Chitosan nanoparticle-based delivery of fused NKG2D–IL-21 gene suppresses colon cancer growth in mice

Nanoparticles can be loaded with exogenous DNA for the potential expression of cytokines with immune-stimulatory function. NKG2D identifies major histocompatibility complex class I chain-related protein in human and retinoic acid early induced transcript-1 in mouse, which acts as tumor-associated antigens. Biologic agents based on interleukin 21 (IL-21) have displayed antitumor activities through lymphocyte activation. The NKG2D-IL-21 fusion protein theoretically identifies tumor cells through NKG2D moiety and activates T cells through IL-21 moiety. In this study, double-gene fragments that encode the extracellular domains of NKG2D and IL-21 genes were connected and then inserted into the pcDNA3.1(-) plasmid. PcDNA3.1-dsNKG2D-IL-21 plasmid nanoparticles based on chitosan were generated. Tumor cells pretransfected with dsNKG2D-IL-21 gene nanoparticles can activate natural killer (NK) and CD8+ T cells in vitro. Serum IL-21 levels were enhanced in mice intramuscularly injected with the gene nanoparticles. DsNKG2D-IL-21 gene nanoparticles accumulated in tumor tissues after being intravenously injected for ~4-24 h. Treatment of dsNKG2D-IL-21 gene nanoparticles also retarded tumor growth and elongated the life span of tumor-bearing mice by activating NK and T cells in vivo. Thus, the dsNKG2D-IL-21 gene nanoparticles exerted efficient antitumor activities and would be potentially used for tumor therapy.