纳米刀消融通过METTL3-m6A-CCL2募集MDSC促进肝癌复发的作用及机制研究

以不可逆电穿孔为原理的纳米刀消融（IRE）广泛应用于肝癌治疗，但复发率高。申请人已报道IRE可在早期激活CD8+T细胞抗肿瘤免疫（Cancer Lett 2021），但后续未能完全抑制复发，机制不明。预实验发现IRE治疗后的复发肿瘤中髓源性抑制细胞MDSC浸润增加，伴IFN-γ+CD8+ T细胞减少；中和抗体阻断MDSC作用可减少复发。进一步研究发现：IRE后介导MDSC募集的CCL2表达增加，伴随mRNA m6A修饰及其催化酶METTL3水平升高；敲低METTL3可抑制CCL2翻译，减少MDSC浸润。项目拟解决“IRE如何通过m6A修饰调控MDSC促进肝癌复发”的关键科学问题，验证科学假说：IRE通过上调肝癌细胞METTL3表达，增加CCL2 mRNA m6A修饰，促进其翻译来募集MDSC抑制抗肿瘤免疫，导致复发。本项目拟阐明肝癌IRE后复发的免疫抑制机制，为防治复发提供新思路。

Nanoknife ablation is widely used in the treatment of hepatocellular carcinoma (HCC) based on the principle of irreversible electroporation (IRE) , however, the tumor recurrence rate after IRE is high. The applicant has uncovered that IRE can activate CD8+ T cell anti-tumor immunity at the early stage (Cancer lett 2021), nevertheless, the positive immune effect fails to completely inhibit the recurrence, and the underlying mechanism remains unknown. Through the preliminary experiments, the applicant found an increased infiltration of the myeloid-derived suppressor cell (MDSC) in the recurrent HCC after IRE, while the level of IFN-γ+CD8+ T cells is significantly decreased. Blocking MDSC by the neutralizing antibody can reduce the tumor relapse. Further researches revealed that the expression level of chemokine CCL2 that recruits MDSC is increased after the insufficient IRE, accompanied by the elevated level of mRNA m6A modification and its catalyzing enzyme METTL3. METTL3 knockdown can inhibit CCL2 translation and reduce the MDSC infiltration. This project aims to answer the key scientific question “How does the IRE regulate MDSC to promote the HCCrecurrence via m6A modification?”, hence verifying the hypothesis that “The IRE ablation upregualtes the METTL3 expression in the HCC cells to increase the m6A modification of CCL2 mRNA, which enhances the CCL2 translation and triggers the MDSC aggregation to inhibit the anti-tumor immune response, thus resulting in the HCC recurrence”. This project intends to clarify the immunosuppressive mechanism leading to the HCC recurrence after IRE ablation, and to provide new ideas for preventing and treating the HCC recurrence.