The products of oncogenes frequently localize to the cell membrane and may possess extracellular domains. Transformation of cells by high molecular weight DNA from human tumors can be mediated by oncogenes and may also result in the expression of new tumor-associated antigens. The use of NIH 3T3 cells transfected with high molecular weight DNA from human tumors as an immunogen has resulted in the production of human tumor--selective monoclonal antibodies. The purpose of this proposal is to investigate the immunotherapeutic potential of monoclonal antibody 45-2D9 produced by immunizing with the NIH 3T3 tertiary c-Ha-ras transformant 45-342. The NIH 3T3 tertiary transfectant 45-342 expresses a 74,000 Mr glycoprotein recognized by the 45-2D9 antibody. The 45-2D9 has been conjugated to the A chain of ricin. The stability, structural integrity, and biologic activity of the 45-2D9 immunotoxin will be deterimined in vivo and compared to the native antibody. Structural half-life and clearance studies of 125I labeled 4502D9 and immunotoxin will be performed will monitoring by SDS- polyacrylamide gel electrophorsis. Biodistribution studies of radiolabeled 4502D9 antibody and immunotoxin will be performed in mice bearing 45-342 cells will spontaneously metastasize in nu/nu mice and form lung colonies in irradiated Balb/c mice following intravenous injection. Thus an ideal model of established disseminated solid tumor is available to study the therapeutic potential of the 45-2D9 antibody and immunotoxin. Potentiators of immunotoxin activity and biochemical modifications of the immounotoxin will be evaluated in vivo. Modification of the immunotoxin by use of antibody fragments, more stable toxin-antibody linkages, and antibody-potentiator conjugates will be studied. Possible synergistic therapeutic effects between the antibody, immunotoxin, and other biologic response modifiers (IL-2, LAK cells, interferon) will be investigated.
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