Our central hypothesis is that glycoproteins and proteoglycans in the pericellular matrix of human gliomas mediate tumor-host cellular interactions and play a central role in tumor growth and invasion. The objective is to identify and characterize these macromolecules by monoclonal antibody analysis of the pericellular matrix of cultured human glioma cells. Antibodies will be raised against two human glioma-derived cell lines, U- 251MG and N-31MG. Both lines express glial fibrillary acidic protein. Secreting hybridomas will be screened by enzyme-linked immunoassay on intact cells and isolated glioma cell matrix and by avidin-biotin immunohistochemistry on frozen sections of a human glioma. Antibodies showing specificity for glioma pericellular matrix will be used in the purification of antigen from cell cultures by immunoaffinity chromatography or immunoprecipitation. Radiolabelled antigen will be isolated from conditioned medium or solubilized matrix and analyzed by SDS- polyacrylamide gel electrophoresis and autofluorography. The ability of isolated antigen to enhance or inhibit glioma cell adhesion in vitro will be determined. In vivo localization of pericellular matrix antigens will be determined by immunohistochemical analysis of frozen sections of a broad spectrum of primary human neuropithelial tumors, normal tissue, and extra-neural neoplasms. Formalin-fixed, paraffin-embedded tumors will also be examined if the epitopes are resistant to such treatment. Human glioma xenografts will be produced in athymic (nude) mice by stereotaxic intracerebral tumor cell transplantation. The localization of pericellular matrix antigens in glioma xenografts will be determined immunohistochemically. Ultrastructural localization of antigens in both human brain tumors and intracerebral xenografts will be determined by immunoelectron microscopy. A specific monoclonal antibody- defined glioma-mesenchymal extracellular matrix antigen (2A6) will be studied in detail. Correlations between antigen expression, tumor morphology, and tumor invasiveness will be assessed with the goal of identifying the biologic and potential diagnostic or prognostic significance of antigen expression.
| McComb, R D; Miller, K A; Carson, S D (1991) Tissue factor antigen in senile plaques of Alzheimer's disease. Am J Pathol 139:491-4 |
