Development of a Novel Human Single Chain Antibody Against EGFRVIII Antigen by Phage Display Technology

摘要

Purpose: EGFRvIII as the mostcommon mutant variant of the epidermal growth factor receptor is resulting fromdeletion of exons 2–7 in the coding sequence and junction of exons 1 and 8through a novel glycine residue. EGFRvIII is highly expressed in glioblastoma, carcinoma of thebreast, ovary, and lung but not in normal cells. The aim of the presentstudy was identification of a novelsingle chain antibody against EGFRvIII as a promising targetfor cancer therapy.Methods: Inthis study, asynthetic peptide corresponding to EGFRvIII protein was used for screening a naive human scFv phagelibrary. A novel five-round selection strategy was used for enrichment of rarespecific clones.Results: After five rounds of screening, six positivescFv clones against EGFRvIII were selected using monoclonal phage ELISA, amongthem, only three clones had expected size in PCR reaction. The specificinteraction of two of the scFv clones with EGFRvIII was confirmed by indirectELISA. One phage clone with higher affinity in scFv ELISA was purified forfurther analysis. The purity of the produced scFv antibody was confirmed usingSDS-PAGE and Western blotting analyses.Conclusion: In the present study, a human anti- EGFRvIII scFv withhigh affinity was first identified from a scFv phage library. This study can bethe groundwork for developing more effective diagnostic and therapeutic agentsagainst EGFRvIII expressing cancers.