Hirata Yuichi, Sudoh Masayuki, Kohara Michinori
Department of Microbiology and Cell biology, The Tokyo metropolitan Institute of Medical Science, Tokyo, Japan.
Yakugaku Zasshi. 2010 Feb;130(2):157-61. doi: 10.1248/yakushi.130.157.
Hepatitis C virus (HCV) persists chronically in most infected patients, eventually causing chronic hepatitis, liver cirrhosis, and in some cases hepatocellular carcinoma. The combination therapy of PEG-IFN and ribavirin improves efficacy in many patients, although it does not lead to sufficient achievements in genotype 1b patients. To aid in invention of new anti-HCV reagents, we focused on host factors that contributed to HCV lifecycle. We identified serine palmitoyltransferase inhibitor as an anti-HCV reagent through high-throughput screening using HCV replicon cells. We investigated the mechanism of anti-HCV effect of SPT inhibitor. It has been reported that sphingolipids and cholesterol compose the lipid raft where replication of HCV occurs. We investigated the influence of SPT inhibitor to lipid rafts by analyzing the detergent-resistant membrane (DRM). The analysis showed that SPT inhibitor moved HCV RNA-dependent RNA polymerase (NS5B) to detergent-soluble fraction from DRM, and Biacore analysis indicated binding of sphingomyelin to NS5B. These results suggest that SPT inhibitor disrupts the interaction between NS5B and sphingomyelin. Moreover, we evaluated the anti-HCV effect of SPT inhibitor in vivo with humanized chimeric mice. SPT inhibitor led to rapid decline in serum HCV-RNA of about 1-2 log within 8 days. Furthermore, combination therapy of SPT inhibitor and PEG-IFN achieved about 3 log reduction in serum HCV-RNA.
丙型肝炎病毒（HCV）在大多数受感染患者中持续慢性感染，最终导致慢性肝炎、肝硬化，在某些情况下还会引发肝细胞癌。聚乙二醇干扰素（PEG-IFN）和利巴韦林的联合疗法在许多患者中提高了疗效，尽管在基因1b型患者中并未取得足够的成果。为了有助于新型抗HCV试剂的研发，我们聚焦于对HCV生命周期有贡献的宿主因子。我们通过使用HCV复制子细胞进行高通量筛选，确定丝氨酸棕榈酰转移酶抑制剂为一种抗HCV试剂。我们研究了SPT抑制剂的抗HCV作用机制。据报道，鞘脂和胆固醇构成了HCV发生复制的脂筏。我们通过分析耐去污剂膜（DRM）来研究SPT抑制剂对脂筏的影响。分析表明，SPT抑制剂将HCV RNA依赖性RNA聚合酶（NS5B）从DRM转移到去污剂可溶部分，并且生物传感器分析表明鞘磷脂与NS5B结合。这些结果表明，SPT抑制剂破坏了NS5B与鞘磷脂之间的相互作用。此外，我们用人源化嵌合小鼠在体内评估了SPT抑制剂的抗HCV作用。SPT抑制剂在8天内使血清HCV-RNA迅速下降约1-2个对数。此外，SPT抑制剂与PEG-IFN的联合疗法使血清HCV-RNA降低了约3个对数。
