Particle-based drug delivery systems target active compounds to the hair follicle and may result in a better penetration and higher efficiency of compound uptake by skin resident cells. As previously proposed, such delivery systems could be important tools for vaccine delivery. In this study, we investigated the penetration of solid fluorescent 40 or 200 nm polystyrene nanoparticles (NPs) as well as virus particles in murine skin to further investigate the efficacy of transcutaneously (TC) applied particulate vaccine delivery route. We demonstrated that 40 and 200 nm NPs and modified vaccinia Ankara (MVA) expressing the green-fluorescent protein penetrated deeply into hair follicles and were internalized by perifollicular antigen-presenting cells (APCs). Fibered-based confocal microscopy analyses allowed visualizing in vivo particle penetration along the follicular duct, diffusion into the surrounding tissue, uptake by APCs and transport to the draining lymph nodes. The application of small particles, such as ovalbumin coding DNA or MVA, induced both humoral and cellular immune responses. Furthermore, TC applied MVA induced protection against vaccinia virus challenge. Our results strengthen the concept of TC targeting of cutaneous APCs by hair follicles and will contribute to the development of advanced vaccination protocols using NPs or viral vectors.

BackgroundCurrent conventional vaccination approaches do not induce potent CD8 T-cell responses for fighting mostly variable viral diseases such as influenza, avian influenza viruses or HIV. Following our recent study on vaccine penetration by targeting of vaccine to human hair follicular ducts surrounded by Langerhans cells, we tested in the first randomized Phase-Ia trial based on hair follicle penetration (namely transcutaneous route) the induction of virus-specific CD8 T cell responses.Methods and FindingsWe chose the inactivated influenza vaccine – a conventional licensed tetanus/influenza (TETAGRIP®) vaccine – to compare the safety and immunogenicity of transcutaneous (TC) versus IM immunization in two randomized controlled, multi-center Phase I trials including 24 healthy-volunteers and 12 HIV-infected patients. Vaccination was performed by application of inactivated influenza vaccine according to a standard protocol allowing the opening of the hair duct for the TC route or needle-injection for the IM route. We demonstrated that the safety of the two routes was similar. We showed the superiority of TC application, but not the IM route, to induce a significant increase in influenza-specific CD8 cytokine-producing cells in healthy-volunteers and in HIV-infected patients. However, these routes did not differ significantly for the induction of influenza-specific CD4 responses, and neutralizing antibodies were induced only by the IM route. The CD8 cell response is thus the major immune response observed after TC vaccination.ConclusionsThis Phase Ia clinical trial (Manon05) testing an anti-influenza vaccine demonstrated that vaccines designed for antibody induction by the IM route, generate vaccine-specific CD8 T cells when administered transcutaneously. These results underline the necessity of adapting vaccination strategies to control complex infectious diseases when CD8 cellular responses are crucial. Our work opens up a key area for the development of preventive and therapeutic vaccines for diseases in which CD8 cells play a crucial role.Trial RegistrationClinicaltrials.gov NCT00261001

Induction of T cell responses has become one of the major goals in therapeutic vaccination against viral diseases and cancer. The use of the skin as target organ for vaccine has been spurred by recent implication of epithelial dendritic cells in CD8 cell cross-priming and suggests that vaccination via the transcutaneous (TC) route may be relevant in the induction of cellular immune responses. We have previously shown that TC application of nanoparticles, on human skin explants, allows targeting of epidermal dendritic cells, possibly via hair follicles. In this study, we have investigated cellular immune responses against an influenza protein-based vaccine by TC vaccination, compared with i.m. vaccination in humans. In this study on 11 healthy volunteers, we found that a newly developed protocol based on cyanocrylate skin surface stripping induced a significant increase in IFNγ-producing T cells specific for influenza vaccine by ELISPOT assays. Interestingly, TC vaccination induced both effector CD4 and CD8 T cell responses, whereas i.m. injection induced strong effector CD4 in the absence of CD8 T cells, as assessed by intracellular cytokine staining and tetramer analyses. This study proposes new perspectives for the development of vaccination strategies that trigger T cell immune responses in humans.