Combination Adjuvants Enhance Recombinant Protein Vaccine Protection against Fungal Infection

Wüthrich, Marcel; Dobson, Hannah E.; Ledesma Taira, Cleison; Okaa, Uju Joy; dos Santos Dias, Lucas; Isidoro-Ayza, Marcos; Petrovsky, Nikolai; Klein, Bruce S.

The development of effective vaccines against fungal infections requires the induction of protective, pathogen-specific cell-mediated immune responses. Here, we asked whether combination adjuvants based on delta inulin (Advax) formulated with Toll-like receptor (TLR) agonists could improve vaccine protection mediated by a fungal recombinant protein, Bl-Eng2 (i.e., Blastomyces endoglucanase 2), which itself harbors an immunodominant antigen and dectin-2 agonist/adjuvant. We found that Bl-Eng2 formulated with Advax3 containing TLR9 agonist or Advax8 containing TLR4 agonist provided the best protection against pulmonary infection with Blastomyces dermatitidis, being more effective than complete Freund’s adjuvant or Adjuplex. Advax3 was most efficient in inducing gamma interferon (IFN-γ)- and interleukin-17 (IL-17)-producing antigen-specific T cells that migrated to the lung upon Blastomyces dermatitidis infection. Mechanistic studies revealed Bl-Eng2/Advax3 protection was tempered by neutralization of IL-17 and particularly IFN-γ. Likewise, greater numbers of lung-resident T cells producing IFN-γ, IL-17, or both IFN-γ and IL-17 correlated with fewer fungi recovered from lung. Protection was maintained after depletion of CD4(+) T cells, partially reduced by depletion of CD8(+) T cells, and completely eliminated after depletion of both CD4(+) and CD8(+) T cells. We conclude that Bl-Eng2 formulated with Advax3 is promising for eliciting vaccine-induced antifungal immunity, through a previously uncharacterized mechanism involving CD8(+) and also CD4(+) T cells producing IFN-γ and/or IL-17. Although no licensed vaccine exists as yet against any fungal disease, these findings indicate the importance of adjuvant selection for the development of effective fungal vaccines.