Construction of Hepatoma-targeting Recombinant Adenovirus Vector of Staphylococcal Enterotoxin A (SEA) and Identification of Its Biological Activity

[Purpose]To construct hepatoma-targeting recombinant adenovirus vector of staphylococcal enterotoxin A(SEA)gene.[Methods] A new transfer plasmid pShuttle2 was constructed with polyA signal sequence without CMV enhancer/promoter using the existing adenovirus transfer plasmids pShuttle and pShuttle-CMV.AFP enhancer,promoter and SEA gene was subcloned into pShuttle2 from the vectors pKS-EP or pMD18-T-SEA respectively.Then the recombinant plasmid was co-transformed into E.coli BJ5183 with backbone vector pAdEasy-1 to obtain recombinant adenovirus DNA.The recombinant adenovirus DNA was transfected into HEK293 cells to prepare adenovirus.After AFP-producing cell line Hepa1-6 and AFP-nonproducing cell lines B16 and NIH3T3 were infected by recombinant adenovirus,the expression of SEA on the surface of cell was detected using indirect immunofluorescent staining by fluorescent microscope and flow cytometry(FCM).The in vitro activity of membrane SEA to stimulate proliferation of lymphocytes was detected by 3H-TDR.[Results] SEA targeting expresion on the surface of AFP-producing Hepa1-6 cells could stimulate proliferation of lymphocytes in vitro but not on AFP-nonproducing B16 and NIH3T3 cells.[Conclusion] Hepatoma-targeting recombinant adenovirus vector of SEA gene is successfully constructed,which lays the foundation for further research on application of SEA in targeted genetherapy and immunological mechanisms for hepatoma.