Suppression of Progression and Metastasis of Established Colon Tumors in Mice by Intravenous Delivery of Short Interfering RNA Targeting KITENIN, a Metastasis-Enhancing Protein

Abstract

KITENIN promotes invasion of mouse colon adenocarcinoma (CT-26) cells in vivo. Here, we studied the effects of in vivo KITENIN ablation on established tumors by using pSUPER vectors (pSUPER-KITENIN) producing short interfering RNA (siRNA). When pSUPER-KITENIN was given weekly or semiweekly for 1 month into tail vein of syngeneic mice that have established colon tumors, tumor size regressed markedly and metastases were inhibited. In mice injected with pSUPER-KITENIN, serum interleukin-2 (IL-2) and IFN-gamma increased and CD4+ and CD8+ T cells infiltrated in the regressed tumor tissues. These effects, observed beginning 2 days after i.v. injection, imply that immune response is involved in the antitumor action of pSUPER-KITENIN. Using a yeast two-hybrid assay, we identified two KITENIN-interacting proteins for the possible mediators of these actions: 90K protein, a known immune modulatory glycoprotein, and protein kinase C inhibitor (PKCI). 90K was increased in the culture medium from CT-26/antisense KITENIN/90K cells. Double culture of accessory cells with CT-26/antisense KITENIN/90K cells revealed increased secretion of IL-1 and IL-6. Overexpression of 90K in CT-26/antisense KITENIN cells further delayed tumor growth compared with that of CT-26/antisense KITENIN cells. Actin arrangement was distorted in CT-26/antisense KITENIN and CT-26/antisense PKCI cells, whereas overexpression of PKCI resulted in increased invasiveness to fibronectin. Thus, antitumor effects of KITENIN siRNA derives from both the generation of a tumor-specific immune response in vivo through increased 90K secretion from tumor cells and the suppression of tumor invasion in which PKCI is related to increased invasiveness. Moreover, siRNA targeting of KITENIN can function as a chemotherapeutic strategy against colon cancer.