Lu Jianfeng, Leng Xisheng, Peng Jirun, Mou Dongcheng, Pang Xuewen, Shang Xiaoying, Chen Weifeng
Department of Hepatobiliary Surgery, Peking University People's Hospital, Beijing 100044, China.
Chin Med J (Engl). 2002 Jul;115(7):1002-5.
To investigate the possibility of using melanoma antigen-1 (MAGE-1) peptide as a tumor vaccine to treat hepatocellular carcinoma (HCC).
The expressions of MAGE-1 in 8 HCC cell lines and in liver cancer tissue from a patient were detected using RT-PCR. The type of human leucocyte antigen I(HLA I) of both 8 HCC cell lines and peripheral blood mononuclear cells of the patient was detected using a microcytotoxicity method to screen out target cell lines for the cytotoxicity assay. Peripheral blood mononuclear cells from the HCC patient pulsed with an MAGE-1 peptide (NYKCRFPEI) were used as antigen presenting cells. Autogenous peripheral blood mononuclear cells were stimulated with antigen presenting cells every 7 days for 4 times to elicit cytotoxic T lymphocytes. The phenotype of effector cells was analyzed using flow cytometry. The cytotoxicity of effector cells was detected with a lactate dehydrogenase releasing assay.
The expressions of both MAGE-1 and HLA-A24 were detected in BEL7405 cell line which were used as the positive target cell line in the cytotoxicity assay. The expression of MAGE-1 alone was detected in HLE, BEL7402, BEL7404, QGY7703 and SMMC7721 cell lines, and the expression of neither MAGE-1 nor HLA-A24 was shown in QGY 7701 and HpG2 cell lines. The last 7 cell lines could be used as negative target cell lines in the cytotoxicity assay. Peripheral blood mononuclear cells expanded 32 folds during 28-day culture. The ratio of CD3(+) T cells increased by 16% (from 54% to 70%), and the ratio of CD8(+) T cells increased by 20% (from 36% to 56%) during 28-day culture. When the ratio of effector cells to target cells was 10:1, effector cells exhibited 62.5% cytotoxicity against autogenous lymphoblasts pulsed with the peptide (NYKCRFPEI) of MAGE-1 antigen, 40.25% cytotoxicity against BEL7405 cells, compared with 17.88% cytolysis observed against autogenous lymphoblasts, 19.55% against HLE cells, and 1.6% against QGY7701 cells. When the ratio of effector cells to target cells was 3.3:1, the cytotoxicity of effector cells against the peptide pulsed autogenous lymphoblasts was 53.6%, which was much higher against autogenous lymphoblasts, HLE cells and QGY7701 cells at 15.6%, 13% and 1%, respectively.
The results demonstrate that cytotoxic T lymphocytes with the ability to specifically lyse target cells expressing both MAGE-1 and HLA-A24 could be successfully induced by the MAGE-1 peptide NYKCRFPEI in vitro. This indicates that a good result might be anticipated if this peptide is used as a tumor vaccine to treat HLA-A24 HCC patients.
探讨使用黑色素瘤抗原-1（MAGE-1）肽作为肿瘤疫苗治疗肝细胞癌（HCC）的可能性。
采用逆转录聚合酶链反应（RT-PCR）检测8种肝癌细胞系及1例患者肝癌组织中MAGE-1的表达。采用微量细胞毒性法检测8种肝癌细胞系及患者外周血单个核细胞的人类白细胞抗原I（HLA I）类型，筛选出用于细胞毒性试验的靶细胞系。用MAGE-1肽（NYKCRFPEI）脉冲处理的肝癌患者外周血单个核细胞作为抗原呈递细胞。每7天用抗原呈递细胞刺激自身外周血单个核细胞4次，以诱导细胞毒性T淋巴细胞。采用流式细胞术分析效应细胞的表型。用乳酸脱氢酶释放试验检测效应细胞的细胞毒性。
在BEL7405细胞系中检测到MAGE-1和HLA-A24的表达，其作为细胞毒性试验的阳性靶细胞系。在HLE、BEL7402、BEL7404、QGY7703和SMMC7721细胞系中仅检测到MAGE-1的表达，而在QGY 7701和HpG2细胞系中未检测到MAGE-1和HLA-A24的表达。后7种细胞系可作为细胞毒性试验的阴性靶细胞系。外周血单个核细胞在28天培养期间扩增了32倍。在28天培养期间，CD3(+) T细胞比例增加了16%（从54%增至70%），CD8(+) T细胞比例增加了20%（从36%增至56%）。当效应细胞与靶细胞比例为10:1时，效应细胞对用MAGE-1抗原肽（NYKCRFPEI）脉冲处理的自身淋巴母细胞的细胞毒性为62.5%，对BEL7405细胞的细胞毒性为40.25%，而对自身淋巴母细胞、HLE细胞和QGY7701细胞的细胞溶解率分别为17.88%、19.55%和1.6%。当效应细胞与靶细胞比例为3.3:1时，效应细胞对肽脉冲处理的自身淋巴母细胞的细胞毒性为53.6%，对自身淋巴母细胞、HLE细胞和QGY7701细胞的细胞毒性分别为15.6%、13%和1%，明显更高。
结果表明，MAGE-1肽NYKCRFPEI在体外可成功诱导出具有特异性裂解表达MAGE-1和HLA-A24靶细胞能力的细胞毒性T淋巴细胞。这表明该肽作为肿瘤疫苗治疗HLA-A24肝癌患者可能会取得良好效果。
