1-2-dioleoyloxy-3-(trimethylammonium)propane and precirol

In the present work, we evaluated the potential utility for gene delivery of three oligochitosans (OligoCh) that differs in the M(n) (OligoChA: 6.1 kDa, OligoChB: 11.5 kDa, and OligoChC: 13.7 kDa), with deacetylation degree of 85%. OligoCh were complexed directly with the pCMS-EGFP plasmid to form OligoCh-DNA carriers. Taking into account the features and benefits of both Ch and SLNs, we also combined the OligoCh with SLNs. The three OligoCh presented a great ability to condense and protect the DNA. The OligoCh of highest M(n) (OligoChC) complexed with SLNs at a OligoChC:DNA:SLN ratio 2.5:1:5 induced the highest transfection level in HEK-293 cells at day 3; being transfection 2-fold higher at day 7. After the intravenous administration to mice, OligoChC-DNA and OligoChC-DNA-SLN vectors were able to induce the expression of EGFP in the spleen, lung and liver, which was maintained for at least 7 days. In spite of the difference in the "in vitro" transfection levels between both vectors, no difference was detected in transfection after "in vivo" administration. Moreover, the OligoChC improved the "in vivo" transfection efficacy of the DNA-SLN vector. This work shows the potential utility of the combination of SLNs and OligoCh for the development of new non-viral vectors for gene therapy.

Topics: Animals; Chitin; Chitosan; Diglycerides; DNA; Fatty Acids, Monounsaturated; Female; Green Fluorescent Proteins; HEK293 Cells; Humans; Liver; Lung; Mice; Mice, Nude; Nanoparticles; Oligosaccharides; Plasmids; Polysorbates; Quaternary Ammonium Compounds; Spleen; Transfection

Naked plasmid DNA is a powerful tool for gene therapy, but it is rapidly eliminated from the circulation after intravenous administration. Therefore, the development of optimized DNA delivery systems is necessary for its successful clinical use. Solid lipid nanoparticles (SLNs) have demonstrated transfection capacity in vitro, but their application for gene delivery has not been conveniently investigated in vivo. We aimed to evaluate the capacity of SLN-DNA vectors to transfect in vivo after intravenous administration to mice. The SLNs, composed of Precirol ATO 5, DOTAP and Tween 80 were complexed with the plasmid pCMS-EGFP which encodes the enhanced green fluorescent protein (EGFP). The resulting systems were characterized in vitro showing a mean particle size of 276 nm, superficial charge of +28 mV, the ability to protect the plasmid and transfection capacity in culture cells. The intravenous administration in mice led to transfection in hepatic tissue and spleen. Protein expression was detected from the third day after administration, and it was maintained for at least 1 week. This work shows for the first time the capacity of SLN-DNA vectors to induce the expression of a foreign protein after intravenous administration, supporting the potential of SLNs for gene therapy.

Topics: Animals; Cell Line; Diglycerides; Fatty Acids, Monounsaturated; Female; Genetic Therapy; Green Fluorescent Proteins; Humans; Injections, Intravenous; Lipids; Liver; Mice; Mice, Inbred BALB C; Microscopy, Fluorescence; Nanoparticles; Particle Size; Plasmids; Polysorbates; Quaternary Ammonium Compounds; Spleen; Surface Properties; Time Factors; Transfection