- 學位論文
石蓯蓉暨奈米化水飛薊賓之抗C型肝炎病毒研究
The investigation of Limonium sinense (Girard) Kuntze and silibinin nano-formulation against hepatitis C virus
並列摘要
A preventive vaccine against hepatitis C virus (HCV) infection remains unavailable and newly developed drugs against viral replication are complicated by potential drug-resistance and high cost. These issues justify the need to develop alternative antiviral agents and expand the scope of strategies for the treatment of hepatitis C, such as targeting viral entry. In this study, we explored the bioactivity of Limonium sinense (L. sinense) and its purified constituents against HCV life cycle using subgenomic replicon and infectious HCV culture systems. Data indicated that the water extract from the underground part of L. sinense (LS-UW) exhibited potent inhibitory activity against HCV at non-cytotoxic concentrations. LS-UW targeted early HCV infection without affecting viral replication, translation, and cell-to-cell transmission, and blocked viral attachment and post-attachment entry/fusion steps. Bioactivity analysis of major constituents from LS-UW through viral infectivity/entry assays revealed that gallic acid (GA) also inhibits HCV entry. Furthermore, both LS-UW and GA could suppress HCV infection of primary human hepatocytes. Due to their potency and ability to target HCV early viral entry, LS-UW and GA may be of value for further development as prospective antivirals against HCV. Silibinin, a flavonolignan well established for its robust anti-HCV activity, has undergone several clinical trials for the management of hepatitis C patients including those in liver transplant setting. Despite its potency, silibinin suffers from poor solubility and bioavailability, making intravenous injection with chemically hydrophilized silibinin the only viable route for administration. To overcome this limitation and to increase its clinical potential, we developed silibinin nanoparticles (SB-NP) using a simple nano-emulsification technique. SB-NP was derived as spherical particles with average size of < 200 nm, consisting of amorphous silibinin encapsulated at > 97% efficiency. The nano-formulation had significantly enhanced solubility with a > 75% increase in its solubility profile at pH 7.4. Experiments with infectious HCV culture systems demonstrated that SB-NP retained its robust anti-HCV activity, efficiently restricted spread of virus from cell to cell, and showed antioxidant function against virus-induced oxidative stress. Oral administration of SB-NP in rodents produced no apparent in vivo toxicity, with animal survival, liver function, blood biochemistry, and erythrocyte stability relatively unaffected. More importantly, pharmacokinetic studies revealed that SB-NP delivered orally was highly bioavailable, producing enhanced serum levels and superior biodistribution to the liver, compared to non-modified silibinin. Finally, SB-NP efficiently reduced HCV infection of primary human hepatocytes. In conclusion, we have successfully generated highly bioavailable silibinin nanoparticles which could be useful for the treatment of hepatitis C. We suggest that SB-NP merits further evaluation for applications in the clinical setting.