Hepatocyte-specific gene expression by a recombinant adeno-associated virus vector carrying the apolipoprotein E enhancer and 1-antitrypsin promoter Research Article

An adeno-associated virus vector was constructed to express exogenous genes to the l iver. The original plasmid construct carried two expression units; a neomycin resistant gene and human 1antitrypsin cDNA under the control of hepatocyte specific transcription elements. Cel ls were transfected with the constructed plasmid DNA with another packaging plasmid, and recombinant adeno-associated viruses (rAAV) were then recovered after adenovirus infection. Alternatively, rAAV were recovered by transduction of DNAs of the packaging plasmid and adenovirus into preselected cells carrying constructed proviral DNA. When the transducing abil it ies were evaluated based on G418 res is tant colony formation on HeLa ce l l s , the lat ter method was found to give almost 10-fold more rAAV. We then isolated G418 resistant colonies and established several independent clones for the HeLa and Hepa1A cells infected with the rAAV. All of the eight clones derived from Hepa1A cells produced significant amounts of the human 1-antitrypsin protein. In contrast, none of the f ive clones derived from HeLa cel ls produced a detectable level of 1antitrypsin. Our results suggest that liver-specific promoter and enhancer maintain the tissue specificity in the rAAV construct, and that the rAAV vector system would be useful in hepatocyte directed gene therapy.