Development of novel gene-therapy using IL-12 and DNA nanoassociate bearing hyaluronan-glycocalyx for recurrent hepatocellular carcinoma after liver transplantation

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In this project, we investigated the effectiveness of gene therapy using IL-12 and DNA nanoassociate bearing hyaluronan-glycocalyx argeting the sinusoidal endothelial cells for the treatment of recurrent hepatocellular carcinoma after liver transplantation. First of all, mIL-12 plasmid DNA was purified. To develop a system for targeting foreign DNA to SECs, comb-type polycations having HA side chains were prepared by coupling HA to poly(L-lysine) (PLL). The HA-grafted-PLL copolymer (PLL-g-HA) thus formed was mixed with DNA in 154 mM NaCl to form soluble nanoassociates bearing hydrated hyaluronate shells. Agarose gel retardation assays revealed selective interaction of the PLL backbone with DNA despite the presence of polyanionic HA side chains.PLL-g-HA. Polymerization of PLL-g-HA with mIL-12 plasmid DNA was then confirmed. However, stable production and supply of PLL-g-HA/pCAGGS-mIL-12 was difficult due to the lack of PLL-g-HA. C3H mice (H-2(k)) were s.c. implanted with 2.5 x 10(6) MH134 cells (H-2(k)) and we treated the established HCC with electroporation-mediated gene therapy using mIL-12 plasmid DNA. Intratumoral gene transfer of mIL-12 elevated intratumoral mIL-12, IFN-gamma, and IFN-gamma-inducible protein-10, significantly inhibited the growth of HCC, compared with HCC-transferred control pCAGGS plasmid.