BackgroundAntitumor therapeutic vaccines are generally based on antigenic epitopes presented by major histocompatibility complex (MHC-I) molecules to induce tumor-specific CD8⁺T cells. Paradoxically, continuous T cell receptor (TCR) stimulation from tumor-derived CD8⁺T-cell epitopes can drive the functional exhaustion of tumor-specific CD8⁺T cells. Tumor-specific type-I helper CD4⁺T (T_H1) cells play an important role in the population maintenance and cytotoxic function of exhausted tumor-specific CD8⁺T cells in the tumor microenvironment. Nonetheless, whether the vaccination strategy targeting MHC-II-restricted CD4⁺T-cell epitopes to induce tumor-specific T_H1 responses can confer effective antitumor immunity to restrain tumor growth is not well studied. Here, we developed a heterologous prime-boost vaccination strategy to effectively induce tumor-specific T_H1 cells and evaluated its antitumor efficacy and its capacity to potentiate PD-1/PD-L1 immunotherapy.MethodsListeria monocytogenesvector and influenza A virus (PR8 strain) vector stably expressing lymphocytic choriomeningitis virus (LCMV) glycoprotein-specific I-A^b-restricted CD4⁺T cell epitope (GP_61–80) or ovalbumin-specific CD4⁺T cell epitope (OVA_323-339) were constructed and evaluated their efficacy against mouse models of melanoma and colorectal adenocarcinoma expressing lymphocytic choriomeningitis virus glycoprotein and ovalbumin. The impact of CD4⁺T cell epitope-based heterologous prime-boost vaccination was detected by flow-cytometer, single-cell RNA sequencing and single-cell TCR sequencing.ResultsCD4⁺T cell epitope-based heterologous prime-boost vaccination efficiently suppressed both mouse melanoma and colorectal adenocarcinoma. This vaccination primarily induced tumor-specific T_H1 response, which in turn enhanced the expansion, effector function and clonal breadth of tumor-specific CD8⁺T cells. Furthermore, this vaccination strategy synergized PD-L1 blockade mediated tumor suppression. Notably, prime-boost vaccination extended the duration of PD-L1 blockade induced antitumor effects by preventing the re-exhaustion of tumor-specific CD8⁺T cells.ConclusionCD4⁺T cell epitope-based heterologous prime-boost vaccination elicited potent both tumor-specific T_H1 and CTL response, leading to the efficient tumor control. This strategy can also potentiate PD-1/PD-L1 immune checkpoint blockade (ICB) against cancer.