RECOMBINANT BIFUNCTIONAL MOLECULE FV/IFN-GAMMA POSSESSES THE ANTITUMOR FV AS WELL AS THE GAMMA-INTERFERON ACTIVITIES

Recombinant DNA techniques were used to clone, construct and express the bifunctional molecule FV/IFN-gamma. The FV/IFN-gamma is a single-chain 42KD fusion protein expressed in E. coli under control of the strong T7 bacteriophage promoter in the expression vector pT7-7-FV-IFN-gamma. The fused gene fragment FV-IFN-gamma containing a single-chain anti-TAG72 FV gene fragment as well as the human recombinant cDNA fragment of IFN-gamma molecule. The renatured soluble form of FV/IFN-gamma was purified from E. coli inclusion bodies using HTPT chromatography. The yield of this fusion protein was estimated at 10mg/L. Our data showed that the FV/IFN-gamma molecule retained the TAG72 antigen-binding specificity and the IFN-gamma activity as measured in ELISA, Western blotting and up-regulation of CEA expression by IFN-gamma. Therefore, it may prove to be useful in targeting the biological effect of IFN-gamma to tumor cells and stimulating its immune destruction.