The goals of this proposal are to define bladder tumor associated antigens through the development or murine and human monoclonal monoclonal and to determine if a monoclonal antibody defined phenotype can separate functionally distinct but histologically similar malignant tumors. Murine monoclonal antibodies reactive with bladder tumor associated antigens will be produced using bladder cancer cell lines as the source of antigen. Cell membrane preparations and other means of purification, e.g., lectins, will be utilized for immunization prior to fusion. A variety of immunization schedules will be employed to enhance the production of antibodies to bladder tumor associated antigens. Human monoclonal antibodies will be produced by the fusion of human lymphocytes with the human myeloma cell line KR12. To increase the yield of monoclonal antibody with high specificity, lymphocytes for fusion will be obtained from regional lymph nodes of patients undergoing cystectomy or from the blood of patients receiving intravesical immunotherapy with bacillus Calmette Guerin. In addition, in vitro immunization of the lymphocytes prior to fusion will be performed. When possible, patient's lymphocytes will be immunized in vitro with autologous cultured tumor cells. This should significantly decrease the problem of immunization with alloantigens. Murine and human monoclonal antibodies that bind to bladder cancer cells will be evaluated as prognostic indicators in patients with superficial bladder tumors. This will be accomplished by correlating antibody binding to the patient's clinical courses. Antibody binding will be determined by immunoperoxidase assay on histologic and cytologic preparations. The assays will be evaluated both with standard light microscopy and with microspectrophotometry. The latter technique can quantify the degree of immunoperoxidase staining and can correlate that with the amount of DNA in the same cell. The experiments described above will define new bladder tumor associated antigens and develop new classifications of bladder cancer based on antigenic phenotype. This will lead to an increased understanding of bladder cancer and facilitate both the selection of appropriate therapy in individual patients and the development of new methods of treatment.
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