Short interfering RNA-mediated gene silencing of vascular endothelial growth factor: Effects on cellular proliferation in colon cancer cells

Hypothesis: By using short interfering RNA (siRNA) to inhibit the in vitro expression of vascular endothelial growth factor (VEGF) A, we hope to further investigate the presence of an autocrine loop in colon cancer cells. We hypothesize that VEGF inhibition will result in decreased cellular proliferation. Design: Human colon cancer cells were evaluated for the expression of VEGF and VEGF receptor 2 (VEGFR-2). In vitro assessments were then made of the ability of anti-VEGF siRNA to knock down expression of VEGF and the subsequent effect this decreased expression had on colon cancer cell proliferation. Setting: Surgical oncology research laboratory. Interventions: Human colon cancer cells from the RKO cell line were transfected with siRNA targeting the coding region of VEGF. Main Outcome Measures: Enzyme-linked immunosorbent assay, Northern blot analysis, and real-time quantitative polymerase chain reaction were performed to establish the ability of siRNA to decrease VEGF production. Proliferation assays were run on transfected and wildtype cells to establish concomitant decrease in VEGF expression and cellular proliferation. Results: The RKO colon cancer cells expressed both VEGF and VEGFR-2. Those cells transfected with siRNA targeting VEGF showed a 94% knockdown in VEGF expression and a 67% decrease in cellular proliferation. Conclusion: Colon cancer cells expressing VEGF and VEGFR-2 may possess an autocrine growth pathway that can be effectively targeted using RNA interference as an antiangiogenic therapy.