NY-ESO-1致敏树突状细胞诱导的CTL对肝癌细胞株的特异性杀伤作用  

CTL induced by NY-ESO-1-impulsed dendritic cells specifically kills NY-ESO-1 positive human hepatocellular caner cells

作　　者：张文敏[1] 陈裕庆[1] 张萌[2] 凌航[1] 文剑明[2] 

机构地区：[1]福建医科大学基础医学院病理学系,福建医科大学肿瘤研究所,福州350004 [2]中山大学附属第一医院病理科,广州510080

出　　处：《中国肿瘤生物治疗杂志》2009年第1期18-23,共6页Chinese Journal of Cancer Biotherapy

基　　金：福建省自然科学基金资助项目(No.C0610023);广州市科技计划资助项目(No.2003J1-C0221)~~

摘　　要：目的:探讨肿瘤睾丸抗原NY-ESO-1(New York-esophageal-1)致敏树突状细胞体外诱导特异性CTL对肝癌细胞株的杀伤作用。方法:重组质粒pGEX-ESO1经原核诱导表达并纯化GST-ESO1融合蛋白肽。重组人粒细胞-巨噬细胞集落刺激因子(rhGM-CSF)和白细胞介素4(rhIL-4)诱导培养人外周血来源的树突状细胞(dendritic cells,DCs),经GST-ESO1融合蛋白肽致敏后诱导特异性CTL增殖。以此CTL为效应细胞,分别以NY-ESO-1阳性表达的肝癌细胞株HepG2和不表达NY-ESO-1的肝癌细胞株H2P为靶细胞,MTT法检测CTL对肝癌细胞株的杀伤作用。结果:重组质粒pGEX-ESO1经IPTG诱导,在大肠杆菌中表达相对分子质量约36 000的GST-ESO1融合蛋白肽,纯化后的质量浓度为50μg/ml;经rhGM-CSF和rhIL-4联合诱导成功培养人外周血DCs,其表型分子HLA-DR为91.4%、CD86为70.5%、CD83为71.2%、CD80为55.3%。NY-ESO-1致敏的DCs能明显诱导CTL增殖,此CTL对肝癌细胞株HepG2的杀伤率显著高于GST刺激组、未致敏DC组和无DC刺激组(均P<0.05),效靶比为50∶1时杀伤效应达到最高峰[(53.23±3.78)%,P<0.01];相同条件下CTL对H2P细胞无特异性杀伤作用。结论:NY-ESO-1抗原致敏的DCs在体外可诱导同种CTL产生和增殖,后者对NY-ESO-1阳性肝癌细胞株具有特异性杀伤效应,该方法为肝癌免疫治疗提供了一条新思路。Objective ：To study the antigen specific anti-tumor effect of cytotoxic T lymphocyte （CTL）, which was induced by cancer testis antigen NY-ESO-l-impulsed dendritic cells （DCs）, against human hepatocellular carcinoma （HCC）. Methods： GST-ESO1 fusion protein was induced in recombinant pGEX-ESO1 vector transformed bacteria by IPTG, and the GST-ESO1 fusion protein was purified. DCs were induced with granulocyte/macrophage colony-stimulating factor （GM-CSF） and interleukin-4 （IL-4） from human peripheral blood mononuclear cells. DCs impulsed with GST-ESO protein peptide were co-cultured with T lymphocytes, and the resultant CTLs were used as effector cells. NY-ESO-1 posi- tive hepatocellular carcinoma HepG2 cells and NY-ESO-1 negative H2P cells were used as target cells to test the specific anti-tumor effect of CTL using MTF. Results： Escherichia coli Bl221 expressed fusion protein peptide GST-ESO1 （Mr 36 000） after transfection with recombinant pGEX-ESO1 vector. The concentration of GST-ESO1 peptide was 50μg/ml after purification. DCs were successfully induced with GM-CSF and IL-4 from human peripheral blood mononuclear cells. DCs-impulsed with NY-ESO1 had high expression of surface molecule such as HLA-DR（91.4% ）, CD86 （70.5%）, CD83（71.2% ） and CD80（55.3% ）. DCs-impulsed with NY-ESO-1 induced production and proliferation of CTL, and this CTL specifically killed NY-ESO-1 positive HepG2 cells. CTL induced by NY-ESO-1 had stronger cytotoxic effect against HepG2 cells compared with GST-impulsed DCs, un-impulsed DCs （P 〈 0.05）. Highest anti-tumor activity was found when the ratio of effector： target was 50 ： 1 （53.23 ± 3.78, P 〈 0.01 ）. Conclusion： DCs-impulsed with NY-ESO- 1 can induce production and proliferation of allogenic CTLs, which show antigen specific anti-tumor effect against NY-ESO-1 positive HCC cells. This result casts new lights on immunotherapy of HCC.

关 键 词：肝细胞癌 NY-ESO-1 树突状细胞 细胞毒性T淋巴细胞 

分 类 号：R735.7[医药卫生—肿瘤] R730.5[医药卫生—临床医学]