Delivery of small interfering RNA using epoxide-conjugated polyamine nanoparticles for the potential treatment of hepatitis C

다국어 초록 (Multilingual Abstract)

The specific silencing of gene expression through RNA interference (RNAi) has tremendous potential to address previously untreatable diseases. RNAi through silencing the targets related to viral replication has recently been highlighted for curing viral diseases. However, the safe and effective delivery of small interfering RNA (siRNA) still remains challenging. Here I developed epoxide-conjugated polyamine lipid-like materials for efficient siRNA delivery to liver, which could be useful for treating hepatitis C viral infection. The epoxide-conjugated polyamine materials were synthesized by reacting an epoxide with 10-carbon-alkyl chain (C12) and four types of polyamines via ring-opening reaction. The positively charged amines in synthesized epoxide-polyamine conjugates facilitated the complexation with negatively charged siRNAs via electrostatic interaction. One of these synthesized materials, C12-conjugated spermine (C12-SPM) has shown minimal toxicity and high housekeeping protein (glyceraldehydes-3-phosphate dehydrogenase, GAPDH) silencing efficiency in human hepatic cells (Huh-7). To apply this siRNA delivery sytem to hepatitis C treatment, I choose protein kinase C-related kinase 2 (PRK2) as a siRNA target. PRK2 is a host kinase which helps replication of haptitis C virus (HCV) by specifically phosphorylating NS5B, HCV RNA polymerase. Two days after PRK2 siRNA transfection using C12-SPM, significant silencing of PRK2 expression was achieved Huh-7 cells with ~70% of PRK2 silencing efficiency. Interestingly, HCV subgenome level in human hepatic cells which were modified to express HCV subgenomic RNA was greatly reduced by PRK2 siRNA delivery using C12-SPM. For in vivo application, C12-SPM-based liposomal nanoparticle was prepared to encapsulate siRNAs. This nanoparticle shows no evidence of toxicity in blood tests. When the C12-SPM liposomal nanoparticles systemically delivered into mouse liver via intravenous administration, the PRK2 expression was reduced about 46% in perfused hepatocytes from mouse liver. Here I demonstrate the potential of epoxide-conjugated lipid-like nanoparticles as efficient siRNA delivery materials with minimal cytotoxicity. This study suggests that epoxide-conjugated polyamine materials developed in this study could provide a potential treatment of hepatitis C by delivery of anti-viral RNAi therapeutics.