Synthetic Messenger RNA and Cell Metabolism Modulation pp 247-274

Part of the Methods in Molecular Biology book series (MIMB, volume 969)

Mannosylated and Histidylated LPR Technology for Vaccination with Tumor Antigen mRNA

Protocol

Abstract

mRNA-based vaccines are currently being developed for treating various diseases including cancers. For this purpose, synthetic or in vitro transcribed (IVT) mRNA encoding tumor antigen offers several advantages over plasmid DNA encoding tumor antigen including better delivery and security. In this chapter, we report the preparation of mannosylated mRNA nanoparticles termed mannosylated lipopolyplexes or Man-LPR loaded with mRNA encoding a melanoma antigen. This formulation enhances the transfection of dendritic cells (DCs) in vivo and the anti-B16F10 melanoma vaccination in mice. The mRNA is formulated with histidylated liposomes and a histidylated polymer. Those pH-sensitive vectors promote membrane destabilization in endosomes upon the protonation of their histidine groups, allowing nucleic acid delivery in the cytosol. To favor DCs targeting via the mannose receptor, a mannose lipid is incorporated in the liposomes. Here, we provide protocols for the preparation of mannosylated liposomes, the synthesis of mRNA, mice immunization based on systemic injection, measurement of the cellular immune response and determination of the number of transfected splenic DC.

Key words

mRNA Transfection Dendritic cells Vaccine Histidine Liposomes Polymer 

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Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  1. 1.Centre de Biophysique Moléculaire, CNRS UPR4301OrléansFrance
  2. 2.Université d’OrléansOrléansFrance

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