A hepatocyte-directed vector has been developed; it includes several key features thought to favor in vivo gene delivery to the liver: electrostatically neutral particles which avoid nonspecific binding to other cells, to the extracellular matrix, and to complement proteins; asialoglycoprotein receptor-mediated endocytosis which may address the complexes to the perinuclear region; and polyethylenimine (PEI)-mediated endosome buffering and swelling as an escape mechanism to the cytoplasm. This system is based on a 5% galactose-bearing polyethylenimine (PEI-gal) polymer which is condensed with plasmid DNA to neutrality. Murine (BNL CL.2) and human (HepG2) hepatocyte-derived cell lines were transfected 10(4)-10(5)-fold more efficiently than murine fibroblasts (3T3), whether transfection was assessed globally (luciferase expression from the cell extract) or following histochemical staining (beta-galactosidase). Under these conditions, over 50% of the hepatocytes were selectively transfected in the presence of 10% serum. Transfection was suppressed by removal of the targeting galactose residues, by their replacement with glucose, or by the addition of excess asialofetuin. Thus, results from comparative and competitive experiments indicate the asialoglycoprotein receptor is involved in transfection of hepatocytes with neutral PEI-gal/DNA complexes.

A new class of phosphoruscontaining dendrimers (abbreviated as P-dendrimer) with protonated or methylated terminal tertiary amines has been prepared and characterized. Five different generations of protonated dendrimers (2-[G 1 ] to 2-[G 5 ]) and the corresponding methylated series (5-have been examined as transfecting agents of the luciferase gene within 3T3 cells. The capability of these dendrimers to transfect cells appears to depend on the size of the dendrimers (the third, fourth, and fifth generation being the most efficient ones) and the chemical nature of the terminal tertiary amines (the protonated forms being more efficient than the methylated ones). The most efficient representatives of this series of P-dendrimers have a transfection activity level comparable with linear PEI with only five equivalents of tertiary amine per nucleotide. Unexpectedly and fortunately, these dendrimers have a better transfection efficiency in the presence of serum than without.

P‐dendrimers appear to be excellent candidates as transfecting agents after positively charged functions are grafted at their periphery. Five different generations of protonated dendrimers (1‐[G1] to 1‐[G5]) and the corresponding methylated series (2‐[G1] to 2‐[G5]) (two examples are shown here) have been examined as transfecting agents of the luciferase gene within 3T3 cells.