miR-374b靶向PD-1/PD-L1信号通路增强CIK杀伤肿瘤细胞效应
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摘要
目的探讨miR-374b在增强细胞因子诱导杀细胞(CIK)抗肿瘤中的作用及机制。方法采用传统方法培养CIK,用流式细胞确定CIK细胞群且分选出CIK;流式分析Hep G2、PLC、H1299、A549的程序性坏死蛋白(PD)-L1表达;采用双荧光素酶系统检测miR-374b在PD-1上的结合,siRNA技术干扰降低miR-374b基因的表达;酶联免疫吸附(ELISA)检测CIK的γ干扰素(IFN-γ)表达;乳酸脱氢酶(LDH)释放法检测CIK体外杀伤肿瘤细胞的能力;过继回输检测CIK体内杀伤肿瘤细胞的能力。结果双荧光素酶系统显示miR-374b可以直接作用于PD-1基因上;Hep G2、PLC、H1299、A549肿瘤细胞系都有比较高的PD-L1表达;miR-374b抑制剂会降低CIK分泌IFNr的水平,抑制CIK在体内和体外对肿瘤细胞的杀伤能力;PD-1 siRNA会提高CIK分泌IFN-γ的水平,增强CIK在体内和体外对肿瘤细胞的杀伤能力。结论 miR-374b可以直接作用于PD-1基因的表达,抑制miR-374b可以增强CIK杀伤肿瘤能力,干扰PD-1表达可以提高CIK抑制肿瘤能力。
引文
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2 Benson DM Jr,Bakan CE,Mishra A,et al.The PD-1/PD-L1 axis modulates the natural killer cell versus multiple myeloma effect:a therapeutic target for CT-011,a novel monoclonal anti-PD-1 antibody[J].Blood,2010;116(13):2286-94.
3 Xie J,Tan ZH,Tang X,et al.miR-374b-5p suppresses RECK expression and promotes gastric cancer cell invasion and metastasis[J].World J Gastroenterol,2014;20(46):17439.
4 Jiang JT,Shen YP,Wu CP,et al.Increasing the frequency of CIK cells adoptive immunotherapy may decrease risk of death in gastric cancer patients[J].World J Gastroenterol,2010;16(48):6155-62.
5 Liu P,Chen L,Huang X.The antitumor effects of CIK cells combined with docetaxel against drug-resistant lung adenocarcinoma cell line SPC-A1/DTX in vitro and in vivo[J].Cancer Biother Radiopharm,2009;24(1):91-8.
6 Yun YS,Hargrove ME,Ting CC.In vivo antitumor activity of anti-CD3 induced activated killer cells[J].Cancer Res,1989;49(17):4770-4.
7 Spranger S,Koblish HK,Horton B,et al.Mechanism of tumor rejection with doublets of CTLA-4,PD-1/PD-L1,or IDO blockade involves restored IL-2 production and proliferation of CD8(+)T cells directly within the tumor microenvironment[J].J Immunother Cancer,2014;2(1):3.
8 Iwai Y,Terawaki S,Honjo T.PD-1 blockade inhibits hematogenous spread of poorly immunogenic tumor cells by enhanced recruitment of effector Tcells[J].Int Immunol,2005;17(2):133-44.
9 Miller PC,Clarke J,Koru-Sengul T,et al.A novel MAPK-microRNA signature is predictive of hormone-therapy resistance and poor outcome in ER-positive breast cancer[J].Clin Cancer Res,2015;21(2):373-85.