Enhancement of anti-programmed cell death protein-1 immunotherapy in non-small cell lung cancer using arginine and citrulline supplementation

BACKGROUND: Immune checkpoint inhibitors (ICIs) have substantially improved outcomes in previously untreatable cancer cases, but their effectiveness as monotherapies is limited. In this study, we examined the role of citrulline in the lung cancer microenvironment and its potential synergistic effects with immunotherapy. METHODS: Murine lung cancer cells (CMT167) were subcutaneously implanted in mice to establish tumor models, which were then treated with the anti-programmed death 1 (PD-1) antibody either alone or in combination with arginine or arginine and citrulline. Tumor growth, survival rate, cytokine levels, immune cell populations, and metabolic marker expression were assessed using histological, immunostaining, flow cytometry, and serum analyses. RESULTS: Mice in the treatment groups exhibited significantly lower tumor volumes than those in the control group (control, 1,161.59±294.73 mm(3); anti-PD-1, 427.38±355.34 mm(3); anti-PD-1 plus arginine, 452.10±332.04 mm(3); anti-PD-1 plus arginine and citrulline, 198.45±236.22 mm(3); P<0.001). Furthermore, the anti-PD-1 plus arginine and citrulline group exhibited significantly improved progression-free survival compared to the control group (P<0.001). The anti-PD-1 plus arginine and citrulline group also showed a significantly higher number of tumor-infiltrating CD8(+) lymphocytes per high-power field (hpf) than the control group (control, 24.22±9.13 cells/hpf; anti-PD-1, 29.20±9.41 cells/hpf; anti-PD-1 plus arginine, 34.33±8.81 cells/hpf; anti-PD-1 plus arginine and citrulline, 46.56±10.01 cells/hpf). CONCLUSIONS: Arginine and citrulline supplementation facilitated CD8(+) lymphocyte infiltration into the tumor microenvironment, thereby augmenting the efficacy of lung cancer immunotherapy.