Firstly, we will continue to conduct fusions with hyperimmunized rats to determine whether or not a monoclonal antibody that recognizes a common glioma antigen can be recovered. At the same time, we will proceed with the characterizations that we have begun with those six rat monoclonals that appear to have partial cross-reactivity to rat glioma cells. These will include studies to determine: (i) specificity against a variety of normal and malignant tissues, and (ii) capacity to exert a tumor-specific cytotoxic effect on rat glioma cells in concert with complement or lymphoid cells. Secondly, we will conduct some preliminary investigations into the nature of the antigen(s) recognized by these monoclones. Glioma cells will be radioiodinated by the lactoperoxidase technique, solubilized, and the membrane components that are recognized will be immunoprecipitated with each antibody. These will be electrophoresed in 7-12% polyacrylamide slab gels and the molecular weights estimated from fluorographs prepared using the dried gels. This will provide a preliminary estimation of the range of membrane components recognized by the antibodies and, together with the specificity data will assist in our selection of those monoclones that will comprise the panel for the in vivo localization studies. Thirdly, a single lot of several hundred milligrams of each antibody is being purified from culture supernates by affinity chromatography using rabbit anti-rat antibody conjugated to Sepharose 4B. Each will be radiolabeled with 125Iodine and its known binding characteristics re-evaluated. If radioiodination seriously interferes with antibody activity, internally-radiolabeled (75) Selenomethionine) antibodies will be generated and tested. However, preliminary studies with radioiodinated rat anti-rabbit erythrocyte monoclonals show little, if any, qualitative or quantitative difference in antibody activity. Finally, we will begin preliminary in vivo localization studies with normal rats to determine whether or not undetected specificities for normal tissue antigens are revealed by these in vivo studies. These in vivo studies will prepare the foundation for our efforts in the third year to: (i) determine the extent, and persistence, of in vivo localization of each antibody in rats bearing primary, autochthonous, avian sarcoma virus-induced gliomas, and (ii) to investigate the safety and efficacy of using these antibodies singly or in combinations with conventional therapies (radiation and chemotherapy) that have demonstrated value in significantly prolonging the survival of tumor-bearing rats.
| Malouf, N N; Coronado, R; McMahon, D et al. (1987) Monoclonal antibody specific for the transverse tubular membrane of skeletal muscle activates the dihydropyridine-sensitive Ca2+ channel. Proc Natl Acad Sci U S A 84:5019-23 |
| Malouf, N N; Taylor, S; Gillespie, G Y et al. (1986) Monoclonal antibody specific for the T-tubule of skeletal muscle. J Histochem Cytochem 34:347-55 |
| Takiguchi, M; Ting, J P; Buessow, S C et al. (1985) Response of glioma cells to interferon-gamma: increase in class II RNA, protein and mixed lymphocyte reaction-stimulating ability. Eur J Immunol 15:809-14 |
| Loeffel, S C; Gillespie, G Y; Mirmiran, S A et al. (1985) Cellular immunolocalization of S100 protein within fixed tissue sections by monoclonal antibodies. Arch Pathol Lab Med 109:117-22 |
