Different behavior of branched and linear polyethylenimine for gene delivery in vitro and in vivo

Background Efficient gene transfer is a major challenge for non-viral gene therapy. Understanding how non-viral vectors initiate gene expression could lead to the development of new future vectors with enhanced efficacy. Methods Linear or branched polyethylenimine (PEI)/DNA complexes were genera ted in varying salt conditions and their transfection efficiencies were com pared in vitro and in vivo using reporter genes, luciferase and green fluor escent protein, and rhodamine labeled DNA (pGeneGrip(TM)). Results The transfection efficiency of linear PEI22/DNA in vitro was genera lly greater than that of branched PEI/DNA when complexes were generated in salt containing buffer. However, PEI complexes generated under salt-free co nditions generally had low transfection activity in vitro. In contrast, PEI 22/DNA salt-free complexes were highly active in vivo. Branched PEI/DNA and salt containing PEI22/DNA complexes were generally 10-100-fold less active than the salt-free PEI22/DNA complexes. Salt-free PEI22/DNA complexes were small, but subsequently grew into aggregates when salt was added. In contr ast, PEI25/DNA complexes remained small even after salt was added under the same conditions. Furthermore, PEI22/pGeneGrip(TM) complexes formed large a ggregates associated with the cell membrane, cytoplasm and nucleus, while b ranched PEI complexes remained as small distinct particles associated with the cell membrane or in the cytoplasm. Conclusions Branched and linear PEI/DNA complexes differ in their ability t o transfect cells. The greater efficiency of linear PEI might be due to an inherent kinetic instability under salt conditions. Understanding how to em ploy this kinetic instability of linear PEI could help in designing future vectors with greater flexibility and transfection efficiency in vivo. Copyr ight (C) 2001 John Wiley & Sons, Ltd.