Background For peptide-based cancer vaccines, successful eradication of tumors relies on the effective and persistent delivery of antigenic peptides to antigen presenting cells (APCs) to prime potent antigen-specific, cytotoxic T cells. Peptide-based vaccines have historically been either; (1) water-based formulations, which provide short exposure of peptides to immune cells; or (2) oil-in-water emulsions that provide longer peptide exposure but can elicit dysfunctional and exhausted T cell phenotypes. By contrast, the DPX^® technology is a non-aqueous, lipid-in-oil, immune-educating therapeutic delivery platform. Antigenic peptides formulated in DPX elicit a robust, targeted, and persistent tumor antigen-specific T cell response that for our lead DPX product, Maveropepimut-S, has translated into clinical benefit in multiple cancer indications, including DLBCL and ovarian cancer.

Methods Herein, we compare DPX to aqueous and emulsion-based formulations for the dynamics of immune cell recruitment to the site of injection (SOI), peptide antigen consumption, and trafficking by immune cells. Immune cell composition and antigen uptake at the SOI were assessed by multi-parameter flow cytometry and confocal microscopy using model peptide antigens administered in C57/Bl6 mice. Antigen-specific immune responses were assessed in draining lymph nodes and/or spleens by IFN-γ ELISPOT.

Results These data reveal that aqueous formulations were poorly able to retain lymphocytes at SOIs and consequently did not elicit a detectable IFN-γ ELISPOT response. Both formulations containing an oil component (DPX and emulsion) were superior in recruiting APCs cells to the SOIs and inducing antigen-specific immune responses. Significant increases in immune cell infiltration were detected as early as 2 days post DPX injection. Antigen uptake was confirmed using confocal microscopy. Both DPX and emulsion platforms induced a prompt increase in antigen presentation in the context of MHC. However, antigen presentation driven by the DPX platform had a distinct profile enriched in CD11b⁺CD11c⁺MHCII⁺ APCs co-expressing the CD80, CD86, and CD40 activation/costimulatory markers. The recruitment and activation of this subset was evident regardless of whether a peptide was present in DPX. By contrast, the emulsion incites a distinctly different CD11b^-CD11c^- population. Interestingly, CD11b⁺CD11c⁺ cells have tendency to express higher number of peptide-MHC complexes per cell compared to the CD11b^-CD11c^- population.

Conclusions Collectively these findings highlight quantitative, qualitative, and temporal differences in immune cell recruitment amongst three delivery platforms and show the unique character of the immune response triggered by the DPX platform typified by the recruitment of CD11b⁺CD11c⁺ APCs that have intrinsically higher capacity for antigen uptake, presentation, and activation.

Ethics Approval Experiments were conducted in accordance with ethics protocols approved by the University Committee on Laboratory Animals at Dalhousie University, Halifax, N.S., Canada.