(483b) Succinate Based Adjuvant-Less Cancer Vaccine Modifies Immunometabolism and Prevent Melanoma Growth in Mice

Introduction: Numerous intracellular as well as extracellular signals regulate dendritic cell (DC) metabolism with multiple metabolic pathways being integral for its survival, growth and functions. These activated DCs play a vital role in helping mount a robust response, essential in vaccine-mediated tumor responses. Hence, direct targeting of DC metabolism can be considered a viable strategy to increase the immunogenicity of cancer vaccines. Adjuvants – important component of cancer vaccines, are also able to directly modify DC metabolism leading to immune activation increasing vaccine efficacy. However, in the past 100 years, only five adjuvants have been tested in humans, due to unknown toxicity profiles. Therefore, providing metabolites that are able to modulate these pathways instead of adjuvants, can potentially be a preferred alternative to adjuvants and lead to better control of immune activation. We hypothesize that, DC activation by direct delivery of metabolites along with an antigen can cause significant DC activation to prime antigen specific T cells for mounting a robust immune response against cancer. Herein, we report an adjuvant-less vaccine strategy using succinate, a vital central-carbon metabolite in TCA cycle, that is able to activate DCs and subsequently upregulate antigen specific T cells (T_C1 and T_C17) in melanoma (B16F10) mouse model.

Methods: Metabolite-based polymeric particles were generated by oil in water emulsions. These particles were characterized using dynamic light scattering (DLS), scanning electron microscopy (SEM), nuclear magnetic resonance (NMR) and release kinetics. Seahorse assay determined the extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) of dendritic cells (DCs) in presence of glutaminase inhibitor (CB839) and the synthesized microparticles. Flow cytometry and enzyme-linked immunosorbent assay (ELISA) determined the modulation of DCs and adaptive immunity by microparticles in vitro. Immunocompetent mice were injected with B16F10 melanoma cells and treated along with TRP2 (antigen against melanoma). On day 16, lymph nodes, tumor and spleen were harvested and used for further immunological studies.

Results: Succinate-based polymers were synthesized and subsequently formed into polymeric-microparticle (1-3 µm) for efficient cellular delivery. eadily phagocytose these microparticles and significantly changed the intracellular metabolite profile as observed by the metabolomics studies as compared to other groups. Notably, these particles also upregulated glycolysis and mitochondrial respiration in bone marrow derived DCs (BMDCs), as observed by up-regulated extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) suggesting higher DC activation. Moreover, cell surface activation indicators like upregulation in costimulatory markers (MHCII⁺CD86^Hi) and increased pro-inflammatory cytokine production (TNFa, IL-12) were observed, in vitro. Interestingly, administration of TRP-2 peptide (without any adjuvant) along with the succinate based microparticles led to significant reduction of subcutaneous melanoma tumor (B16F10) growth leading to increased survival in immunocompetent mice for up to a month. Furthermore, among tumor infiltrating T cell populations, a 3-fold increase in Tc17 population along with significant downregulation of Tregs and proliferating Tregs was observed.

Conclusion: DC modulation by direct delivery of metabolites along with an antigen can cause significant DC activation. These results demonstrate that succinate based microparticles, in absence of any adjuvants are capable of activating immune system in mice and is a viable strategy against melanoma.