title: The first constant domain (C(H)1 and C(L)) of an antibody used as heterodimerization domain for bispecific miniantibodies.
abstract: Bispecific miniantibodies were constructed by genetically fusing the C(H)1 domain of an IgG1 to the C-terminus of a single-chain Fv fragment (scFv-425), specific for the EGF receptor, and fusing the C(L) domain of a kappa light chain to the C-terminus of a scFv specific for CD2 (scFv-M1). An efficient dicistronic gene arrangement for functional expression in Escherichia coli was constructed. Immunoblots demonstrated correct domain assembly and the formation of the natural C(H)1-C(L) disulfide bridge. Gel filtration confirmed the correct size, sandwich ELISAs demonstrated bispecific functionality, and SPR biosensor measurements determined binding to EGF-R in comparison to bivalent constructs. Bispecific anti-EGF-R/anti-CD2 miniantibodies are candidates for the immunotherapy of cancer.
authors.affiliation: Biochemisches Institut, Universität Zürich, Switzerland.
authors.forename: K M
authors.initials: KM
authors.lastname: Müller
authors.forename: K M
authors.initials: KM
authors.lastname: Arndt
authors.forename: W
authors.initials: W
authors.lastname: Strittmatter
authors.forename: A
authors.initials: A
authors.lastname: Plückthun
mesh_terms: D000595:Amino Acid Sequence; D018033:Antibodies, Bispecific / Q000096:biosynthesis / Q000737:chemistry*; D000918:Antibody Specificity; D001483:Base Sequence; D001666:Binding Sites, Antibody; D018801:CD2 Antigens / Q000276:immunology*; D003001:Cloning, Molecular; D019281:Dimerization; D066246:ErbB Receptors / Q000276:immunology*; D004926:Escherichia coli; D005803:Genes, Immunoglobulin; D006801:Humans; D007127:Immunoglobulin Constant Regions / Q000096:biosynthesis / Q000737:chemistry*; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000737:chemistry*; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis / Q000737:chemistry*; D007145:Immunoglobulin kappa-Chains / Q000096:biosynthesis / Q000737:chemistry*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D015183:Restriction Mapping
chemical_list: D018033:Antibodies, Bispecific; D018801:CD2 Antigens; D007127:Immunoglobulin Constant Regions; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007145:Immunoglobulin kappa-Chains; D011994:Recombinant Proteins; D066246:ErbB Receptors