Construction of dual function plasmid co-expressing ENDO-VEGI_(151) and survivin-siRNA and its anti-tumor activity

Objective: To construct a dual function expressional plasmid pCDNA3.1-ENDO-VEGI151/survivin-shRNA(pEV/si-survivin),and study its effect on the proliferation and apoptosis of breast cancer MDA-MB-231 cells and vascular endothelial cells(HUEVCs),so as to evaluate its feasibility for gene therapy of cancer.Methods: The efficient siRNA sequences targeting survivin was screened in MDA-MB-231 cells;the pEV/si-survivin expression vector was constructed and transfected into MDA-MB-231 and HUEVC cells,and the expression levels of ENDO-VEGI151 and survivin were detected by the real-time PCR and Western blotting analysis;MTT assay was used to detect the proliferation inhibition in the cells of the two groups after transfection;and flow cytometry was used to detect the changes of cell cycles and apoptosis.Results: The dual function recombinant plasmid pEV/si-survivin was successfully constructed and it was correctly expressed in both MDA-MB-231 and HUEVC cells.The plasmid significantly inhibited the expression of survivin and the cell proliferation(inhibition rate being [39.36±4.16]% at 48 h and [48.43±3.49)% at 72 h);it also significantly promoted cell apoptosis([18.33±1.48]% vs [4.80±1.01]%,P0.01) and induced cell cycles arrest(P0.05) in MDA-MB-231 cells.The plasmid also significantly inhibited cell proliferation(inhibition rate being [38.16±3.37]%) at 48 h and [53.75±4.53]% at 72 h),promoted apoptosis,and arrested the cell cycles(P0.05) in HUEVC cells.Conclusion: The dual function expressional plasmid pEV/si-survivin possess both angiogenesis inhibition and apoptosis promotion functions,and is expected to exert synergistic effect in vivo to improve the therapeutic outcome for patients with cancer.