Separation of the Tumor Rejection Antigen of Rous Sarcoma Virus-induced Murine Fibrosarcoma

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Abstract
The tumor antigen capable of inducing tumor resistance (tumor rejection antigen; TRA) was separated and some of its physicochemical properties were characterized. Cytosol and plasma membrane fractions were separated from Rous sarcoma virus (RSV)-induced CSA1M tumor cells. Immunization with membrane but not cytosol fraction of these tumor cells together with complete Freund''s adjuvant resulted in complete protection against subsequent challenge with viable CSA1M cells. The TRA activity contained in the membrane fraction was recovered in the sodium dodecyl sulfate (SDS)-solubilized fraction after the SDS-extraction of CSA1M membranes. This CSA1M SDS-solubilized preparation gave protection against syngeneic RSV-induced CSA9F tumor cells as well as the homologous tumor cell type, but failed to induce resistance to RSV-unrelated tumor cells. The membrane or SDS-solubilized fraction from RSV-unrelated tumor cells was unable to generate anti-CSA1M protective immunity. Physicochemical analyses have demonstrated that TRA activity in the SDS-solubilized fraction was completely abolished by treatment with proteinase K but was only marginally affected after treatment with glycosidase mixture. When the SDS-solubilized preparation was applied to a Sephacryl S-300 superfine column, TRA activity was recovered in the range of molecular weight of 50-90 kD. Further fractionation of this TRA-positive fraction by SDS-polyacrylamide gel electrophoresis revealed that the molecular size of TRA is 56-68 kD. These results indicate that membrane proteins which were isolated from CSA1M tumor cells and have a molecular size of about 60 kD are capable of inducing RSV-induced tumor-specific in vivo protective immunity.