RESUMO
We have shown previously that human monoclonal antibodies are not very immunogenic in rhesus monkeys, with only one monkey out of five mounting an anti-monoclonal antibody response. Two additional monkeys have been injected multiple times with much larger amounts of one human monoclonal antibody. No anti-antibody response has been detected in these monkeys. Structural changes that occur in the monoclonal antibodies over time in vivo have been investigated by Western Blots using anti-idiotypic antisera. Sodium dodecyl sulfate gel electrophoresis reveals that very little antibody has altered molecular weight. Isoelectric focusing patterns change more dramatically. Forms of the antibodies with lower isoelectric points appear in the serum. These forms have a similar in vivo half-life as the freshly prepared antibody. Very low pI forms of the monoclonal antibodies are not detected in the serum. Isoelectric focusing can be used to determine the in vivo or in vitro condition of a monoclonal antibody preparation. Finally, the monkey anti-human IgG that arose in the single monkey studied previously has been further characterized.
Assuntos
Anticorpos Monoclonais/administração & dosagem , Animais , Anticorpos Anti-Idiotípicos/biossíntese , Anticorpos Monoclonais/isolamento & purificação , Meia-Vida , Humanos , Imunoglobulinas/metabolismo , Focalização Isoelétrica , Macaca mulatta , Peso MolecularRESUMO
Five rhesus monkeys were injected multiple times over several months with two different human IgG1 monoclonal antibodies, both directed against human cytomegalovirus. Three monkeys each injected four times with monoclonal antibody EV2-7 for over 200 days showed no response other than a normal decay in antibody level. The in vivo half life of this antibody was substantially longer when measured with an idiotype-specific two site immunoassay than with radiolabeled antibody, indicating that the iodination procedure greatly affected the stability of the antibody. Although there was considerable individual variation in the half-life of EV2-7, from 8.9 to 30.5 days, the half-life was fairly long, especially considering the size of the monkeys. Two monkeys were injected with monoclonal antibody EV1-15. One monkey has responded in a similar manner to the EV2-7-injected monkeys. However, the other monkey has produced an anti-idiotypic antibody (or antibodies) of high affinity. It is possible that this response was triggered by the unusual physical nature of antibody EV1-15 or the effect of the species difference between human and rhesus monkey. In any case, the results from these five monkeys indicate that human monoclonal antibodies should have a significant advantage over mouse monoclonal antibodies for in vivo therapeutic applications.
Assuntos
Anticorpos Monoclonais/imunologia , Macaca mulatta/imunologia , Macaca/imunologia , Animais , Anticorpos Anti-Idiotípicos/imunologia , Humanos , Alótipos de Imunoglobulina/imunologia , Idiótipos de Imunoglobulinas/imunologia , Taxa de Depuração Metabólica , Especificidade da Espécie , Fatores de TempoRESUMO
Four hybridoma cell lines stably secreting human monoclonal antibodies directed against hepatitis B surface antigen have been isolated. The monoclonal antibodies have been characterized by determining several allotypes, measuring affinity for HBsAg, binding to two HBsAg subtypes, and kinetics of antibody binding to solid adsorbed antigen. In addition, the relative position of the epitopes have been determined by competition in binding to HBsAg. The results indicate that human anti-HBsAg monoclonal antibodies of quite different properties can be isolated. Pharmacokinetics of one antibody have been measured in two rhesus monkeys. This monoclonal antibody, OST 577, has been compared to hyperimmune gamma globulins in a Bureau of Biologics approved radioimmunoassay. It is expected that these antibodies will be useful in preventing and treating hepatitis B.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos de Superfície da Hepatite B/imunologia , Animais , Afinidade de Anticorpos , Reações Antígeno-Anticorpo , Ligação Competitiva , Ensaio de Imunoadsorção Enzimática , Epitopos , Humanos , Hibridomas , Isoanticorpos , Macaca mulatta , MasculinoRESUMO
Human monoclonal antibodies, owing to their decreased immunogenicity, are expected to be an improvement over mouse monoclonal antibodies for in vivo therapy. Human and primate monoclonal antibodies are best produced with a human x mouse heteromyeloma. Several human chromosomes are stable in the human x (human x mouse) hybrids. Chimpanzee anti-digoxin monoclonal antibodies were prepared and characterized. Because they are structurally very similar to human antibodies, they should be well tolerated in humans. The anti-digoxin antibodies can be used for therapy of extreme overdoses or as an in vivo diagnostic tool for slight overdoses. Because the advantage of using human monoclonal antibodies is their lack of immunogenicity, preparation of the antibody must be scrupulous so as not to introduce extraneous immunogens. Analysis to ensure the purity of the preparation can be complicated by the presence of high concentrations of the antibody and the low levels of contamination that must be detected. We describe a Western blot assay for Protein A that is sensitive even in the presence of human IgG.