title: [Construction of pVAX-WIF-1 Eukaryotic Expression Vector and Its Anti-tumor Effect on Lung Cancer].
abstract: OBJECTIVE
  WIF-1 is an important tumor-suppressing gene in lung cancer, and its encoding protein WIF-1 can reduce proliferation and promote apoptosis by inhibiting Wnt/β-catenin signaling in lung cancer. This study constructs a eukaryotic expression plasmid carrying WIF-1 using FDA-approved clinical plasmid pVAX and explores the anti-tumor effect of pVAX-WIF-1 on A549 lung cancer cells in vitro and vivo.
  
  
  METHODS
  The DNA fragment of human WIF-1 coding sequence was amplified by PCR and was cloned into the multiple cloning sites of eukaryotic expression vector pVAX to construct pVAX-WIF-1. A recombinant plasmid was transfected into lung cancer A549 cells, and the expression of WIF-1 genes was verified by Western blot after transfection. Subsequently, the effect of pVAX-WIF-1 on cell apoptosis and proliferation was identified by MTT assay, staining A549 cells with Hoechst 3235, and flow cytometry. Finally, the A549 subcutaneous xenograft was established to detect the effect of pVAX-WIF-1 on lung tumor growth in vivo.
  
  
  RESULTS
  The results of restriction enzyme digestion, PCR, and sequencing indicated that eukaryotic expression plasmid pVAX-WIF-1 was successfully constructed. The protein expression level of WIF-1 was increased in the transfected A549 cells. Further results showed that transfection with pVAX-WIF-1 significantly inhibited proliferation and promoted apoptosis in A549 cells. Moreover, pVAX-WIF-1 significantly inhibited the tumor growth of the A549 subcutaneous xenograft in vivo.
  
  
  CONCLUSIONS
  The recombinant eukaryotic expression vector pVAX-WIF-1 was successfully constructed. Transfection with pVAX-WIF-1 could significantly inhibit proliferation and promote apoptosis of lung cancer A549 cells and also effectively inhibit the tumor growth of the A549 subcutaneous xenograft in vivo. Our research can contribute to clinical applications of WIF-1 in lung cancer gene therapy.
authors.affiliation: Department of Medical Oncology, Sichuan Academy of Medical Sciences and Sichuan Provincial People' Hospital, 610072 Chengdu, China.
authors.forename: Ning
authors.initials: N
authors.lastname: An
authors.affiliation: State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, 610041 Chengdu, China.
authors.forename: Xinmei
authors.initials: X
authors.lastname: Luo
authors.affiliation: State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, 610041 Chengdu, China.
authors.forename: Sujuan
authors.initials: S
authors.lastname: Ye
authors.affiliation: State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, 610041 Chengdu, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, 610041 Chengdu, China.
authors.forename: Weihan
authors.initials: W
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, 610041 Chengdu, China.
authors.forename: Qianqian
authors.initials: Q
authors.lastname: Jiang
authors.affiliation: State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, 610041 Chengdu, China.
authors.forename: Wen
authors.initials: W
authors.lastname: Zhu
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics* / Q000378:metabolism / Q000627:therapeutic use; D000818:Animals; D017209:Apoptosis; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D005260:Female; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000503:physiopathology / Q000628:therapy*; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D010957:Plasmids / Q000235:genetics* / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism / Q000627:therapeutic use
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D012097:Repressor Proteins; C485569:WIF1 protein, human