ICOS与4-1BB共刺激域增强CAR-T细胞在肺癌中的抗肿瘤活性  

ICOS and 4-1BB costimulatory domain enhance antitumor activity of CAR-T cells in lung cancer

作　　者：张志东 魏海龙 林振海 陈吉柏 彭文 ZHANG Zhidong;WEI Hailong;LIN Zhenhai;CHEN Jibo;PENG Wen(Department of Thoracic and Heart Tumor Surgery,Danzhou People's Hospital,Danzhou 571700,China)

机构地区：[1]儋州市人民医院胸心肿瘤外科,儋州571700

出　　处：《中国免疫学杂志》2022年第22期2753-2757,共5页Chinese Journal of Immunology

基　　金：海南省卫生健康行业科研项目(19A200156)。

摘　　要：目的:探究整合ICOS共刺激结构域的三代CAR-T细胞是否可对肺癌产生显著抗肿瘤活性。方法:采用慢病毒感染法构建二代CAR-T细胞(MSLN-BBz CAR-T)及三代CAR-T细胞(MSLN-ICOSBBz CAR-T),流式细胞术检测CAR-T细胞阳性率、效靶细胞共孵育后T细胞CD69表达以及肿瘤浸润T细胞抗肿瘤表型;荧光显微镜观察效靶细胞共孵育后T细胞增殖情况;活体成像检测肺癌细胞在小鼠体内的生长情况;HE染色检测CAR-T细胞对主要脏器的毒性。结果:流式细胞术结果显示,成功采用慢病毒构建MSLN-BBz CAR-T及MSLN-ICOSBBz CAR-T,阳性率约为40%;效靶细胞共孵育后,MSLN-ICOSBBz CAR-T较MSLN-BBz CAR-T CD69表达显著升高(P<0.001),同时MSLN-ICOSBBz CAR-T靶细胞杀伤能力更强(P<0.001);小鼠移植瘤模型显示,MSLN-ICOSBBz CAR-T比MSLN-BBz CAR-T拥有更强的体内抑瘤能力(P<0.01);肿瘤浸润T细胞分析发现,相较于MSLN-BBz CAR-T,MSLN-ICOSBBz CAR-T IFN-γ表达更高(P<0.0001),PD-1表达则较低(P<0.0001),提示其肿瘤杀伤活性更好;HE染色结果显示,MSLN-BBz CAR-T及MSLN-ICOSBBz CAR-T均未对主要脏器造成明显损伤。结论:ICOS协同4-BB共刺激结构域可显著提高CAR-T细胞对肺癌的抗肿瘤活性,为MSLN-ICOSBBz CAR-T对肺癌治疗的进一步探索提供了依据。Objective:To explore whether third generation of CAR-T cells integrated with ICOS costimulatory domain can produce significant anti-tumor activity against lung cancer.Methods:Second generation CAR-T cells(MSLN-BBz CAR-T)and third generation CAR-T cells(MSLN-ICOSBBz CAR-T)were constructed by lentivirus infection.Positive rate of CAR-T cells,expression of CD69 on T cells after co-incubation of effector and target cells and anti-tumor phenotype of tumor infiltrating T cells were detected by flow cytometry.Proliferation of T cells after co-incubation of effector and target cells was observed by fluorescence microscope.Growth of lung cancer cells in mice was detected by in vivo imaging.Toxicity of CAR-T cells to major organs was detected by HE staining.Results:Flow cytometry showed that MSLN-BBz CAR-T and MSLN-ICOSBBz CAR-T were successfully constructed by lentivirus and positive rates were about 40%.After co-incubation with effector and target cells,expression of CD69 of MSLN-ICOSBBz CAR-T was significantly higher than that of MSLN-BBz CAR-T(P<0.001).Cytotoxicity of MSLN-ICOSBBz CAR-T was stronger than that of MSLN-BBz CAR-T(P<0.001).Model of transplanted tumor in mice showed that MSLN-ICOSBBz CAR-T had stronger tumor inhibition ability in vivo than MSLN-BBz CAR-T(P<0.01).Analysis of tumor infiltrating T cells showed that IFN-γexpression of MSLN-ICOSBBz CAR-T was higher than that of MSLN-BBz CAR-T(P<0.0001),while PD-1 expression was lower(P<0.0001),suggesting that MSLN-BBz CAR-T had better tumor killing activity.HE staining showed that MSLN-BBz CAR-T and MSLN-ICOSBBz CAR-T did not cause significant damage to main organs.Conclusion:ICOS combined with 4-BB costimulatory domain can significantly improve antitumor activity of CAR-T cells against lung cancer,which provides a basis for further exploration of MSLN-ICOSBBz CAR-T in treatment of lung cancer.

关 键 词：肺癌 嵌合抗原受体T细胞 ICOS 免疫治疗 

分 类 号：R392[医药卫生—免疫学]