MDM2 siRNA Inhibits Angiogenesis by Reducing Vascular Endothelial Growth Factor Production in Breast Cancer Cells

Objective  To investigate the inhibitory effects of murine double minute 2 (MDM2) small interfering RNA (siRNA) on vascular endothelial growth factor(VEGF) production in human breast cancer cells and VEGF-mediated angiogenesis in xenograft tissues.

Methods  siRNA sequences of MDM2 were chemically synthesized and then transfected into human breast cancer cell line MDA-MB-468 which overexpressed MDM2. After transfection, the cells were cultured under hypoxic condition for different period of time. VEGF expression in tumor cells was measured by Western blot. The level of VEGF protein secreted in the culture supernatant was quantified by enzyme-linked immunosorbent assay (ELISA). Further, the stably-transfected cells were transplanted into the nude mice. The tumor growth was analyzed. MDM2 protein expression of tumors was determined by Western blot. Serum levels of VEGF were also measured by ELISA. VEGF expression and microvessel density (MVD) were studied by immunohistochemistry.

Results  In MDA-MB-468 cells, gene silencing of MDM2 resulted in the down-regulation of VEGF secretion (P=0.006) . In vivo, tumorigenicity assay showed that MDM2 depletion restrained tumor growth (P=0.008) . Serum levels of VEGF were substantially decreased (P=0.008) . MVD in transplanted tumor tissue was also significantly reduced (P=0.003) .

Conclusion  MDM2 knockdown by siRNA causes a strong reduction of VEGF production in breast cancer cells and inhibition of angiogenesis in tumor transplants. The MDM2/VEGF pathway may open a new approach for anti-tumor angiogenesis and molecular targeted drugs development.