Galactosylated multimodular lipoplexes for specific gene transfer into primary hepatocytes

Background Numerous synthetic cationic vectors have been synthesized and are successfully used for in vitro gene transfer but an excess of positive charges can lead to cytotoxicity and does not enable specific transfection. Methods We decided to develop alternative molecular systems consisting of neutral, colloidally stable bioassemblies equipped with ligands for specific cell targeting. Consequently, we directed our efforts toward the development of a multimodular non-viral gene delivery system consisting of a condensed core of DNA with cationic liposomes of bis(guanidinium)-tren-cholesterol and an external corona of poly(ethylene oxide) stretches harbored by the steric stabilizers used to stabilize lipoplexes colloidally. A ligand capable of cell targeting by receptor-mediated endocytosis was covalently linked at the poly(ethylene oxide) extremity of steric stabilizers. Steric stabilizers were functionalized by a one-step enzymatic galactosylation to develop new supramolecular assemblies of lipoplexes able to target asialoglycoprotein receptors located on primary hepatocytes. Results Cryo-TEM and fluorescence experiments showed that DNA was condensed within lamellar complexes whose size ranged between 100 to 300 nm in diameter. Bis(guanidinium)-tren-cholesterol-DNA lipoplexes, colloidally stabilized by galactosylated steric stabilizers at a galactosylated steric stabilizer/DNA ratio of 300, led to specific transfection of primary hepatocytes whereas ungalactosylated steric stabilizer did not transfect. Conclusions Our findings confirm the receptor-mediated endocytosis pathway of galactosylated multimodular lipoplexes. Thus, we conclude that the fabrication of a multimodular assembly harboring a ligand without nonspecific interaction with cell membranes is possible and a highly promising system to transfect other primary or cultured cells specifically through a receptor-dependent mechanism. Copyright (C) 2008 John Wiley & Sons, Ltd.