(H-314) Macrophage Vaccine as a Potent Initiator of Activation of NK, CD4+, and CD8+ T Cells for Robust Tumor Control

Introduction:: Macrophages, despite their abundance and ubiquitous presence in inflamed and infected tissues, have been largely neglected for their role in antigen presentation in part due to the perception that their key function is in innate immunity and that they are less efficient in antigen presentation compared to dendritic cells. Here, we demonstrate, using a murine model of melanoma, that activated macrophages serve as a potent cancer vaccine. The macrophage vaccine (Mvax) elicited a strong CD8+ T cell response, along with the NK and CD4+ T cell activation, inducing a ‘hot’ tumor transition and robust tumor suppression. Owing to their unique ability to infiltrate tissues, macrophages show a unique in vivo trafficking behavior and orchestrate the modulation of multiple stages of immune responses, providing a new platform for cancer vaccines that could be effective against tumors that have been less responsive to traditional cancer vaccines.

Materials and Methods:: Macrophages are derived from bone marrow harvested from 5-6 weeks old C57BL/6 mice from Jackson Labs. Bone marrow cells were cultured with M-CSF-supplemented media for 7 days to achieve cells with macrophage phenotype. Prior to the administration of macrophages to tumor-bearing mice, cells were activated for 4 hours with a novel activation cocktail which was designed to induce sufficient activation without elevating their systemic toxicity. Two hours through the incubation, tumor-specific antigen peptides were added to the media for peptide loading on the MHC-I molecules. The resulting cells were administered via the tail-vein IV.

Results, Conclusions, and Discussions:: A model tumor antigen peptide, SIINFEKL, was readily loaded on macrophages in vitro, demonstrating a key step in the utility of macrophages as a vaccine (a). Upon IV administration via the tail vein in mice, macrophages efficiently trafficked to the inguinal tumor-draining lymph nodes and tumors (b, c). Antigen-loaded macrophages with or without activation induced a robust systemic CD8+ T cell response, determined by the frequency of antigen-specific CD8+ T cells in peripheral blood using tetramer staining, which was further increased after booster injections (d). However, activated macrophages loaded with antigen (Mvax) led to higher suppression of subcutaneously developed B16F10OVA tumor growth (e) compared to non-activated macrophages or saline controls. In addition to the CD8+ T cells, we observed a 2-fold increase in the frequencies of activated NK cells and effector CD4+ T cells in the systemic circulation (f). These correlated with enhanced NK and CD8+ T cell activation in the TME, along with the M1 transition of tumor-associated macrophages and increased intratumoral IFN-g concentration, all of which are signs of a ‘hot’ tumor, which were not observed from groups treated with Mvax without activation (Mvax w/o activation) or without antigen pulsing (Mvax w/o Ag) (g-i).

Cancer vaccines have traditionally been focusing on activating DCs to modulate adaptive immune responses, primarily the CD8+ T cells. Here, by pulsing macrophages first with an activation cocktail and then with tumor-specific antigen peptides ex vivo, followed by IV administration, we have demonstrated the use of macrophages as a potent cancer vaccine that orchestrates the M1 transition of tumor-associated macrophages and activation of both the innate and adaptive immune cells, including NK, CD4+, and CD8+ T cells. Based on the previous reports demonstrating the benefits of activating multiple immune pathways for achieving better tumor control, we posit that macrophages represent a promising vaccine platform for treating cancers that have been less responsive to conventional treatments.

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