Multifunctional HBc Virus-Like Particles Reprogram Immunosuppressive Macrophages and Potentiate CD8+ T Cell Responses for Enhanced Cancer Immunotherapy

Introduction: Tumor-associated macrophages (TAMs) promote immunosuppression, hindering immune checkpoint blockade and immunotherapy efficacy. To overcome this, we developed a novel multifunctional nanovaccine based on hepatitis B core virus-like particles (HBc VLP) to synergistically remodel the immunosuppressive tumor microenvironment through integrated TAM reprogramming and B7-H3 checkpoint blockade. Methods: The core VLP co-displayed tumor antigen peptide MAGE-A10 and TAM-targeting peptide M2pep via fusion expression. Immunostimulatory CpG oligodeoxynucleotide 1826 (CpG) was encapsulated within VLP. Anti-B7-H3 antibody (αB7-H3) and polyethylene glycol (PEG) were chemically conjugated to the surface for checkpoint blockade and prolonged circulation, forming CpG@VLP-αB7-H3-PEG. Results: Structural characterization using transmission electron microscopy and dynamic light scattering confirmed the hollow spherical self-assembly of VLP. Nanovaccines efficiently targeted TAMs in vitro and in vivo. Following CpG encapsulation (5.60 μg/mg), the nanovaccine reprogrammed M2-like TAMs into an M1-like phenotype. This was achieved by elevating the M1/M2 ratios of CD86/CD206 and MHC II/CD206 to 15.50-fold and 3.11-fold, respectively, as determined by flow cytometry. Further conjugation of αB7-H3 (250 μg/mg) significantly enhanced T-cell activation in TAM-T cell co-culture assays. In B16-F10 melanoma-bearing mice, reprogrammed iNOS + M1-like macrophages triggered robust antitumor immunity, achieving a tumor inhibition rate of 63.47%. These macrophages also function as antigen-presenting cells and increase the proportion of tumor-infiltrating Granzyme B + CD8 + T cells. αB7-H3 conjugation further boosted infiltrating immune cells, M1-like macrophages, activated CD69 + CD4 + /CD8 + T cells, and cytotoxic T lymphocytes. PEGylation amplified systemic tumor-specific immunity and increased tumor inhibition by 80.12%. Conclusion: This HBc VLP-based nanovaccine constitutes a pioneering multifunctional platform designed to overcome TAM-mediated immunosuppression through synergistic integration of three modalities: antigen presentation, TAM phenotype reprogramming, and B7-H3 checkpoint blockade. To the best of our knowledge, this is the first nanovaccine architecture to enable coordinated immunomodulation. Its modular design supports the clinical translation of solid tumors and personalized immunotherapy. Keywords: nanovaccine, HBc VLP, TAM reprogramming, B7-H3 checkpoint blockade, tumor immune microenvironment