CpG/OX40 原位疫苗联合抗血管生成药物增强小鼠全身性抗肿瘤效应

Objective: To investigate the systemic anti-tumor effects and immune mechanisms of CpG oligonucleotide and OX40 agonist antibody in situ vaccine (CpG + OX40) combined with anti-angiogenic drug anlotinib in the treatment of mouse ovarian cancer. Methods: A bilateral (primary and metastatic) ID8 cell mouse ovarian cancer model was established. And tumor-bearing mice were treated with anlotinib, CpG + OX40, or CpG + OX40 + anlotinib (the triple treatment), respectively. The anti-tumor effects of different treatment groups were evaluated by monitoring tumor volume and survival time. The changes of immune cells and cytokines in the tumor microenvironment were detected using flow cytometry and ELISA. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the relative expression levels of molecules reflecting vascular density and maturity in tumor tissues. Results: Compared with other treatment groups, the CpG + OX40 + anlotinib treatment significantly inhibited tumors on the treatment side (primary side) and the untreated side (metastatic side) (P < 0.01), and significantly improved the survival time of tumor-bearing mice (P < 0.05). The flow cytometry analyses showed that the CpG + OX40 + anlotinib treatment significantly increased the proportion of tumor infiltrating CD4⁺ T and CD8⁺ T cells in tumors at both the treated and untreated sides (P < 0.05). Immune cell depletion confirmed that when CD4⁺ T, CD8⁺ T, or NK cells were depleted alone, there was no significant difference in the inhibitory effects of the triple treatment on primary-side tumors, while the anti-tumor effects on metastatic-side tumors were significantly weakened but were still stronger than those of the PBS group (P < 0.01). When all three types of immune cells were depleted simultaneously, there was no statistically significant difference in their tumor suppressive effects compared with the PBS group (P > 0.05). The ELISA results showed that compared with other treatment groups, the triple treatment group showed a significant increase in Th1-related cytokines in the primary-side and metastatic-side tumors (P < 0.05), and a significant decrease in the expression of Th2-related cytokines (P < 0.05). The qRT-PCR results showed that, compared with the control group, the triple treatment group exhibited significantly lower CD31 expression (P < 0.0001) and a significantly higher Ang-1/Ang-2 ratio (P < 0.001) in tumor tissues on both sides. Conclusion: The combination of CpG + OX40 in situ vaccine and anlotinib has stronger systemic anti-tumor effects. Adaptive immunity, innate immunity and vascular density regulation play a crucial role in its anti-tumor effects, which provide potential treatment options for advanced cancer patients.