mIL21-SA双功能融合蛋白锚定治疗膀胱癌实验研究  

Anticancer Evaluation Against Bladder Cancer of mIL21-SA Bifunctional Fusion Protein

作　　者：王菲[1] 胡志明[1] 法萍萍[1] 白莉[1] 石小军[1] 罗满生[1] 李金龙[1] 高基民[2] 

机构地区：[1]南方医科大学生物技术学院生物治疗研究所,广东广州510515 [2]温州医学院浙江省医学遗传学重点实验室/浙江省模式生物技术与应用重点实验室,浙江温州325035

出　　处：《生物技术》2012年第4期83-87,共5页Biotechnology

基　　金：国家"863"计划项目(No.2006AA02Z4C4);广州市白云区科技计划项目(2009-SZ-40)资助

摘　　要：目的:制备链亲合素标记的鼠白细胞介素21融合蛋白(mIL21-SA),治疗小鼠表浅膀胱癌。方法:构建mIL21-SA-pET21质粒,BL21表达,Ni-NTA纯化,透析复性,Western blot鉴定,MTT法检测其对小鼠胸腺细胞的增殖作用,流式检测其对生物素化的MB49锚定修饰率。建立小鼠MB49表浅膀胱癌模型,将mIL21-SA锚定在小鼠膀胱表面进行治疗并观察小鼠存活时间。60d后对mIL21-SA治疗后存活小鼠进行二次攻击。结果:mIL21-SA可以促进T细胞增殖,且具有SA介导的高效结合已生物素化的MB49表面的功能(修饰率98.74%)。膀胱灌注60d后,mIL21-SA组有10只(10/25)存活,二次攻击后,仍有6只(6/10)存活。与对照组比较均有统计学意义(P<0.05)。结论:该实验研制了具有双重活性的mIL21-SA,mIL21-SA锚定修饰治疗表浅膀胱癌是一种有效的免疫治疗方法。Objective:To obtain streptavidin-tagged mIL21 bifunctional fusion protein and evaluate its anticancer activity against superficial bladder cancerl.Method:mIL21-SA-pET21 plasmid was constructed and expressed in BL21 host bacteria.The mIL21-SA fusion protein was purified through Ni-NTA affinity chromatography,refolded by dialysis and verified by Western blotting.The effect of mIL21-SA fusion protein on mouse thymocytes proliferation was evaluated by MTT.Flow cytometry was used to detect mIL21-SA fusion protein on the biotinylated MB49 tumor cells.A mouse orthotopic model of MB49 bladder cancer was established by perfusing MB49 cells into mouse bladders.The mIL21-SA fusion protein was immobilized on the biotinylated mucosal surface of the bladder wall.Treatment and observed survival.After 60 days of the MB49 implantation,the mIL21-SA-treated survivals were intravesical rechallenged with wild-type MB49 cells and monitored for survival.Result:The mIL21-SA fusion protein exhibited bifunctional activities,i.e.both stimulative effect for mouse thymocyte proliferation and SA-mediated high-affinity binding to biotinylated cell surfaces(anchoring modified rate was about 98.74%).After 60 days of the MB49 implantation,there were 10 out of 25 mice survived in mIL21-SA-treated group.More importantly,6 out of 10 mice still survived after a second MB49 tumor challenge.Conclusion:The mIL21-SA bifunctional fusion protein is successfully prepared and it is a effective immunotherapy for superficial bladder cancer.

关 键 词：链亲合素 白细胞介素21 锚定 表浅膀胱癌 

分 类 号：R737.14[医药卫生—肿瘤]