Construction, Expression And Targeting Studies Of Mouse ScFv, Humanized ScFv And Their Fusions To TNFα Against Hepatocellular Carcinoma

Based on the mouse anti-human monoclonal antibody HAb25 single chain Fv fragment with partially deleted light chain variable domain gene,we subtituted the defective gene with it's intact light chain variable domain gene and reconstructed a mouse anti-human hepatocellular carcinoma(HCC) single chain Fv(mscFv) .We linked it with E-tag and obtained high fusion expression in prokaryotic expression vector pET15b.The activity of the purified product showed that mscFv has preserved the specificity and affinity of it's parental antibody quite well.Based on this, we linked mscFv and it's humanized anti-HCC scFv gene (hscFv, constructed by Yuan Qing'an from The Institute of Military Medical Science) with human TNFα gene respectively,and constructed the prokaryotic expression vector of anti-HCC bifunctional antibody pGEX 4T-1 m/hscFv-TNFα with high expression.The activity of purified products confirmed that the m/hscFv-TNFα retained specificity of HAb25.They not only showed selective cytotoxicity to target cells (SMMC-7721),but also induced inhibitary effect on HCC xenografts in nude mice,Among them mscFv-TNFα caused complete remission.