融合蛋白anti-CD19(Fab)-LDM对B细胞淋巴瘤BJAB细胞杀伤作用及其机制研究  

Cytotoxicity of Engineered Fusion Protein Anti-CD19 (Fab)-LDM on B-cell Lymphoma BJAB Cells and Its Mechanism

作　　者：范冬梅 杨圆圆 姜琳琳 熊冬生 杨铭 陶欣勇 周珊 刘红芹 屈浩 韩博文 石釧 FAN Dong-mei;YANG Yuan-yuan;JIANG Lin-Iin;XIONG Dongsheng;YANG Ming;TAO Xin-yong;ZHOU Shan;LIU Hong-qin;QU Hao;HAN Bo-wen;SHI Chuan(State Key Laboratory of Experimental Hematology ,Insitute of Hematology & Hospital of BloodDiseases,Chinese Academy of Medical Sciences & Peking Union Medical College,Tianjin300020,China;Tianjin Tongsheng Shidai Biotechnology Co.Ltd.,Tianjin 300020,China)

机构地区：[1]中国医学科学院北京协和医学院血液病医院血液学研究所实验血液学国家重点实验室,天津300020 [2]天津同生时代生物技术有限公司,天津300020

出　　处：《中国肿瘤》2019年第3期220-226,共7页China Cancer

基　　金：国家科技重大专项(2012ZX09102301)

摘　　要：[目的]探讨融合蛋白anti-CD19(Fab)-LDM对B细胞淋巴瘤细胞株BJAB细胞的杀伤作用及其机制研究。[方法]利用流式细胞分析技术(FACS)和激光共聚焦显微镜技术,检测融合蛋白anti-CD19(Fab)-LDM与BJAB细胞的结合活性。MTT法检测融合蛋白anti-CD19(Fab)-LDM对BJAB细胞的体外杀伤活性。彗星电泳实验检测融合蛋白anti-CD19 (Fab)-LDM对BJAB细胞DNA的损伤。FACS检测不同浓度融合蛋白anti-CD19(Fab)-LDM处理BJAB细胞后细胞周期的变化。[结果] FACS和激光共聚焦显微镜技术实验结果表明,融合蛋白anti-CD19(Fab)-LDM能与BJAB细胞结合。MTT法实验结果表明,融合蛋白anti-CD19(Fab)-LDM对BJAB细胞杀伤活性较单用力达霉素或阿霉素强,IC50值分别为(0.15±0.02)nmol/L、(0.38±0.03)nmol/L和(57.15±2.30)nmol/L。彗星电泳实验结果表明,用5pmol/L的融合蛋白anti-CD19(Fab)-LDM及力达霉素处理BJAB细胞后,可引起细胞不同程度的DNA损伤,由于融合蛋白anti-CD19(Fab)-LDM具有靶向性和细胞内化的特性,对DNA造成的损伤较力达霉素组明显。FACS结果显示,随着融合蛋白anti-CD19(Fab)-LDM浓度的增加,S期的比例从53.78%升高至77.29%,呈剂量依赖性。[结论]融合蛋白anti-CD19 (Fab)-LDM可以靶向杀伤B细胞淋巴瘤细胞株BJAB,引起细胞周期阻滞,在B细胞淋巴瘤生物治疗中具有潜在的应用价值。[Purpose] To investigate the cytotoxicity of the engineered fusion protein anti-CD19 (Fab)-LDM to B-cell lymphoma BJAB cells and its mechanism.[Methods] The antigen binding activity of anti-CD19 (Fab)-LDM on BJAB cells was observed by FACS and confocal microscopy. The cytotoxicity of the engineered fusion protein anti-CD19 (Fab)-LDM to BJAB cells was tested by MTT assay. Comet assay was used to determine the DNA damage in BJAB cells induced by antiCD19 (Fab)-LDM. Changes of cell cycle of BJAB cells treated with anti-CD19 (Fab)-LDM were detected by FACS.[Results] FACS and confocal microscopy showed that the antibody bound specifically to BJAB cells. MTT assay demonstrated that the engineered fusion protein anti-CD19 (Fab)-LDM strongly enhanced the cytotoxicity to BJAB cells compared with adriamycin or lidamycin (LDM). The comet assay showed that cells treated with anti-CD19 (Fab)-LDM induced more DNA damage than cells treated with LDM. BJAB cells treated with anti-CD19 (Fab)-LDM resulted in cell cycle arrest in a concentration dependence manner.[Conclusion] The fusion protein anti-CD19 (Fab)-LDM can target to kill CD19-positive B lymphoma cells and induce cell cycle arrest. These findings may be useful in clinical practice for B-cell lymphoma treatment.

关 键 词：基因工程抗体 融合蛋白 力达霉素 B细胞淋巴瘤 细胞周期 

分 类 号：R733.4[医药卫生—肿瘤]