lncRNA IL21-AS1经外泌体诱导肿瘤免疫逃逸微环境致卵巢癌铂类耐药的机制研究

Chemotherapy is one of the most effective and commonly used treatments for ovarian cancer. However, the development of chemoresistance limits its clinical application in ovarian cancer patients. Ovarian cancer immune microenvironment plays a key role in the development of chemoresistance. However, the mechanisms of platinum resistant cancer cells can confer drug resistance to sensitive cells needs to be elucidated. We found that the overexpression of long noncoding RNA IL21-AS1 in the exosomes of cisplatin-resistance ovarian cancer cell. We further demonstrated exosome-transferred IL21-AS1 could confer the resistant phenotype to recipient ovarian cancer parental cells. IL-21 production were increased when T cells feded with resistant cell derived exosomes in the Skov3/T-cell co-culture model. Interestingly, we found IL21-AS1 sharing the same microRNA response elements with PD-L1, and rasied the PD-L1 expression. In view of this, we made a scientific hypothesis that exosome-transmitted IL21-AS1 promotes tumor immune evasion induced ovarian cancer drug resistance by regulating IL-21 and PD-L1. In this project, we focus on the cross-talk between tumor cells and microenvironment, co-culture tumor microenvironment in vitro model and the mice transplanted tumor model will be used. We will use ELISA, flow cytometry, luciferase reporter gene system, RNA immune co-precipitation (RIP), FISH, tail intravenous injecting exosomes technology, discussed the immune regulation function of exosome-transmitted IL21-AS1 in tumor microenvironment, promoting ovarian cancer of platinum resistance mechanisms. We will identify a novel mechanism by which IL21-AS1 influences IL-21 and PD-L1 immune checkpoint signalling pathways to treat chemoresistant ovarian cancer.

卵巢癌耐药是制约生存率的关键因素，肿瘤免疫微环境在耐药性的形成中起重要作用，然而肿瘤细胞传播耐药表型机制不详。申请者前期发现：lncRNA IL21-AS1在卵巢癌铂类耐药株外泌体中高表达；摄入耐药细胞外泌体后，敏感细胞产生耐药性，T细胞分泌IL-21减少。进一步研究表明IL21-AS1与PD-L1间存在相同的miRNA应答元件，并上调PD-L1表达。由此提出外泌体携带IL21-AS1调控IL-21和PD-L1表达，诱导免疫逃逸微环境并传递耐药表型，产生殊途同归的促进卵巢癌耐药机制的科学假说。本项目将以体外细胞共培养肿瘤微环境模型、小鼠移植瘤模型为研究对象，运用尾静脉注射外泌体和基因操作双干预手段，通过流式细胞术、荧光素酶报告基因系统、RNA免疫共沉淀、FISH等技术，结合临床标本验证，从外泌体介导肿瘤细胞与微环境交互作用层面深入阐释内在机制，为卵巢癌铂类耐药建立新的科学认识。

卵巢癌耐药是制约生存率的关键因素，肿瘤免疫微环境在耐药性的形成中起重要作用，然而肿瘤细胞传播耐药表型机制不详，因此揭示卵巢癌免疫微环境特征及耐药形成机制是卵巢肿瘤研究的核心问题，对能否控制卵巢癌、提高患者生存率以及提高免疫治疗效果至关重要。在本研究中我们通过卵巢癌外泌体高通量测序分析发现lncRNA IL21-AS1在卵巢癌铂类耐药株外泌体中高表达；摄入耐药细胞外泌体后，敏感细胞产生耐药性，T细胞分泌IL-21减少。进一步研究表明IL21-AS1与PD-L1间存在相同的miRNA应答元件，并上调PD-L1表达。进一步细胞研究发现IL21-AS1可以促进卵巢癌细胞系的增殖能力，并调控诱导肿瘤干细胞不等向分化。通过进一步机制研究发现，IL21-AS1可以调控PI3K/AKT及Nanog/Sox信号通路相关基因表达。从外泌体介导肿瘤细胞与微环境交互作用层面深入阐释内在机制，为卵巢癌铂类耐药建立新的科学认识。此外，我们也通过生物信息学分析进一步揭示卵巢癌免疫微环境的分型，挖掘潜在免疫相关的风险因子，为后续深入研究卵巢癌免疫微环境的调控夯实基础。