A small bispecific antibody construct expressed as a functional single-chain molecule with high tumor cell cytotoxicity.
Construction of a bispecific single-chain antibody derivative is described that consists of two different single-chain Fv fragments joined through a Gly-Ser linker. One specificity of the two Fv fragments is directed against the CD3 antigen of human T cells and the other is directed against the epithelial 17-1A antigen; the latter had been found in a clinical trial to be a suitable target for antibody therapy of minimal residual colorectal cancer. The construct could be expressed in CHO cells as a fully functional protein, while its periplasmic expression in Escherichia coli resulted in a nonfunctional protein only. The antigen-binding properties of the bispecific single-chain antibody are indistinguishable from those of the corresponding univalent single-chain Fv fragments. By redirecting human peripheral T lymphocytes against 17-1A-positive tumor cells, the bispecific antibody proved to be highly cytotoxic at nanomolar concentrations as demonstrated by 51Cr release assay on various cell lines. The described bispecific construct has a molecular mass of 60 kDa and can be easily purified by its C-terminal histidine tail on a Ni-NTA chromatography column. As bispecific antibodies have already been shown to be effective in vivo in experimental tumor systems as well as in phase-one clinical trials, the small CD3/17-1A-bispecific antibody may be more efficacious than intact antibodies against minimal residual cancer cells.
Mack
M
M
Institut für Immunologie, Munich, Germany.
Riethmüller
G
G
Kufer
P
P
D000595:Amino Acid Sequence; D000918:Antibody Specificity; D000970:Antineoplastic Agents / Q000633:toxicity*; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D007128:Immunoglobulin Fragments / Q000235:genetics / Q000633:toxicity*; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000633:toxicity*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000633:toxicity; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; C055966:immunoglobulin Fv