GENETICS AND BIOCHEMISTRY OF MYELOMA IG PRODUCTION

The ability to produce antibody molecules using genetic engineering and gene transfection has revolutionized our capacity to utilize antibodies as diagnostic and therapeutic agents. However, in order to be able to most effectively exploit this technology, it is critically important that we understand what structures on the antibody molecule determine its biologic properties. Only when we possess this knowledge will we be able to produce antibodies with the optimal combinations of properties. Therefore, we propose the following experiments designed to define the structure(s) determining the functional properties of human IgG, IgD and IgE. Cellular receptor binding is important in determining the effector functions and trafficking patterns of antibodies. Therefore we will define the residues in IgG binding the low affinity receptors, FcgammaRII and FcgammaRIII, which are important for mobilizing cytotoxic cells. We will also identify the structures important for binding the MHC like Fc receptor, FcRn. It is also possible to use genetically engineered antibodies as antigens. We will use our recombinant Abs to define the epitope recognized by monoclonal rheumatoid factors (RFs) to investigate the hypothesis that RFs from individuals with rheumatoid arthritis (RA) recognize different epitopes than do non-disease associated RFs. The half-life of a protein is an important factor in determining its in vivo effectiveness. IgG is unusual in that its catabolic rate is determined both by its structure and its serum concentration. We will use exon-shuffling and site-directed mutagenesis to identify the residues on IgG which contribute to these properties. Complement activation is a critical antibody function. We will extend our studies of complement activation to include an analysis of the contribution of antibody structure to steps in the complement cascade subsequent to the binding of Clq. We can now produce antigen specific IgE and IgD in large quantities. IgE is well recognized for its role in immediate hypersensitivity and we will now use our expression techniques to produce and characterize the newly described isoforms of IgE. We will also investigate the role of the carbohydrate present on IgD in both receptor binding and in facilitating the assembly of the H/2 L/2, IgD molecule.