Targeted delivery of siRNAs against hepatocellular carcinoma-related genes by a galactosylated polyaspartamide copolymer

Given the lack of effective treatments for Hepatocellular carcinoma (HCC), the development of novel therapeutic approaches is very urgent. Here, siRNAs were delivered to HCC cells by a synthetic polymer containing α,β-poly-(N-2-hydroxyethyl)-D,L-aspartamide-(PHEA) derivatized with diethylene triamine (DETA) and bearing in the side chain galactose (GAL) linked via a polyethylene glycol (PEG) to obtain (PHEA-DETA-PEG-GAL, PDPG). The GAL residue allows the targeting to the asialo-glycoprotein receptor (ASGPR), overexpressed in HCC cells compared to normal hepatocytes. Uptake studies performed using a model siRNA or a siRNA targeted against the enhanced green fluorescence protein, demonstrated the PDPG specific delivery of siRNA to HuH7 cells, a human cellular model of HCC. GAL-free copolymer (PHEA-DETA-PEG-NH2, PDP) or the chemical block of ASGPR, impaired PDPG targeting effectiveness in vitro. The specificity of PDPG delivery was confirmed in vivo in a mouse dorsal skinfold window chamber assay. Functional studies using siRNAs targeting the mRNAs of HCC-related genes (eEF1A1, eEF1A2 and E2F1) delivered by PDPG, significantly decreased HuH7 vitality/number and down regulated the expression of the target genes. Only minor effectiveness was in contrast observed for PDP. In IHH, a human model of normal hepatocytes with reduced ASGPR expression, PDPG barely reduced cell vitality. In a subcutaneous xenograft mouse model of HCC, PDPG-siRNAs reduced HCC tumor growth compared to controls without significant toxic effects. In conclusion, our study demonstrates the valuable potentials of PDPG for the specific delivery of siRNAs targeting HCC-related genes.