Lentiviral vector construction of RGD- TAT-KDR siRNA fusion gene and assessment of its antitumor activity

ObjectiveTo construct a lentiviral vector of transcriptional activator protein-kinase insert domain receptor(TAT-KDR) siRNA fusion gene mediated by arginyle-glycyl-d-aspartic acid(RGD) and to assess its effects on human lung cancer A549 cells.MethodsTwo oligonucleotide chains of coding RGD were synthetized and then cloned into downstream of thrombin-antithrombin III complex(TAT) in pGC/TAT-KDR siRNA vector to construct TAT-RGD-KDR siRNA fusion gene vector.Lentivirus was packaged and used to infect A549 cells.The expression level of KDR gene in A549 cells was detected with Western blot and real-time PCR.The proliferation,apoptosis and invasion ability of A549 cells were measured with 3-4,5-dimethylthiazol-2-yl-2,5 diphenyltetrazolium bromide(MTT) method,dual color flow cytometry,and transwell invasion assay.ResultsSequencing and Basic Local Alignment Search Tool(BLAST) alignment confirmed that the RGD sequence was consistent with the design of the recombinant vector and the vector could reduce expression level of KDR mRNA and protein in A549 cells and inhibit the proliferation and invasiveness and promote apoptosis of A549 cells in vitro.ConclusionThe lentiviral vector of TAT-RGD-KDR siRNA fusion gene can inhibit the proliferation and invasion and promote the apoptosis of A549 cell line by inhibiting the expression level of KDR mRNA and protein.