Tumor neoantigens, defined as tumor-specific antigens arising from somatic mutations, have shown great potential as targets for cancer vaccines in clinical studies. However, the number of neoantigens capable of effectively activating immune responses is quite limited. Over the past few decades, tumor neoantigen vaccines based on MHC-I epitopes that activate CD8+ T cells have been extensively studied. However, growing evidence suggests that CD4+ T cells are important in cancer immunotherapy. In contrast to CD8+ T cells, the receptors on CD4+ T cells exhibit a wider range of antigen peptide-MHC recognition, which can detect more tumor mutation antigens. In our earlier studies, a nitrated CD4+ T-cell epitope (NitraTh) was constructed as a novel CD4+ T-cell epitope that can enhance the immunogenicity of multiple tumor antigens. Therefore, we designed vaccines targeting MHC-II neoantigen epitopes using the nitrated T-cell epitope containing immunogenic amino acids. We found that vaccines conjugated with NitraTh exhibited enhanced immunogenicity. Crucially, the NitraTh-modified MHC-II tumor neoantigen vaccines increased the proportion of CD4+ T cells that infiltrate tumors and the spleen, elevated the expression of several cytokines with antitumor effects and facilitated the transformation of CD4+ T cells into Th1 cells, thereby reducing tumor growth. Additionally, the nitrated epitope has been shown to transform naïve CD4+ T cells into effector memory cells, thus facilitating enduring antitumor actions. The strategy of combining nitrated epitopes with MHC-II neoantigen epitopes confirms the significance of CD4+ T-cell immunity in cancer and may provide a novel approach for cancer vaccine design. SIGNIFICANCE STATEMENT: This study presents a novel design paradigm for tumor vaccines-combining MHC-II epitopes with nitrated CD4+ T-cell epitopes. This approach promotes the differentiation of CD4+ T cells toward a Th1 phenotype and generates long-lasting effector memory CD4+ T cells. Under the enhanced effects of CD4+ T cells, the vaccines we designed achieved superior antitumor efficacy and improved the immunosuppressive tumor microenvironment.
