Specific gene transfer mediated by galactosylated poly-L-lysine into hepatoma cells.
Plasmid DNA/galactosylated poly-L-lysine(GalPLL) complex was used to transfer luciferase reporter gene in vitro into human hepatoma cells by a receptor-mediated endocytosis process. DNA was combined with galPLL via charge interaction (DNA:GalPLL:fusogenic peptide, 1:0.4:5, w/w/w) and the resulting complex was characterized by dynamic light scattering, gel retardation assay and zeta potential analyzer to determine the particle size, electrostatic charge interaction, and apparent surface charge. The complex was tested for the efficiency of gene transfer in cultured human hepatoblastoma cell line Hep G2 and fibroblast cells NIH/3T3 in vitro. The mean diameter of the complex (DNA:GalPLL=1:0.4, w/w) was 256+/-34.8 nm, and at this ratio, it was positively charged (zeta potential of this complex was 10.1 mV). Hep G2 cells, which express a galactose specific membrane lectin, were efficiently and selectively transfected with the RSV Luc/GalPLL complex in a sugar-dependent manner. NIH/3T3 cells, which do not express the galactose-specific membrane lectin, showed only a marginal level of gene expression. The transfection efficiency of GalPLL-conjugated DNA complex into Hep G2 cells was greatly enhanced in the presence of fusogenic peptide that can disrupt endosomes, where the GalPLL-DNA complex is entrapped with the fusogenic peptide. With the fusogenic peptide KALA, the luciferase activity in Hep G2 cells was ten-fold higher than that of cells transfected in the absence of the fusogenic peptide. Our gene transfer formulation may find potential application for the gene therapy of liver diseases.
Gene Therapy Research Unit, Korea Research Institute of Bioscience and Biotechnology, Yusong, P.O. Box 115, Taejon, 305-600, South Korea.
J
J
Han
Y
Y
Il Yeom
D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000235:genetics*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D010957:Plasmids / Q000235:genetics; D011107:Polylysine / Q000235:genetics*; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D014162:Transfection / Q000235:genetics; D014407:Tumor Cells, Cultured
D011956:Receptors, Cell Surface; C039707:galactose receptor; D011107:Polylysine; D004247:DNA