ABSTRACT
<p><b>OBJECTIVE</b>To investigate the pro-apoptotic effect of Her-2 targeted recombinant caspase-6 fusion protein on osteosarcoma SOSP-9607 cells.</p><p><b>METHODS</b>Recombinant immunocasp-6 was generated by sequential fusion of the genes of a signal peptide, a single-chain Her-2 antibody (e23sFv), a PEA translocation domain (PEA aa253-364) and an active caspase-6. The immunocasp-6 gene was cloned into pCMV plasmid to construct a kind of eukaryotic expression vector, i.e. pCMV-e23sfv-PE II-caspase-6 (abbr. pCMV-6) and transfected into SOSP-9607 cells. Murine xenograft models were randomly divided into two groups that received i.m. injections of liposome encapsulated pCMV-6 or pCMV alone. The tumor volume and weight of the nude mice and the tumor weight of the cured mice were observed and statistically analyzed. The morphological changes of the tumors were examined with HE staining, apoptotic morphology of the tumor was observed by TUNEL staining and the gene expression was analyzed by immunohistochemical staining.</p><p><b>RESULTS</b>The tumor growth of the mice in the treatment group was significantly slower than that of the control group (P = 0.001). The weight of the nude mice in the treatment group was significantly higher than that of the control group (P = 0.0002). The tumor weight of the mice in the treatment group was significantly lower than that of the control group (P = 0.0006). HE and TUNEL staining of the tumor of nude mice in the treatment groups showed typical characteristics of apoptosis, while normal structure was found in the control group. Furthermore, caspase-6 was not found in the tumor and muscle tissues in the control group, but only in the treatment group by immunohistochemistry.</p><p><b>CONCLUSION</b>Immunocasp-6 can selectively recognize and bind to and kill HER-2 positive osteosarcoma cells, therefore, to offer some foundation for the clinical treatment of osteosarcoma.</p>
Subject(s)
Animals , Humans , Mice , ADP Ribose Transferases , Genetics , Apoptosis , Bacterial Toxins , Genetics , Bone Neoplasms , Metabolism , Pathology , Caspase 6 , Genetics , Metabolism , Cell Line, Tumor , Exotoxins , Genetics , Mice, Inbred BALB C , Mice, Nude , Neoplasm Transplantation , Osteosarcoma , Metabolism , Pathology , Plasmids , Random Allocation , Receptor, ErbB-2 , Genetics , Recombinant Fusion Proteins , Genetics , Metabolism , Transfection , Tumor Burden , Virulence Factors , GeneticsABSTRACT
Objetive: To investigate whether apoptosis of SGC7901 cells can be induced by the expression of the recombinant gene of anti-HER2 ScFv/tBid. Methods: The recombinant anti-HER2 ScFv/tBid gene was cloned into vector pCMV and the recombinant plasmid was transfected into SGC7901 cells. The gene expression was detected by RT-PCR and immunofluorescent staining. Cell counting was carried out to show the effect of the gene transfection on cell growth. At the same time, significant apoptotic peak was detected by flow cytometry in recombinant anti-HER2 ScFv/tBid gene transfected cells. Results: The fusion protein of anti-HER2 ScFv/tBid was observed in the cytoplasm of transfected SGC7901 cells. The transfected cells displayed typical cell growth inhibition and apoptosis. Conclusion: Fusion protein of anti-HER2 ScFv/tBid can induce apoptosis of SGC7901.