RNA and hAAT protein in vitro were tested in
vivo by transducing regenerating rat livers. Long-term serum expression of
the hAAT reporter gene was normalized to retroviral transduction efficiency
as determined by using a polymerase chain reaction-based assay of genomic
DNA from transduced rat livers. The hAAT, mAIb, and rPEPCK promoters were,
respectively, 35- , 8-, and 0.02-fold as strong as the previously studied
constitutive Pol-II promoter. We conclude that the hAAT promoter resulted
in the highest expression from a retroviral vector and may result in
therapeutically significant expression of other clinically significant
blood proteins.
Volume 84,
Issue 10,
pp. 3394-3404,
11/15/1994
Copyright © 1994 by The American Society of Hematology
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