Hepatocyte-targeted in vivo gene expression by intravenous injection of plasmid DNA complexed with synthetic multi-functional gene delivery system.
To achieve hepatocyte-targeted in vivo gene expression, a carrier that controls both the tissue and intracellular distribution of DNA was designed and synthesized. A cationic polymer, poly(L-ornithine) (pOrn), was modified first with galactose, then with a fusigenic peptide (mHA2) to obtain Gal-pOrn-mHA2. When applied with Gal-pOrn-mHA2 to asialoglycoprotein receptor-positive cells, fluorescein-labeled DNA showed a diffuse profile, suggesting the release of DNA from endosomes and/or lysosomes by the carrier. Then the biodistribution and gene expression after intravenous injection of DNA complexes (10 microg DNA per mouse) were examined. After injection of [32P]DNA/Gal-pOrn-mHA2, about 60% of the radioactivity was recovered in the liver, mostly in parenchymal cells. A large amount (81 ng/g tissue) of transgene product (luciferase) was detected in the liver of mice injected with DNA/Gal-pOm-mHA2, which was 280-fold greater than that obtained with DNA/DOTMA:Chol liposomes (50 microg DNA). Prior administration of galactosylated albumin reduced the gene expression to 1/100, indicating the asialoglycoprotein receptor-mediated gene transfer in liver parenchymal cells, ie hepatocytes. The luciferase activity in hepatocytes contributed more than 95% of the total activity in all the tissues examined. Thus, hepatocyte-targeted in vivo gene expression was achieved by the intravenous injection of DNA complex with the multifunctional gene carrier.
Nishikawa
M
M
Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University, Japan.
Yamauchi
M
M
Morimoto
K
K
Ishida
E
E
Takakura
Y
Y
Hashida
M
M
D000818:Animals; D037263:Asialoglycoprotein Receptor; D005260:Female; D015870:Gene Expression; D018390:Gene Targeting / Q000379:methods*; D005818:Genetic Engineering / Q000379:methods; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage*; D006801:Humans; D007275:Injections, Intravenous; D008099:Liver / Q000378:metabolism*; D008114:Liver Neoplasms, Experimental / Q000628:therapy; D008156:Luciferases / Q000235:genetics; D051379:Mice; D008813:Mice, Inbred ICR; D018613:Microscopy, Confocal; D010957:Plasmids / Q000008:administration & dosage; D011956:Receptors, Cell Surface / Q000378:metabolism*; D014162:Transfection / Q000379:methods*; D014407:Tumor Cells, Cultured
D037263:Asialoglycoprotein Receptor; D011956:Receptors, Cell Surface; D008156:Luciferases