PEI/DNA complexes

Bettinger, T., Remy, J.S. and Erbacher, P. (1999) Size reduction of galactosylated PEI/DNA complexes improves lectin-mediated transfer into hepatocytes. Bioconjug. Chem., 10, 558-561. 

Figuer 17.3 Tracking of labeled PEI/DNA complexes (green) in cells with labeled lysosomes (red). Each bar indicates 10 pm. Times post-transfection are as follows (A) two hours—clumps of PEI/DNA complexes are present on the plasma membrane, and lysosomes are scattered throughout the cytoplasm (B) three hours—PEI/DNA complexes are located in endocytotic vesicles and are somewhat surrounded by lysosomes. There is no overlap of PEI and lysosomal probes (C) five hours—PEI/DNA complexes have entered the nucleus (shown by arrow) while cytoplasmic vesicles containing PEI/DNA complexes still have not fused with lysosomes. (Reproduced with permission from Godbey el al., 2000a). (see Color Plate 12)... 

Color Plate 12 Tracking of labeled PEI/DNA complexes (green) in cells with labeled lysosomes 342... 

Kunath K, von Harpe A, Fischer D et al (2003) Galactose-PEI-DNA complexes for targeted gene delivery degree of substitution affects complex size and transfection efficiency. J Control Rel 88(1) 159-172... 

Whether the components of the gene carriers actually remain associated during import into the nucleus or enter individually cannot be answered by optical methods as their resolution is limited. A possible technique to study the complexation of DNA within cells is fluorescence correlation spectroscopy (FCS). Clamme et al. studied the intracellular fate of PEI after transfection with polyplexes by two-photon fluorescence FCS [54]. They showed that PEI binds to the inner membrane of endosomes and lysosomes and shows free diffusion in the cytosol as well as the nucleus. However, they did not detect any PEI/DNA complexes inside the nucleus. 

R. Kircheis, and E. Wagner. 2001. Different strategies for formation of pegylated EGF-conjugated PEI/DNA complexes for targeted gene delivery. Bioconjug. Chem. 12 529-537. 

Interestingly, larger PEI/DNA complexes were more effective transfection agents than smaller particles over a range of PEI molecular weights in one study... 

Enhanced transfection with larger vectors was also seen with lipid-DNA lipoplexes in CHO cells [113]. However, the opposite was found with linear PEI (L-PEI) in vivo. Smaller complexes led to higher levels of gene expression in adult and newborn mice, which correlated to their diffusivity through tissue [100]. In this study, L-PEI DNA complexes were shown to cross the endothelial cell barrier following intravenous administration and preferentially transfect pulmonary cells [92]. Cationic lipids, on the other hand, show some expression in pulmonary cells following intravenous administration but preferentially transfected endothelial cells [51,53,114,115], perhaps due to their reduced stability compared to polyplexes. 

Stability. Current protocols for nonviral delivery systems call for the preparation of the vector at the bedside, due to their aggregation over time [127-129]. Specifically, aggregation of some lipoplex formulations stored as liquids reduced their transfection efficiency [130,131], Electron microscopy was used to show that PEI/DNA complexes did not aggregate, but vectors made with poly-L-lysine and dendrimers did in one study [103],... 

Gautam A, Densmore CL, Golunski E, Xu B, Waldrep JC. Transgene expression in mouse airway epithelium by aerosol gene therapy with PEI-DNA complexes. Mol Ther 2001, 3, 551-556. 

Goula D, Remy JS, Erbacher P, Wasowicz M, I evi G, Abdalkan B, Demeneix BA. Size, diffusibility and transfection performance of linear PEI/DNA complexes in the mouse central nervous system. Gene Therapy 1998, 5, 712-717. 

Owing to its free amine functions, chitosan may also be protonated (pfCa = 5.6). In in vitro studies with Hela-cells, chi-tosan/DNA complexes showed a gene-transfection potency similar to that reported for PEI/DNA complexes. Plain chitosan, however, is almost insoluble in water at neutral pH (but soluble at acidic pH), for this reason, trimethylated, quaternary chitosan derivatives have been produced that are sufficiently soluble under physiological conditions and easily complex DNA molecules. In in vitro experiments with COS-1 and CaCo-2 cells, these innovative chitosan derivatives proved superior to nontreated chitosan polymers, particularly as they showed no unspecific cytotoxicity [87]. 

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