Objective To investigate the efficacy of histone deacetylase (HDAC) inhibitor chidamide combined with the PD-1 inhibitor on CD8⁺ T cells anti-cancer function in OVA-expressing MC38 (MC38-OVA) colorectal-bearing mice. Methods Animal experiments: C57BL/6 tumor models were constructed by subcutaneously injecting MC38-OVA colorectal cancer cells into the back of mice. We randomized mice into control group, chidamide group, anti-PD-1 group and chidamide+anti-PD-1 group (20 each group). We monitored the tumor growth and animal survival rate of each group; we employed a flow-based method to detect the number and ratio of tumor-infiltrating CD8⁺ T cells, CD8⁺IFN-γ⁺ T cells, OVA antigen-specific CD8⁺ T cells, and the expression changes of regulatory T cells (Treg), myeloid-derived suppressor cells (MDSC), and tumor-associated macrophages (TAM). Cell experiments: We used a flow-based method to detect the apoptosis of CD8⁺ T cells and MC38-OVA tumor cells after 0, 10, 25, 50, 100, or 200 nmol/L chidamide treatment. The proliferation of CD8⁺ T cells and MC38-OVA tumor cells treated with 0 and 100 nmol/L chidamide was detected by Ki-67 antibody labeling and cell counting. To evaluate CD8⁺ T cell killing ability, we treated CD8⁺ T cells with various conditions (control group, chidamide group, anti-PD-1 group and chidamide+anti-PD-1 group) followed by co-culture with MC38-OVA tumor cells, using the flow-based method. In the condition that CD8⁺ T cells treated with 0 and 100 nmol/L chidamide co-cultured with the same number of MC38-OVA tumor cells, the expression of CD107a was detected by flow cytometry. Results Compared with control group, the tumor growth was inhibited (P<0.05) while the survival rate was improved (P<0.01) in chidamide+anti-PD-1 group. The number of tumor-infiltrating CD8⁺ T cells was significantly higher in chidamide group, anti-PD-1 group and chidamide+anti-PD-1 group than that in control group (P<0.05). Nonetheless, the ratio and levels of CD8⁺IFN-γ⁺ and OVA antigen-specific CD8⁺ T cells were significantly higher in chidamide+anti-PD-1 group than those in other groups (P<0.05). The in vitro experiment results showed that chidamide could enhance the killing ability of CD8⁺ T cells and the expression of CD107a. Conclusion Chidamide combined with PD-1 inhibitor significantly enhanced the number and function of tumor-infiltrating CD8⁺ T cells and increased antigen-specific CD8⁺ T cells, which will provide a theoretical and experimental basis for the combination of chidamide in clinical solid tumor immunotherapy.