Elazar Menashe, Liu Michael, McKenna Sean A, Liu Ping, Gehrig Elizabeth A, Puglisi Joseph D, Rossignol Jean-François, Glenn Jeffrey S
Division of Gastroenterology and Hepatology, Department of Medicine, Stanford University School of Medicine, Palo Alto, California, USA.
Gastroenterology. 2009 Nov;137(5):1827-35. doi: 10.1053/j.gastro.2009.07.056. Epub 2009 Aug 4.
BACKGROUND & AIMS: New therapies are needed to treat patients infected with hepatitis C virus (HCV), a major worldwide cause of chronic liver disease. Nitazoxanide (NTZ), originally used to treat cryptosporidiosis infection, recently was shown to have unexpected antiviral activity in the HCV replicon system and in chronically infected patients. A pilot clinical study suggested that NTZ can augment the antiviral effect of interferon (IFN), although the molecular basis for its effect was unknown.
We analyzed the effects of NTZ on the regulation of eukaryotic initiation factor-2alpha (eIF2alpha) and its IFN-induced kinase, protein kinase activated by double-stranded RNA (PKR), in cells that support HCV RNA replication and in vitro biochemical assays.
NTZ increased eIF2alpha phosphorylation, a modification known to mediate host cell antiviral defenses. The addition of IFN to cell cultures increased NTZ-induced eIF2alpha phosphorylation. NTZ also increased PKR phosphorylation. In vitro, NTZ promoted PKR autophosphorylation, a key step in activating PKR's kinase activity for eIF2alpha. Finally, NTZ-induced eIF2alpha phosphorylation was reduced in the presence of specific inhibitors of PKR autophosphorylation.
An important mechanism of NTZ's action involves activation of PKR, a key kinase that regulates the cell's innate antiviral response. These observations could explain the clinical antiviral effect of NTZ. NTZ might represent a new class of small molecules capable of potentiating and recapitulating important antiviral effects of IFN.
丙型肝炎病毒（HCV）感染是全球慢性肝病的主要病因，因此需要新的治疗方法来治疗HCV感染患者。硝唑尼特（NTZ）最初用于治疗隐孢子虫感染，最近在HCV复制子系统和慢性感染患者中显示出意外的抗病毒活性。一项初步临床研究表明，NTZ可增强干扰素（IFN）的抗病毒作用，但其作用的分子基础尚不清楚。
我们在支持HCV RNA复制的细胞和体外生化试验中，分析了NTZ对真核起始因子-2α（eIF2α）及其IFN诱导激酶——双链RNA激活的蛋白激酶（PKR）调控的影响。
NTZ增加了eIF2α磷酸化，这种修饰已知可介导宿主细胞的抗病毒防御。在细胞培养物中添加IFN可增加NTZ诱导的eIF2α磷酸化。NTZ还增加了PKR磷酸化。在体外，NTZ促进了PKR自身磷酸化，这是激活PKR对eIF2α激酶活性的关键步骤。最后，在存在PKR自身磷酸化特异性抑制剂的情况下，NTZ诱导的eIF2α磷酸化减少。
NTZ作用的一个重要机制涉及PKR的激活，PKR是调节细胞固有抗病毒反应的关键激酶。这些观察结果可以解释NTZ的临床抗病毒作用。NTZ可能代表一类能够增强和重现IFN重要抗病毒作用的新型小分子。
