Immunization using the skin : using topical Toll-like receptor 9 agonists as a method to increase vaccine efficacy

Vaccination is a cost-effective method to prevent diseases. Yet, vaccine effectiveness remains lower than anticipated and infectious disease is still a leading cause of illness and death. Immunostimulatory oligodeoxynucleotides containing CpG motifs (CpG ODNs) activate Tolllike receptor 9 (TLR9), bridging innate and adaptive immune responses. CpG ODNs have been studied as adjuvants, agents to improve vaccine efficacy. However, split administration of topical TLR9 agonists with parenteral vaccines has been less explored. I hypothesized that topical CpG ODN 1826 activates the skin immune system in a unique fashion, increasing the efficacy of locally administered protein-based vaccines. I found that an effective strategy to administer CpG ODN was single dose topically at time of local parenteral vaccine delivery. The generation of antigen-specific antibodies, CD4⁺ T helper 1 cells and CD8⁺ T cells were augmented. In murine Listeria monocytogenes and influenza A infection models, application of topical CpG ODN at time of parenteral protein immunization reduced bacterial and viral burden. The mechanisms whereby topical CpG adjuvant improves vaccine outcomes were investigated. Using bone marrow chimeric mice, I found that TLR9 expression in the hematopoietic compartment was necessary while expression in the stromal compartment also contributed to the enhancement of specific CD8⁺ T cell generation. Keratinocytes responded to topical CpG ODN partly by up-regulating TLR9 expression. Topical CpG ODNs were internalized by CD11c⁺ cells and were detected within the skin draining lymph nodes (SLNs). Topical compared to subcutaneous administration of CpG ODN differentially modulated cytokine and chemokine gene expressions in the SLNs and induced a higher proportion of specific CD4⁺ T cells to express tissue-homing molecules. In the influenza model, topical CpG adjuvant increased the proportion of specific CD8⁺ memory T cells at the site of infection. This work demonstrates the effectiveness of a novel immunization strategy that separates topical CpG adjuvant from a protein-based vaccine in enhancing rapid and long-lasting protective humoral and cellular vaccine responses. These improved responses are the consequences of the activation of stromal and hematopoietic cells in the skin, which subsequently modulate the microenvironment of the SLNs and alter the migratory ability of the T cells to tissues.Medicine, Faculty ofMedicine, Department ofExperimental Medicine, Division ofGraduat