基于4-1BBL和CD3双信号的融合蛋白协同抗肿瘤作用研究  

Anti-tumor Effects of the Combined Administration of Fusion Protein Based on 4-1BBL and CD3 Signals

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作  者:姜文国[1] 刘荣[2] 刘芳[2] 王金宏[2] 邵晓枫[2] 舍鸣[2] 朱慧芳[2] 

机构地区:[1]滨州医学院药理学教研室药物研究所,滨州256603 [2]北京协和医学院中国医学科学院血液学研究所实验血液学国家重点实验室,天津300020

出  处:《生物技术通报》2010年第10期205-209,共5页Biotechnology Bulletin

基  金:山东省自然科学基金(Q2006C02)

摘  要:旨在研究4-1BBL/CD20融合蛋白增强抗CD3/抗CD20 diabody介导的特异性靶向杀伤活性。采用亲和层析法纯化本室构建的抗-CD3/抗-CD20 diabody和4-1BBL/CD20融合蛋白可溶性表达产物;采用calcein释放试验测定其介导的体外靶向杀伤活性;采用人B淋巴瘤细胞系Raji裸鼠移植瘤模型测定其介导的体内靶向杀伤活性。纯化4-1BBL/CD20融合蛋白在体外能增强抗-CD3/抗-CD20 diabody介导激活的T细胞杀伤Raji细胞;在人B淋巴瘤细胞系Raji裸鼠移植瘤模型联合人T淋巴细胞4-1BBL/CD20融合蛋白增强抗-CD3/抗-CD20 diabody高效抑制Raji细胞裸鼠移植瘤的生长,明显延长荷瘤裸鼠的生存时间。在体外和体内4-1BBL/CD20融合蛋白均能增强抗-CD3/抗-CD20 diabody介导激活的T细胞杀伤表达CD20抗原的肿增细胞,是一个有望用于B细胞恶性肿瘤临床治疗的特异性融合蛋白。This study was to examine a recombinant human 4-1BB ligand (4-1BBL) /anti-CD20 fusion protein activity, alone and in combination with an anti-CD3/anti-CD20 bispecific diabody. Human peripheral blood lymphocytes (PBLs) were isolated by Ficoll-Hypaque density-gradient centrifugation. The Annexin-V assay was performed to study the inhibition of PBL apoptosis by 4-1BBL/ CD20. Transcript expression of Bcl-xL and Bfl-1 was investigated by RT-PCR. Cytotoxicity analysis was performed by using calcein- AM retention assays. Nude mice bearing a xenotransplanted Raji Burkitt' s lymphoma were used to observe in vivo. The results showed that combination treatment with the 4-1BBL/anti-CD20 fusion protein and the anti-CD3/anti-CD20 diabody led to significantly in- creased T cell cytotoxicity to human B lymphoma cells in vitro and drastically more potent tumor inhibitory activity in vivo in xenografted B-cell lymphoma in nude mice. Mechanistic studies revealed that the combination treatment remarkably inhibited apoptosis of human peripheral blood lymphocytes, accompanied by up-regulation of Bcl-XL and Bfl-1. The combined administration of 4-1BBL/CD20 and diabodycould strongly potentiate the antitumor activity of the diabody, thus may have significant clinical application in the treatment of human CD20-positive B cell malignancies.

关 键 词:4—1BBL CD20 靶向治疗 融合蛋白 

分 类 号:R730.5[医药卫生—肿瘤]

 

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