Analyzing the contribution of intracellular localization of hepatitis C virus replication complex for the drug resistance.

本研究旨在揭示丙型肝炎病毒（HCV）复制复合体的细胞内定位与抗病毒药物如干扰素-α的作用之间的关系，并通过了解复制复合体中病毒的复制机制，最终开发新的抗HCV策略。富含鞘磷脂和胆固醇的细胞器膜的微结构域以“脂筏”的形式被认为是HCV复制复合物形成的支架，提示脂筏可能成为新的治疗靶点。在本研究中，我们调查的影响myriocin，鞘磷脂合成的抑制剂，HCV复制使用HCV亚基因组复制子系统，以及感染性HCV培养系统。我们还研究了多球菌素加α干扰素或多球菌素加辛伐他汀对HCV复制的联合作用。多孢菌素以时间和剂量依赖性方式抑制基因型Ib亚基因组HCV复制子RNA和基因型2a感染性HCV RNA的细胞内RNA复制（亚基因组HCV-Ib，最大值）。120小时时70%;基因组HCV-2a，最大值96小时时为60%）。多球壳菌素加IFN或辛伐他汀联合治疗协同减弱细胞内HCV-RNA复制，我们的数据表明，鞘磷脂合成抑制剂和HMG-CoA抑制剂强烈抑制HCV复制，无论基因型，表明宿主的脂质代谢可能是一个新的HCV治疗靶点。

This research is aimed to disclose the relationship between intracellular localization of hepatitis C virus (HCV) replication complex and the effect of anti-viral drugs such as interferon-alpha, and finally to develop new anti-HCV strategies through understanding of viral replication mechanism(s) in the replication complex.Recently, micro-domains of organelle membranes rich in sphingomyelin and cholesterol in the form of "lipid rafts" have been considered to act as a scaffold for HCV replication complex formation, indicating the possibility that the lipid raft could be a new therapeutic target. In the present study, we investigated the effect of myriocin, an inhibitor of sphingomyelin synthesis, on HCV replication using an HCV-subgenomic replicon system, as well as the infectious HCV culture system. We also investigated the combined effect of myriocin plus interferon-alpha, or myriocin plus simvastatin, on HCV replication. Myriocin suppressed intracellular RNA replication of both genotype lb subgenomic HCV replicon RNA and genotype 2a infectious HCV RNA in a time-and dose-dependent manner (subgenomic HCV-lb, max. 70% at 120 hours ; genomic HCV-2a, max. 60% at 96 hours). Combination treatment with myriocin plus IFN or simvastatin attenuated intracellular HCV-RNA replication synergistically.Our data demonstrate that both the sphingomyelin synthesis inhibitor and HMG-CoA inhibitor strongly suppress HCV replication, irrespective of genotype, indicating that the lipid metabolism of the host could be a novel target for HCV therapy.