The Radioimmune Effects of Carbon Ion Improved by PD-1 antibody via STING pathway to inhibit immune evasion in Osteosarcoma

Background Osteosarcoma is an aggressive malignancy characterized by therapy resistance, radioresistance, and immune evasion, contributing to poor prognosis. Carbon ion irradiation (CIR) offers superior physical and biological advantages for radioresistant tumors, yet recurrence persists. Given its potential to enhance immune activation, we investigated CIR immunomodulatory effects in osteosarcoma and evaluated its therapeutic synergy with immune checkpoint blockade. Methods Immunogenic cell death (ICD) markers were assessed via flow cytometry, ELISA, and chemiluminescence. LM8/C3H and K7M2/Balb/c subcutaneous osteosarcoma mouse models were established. Immune cell infiltration and exhaustion markers were analyzed by flow cytometry. Immunofluorescence, Western blot and qPCR were employed to assess cGAS-STING pathway activation. PD-L1 expression was validated by immunohistochemistry. STING-knockdown cell lines were constructed to investigate its role in PD-L1, ICD, MHC-I. Results CIR induced stronger ICD and upregulated MHC-I expression. CIR triggered cytoplasmic dsDNA release and robustly activated the cGAS-STING pathway. Both irradiation upregulated MHC-I and CRT via STING-independent pathways, while CIR specifically upregulated PD-L1 via STING. CIR promoted CD8 ⁺ T-cell infiltration and PD-1 expression. CIR alone showed stronger tumor growth inhibition than photons, but tumor regrowth accelerated after 24 days in both monotherapy groups. CIR combined with anti-PD-1 significantly suppressed tumor growth. Conclusion CIR enhances tumor antigen release, antigen presentation, and CD8 ⁺ T-cell infiltration in osteosarcoma while upregulating PD-1 expression. CIR activates the cGAS-STING pathway but concurrently induce immunosuppression via STING-mediated PD-L1 upregulation. The combination of CIRT and anti-PD-1 therapy achieves superior tumor control.