title: Superantigen-SEA gene modified tumor vaccine for hepatocellular carcinoma: an in vitro study.
abstract: OBJECTIVE
  To construct an eukaryotic superantigen gene expression vector containing the recombinant gene of SEA and CD80 molecule transmembrane region (CD80TM), and to express staphylococcus enterotoxin A (SEA) on the membrane of hepatocellular carcinoma (HCC) cell to form a superantigen gene modified tumor vaccine for HCC.
  
  
  METHODS
  SEA and linker-CD80TM gene were amplified through PCR from plasmid containing cDNA of SEA and CD80. Gene fragments were then subcloned into the multiple cloning sites of retroviral vector pLXSN. Recombinant plasmid was transferred into HepG2 cells mediated with lipofectamine, positive clones were selected in culture medium containing G418. RT-PCR and indirect immunofluorescence studies confirmed that SEA was expressed specifically on HCC cell membrane. INFgamma-ELISPOT study demonstrated that SEA protein was expressed on the membrane of HCC cells. Cytotoxicity of HepG2-SEA primed CTLs (SEA-T) was analyzed by (51)Cr release assay. T cells cultured with rhIL-2 (IL-2-T) were used as control.
  
  
  RESULTS
  Restriction digestion and sequence analyses confirmed the correctness of length, position and orientation of inserted fusion genes. SEA was expressed on the surface of HepG2 cells, HepG2-SEA had strong stimulating effect on production of HepG2 specific CTL (P<0.001). SEA-T had enhanced cytotoxicity to HepG2 cells (P<0.05).
  
  
  CONCLUSIONS
  Tumor cell membrane expressed superantigen can be used to reinforce the immune effect of tumor cell vaccine for HCC, which provides a new method of the enhanced active immunotherapy for HCC.
authors.affiliation: Department of Pathology, Fourth Military Medical University, Xi'an 710032, China. suiyanf@fmmu.edu.cn
authors.forename: Shao-Ying
authors.initials: SY
authors.lastname: Lu
authors.forename: Yan-Fang
authors.initials: YF
authors.lastname: Sui
authors.forename: Zeng-Shan
authors.initials: ZS
authors.lastname: Li
authors.forename: Jing
authors.initials: J
authors.lastname: Ye
authors.forename: Hai-Long
authors.initials: HL
authors.lastname: Dong
authors.forename: Ping
authors.initials: P
authors.lastname: Qu
authors.forename: Xiu-Min
authors.initials: XM
authors.lastname: Zhang
authors.forename: Wen-Yong
authors.initials: WY
authors.lastname: Wang
authors.forename: Yu-Song
authors.initials: YS
authors.lastname: Li
mesh_terms: D018122:B7-1 Antigen / Q000235:genetics; D019496:Cancer Vaccines / Q000235:genetics*; D006528:Carcinoma, Hepatocellular; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000502:physiology; D004768:Enterotoxins / Q000235:genetics*; D015870:Gene Expression; D006801:Humans; D066298:In Vitro Techniques; D008113:Liver Neoplasms; D018089:Superantigens / Q000235:genetics*; D014162:Transfection
chemical_list: D018122:B7-1 Antigen; D019496:Cancer Vaccines; D004768:Enterotoxins; D018089:Superantigens; C026066:enterotoxin A, Staphylococcal