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| - Abstract Transcutaneous immunization (TCI) is a non-invasive and easy-to-use vaccination method. We demonstrated the efficacy and safety of a transcutaneous vaccine formulation using a hydrogel patch in animal experiments. In the present study, we performed a clinical study to apply our TCI formulation for vaccination against tetanus and diphtheria in human. The TCI device was a hydrogel patch (antigen-free) applied to the left brachial medial skin of 22 healthy volunteers for 48h. Next, the hydrogel patch, containing 2mg tetanus toxoid (TT) and 2mg diphtheria toxoid (DT) as the TCI formulation, was applied to 27 healthy volunteers for 24h and some volunteers were vaccinated again by TCI formulation. For safety assessment, the patch application site was observed to assess local adverse events, and systemic adverse events were determined by a blood test. The antigen-free hydrogel patch and TCI formulation containing TT and DT did not induce local or systemic severe adverse events. For vaccine efficacy estimation, toxoid-specific serum antibody titers were determined by ELISA and the toxin-neutralizing activity of the induced antibody was evaluated in a passive-challenge experiment. The anti-TT IgG titer and the anti-DT IgG titer increased, and a significant effect was detected by paired t-test. The antibody titers were maintained at higher level than that before vaccination for at least 1 year. Moreover, toxoid-specific antibodies were produced by the second vaccination in some subjects. Antibodies induced by application of the TCI formulation neutralized the toxin and prevented toxic death in mice. In addition, changes in the skin condition due to application of the TCI formulation were observed under in vivo confocal Raman spectroscopy. The amount of water and patch components in the stratum corneum increased after application of the TCI formulation, suggesting that the change in the skin condition was related to antigen penetration. These data indicate that this easy-to-use TCI system induces an immune response without severe adverse reactions in humans. This easy-to-use and safe TCI formulation enables mass treatment in an outbreak setting and increased vaccination rates in developing countries, and will greatly contribute to worldwide countermeasures against infectious diseases.
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