Hepatocyte-targeted in vivo gene expression by intravenous injection of plasmid DNA complexed with synthetic multi-functional gene delivery system

Abstract

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 GalpOrn-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 g DNA per mouse) were examined. After injection of [ 32 P]DNA/Gal-pOrn-mHA2, about 60% of the radioactivity was recovered in the liver, mostly in paren-

Intracellular trafficking of nonviral vectors

“…161 When incorporated into an ASOR-targeted gene delivery polymer, the HA2 peptide enhanced gene transfer to the liver 300-fold higher than complexes lacking the HA2 peptide. 162 The authors reported observations that the peptide induced diffusion of the complex into the cytoplasm, suggesting that gene transfer enhancement was due to HA2-induced endosomolysis.…”

Glycosylation-mediated targeting of carriers

“…Of these, the release of pDNA from endosomes/lysosomes into the cytoplasm after internalization is a major obstacle for efficient gene expression in hepatocytes. Therefore, manipulation of this process by functional peptides or the use of PEI with pH buffering functions in lysosomes [53] by galactosylated macromolecules might allow high transfection rates in vivo [40,49,[54][55][56]. Our group developed a mannosylated macromolecular carrier system for cell-selective targeting based on dextran, which has high solubility, an abundance of hydroxyl groups for chemical modification, low immunogenicity, and is used in the clinic as a plasma expander [2].…”

Nonviral Approaches Satisfying Various Requirements for Effective in Vivo Gene Therapy.

“…Separation of the liver cells revealed that such complexes were preferentially taken up by hepatocytes, the cells possessing asialoglycoprotein receptors. When the transfection efficiency of the pDNA/galactosylated polymer complex was boosted by the use of a fusogenic peptide (an acid-sensitive peptide designed based on the amino-terminal of influenza virus hemagglutinin subunit HA-2 or mHA2), the amount of transgene product in the hepatocytes accounted for over 95% of the total amount in all the tissues examined, 23) indicating the success of this target cell-specific gene transfer in vivo. Figure 3 summarizes the tissue disposition of pDNA and transgene expression following intravenous injection of pDNA complexed with a hepatocyte-targeted polymeric carrier in mice.…”

“…The transfection efficiency into mouse liver was enhanced by the covalent binding of a fusogenic peptide to a hepatocyte-specific pDNA complex. 23) Another approach involves the use of a vector with a high buffering capacity and the ability to swell when protonated. Such a system, e.g.…”

“…Figure 3 summarizes the tissue disposition of pDNA and transgene expression following intravenous injection of pDNA complexed with a hepatocyte-targeted polymeric carrier in mice. 23) Tissue (Cell)-Specific Promoters Use of a tissue-specific promoter is another strategy for achieving target cellspecific gene transfer. Some promoters are active only in a specific type of cells, which offers high specificity of transgene expression following in vivo administration of pDNA.…”