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MAbs ; 7(1): 129-37, 2015.
Article in English | MEDLINE | ID: mdl-25523454

ABSTRACT

Immunization of mice or rats with a "non-self" protein is a commonly used method to obtain monoclonal antibodies, and relies on the immune system's ability to recognize the immunogen as foreign. Immunization of an antigen with 100% identity to the endogenous protein, however, will not elicit a robust immune response. To develop antibodies to mouse proteins, we focused on the potential for breaking such immune tolerance by genetically fusing two independent T-cell epitope-containing sequences (from tetanus toxin (TT) and diphtheria toxin fragment A (DTA)) to a mouse protein, mouse ST2 (mST2). Wild-type CD1 mice were immunized with three mST2 tagged proteins (Fc, TT and DTA) and the specific serum response was determined. Only in mice immunized with the T-cell epitope-containing antigens were specific mST2 serum responses detected; hybridomas generated from these mice secreted highly sequence-diverse IgGs that were capable of binding mST2 and inhibiting the interaction of mST2 with its ligand, mouse interleukin (IL)-33 (mIL-33). Of the hundreds of antibodies profiled, we identified five potent antibodies that were able to inhibit IL-33 induced IL-6 release in a mast cell assay; notably one such antibody was sufficiently potent to suppress IL-5 release and eosinophilia infiltration in an Alternaria alternata challenge mouse model of asthma. This study demonstrated, for the first time, that T-cell epitope-containing tags have the ability to break tolerance in wild-type mice to 100% conserved proteins, and it provides a compelling argument for the broader use of this approach to generate antibodies against any mouse protein or conserved ortholog.


Subject(s)
Antibodies, Monoclonal, Murine-Derived/immunology , Antibody Specificity , Epitopes, T-Lymphocyte/immunology , Receptors, Interleukin/immunology , Animals , Antibodies, Monoclonal, Murine-Derived/pharmacology , Asthma/drug therapy , Asthma/immunology , Asthma/pathology , Cell Line, Transformed , Diphtheria Toxin/chemistry , Diphtheria Toxin/immunology , Epitopes, T-Lymphocyte/chemistry , Female , Humans , Interleukin-1 Receptor-Like 1 Protein , Mice , Mice, Inbred BALB C , Rats , Receptors, Interleukin/chemistry , Tetanus Toxin/chemistry , Tetanus Toxin/immunology
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