title: humMR1, a highly specific humanized single chain antibody for targeting EGFRvIII.
abstract: Production of an efficient humanized single chain antibody is reported here to specifically target EGFRvIII, a truncated receptor expressed in a wide variety of human cancers. CDR loops of MR1, a phage display-derived murine single chain antibody developed against this mutant receptor, were grafted on human frameworks that had been selected based on similarity to MR1 in terms of two distinct parameters, variable domain protein sequence and CDR canonical structures. Moreover, two point mutations were introduced in CDR-H2 and CDR-H3 loops of the humanized antibody to destroy its cross-reactivity to wild-type EGFR. The resultant antibody, referred to as humMR1, was found by MTT assay, ELISA and western blot techniques to be highly specific for EGFRvIII. The affinity of this antibody for EGFRvIII-specific 14-amino acid synthetic peptide and HC2 cells were measured to be 1.87 × 10(10) and 2.17 × 10(10)/M respectively. This humanized antibody leads to 78.5% inhibition in proliferation of EGFRvIII-overexpressing cells.
authors.affiliation: Biotechnology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran.
authors.forename: Yaghoub
authors.initials: Y
authors.lastname: Safdari
authors.affiliation: Biotechnology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran. Electronic address: farajnias@tbzmed.ac.ir.
authors.forename: Safar
authors.initials: S
authors.lastname: Farajnia
authors.affiliation: Paramedical Faculty, Tabriz University of Medical Sciences, Tabriz, Iran.
authors.forename: Mohammad
authors.initials: M
authors.lastname: Asgharzadeh
authors.affiliation: Biosensor Research Center, Endocrinology and Metabolism Molecular-Cellular Sciences Institute, Tehran University of Medical Sciences, Tehran, Iran.
authors.forename: Kobra
authors.initials: K
authors.lastname: Omidfar
authors.affiliation: Pharmaceutical Sciences Research Center, School of Pharmacy, Mazandaran University of Medical Science, Sari, Iran.
authors.forename: Masoumeh
authors.initials: M
authors.lastname: Khalili
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D066246:ErbB Receptors / Q000276:immunology*; D015395:Histocompatibility Antigens Class I / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D006801:Humans; D015778:Minor Histocompatibility Antigens; D008969:Molecular Sequence Data; D009154:Mutation; D011487:Protein Conformation; D016415:Sequence Alignment; D057127:Single-Chain Antibodies / Q000494:pharmacology*
chemical_list: D015395:Histocompatibility Antigens Class I; C094840:MR1 protein, human; D015778:Minor Histocompatibility Antigens; C490628:Mr1 protein, mouse; D057127:Single-Chain Antibodies; C410723:epidermal growth factor receptor VIII; D066246:ErbB Receptors
keywords: EGFRvIII; Humanization; Single chain antibody; Specificity