1. Therapeutic human antibodies derived from PCR amplification of B-cell variable regions.
- Author
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Larrick JW, Wallace EF, Coloma MJ, Bruderer U, Lang AB, and Fry KE
- Subjects
- Amino Acid Sequence, Animals, Antibodies, Bacterial genetics, Antibodies, Bacterial therapeutic use, Antibodies, Monoclonal genetics, Antibodies, Monoclonal immunology, Antibodies, Monoclonal therapeutic use, Base Sequence, CHO Cells, Cell Line, Transformed, Cricetinae, DNA genetics, Enzyme-Linked Immunosorbent Assay, Humans, Hybridomas immunology, Immunoglobulin Heavy Chains genetics, Immunoglobulin Heavy Chains immunology, Immunoglobulin Variable Region immunology, Mice, Molecular Sequence Data, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins therapeutic use, Transfection, Antibodies, Bacterial biosynthesis, Antibodies, Monoclonal biosynthesis, Genes, Immunoglobulin, Immunoglobulin Variable Region genetics, Polymerase Chain Reaction, Recombinant Fusion Proteins biosynthesis, Tetanus Toxoid immunology
- Abstract
Despite advances in the in vitro immunization of human B cells (Borrebaeck et al. 1988) and the development of immunodeficient mice (McCune et al. 1988) for the reconstitution of the human immune system ex vivo, immortalization of antigen-specific human B cells remains the limiting step in the generation of human monoclonal antibodies. Typically this is performed with the aid of Epstein-Barr virus transformation followed by subcloning, confirmation of antigen binding and hybridization of the B lymphoblasts to a suitable fusion partner such as GLI-H7. This general approach is effective and widely used; however, it is time-consuming with erratic results. These were the immediate reasons we and others devised methods to directly obtain the variable regions from small numbers of human B cells (Larrick et al. 1987). The success of the PCR-based approach is illustrated above. In the present studies we successfully captured and stably produced antibodies from the V regions of two potent human anti-tetanus antibodies secreted by heteromyelomas that were too unstable for scale-up production. Although further preclinical evaluation of these antibodies is in progress, results to date indicate that the recombinant antibodies produced in myeloma-based cell lines or CHO cells are equivalent in binding specificity and activity to the native heteromyeloma-derived antibodies. Recent studies from this laboratory indicate that effective anti-tetanus protection will require a cocktail of anti-tetanus antibodies. Details of this work will be the subject of a future communication (Lang et al., in preparation).
- Published
- 1992
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