CCL4调控肿瘤浸润Tregs表型与功能影响EGFR-TKI获得性耐药非小细胞肺癌抗PD-1/PD-L1疗效的机制

EGFR突变晚期NSCLC抗PD-(L)1治疗有效率差，但机制未明。课题组前期发现EGFR-TKI获得性耐药NSCLC对免疫治疗具有原发耐药特征；其肿瘤微环境中CD39+Treg浸润显著增多，CD8+T细胞浸润减少。scRNA-seq分析发现CCL4在EGFR-TKI获得性耐药NSCLC肿瘤中表达上调。结合既往文献我们推测：EGFR-TKI获得性耐药NSCLC通过CCL4激活Treg细胞内JAK/STAT通路上调CD39表达，进而影响抗PD-(L)1疗效。本项目拟通过EGFR转基因小鼠模型、Treg清除实验、ATP水解实验等，采取流式细胞术、IHC、RT-PCR、ELISA等分子生物学方法阐明EGFR-TKI获得性耐药NSCLC通过CCL4影响肿瘤微环境的细胞与分子机制，并在临床样本中验证。本项目的开展对EGFR-TKI获得性耐药NSCLC患者后线免疫联合治疗策略的开发具有重要的指导意义。

The efficacy of anti-PD-(L)1 based immunotherapy in advanced NSCLC with EGFR mutation is poor, and the mechanism is unclear. Our group previously found that advanced NSCLC patients with acquired resistance to EGFR-TKI showed resistance to immunotherapy, and there was significantly increased infiltration of CD39+Treg and decreased infiltration of CD8+ T cells in their tumor microenvironment. scRNA-seq analysis found that CCL4 was upregulated in the tumor cells of those patients with acquired resistance to EGFR-TKI. Based on our findings and previous reports, we infer that advanced NSCLC after acquiring resistance to EGFR-TKI upregulates the expression of CD39 on Treg by secreting CCL4 through JAK/STAT pathway, which in turn affects the efficacy of anti-PD-(L)1 treatment. Our project aims to clarify the specific cellular and molecular mechanisms of how advanced NSCLC with EGFR-TKI acquired resistance remodels the tumor microenvironment, and experiments and methods including EGFR transgenic mouse model, Treg depletion, ATP hydrolysis, flow cytometry, IHC, RT-PCR and so on will be adopted. The findings will be verified in clinical tissue samples. This project will provide rationality for the anti-PD-1/PD-L1 antibody based combinational therapies in advanced NSCLC patients with EGFR mutation and resistance to EGFR-TKI.