Research ArticleEnhancement of dendritic cells transfection in vivo and of vaccination against B16F10 melanoma with mannosylated histidylated lipopolyplexes loaded with tumor antigen messenger RNA
Graphical Abstract
Mannosylated lipopolyplexes loaded with tumor antigen mRNA (Man-LPR) enhance the transfection of dendritic cells in vivo and inhibition of B16F10 melanoma growth.
Section snippets
Methods
All reagents were purchased from Sigma-Aldrich (St. Quentin Fallavier, France) unless stated otherwise. PEGylated histidylated poly-l-lysine (PEG-HpK) was synthesized as described.16, 21 O,O-dioleyl-N-[3N-(N-methylimidazolium iodide)propylene] phosphoramidate (lipid 1) and O,O-dioleyl-N-histamine phosphoramidate (lipid 2) were synthesized as described (Supplementary Figure S1).18, 19 β-d-mannopyranosyl-N-dodecylhexadecanamide (Man-lipid) was synthesized as described.22
In vitro transfection of DCs with mRNA LPR100
mRNA and pDNA encoding EGFP were formulated as lipopolyplexes. Nucleic acids (5 μg) were first condensed with the PEG-HpK (15 μg) before adding 10 μg of either Lip100 or Man11-Lip100. The electrophoretic mobility shift assays showed that mRNA and pDNA did not migrate in the agarose gel, indicating that they were in complex with LPR100, LPD100, Man11-LPR100, and Man11-LPD100 at the ratio used (Supplementary Figs. S2 and S3, which can be found in the online version of this article). The nucleic
Discussion
In the present study we have incorporated mannose motifs into liposomes to increase the delivery and the transfection efficiency of mRNA lipopolyplexes into DCs. From an experimental point of view, EGFP or MART-1 mRNA (∼900 nt) was first condensed with a PEG-HpK. Next, a liposomal formulation comprising three lipids (a mannosylated lipid, an imidazole lipophosphoramidate, and an imidazolium lipophosphoramidate), was added. Lipophosphoramidates have a chemical structure that is bio-inspired from
Supplementary data
Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.nano.2010.12.010.
The following is the supplementary material related to this article
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This work was supported by Ligue contre le Cancer (Région Centre) and Canceropole Grand Ouest. F.P. was a recipient of a fellowship from Ligue Nationale contre le Cancer.