pGEX┐e23（scFv）PE40融合基因的构建及其在大肠杆菌中的表达论文.docVIP

　　pGEX┐e23（scFv）PE40融合基因的构建及其在大肠杆菌中的表达论文 .freelembrane receptor（HER2）that belongs to the epidermal groily and has intrinsic tyrosine kinase activity ［1-4］ .HER-2gene is activated by amplifica-tion and overexpression of its protein product，origenic or transforming activity ［5-6］ .It has been reported that HER2is overexpressed in25%～30%of breast cancers，urinary bladder cancer and gastric carcino-ma.The study of breast cancer suggests that c-erbB-2has a direct role in the pathogenesis and clini-cal aggressiveness of c-erbB-2overexpressing tu-mors ［7，8］ .The patients etastasis and recurrence，shorter non-tumor life span and total life span ［9-13］ .The anti-p185humanized monoclonal antibody Herceptin can effectively kill several c-erbB-2positive cell lines and dramatically inhibit the groouse.As a re-sult，Herceptin had been approved by FDA of U S A to be used in the first line chemotherapy as an ad-junctive agent in c-erbB-2positive metastasis breast cancers in September，1998 ［14］ .In our study，eric immunotoxin termed e23（scFv）PE40.In e23（scFv）PE40，the variable domain of the light chain of mAb e23ain of the heavy chain，ainⅡandⅢof PE.Then munotoxin successfully in a prokaryotic fusion protein expression vector pGEX-4T. MATERIALS AND METHODS Materials pLNCX-e23（scFv）PE40that consisted of e23，an anti-p185single chain variable fragment（scFv）and PE40 Promega pany.NucleoTrap Gel Purification Kit Clontech pany.Restriction endonuclease and enzymes used in DNA cloning Takara.Anti-PE40serum a.Other agents ents of the VL5′sequence of e23（scFv）ited pany and an EcoRI recognition site an diploid fibroblasts is associated al groal model of human gas-tric cancer using a bination of anti-erbB-2monoclonal anti-bodies ［J］.Cancer Res，1992;52:2771-2776. ［3］Michael FP，Carlos CC.Expression of the HER2/neu proto-oncogene in normal human adult and fetal tissues ［J］.Onco-gene，1990;5:953-962. ［4］Chen SY，Yang AG.Potent antitumour activity of a neor-specific killer