Xu Jie-Jie, Yao Kun, Yu Chen-Jie, Chen Xi, Lu Mei-Ping, Sun Hua, Li Bai-Zhou, Ding Chuan-Ning, Zhou Feng
Department of Microbiology and Immunology, Nanjing Medical University, Nanjing, China.
Auris Nasus Larynx. 2006 Dec;33(4):441-6. doi: 10.1016/j.anl.2006.05.019. Epub 2006 Aug 24.
Adoptive immunotherapy with specific cytotoxicity T lymphocytes (CTLs) induced by dendritic cells (DCs) pulsed with tumor antigens plays a crucial role in the immunity against tumor. The purpose of this study was to assess the feasibility and efficacy of latent membrane protein 2A (LMP2A)-specific CTLs immunity against nasopharyngeal carcinoma (NPC) in vitro and in vivo.
DCs were generated by culturing the monocytes purified from human peripheral blood mononuclear cells (PBMCs) in cytokine cocktail containing granulocyte/macrophage colony-stimulating factor (GM-CSF), interleukin-4 (IL-4), and tumor necrosis factor-alpha (TNF-alpha). The phenotype of mature DCs was analyzed by fluorescence activated cell sorter (FACS). Mature DCs transfected with EBV-LMP2A recombinant adenovirus were co-cultured with autologous PBMCs to induce LMP2A-specific CTLs. The expression of the surface antigens such as CD3, CD4, CD8 and CD56 on CTLs were detected by FACS. The specific cytotoxicity of CTLs on target CNE cells expressing EBV-LMP2A was confirmed using cytotoxicity assay. The anti-tumor effect of LMP2A-specific CTLs in vivo was assessed in the mice tumor models implanted with CNE cells expressing EBV-LMP2A.
Mature DCs expressed typically morphologic characteristics and high level of surface markers (CD1a, CD83, CD86, CD80 and HLA-DR). LMP2A-transfected DCs could induce LMP2A-specific CTLs consisting of a majority of CD4+ and CD8+ T cells. Cytotoxicity assay confirmed that the LMP2A-specific CTLs displayed significant cytotoxicity on target CNE cells compared with the controls. The study in vivo demonstrated that the treatment using these specific CTLs retarded the growth of established tumor in the treated mice.
These findings suggest that DCs transfected with EBV-LMP2A recombinant adenovirus can elicit LMP2A-specific CTLs that have a specific killing effect on NPC in vitro and in vivo. The results provide experimental basis for the further immunotherapy of NPC in clinical trails.
用肿瘤抗原脉冲处理的树突状细胞（DCs）诱导的特异性细胞毒性T淋巴细胞（CTLs）进行过继性免疫治疗在抗肿瘤免疫中起关键作用。本研究的目的是评估潜伏膜蛋白2A（LMP2A）特异性CTLs免疫在体外和体内抗鼻咽癌（NPC）的可行性和疗效。
通过在含有粒细胞/巨噬细胞集落刺激因子（GM-CSF）、白细胞介素-4（IL-4）和肿瘤坏死因子-α（TNF-α）的细胞因子混合物中培养从人外周血单个核细胞（PBMCs）中纯化的单核细胞来生成DCs。通过荧光激活细胞分选仪（FACS）分析成熟DCs的表型。用EBV-LMP2A重组腺病毒转染的成熟DCs与自体PBMCs共培养以诱导LMP2A特异性CTLs。通过FACS检测CTLs上CD3、CD4、CD8和CD56等表面抗原的表达。使用细胞毒性测定法确认CTLs对表达EBV-LMP2A的靶CNE细胞的特异性细胞毒性。在植入表达EBV-LMP2A的CNE细胞的小鼠肿瘤模型中评估LMP2A特异性CTLs在体内的抗肿瘤作用。
成熟DCs表现出典型的形态特征和高水平的表面标志物（CD1a、CD83、CD86、CD80和HLA-DR）。LMP2A转染的DCs可诱导由大多数CD4 +和CD8 + T细胞组成的LMP2A特异性CTLs。细胞毒性测定证实，与对照相比，LMP2A特异性CTLs对靶CNE细胞显示出显著的细胞毒性。体内研究表明，使用这些特异性CTLs进行治疗可延缓治疗小鼠中已建立肿瘤的生长。
这些发现表明，用EBV-LMP2A重组腺病毒转染的DCs可引发LMP2A特异性CTLs，其在体外和体内对NPC具有特异性杀伤作用。这些结果为NPC在临床试验中的进一步免疫治疗提供了实验依据。
