Therapeutic human antibodies derived from PCR amplification of B-cell variable regions.
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).
Larrick
J W
JW
Palo Alto Institute of Molecular Medicine, Mountain View, California 94043.
Wallace
E F
EF
Coloma
M J
MJ
Bruderer
U
U
Lang
A B
AB
Fry
K E
KE
D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000096:biosynthesis* / Q000235:genetics / Q000627:therapeutic use; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000627:therapeutic use; D001483:Base Sequence; D016466:CHO Cells; D002461:Cell Line, Transformed; D006224:Cricetinae; D004247:DNA / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D005803:Genes, Immunoglobulin; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000235:genetics* / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000627:therapeutic use; D013745:Tetanus Toxoid / Q000276:immunology*; D014162:Transfection
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins; D013745:Tetanus Toxoid; D004247:DNA