Production of Human Anti-Peptide Antibody for Clinical Use by In Vitro Immunization

Abstract

Human monoclonal antibodies have a large potential to be used for diagnosis and therapy of various diseases, such as cancer, allergy, and so on. However, there exist some problems on sensitizing antigens to the human in order to obtain the antigenspecific human antibodies because of ethical and moral reasons. In vitro immunization method (IVI), in which primary B cells are activated to produce antigen-specific B lymphocytes, was established to solve this problem. In our previous study, we have established an IVI of human peripheral blood lymphocytes (PBL) against soluble proteins such as Cholera toxin B subunit (CTB), KLH or Rice allergic protein (RA) (Ichikawa et al., 1999). Furthermore, using a fragment peptide of CTB or RA, we have demonstrated that peptide antigen can be useful as sensitising antigen on IVI. However, some of proteins were difficult to produce antibodies because of serious effect on PBL on IVI, such as cytotoxicity, autoimmune response, change in the cytokine balance, and so on. TNF-α is a pleiotropic cytokine primarily produced by activated macrophages to response to a variety of inflammatory agents, so it is difficult to use as sensitizing antigen on IVI. Furthermore, Anti-TNF-α chimeric monoclonal antibody Infliximab have been reported to improve symptoms of rheumatoid arthritis and Crohn's disease, and approved for treatment of these disease (Targan et al., 1997, Maini et al., 1999). In this study, we tried to establish an in vitro immunization protocol against peptide fragments derived from tumor necrosis factor-α (TNF-α) for production of antibody for clinical use.