结直肠癌细胞及其外泌体中调控肿瘤免疫分子PD-1/PD-L1表达的关键microRNAs筛选、鉴定及功能验证

Immune escape plays an important role in the occurrence and progression of malignant tumors such as colorectal cancer (CRC). Blocking PD-1/PD-L1 signaling pathway inhibits tumor immune escape and provides a new direction for tumor immunotherapy; The molecular mechanism of PD-1/PD-L1 expression regulation in CRC cells is not fully understood. In view of the previous work basis, we initially proposed that the levels of miRNAs (miR-195-5p, miR-378a-3p, etc.) in CRC cells and their exosomes decreased, causing PD-1 on the surface of tumor cells and CD8+ T lymphocytes. The expression of PD- L1 and PD-1 is up-regulated, which in turn inhibits the killing effect of CD8+ T lymphocytes on tumor cells, and finally the scientific hypothesis of tumor immune escape. This study aims to study the regulation of PD-1/PD-L1 expression and the anti-tumor immune response to T cells by using CRC cell lines and intestinal-specific miRNAs knockout mice as subjects. The impact of CRC extracellular body-associated miRNAs can be used as a new marker for screening PD-1/PD-L1 monoclonal antibody immunotherapy.

免疫逃逸在结直肠癌（CRC）等恶性肿瘤的发生和演进过程中发挥着重要的作用，阻断PD-1/PD-L1信号通路抑制肿瘤免疫逃逸为肿瘤免疫治疗提出了新的方向；然而CRC细胞中PD-1/PD-L1表达调控的分子机制目前尚不完全清楚。鉴于前期的工作基础，我们初步提出“CRC细胞及其外泌体中的miRNAs（miR-195-5p、miR-378a-3p等）水平下降，引起肿瘤细胞及CD8+T淋巴细胞表面的PD-L1和PD-1表达上调，进而抑制CD8+T淋巴细胞对肿瘤细胞的杀伤作用，最终发生肿瘤免疫逃逸” 这一科学假设。本课题拟以CRC细胞株及肠道特异性miRNAs敲除鼠为研究对象，深入探讨肿瘤外泌体相关miRNAs表达失调对PD-1/PD-L1表达的调控、以及对T细胞抗肿瘤免疫反应的影响；并初步明确CRC细胞外泌体相关miRNAs可否作为PD-1/PD-L1单克隆抗体免疫治疗适用人群筛选的新的标志物。

免疫逃逸在结直肠癌（CRC）等恶性肿瘤的发生和演进过程中发挥着重要的作用，阻断PD-1/PD-L1信号通路抑制肿瘤免疫逃逸为肿瘤免疫治疗提出了新的方向；然而CRC细胞中PD-1/PD-L1表达调控的分子机制目前尚不完全清楚。我们发现CRC细胞及其外泌体中miR-195-5p可以特异性靶向肿瘤细胞表面PD-L1及CD8+T细胞的PD-1，miR-195-5p与结直肠癌患者的Dukes分期、TNM分期显著负相关关系，二者表达具有明显相关性，低表达组患者生存时间明显低于高表达组；CRC细胞及外泌体miR-195-5p表达失调会引起肿瘤细胞及CD8+T淋巴细胞表面的PD-L1和PD-1表达上调，进而抑制CD8+T淋巴细胞对肿瘤细胞的杀伤作用，最终发生肿瘤免疫逃逸，促进肿瘤增殖；小鼠miR-195-5p并联合PD-L1免疫检测点抑制剂治疗可以有效降低结直肠癌肿瘤负荷。本项目的顺利完成，不仅可以为PD-1/PD-L1抑制剂抗肿瘤治疗提供新的筛选指标，而且有望为CRC患者的肿瘤免疫治疗提供新的分子靶点。