Targeting Mitochondrial Metabolic Reprogramming: Reversing CD8⁺ T Cell Exhaustion and Improving Cancer Immunotherapy Response

This study explores the reversal effect of targeting mitochondrial metabolic reprogramming on CD8⁺ T cell exhaustion within the tumor microenvironment and its synergistic mechanism when combined with immune checkpoint blockade therapy. B16-OVA melanoma and LLC lung cancer tumor-bearing mouse models were established. Using single-cell RNA sequencing, metabolic flux analysis, Seahorse cellular energy metabolism assay, and flow cytometry, we compared the mitochondrial function, metabolic characteristics, and exhaustion phenotypes of tumor-infiltrating CD8⁺ T cells with CD8⁺ T cells from the spleen/draining lymph nodes. By pharmacologically activating or overexpressing genes to intervene in the PGC-1α-mediated mitochondrial biogenesis pathway, we evaluated CD8⁺ T cell mitochondrial membrane potential, ROS levels, ATP production, oxygen consumption rate, and the ability to secrete IFN-γ, TNF-α, and Granzyme B. Further, combined treatment with anti-PD-1 was applied to observe tumor growth, survival, and the composition of tumor-infiltrating lymphocytes. Tumor-infiltrating CD8⁺ T cells exhibited reduced mitochondrial membrane potential, accumulated ROS, decreased oxygen consumption rate and ATP production, and showed increased expression of PD-1, TIM-3, LAG-3, and TOX, suggesting that metabolic dysfunction is closely related to T cell exhaustion. Targeting mitochondrial metabolic reprogramming can restore mitochondrial adaptability of CD8⁺ T cells, enhance their proliferation, cytokine secretion, and cytotoxicity, while reducing expression of exhaustion-associated molecules. The combination of this intervention with anti-PD-1 antibody significantly inhibited tumor growth, extended survival in tumor-bearing mice, and increased the proportion of stem-like/effector memory CD8⁺ T cells in tumors. Mitochondrial metabolic reprogramming is a key regulatory node for reversing CD8⁺ T cell exhaustion. Targeting this pathway can remodel the tumor immune microenvironment and enhance the response to immune checkpoint therapy, providing a potential new strategy for cancer immunotherapy.