Development of hybrid-type modified chitosan derivative nanoparticles for the intracellular delivery of midkine-siRNA in hepatocellular carcinoma cells

摘要

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Most of the patients with HCC lose the surgical opportunity at the time of diagno-sis. Some novel therapeutic modalities, like gene therapy, are promising for the treatment of HCC. However, the success of gene therapy depends on two aspects: efficient gene materials and gene delivery vectors. The present study was to develop new chitosan-based nanoparticles for a midkine-siRNA (anti-HCC gene drug) delivery. METHODS: The novel gene delivery vector (MixNCH) was syn-thesized by hybrid-type modification of chitosan with 2-chloro-ethylaminehydrochlorideandN,N-dimethyl-2-chloroethylamine hydrochloride. The chemical structure of MixNCH was char-acterized by FT-IR and 1HNMR. The cytotoxicity of MixNCH was determined by MTS assay. The gene condensation ability and size, zeta potential and morphology of MixNCH/MK-siRNA nanoparticles were measured. The in vitro transfection and gene knockdown efficiency of midkine by MixNCH/MK-siRNA nanoparticles was detected by qRT-PCR and Western blotting. Gene knockdown effect at the molecule level on the proliferation of HepG2 in vitro was determined by MTS assay. RESULTS: MixNCH was successfully acquired by aminoalkyl-ation modification of chitosan. The MixNCH could condense MK-siRNA well above the weight ratio of 3. The average size of MixNCH/MK-siRNA nanoparticles was 100-200 nm, and the surface charge was about +5 mV. Morphologically, MixNCH/MK-siRNA nanoparticles were in regular spherical shape with no aggregation. Regarding to the in vitro transfection of nanoparticles, the MixNCH/MK-siRNA nanoparticles reduced MK mRNA level to 14.03%±4.03%, which were comparable to Biotrans (8.94%±3.77%). MixNCH/MK-siRNA effectively inhibited the proliferation of HepG2 in vitro. CONCLUSION: MixNCH/MK-siRNA nanoparticles could be effective for the treatment of hepatocellular carcinoma.