Shielding the cationic charge of nanoparticle-formulated dermal DNA vaccines is essential for antigen expression and immunogenicity

Nanoparticle-formulated DNA vaccines hold promise for the design of in vivo vaccination platforms that target defined cell types in human skin. A variety of DNA formulations, mainly based on cationic liposomes or polymers, has been investigated to improve transfection efficiency in in vitro assays.\ud \ud Here we demonstrate that formulation of DNA into both liposomal and polymeric cationic nanoparticles completely blocks vaccination-induced antigen expression in mice and ex vivo human skin. Furthermore, this detrimental effect of cationic nanoparticle formulation is associated with an essentially complete block in vaccine immunogenicity. The blocking of DNA vaccine activity may be explained by immobilization of the nanoparticles in the extracellular matrix, caused by electrostatic interactions of the cationic nanoparticles with negatively charged extracellular matrix components. Shielding the surface charge of the nanoparticles by PEGylation improves in vivo antigen expression more than 55 fold. Furthermore, this shielding of cationic surface charge results in antigen-specific T cell responses that are similar as those induced by naked DNA for the two lipo- and polyplex DNA carrier systems. These observations suggest that charge shielding forms a generally applicable strategy for the development of dermally applied vaccine formulations. Furthermore, the nanoparticle formulations developed here form an attractive platform for the design of targeted nanoparticle formulations that can be utilized for in vivo transfection of defined cell types\u

Optimization of Intradermal Vaccination by DNA Tattooing in Human Skin

The intradermal administration of DNA vaccines by tattooing is a promising delivery technique for genetic immunization, with proven high immunogenicity in mice and in nonhuman primates. However, the parameters that result in optimal expression of DNA vaccines that are applied by this strategy to human skin are currently unknown. To address this issue we set up an ex vivo human skin model in which DNA vaccine-induced expression of reporter proteins could be monitored longitudinally. Using this model we demonstrate the following: First, the vast majority of cells that express DNA vaccine-encoded antigen in human skin are formed by epidermal keratinocytes, with only a small fraction (about 1%) of antigen-positive epidermal Langerhans cells. Second, using full randomization of DNA tattoo variables we show that an increase in DNA concentration, needle depth, and tattoo time all significantly increase antigen expression (p < 0.001), with DNA concentration forming the most critical variable influencing the level of antigen expression. Finally, in spite of the marked immunogenicity of this vaccination method in animal models, transfection efficiency of the technique is shown to be extremely low, estimated at approximately 2 to 2000 out of 1 × 1010 copies of plasmid applied. This finding, coupled with the observed dependency of antigen expression on DNA concentration, suggests that the development of strategies that can enhance in vivo transfection efficacy would be highly valuable. Collectively, this study shows that an ex vivo human skin model can be used to determine the factors that control vaccine-induced antigen expression and define the optimal parameters for the evaluation of DNA tattoo or other dermal delivery techniques in phase 1 clinical trials

Tc-99m-PEG-Liposomes for the evaluation of colitis in Crohn's disease

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