Amphiphilic block copolymers enhance the cellular uptake of DNA molecules through a facilitated plasma membrane transport

INSERM, U915, l'institut du thorax, Nantes, F-44000, France.
Nucleic Acids Research (Impact Factor: 9.11). 10/2010; 39(4):1610-22. DOI: 10.1093/nar/gkq922
Source: PubMed


Amphiphilic block copolymers have been developed recently for their efficient, in vivo transfection activities in various tissues. Surprisingly, we observed that amphiphilic block copolymers such as Lutrol® do not allow the transfection of cultured cells in vitro, suggesting that the cell environment is strongly involved in their mechanism of action. In an in vitro model mimicking the in vivo situation we showed that pre-treatment of cells with Lutrol®, prior to their incubation with DNA molecules in the presence of cationic lipid, resulted in higher levels of reporter gene expression. We also showed that this improvement in transfection efficiency associated with the presence of Lutrol® was observed irrespective of the plasmid promoter. Considering the various steps that could be improved by Lutrol®, we concluded that the nucleic acids molecule internalization step is the most important barrier affected by Lutrol®. Microscopic examination of transfected cells pre-treated with Lutrol® confirmed that more plasmid DNA copies were internalized. Absence of cationic lipid did not impair Lutrol®-mediated DNA internalization, but critically impaired endosomal escape. Our results strongly suggest that in vivo, Lutrol® improves transfection by a physicochemical mechanism, leading to cellular uptake enhancement through a direct delivery into the cytoplasm, and not via endosomal pathways.

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    • "Synthetic delivery system (SDS) consisting of tetra-functional block copolymers (TFBC) demonstrated of a dramatic enhancement of target protein expression in preclinical models [33] [34] [35], maximizing access of the plasmid DNA to the cytosol, with activation of DNA sensors that trigger an innate immune response. Benefiting of this expertise, we set up a DNA vaccine model using the TFBC 704, which is under regulatory development for the treatment by vaccination of the human hepatocellular-carcinoma, to document the protective role of MA-PLC immunization in CF hosts using F508 (the most frequent CF mutation) mutated mice [36] "
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    • "Female, 8-week-old outbred Swiss mice (BALB/c genetic background) and athymic nude mice were obtained from Harlam (France). Intramuscular injections of expression plasmids were performed as previously described: 8 µg expression plasmid or 2 µg inducible expression plasmid plus 6 µg pQE30 plasmid formulated with the amphiphilic block copolymer 704 (29) were administered into the tibialis anterior muscles of mice. Hydrodynamic injections were prepared in a saline physiological buffer corresponding to 10% body volume of the mouse and were administered over a 5-s period into the tail vein of mice (30). "
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