Selective repopulation of normal mouse liver by hepatocytes transduced in vivo with recombinant adeno-associated virus.
The use of recombinant adeno-associated virus (rAAV) as gene therapy vector for treating liver metabolic diseases is limited by its low transduction efficiency. We describe a strategy for achieving stable and efficient genetic reconstitution in liver after direct administration of rAAV and selective expansion of transduced cells. We have exploited the biology of apoptosis to develop a generic approach for selectively repopulating liver with vector-transduced hepatocytes. Low-level, stable transduction of hepatocytes was achieved by direct injection of rAAV into mouse liver. Expansion of these vector-transduced cells was achieved by incorporating into the construct a minigene expressing Bcl-2 followed by induction of apoptosis in non-vector-containing hepatocytes by systemic administration of a Fas antibody (Ab). Western and Southern blot analysis demonstrated amplification of bcl-2 gene product and viral copy number, respectively, in vector-treated mouse liver when placed under selection. In addition, the percentage of vector transduced cells increased from 2 to 20% after three administrations of Fas Ab, based on immunohistochemical studies.
Chen
S J
SJ
Institute for Human Gene Therapy, Department of Medicine and Molecular and Cellular Engineering, University of Pennsylvania, Philadelphia, PA 19104, USA.
Tazelaar
J
J
Wilson
J M
JM
D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D017690:Cell Transplantation / Q000379:methods*; D000229:Dependovirus / Q000235:genetics*; D016654:Genes, RAG-1 / Q000235:genetics; D019254:Genes, bcl-2; D022781:Hepatocytes / Q000502:physiology* / Q000821:virology; D006801:Humans; D008099:Liver / Q000166:cytology*; D051379:Mice; D008822:Mice, Transgenic; D011994:Recombinant Proteins / Q000378:metabolism; D012641:Selection, Genetic; D019014:fas Receptor / Q000378:metabolism*
D011994:Recombinant Proteins; D019014:fas Receptor