Vascular Endothelial Growth Factor-C siRNA Delivered via Calcium Carbonate Nanoparticle Effectively Inhibits Lymphangiogenesis and Growth of Colorectal Cancer In Vivo

Abstract

A nonviral gene carrier, calcium carbonate (CaCO(3))-nanoparticle, was evaluated for the efficient in vitro and in vivo delivery of siRNA-targeting vascular endothelial growth factor-C (VEGF-C). The chemically synthesized CaCO(3)-nanoparticle has a 58-nm diameter and a + 28.6-mV positive surface charge. It is capable of forming a CaCO(3)-nanoparticle-DNA complex and transferring DNA into targeted cells with high transfection efficiency, while effectively protecting the encapsulated DNA from degradation. Further, the CaCO(3)-nanoparticle-DNA complex has no obvious cytotoxicity for LoVo cells, while a liposome-DNA complex exhibited measurable cytotoxicity. LoVo cells transfected with a VEGF-C-targeted small interfering RNA (siRNA) via the CaCO(3)-nanoparticle exhibits significantly reduced VEGF-C expression, as measured by real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay, whereas no decrease in VEGF-C expression is observed in cells treated by control transfection. Transfection of LoVo cells with VEGF-C siRNA via the CaCO(3)-nanoparticle also dramatically suppresses tumor lymphangiogenesis, tumor growth, and regional lymph-node metastasis in subcutaneous xenografts. Significant downregulation of VEGF-C messenger RNA expression in a subcutaneous xenograft derived from VEGF-C siRNA-treated LoVo cells was confirmed by real-time PCR, as compared to controls. We conclude that the CaCO(3)-nanoparticle is a novel, nonviral system for the effective delivery of siRNA for cancer gene therapy.