Dendritic cells (DCs), a vital component of the innate immune system, are considered to lack antigen specificity and be devoid of immunological memory. Strategies that can induce memory-like responses from innate cells can be utilized to elicit protective immunity in immune deficient persons. Here we utilize an experimental immunization strategy to modulate DC inflammatory and memory-like responses against an opportunistic fungal pathogen that causes significant disease in immunocompromised individuals. Our results show that DCs isolated from protectively immunized mice exhibit enhanced transcriptional activation of interferon and immune signaling pathways. We also show long-term memory-like cytokine responses upon subsequent challenge with the fungal pathogen that are abrogated with inhibitors of specific histone modifications. Altogether, our study demonstrates that immunization strategies can be designed to elicit memory-like DC responses against infectious disease.

Developing an effective and safe recombinant vaccine requires microbe-specific antigens combined with an adjuvant/delivery system to strengthen protective immunity. In this study, we designed and expressed a multivalent, recombinant polypeptide antigen (rCpa1) that consists of three previously identified antigens (i.e., Ag2/Pra, Cs-Ag and Pmp1) and 5 pathogen-derived peptides with high affinity for human MHC II molecules. The purified rCpa1 was encapsulated into four types of yeast cell-wall particles containing various compositions of β-glucan, mannan and chitin or mixed with an oligonucleotide (ODN) containing 2 methylated dinucleotide CpG motifs. This multivalent antigen encapsulated into glucan-chitin particles (GCP-rCpa1) showed a significantly elevated reduction of fungal burden for human HLA-DR4 transgenic mice compared to the other tested adjuvant-rCpa1 formulations. Among the tested adjuvants GCPs and GPs were both capable of stimulating a mixed Th1 and Th17 response. Mice vaccinated with GCP-rCpa1 showed elevated IL-17 production in T-cell recall assays and early lung infiltration of activated Th1 and Th17 cells compared to GP-rCpa1-vaccinated mice. Both C57BL/6 and HLA-DR4 transgenic mice that were vaccinated with GCP-rCpa1 vaccine increased surivial compared to the mice received GCPs alone. Concurringly, GCP-rCpa1 vaccine stimulated enhanced infiltration of macrophages to engulf and process the vaccine for antigen presentation in the injection sites compared to GP-rCpa1 injection. This is the first attempt to systematically characterize the presentation of a multivalent coccidioidomycosis vaccine encapsulated with selected adjuvants which enhance protective cellular immune response to infection.

Development of vaccines against opportunistic infections is difficult as patients most at risk of developing disease are deficient in aspects of the adaptive immune system. Here, we utilized an experimental immunization strategy to induce innate memory in macrophages in vivo. Unlike current trained immunity models, we present an innate memory-like phenotype in macrophages that is maintained for at least 70 days post-immunization and results in complete protection against secondary challenge in the absence of adaptive immune cells. RNA-seq analysis of in vivo IFN-γ primed macrophages revealed a rapid up-regulation of IFN-γ and STAT1 signaling pathways following secondary challenge. The enhanced cytokine recall responses appeared to be pathogen-specific, dependent on changes in histone methylation and acetylation, and correlated with increased STAT1 binding to promoter regions of genes associated with protective anti-fungal immunity. Thus, we demonstrate an alternative mechanism to induce macrophage innate memory in vivo that facilitates pathogen-specific vaccine-mediated immune responses.