重组抗死亡受体5人-鼠嵌合抗体的表达和活性研究  被引量:3

Expression and tumoricidal activity of a human-mouse chimeric anti-DR5 antibody through a Furin/ 2A peptide

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作  者:李蒙[1] 吕付佳[1] 史娟[1] 刘彦信[1] 郑德先[1] 

机构地区:[1]中国医学科学院基础医学研究所医学分子生物学国家重点实验室 ,北京100005

出  处:《中华医学杂志》2011年第10期707-710,共4页National Medical Journal of China

摘  要:目的获得重组抗死亡受体5(DR5)全长人-鼠嵌合抗体表达并研究其生物学活性。方法利用具有自我剪切能力的弗林蛋白酶(Furin)/口蹄疫病毒2A多肽(F/2A)连接人.鼠嵌合抗体的重链和轻链,形成一个开放阅读框(ORF),插入慢病毒表达载体pWPXL,构建重组抗DR5全长人-鼠嵌合抗体表达载体pWPXL—HF2AL;采用Western印迹测定该嵌合抗体的表达和采用酶联免疫吸附试验(ELISA)测定其与抗原的结合特异性;使用四唑氮盐(MTS)比色法分析其对肿瘤细胞的杀伤活性。结果利用F/2A多肽技术表达的人.鼠嵌合抗体可在F/2A处自我剪切,人-鼠嵌合抗体的轻、重链表达对称并具有与其母本鼠源抗体相似的亲和力和杀伤多种肿瘤细胞的活性。如含3μg/ml嵌合抗体的培养基使结肠癌细胞HCT116、肝癌细胞SMMC7721、肺癌细胞A549和神经胶质瘤细胞U251的细胞存活率分别降低至20.6%±2.6%,35.1%±2.7%,76.1%±6.2%和15.6%±2.0%。结论成功实现F/2A多肽介导的全长人-鼠嵌合抗体的表达,为肿瘤的生物治疗提供了新的策略。Objective To express a full-length human-mouse chimeric anti-DR5 antibody from a single open reading frame with tumoricidal activity to various cancer cells. Methods The heavy and light chains of chimeric antibody were joined by the Furin and 2A (F/2A) self-cleavage peptide and cloned into a lentiviral vector of pWPXL. Then the HEK293 cells were infected with the constructed expression vector pWPXL-HF2AL. Western blot, enzyme-linked immunosorbent assay (ELISA) and MTS assay were used to detect the chimeric antibody expression, cleavage, binding affinity to the antigen and tumoricidal activity to various tumor cells. Results The recombinant chimeric antibody was successfully expressed from a single open reading frame in pWPXL-HF2AL construct. And it possessed a similar binding affinity to the parental murine counterpoint and strong tumorieidal activity to various cancer cells. For example, on the concentration of 3 μg/ml, it made the relative cells viability of HCT116, SMMC7721, A549 and U251 down to 20.6% ± 2.6%, 35.1% ± 2.7%, 76.1% ±6.1% and 15.6% ± 2.0% respectively. Conclusions The human-mouse chimeric anti-DR5 antibody of F/2A peptide is successfully expressed. Possessing a strong tumoricidal activity in various cancer cells, it may provide a novel strategy for cancer biotherapy.

关 键 词:肿瘤 治疗 嵌合抗体DR5 Furin/2A多肽 慢病毒载体 

分 类 号:R730.5[医药卫生—肿瘤]

 

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