抗间皮素CAR-T细胞治疗胰腺癌的体外研究  

In vitro study of anti-mesothelin CAR-T cells for pancreatic cancer

作　　者：田静 白天凯 张之勇 闫滨[1] TIAN Jing;BAI Tian-kai;ZHANG Zhi-yong;YAN Bin(College of Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan 250355, China;College of Basic Medicine, Shandong First Medical University, Jinan 250013, China)

机构地区：[1]山东中医药大学药学院,山东济南250355 [2]山东第一医科大学基础医学院,山东济南250013

出　　处：《陕西科技大学学报》2022年第4期67-73,共7页Journal of Shaanxi University of Science & Technology

基　　金：国家卫生部“重大新药创制”科技重大专项子项目(2014ZX09509001001);山东省医药卫生科技发展计划项目(202002050626)。

摘　　要：本研究旨在观察抗间皮素(mesothelin,MSLN)嵌合抗原受体修饰的T细胞(Chimeric antigen receptor-modified T cells,CAR-T)在体外对两种胰腺癌细胞的杀伤作用,以及猪苓多糖(PPS)、人参多糖(GPS)、黄芪多糖(APS)、香菇多糖(LNT)和灵芝多糖(GLP)等调节免疫药物对CAR-T细胞增殖及杀伤肿瘤效应的影响.首先通过构建慢病毒表达载体转染制备稳定表达抗MSLN CAR-T细胞,然后采用实时无标记细胞分析仪(RTCA)和乳酸脱氢酶(LDH)等两种检测方法体外实验验证CAR-T细胞对PANC-1、ASPC-1两种胰腺癌细胞的杀伤能力.最后采用RTCA、CCK8法检测PPS、GPS、APS、LNT和GLP多糖对CAR-T细胞增殖及杀伤肿瘤效应的影响.结果表明,体外构建的MSLN CAR-T细胞对PANC-1、ASPC-1这两种胰腺癌细胞都有较好的杀伤效应,并且对MSLN蛋白高表达的ASPC-1细胞杀伤率显著高于MSLN蛋白低表达的PANC-1细胞(p<0.01);PPS、GPS、APS、LNT以及GLP等调节免疫药物对CAR-T细胞杀伤肿瘤的影响较小,但是与对照组相比,0.156 mg/mL的灵芝多糖和0.625 mg/mL的香菇多糖对CAR-T细胞增殖有一定的促进作用.The aim of this study was to observe the in vitro killing effects of anti-mesothelin(MSLN)chimeric antigen receptor-modified T cells(CAR-T)on two types of pancreatic cancer cells,and the effects of polysaccharides of Polyporus(PPS),ginseng polysaccharides(GPS),Astragalus polysaccharide(APS),lentinan and Ganoderma lucidum polysaccharide(GLP)on the proliferation and tumor-killing effects of CAR-T cells by immunomodulatory drugs.Stably expressed anti-MSLN CAR-T cells were first prepared by transfection with a lentiviral expression vector,and then the killing ability of CAR-T cells against two types of pancreatic cancer cells,PANC-1 and ASPC-1,was verified in vitro using two assays,including real-time label-free cell analyzer(RTCA)and lactate dehydrogenase(LDH).Finally,the effects of PPS,GPS,APS,LNT and GLP polysaccharide on the proliferation and tumor-killing effects of CAR-T cells were examined by RTCA and CCK8 assay.The results showed that the in vitro-constructed MSLN CAR-T cells could effectively kill two types of MSLN-positive pancreatic cancer cells,and the killing rate of ASPC-1 cells with high MSLN protein expression was significantly higher than that of PANC-1 cells with low MSLN protein expression(p<0.01);the regulatory immune drugs such as PPS,GPS,APS,LNT and GLP had a cells had less effect on tumor killing,but 0.156 mg/mL of Ganoderma lucidum polysaccharide and 0.625 mg/mL of Shiitake mushroom polysaccharide had some promotion effect on CAR-T cell proliferation.

关 键 词：嵌合抗原受体T细胞(CAR-T) 胰腺癌 间皮素 体外杀伤 多糖 

分 类 号：R735.9[医药卫生—肿瘤]