抗人类表皮生长因子受体2/表皮生长因子受体双特异性抗体对人胰腺癌细胞PANC-1的靶向治疗  

The targeted therapy of a human epidermal growth factor receptor-2/epidermal growth factor receptor targeted bispecific antibody which against the pancreatic carcinoma in vitro and in vivo

作　　者：黄长山[1] 余伟[1] 王谦[1] 丁月超[1] 马超[1] 黄涛[1] 张浩[2] 

机构地区：[1]郑州大学附属肿瘤医院暨河南省肿瘤医院肝胆胰外科,450003 [2]上海中医药大学附属曙光医院普外科,201203

出　　处：《中华实验外科杂志》2017年第8期1330-1332,共3页Chinese Journal of Experimental Surgery

摘　　要：目的 通过同时靶向人类表皮生长因子受体-2（Her-2）和表皮生长因子受体（EGFR）分子的双特异性抗体抑制人胰腺癌细胞PANC-1体内外增殖.方法 通过基因合成获得曲妥珠单抗和西妥昔单抗的基因序列,经重叠聚合酶链反应（PCR）融合成目的基因,构建入表达载体,转化进毕赤酵母X-33进行表达,获得同时靶向Her-2和EGFR的双特异性抗体CT-BiAb;流式检测CT-BiAb 对胰腺癌细胞系PANC-1的结合能力;MTT法检测CT-BiAb对PANC-1细胞的增殖抑制;通过流式双染法检测给药后PANC-1细胞的凋亡率;建立PANC-1荷瘤小鼠动物模型,检测CT-BiAb的体内抗肿瘤活性.结果 通过毕赤酵母表达及纯化,获得CT-BiAb,经Western blot鉴定验证了CT-BiAb 表达及装配正确.流式细胞术检测CT-BiAb与PANC-1的结合率为50.9%.细胞增殖抑制实验,与PBS组相比,亲本抗体组与CT-BiAb组对PANC-1均有抑制作用,且抑制作用呈浓度依赖性.凋亡实验,CT-BiAb组PANC-1细胞凋亡率[（33.50±7.14）%]高于亲本曲妥珠单抗[（15.86±4.32）%,P=0.022],也高于西妥昔单抗组[（18.64±5.71）%,P=0.048].移植瘤模型实验,CT-BiAb 高剂量组肿瘤抑制率可以达到（73.76±10.21）%,明显优于亲本曲妥珠单抗[（32.55±14.42）%,P=0.001],也明显优于西妥昔单抗组[（52.63±8.47）%,P=0.006].结论 本文构建了CT-BiAb,其具有良好的体内外抗肿瘤活性,有效地抑制PANC-1的体内外增殖,为抗肿瘤治疗提供了新的思路.Objective To improve the anti-tumor effect in vitro and in vivo,a bispecific antibody （CT-BiAb）was constructed.Methods CT-BiAb was obtained by overlapping polymerase chain reaction （PCR） between variable region of Trastuzumab,G4S linker and variable region of Cetuximab.Pichia pastoris was used to express.The binding activity of CT-BiAb to PANC-1 was detected by flow cytometry.Methyl thiazol tetrazolium （MTT） assay was used to detect the inhibition of cell growth.And flow cytometry was used to detect the apoptosis of PANC-1 after treatment with CT-BiAb.Finally,mouse tumor model was generated by endermic injecting PANC-1 cells,and we used this model to detect the antitumor activity of CT-BiAb.Results CT-BiAb was purified and the Western blot result showed that CT-BiAb has a correct molecular weight.CT-BiAb maintained binding activity to PANC-1 and the binding ratio was 50.9%.From the MTT assay,CT-BiAb restrained cell growth.From the apoptosis analysis,we found the cell apoptosis ratio of CT-BiAb （33.50±7.14）%was better than patented Trastuzumab （15.86±4.32）% （P=0.022） and Cetuximab （18.64±5.71）% （P=0.048）.In vivo assay of the PANC-1 cell xenografts in nude mice showed that the tumor inhibitory rate of high dose group （73.76±10.21）% was better than patented Trastuzumab （32.55±14.42）% （P=0.001） and Cetuximab （52.63±8.47）% （P=0.006）.Conclusion We established and express CT-BiAb,which maintained the binding activity of parent antibody.CT-BiAb provide rationale and drug development strategy for dual inhibition of human epidermal growth factor receptor-2 （Her-2） and epidermal growth factor receptor （EGFR） signaling in pancreatic carcinoma.

关 键 词：人类表皮生长因子受体2 表皮生长因子受体 胰腺癌 双特异性抗体 

分 类 号：R735.9[医药卫生—肿瘤]