Heat Shock Protein Synthesis Inhibitor Treatment of Hepatitis C Viral Infection

描述(由申请人提供)：丙型肝炎病毒(丙型肝炎病毒)感染在全球范围内的流行率为3%，是发达国家治疗肝硬变的肝移植的主要实体。在美国，丙型肝炎病毒是最常见的慢性血液传播感染，影响了1.8%的人口，似乎是导致美国肝癌最近翻了一番的主要病因。目前的治疗方法包括聚乙二醇化干扰素-1(PEG化干扰素-1)和利巴韦林(RBV)。在美国，70%的患者感染了持续病毒学应答(SVR)只有42-46%的1型基因。一般来说，所有基因型的治疗都可能伴随着不良反应，治疗的禁忌症并不少见。出于这些原因，有必要开发毒性更低、导致更高SVR的额外疗法，无论是作为辅助疗法还是替代疗法。我们用质谱仪鉴定了丙型肝炎病毒编码蛋白NS5A与热休克蛋白形成的复合体中的热休克蛋白S、热休克蛋白40和热休克蛋白70。我们通过共聚焦显微镜和免疫共沉淀证实了NS5A/HSP的相互作用。HSP40和HSP70基因敲除都减少了丙型肝炎病毒细胞培养系统中传染性病毒颗粒的产生。热休克蛋白合成抑制剂Quercetin和KNK437在无毒浓度下减少了感染性颗粒的产生。这种对病毒产生的显著抑制，加上其已知的低毒性和在先前和正在进行的临床试验中的使用，促使这位医生在床边提议研究用栎素治疗丙型肝炎病毒感染的患者。在这项建议中，我们的目标是进一步了解HSP40和HSP70以及热休克蛋白合成抑制剂对丙型肝炎病毒感染的影响，并在I期临床试验中确定Qercetin在慢性丙型肝炎患者中的安全性和抗病毒活性。为了实现这一目标，我们提出了三个相互关联的具体目标：1.我们将在HCVcc模型中确定热休克蛋白40和70在丙型肝炎病毒产生中的重要性。2.我们将在HCVcc模型中确定热休克蛋白合成抑制剂对丙型肝炎病毒感染的影响。我们将通过I期试验来测试热休克蛋白合成抑制剂Qercetin治疗慢性丙型肝炎病毒感染的临床可行性。进一步了解热休克蛋白和热休克蛋白合成抑制在丙型肝炎病毒感染中的作用，可能有助于成功治疗慢性丙型肝炎，降低肝硬变和肝细胞癌的发生率。 公共卫生相关性：慢性丙型肝炎病毒(丙型肝炎病毒)感染是导致需要肝移植的肝硬变的头号原因，也是最近美国肝细胞癌增加一倍的主要原因。我们已经鉴定了热休克蛋白S 40和70是与丙型肝炎病毒感染有关的细胞蛋白。我们建议在这个实验台上研究HSP40、HSP70和HSP合成抑制物在丙型肝炎病毒感染中的作用，并提出治疗慢性丙型肝炎病毒感染从而降低肝硬变和肝细胞癌发生率的建议。

DESCRIPTION (provided by applicant): Hepatitis C virus (HCV) infection has a worldwide prevalence of 3% and is the main entity responsible for liver transplantation in developed countries for treatment of cirrhosis. In the United States, HCV is the most common chronic blood borne infection affecting 1.8% of the population and appears to be the major etiologic factor responsible for the recent doubling of HCC in the United States. Current therapy consists of pegylated interferon-1 (PEG-IFN) and ribavirin (RBV). 70% of patients in the United States are infected with genotype 1 for which sustained virologic response (SVR) is only 42-46%. Generally, therapy of all genotypes can be accompanied by adverse effects and contraindications to therapy are not infrequent. For these reasons there is the need to develop additional therapies that are less toxic and result in higher SVR either as adjuncts or replacement therapies. We have identified the heat shock proteins (HSP)s HSP40 and HSP70 in complex with the HCV encoded protein NS5A through mass spectrometric analysis. We confirmed an NS5A/HSP interaction by confocal microscopy and coimmunoprecipitation. HSP40 and HSP70 knockdown both reduced infectious viral particle production in a HCV cell culture system. Treatment with the heat shock protein synthesis inhibitors Quercetin and KNK437 reduced infectious particle production at non-toxic concentrations. This striking inhibition of virus production combined with its known low toxicity and use in previous and ongoing clinical trials serves to motivate this bench to bedside proposal to study treat HCV infected patients with Quercetin. In this proposal, our goals are to further understand the impact of HSP40 and HSP70 and heat shock protein synthesis inhibitors on HCV infection and determine Quercetin's safety and antiviral activity in patients suffering from chronic HCV infection in a phase I clinical trial. To achieve this we propose three interrelated specific aims: 1. We will determine the importance of heat shock proteins 40 and 70 in hepatitis C virus production in the HCVcc model. 2. We will determine the impact of heat shock protein synthesis inhibitors on hepatitis C viral infection in the HCVcc model. We will test the clinical feasibility of the heat shock protein synthesis inhibitor Quercetin on patients with chronic hepatitis C viral infection through a phase I trial. Further understanding of heat shock proteins and heat shock protein synthesis inhibition in HCV infection may allow for successful treatment of chronic hepatitis C and reduce the incidence of cirrhosis and hepatocellular carcinoma. PUBLIC HEALTH RELEVANCE: Chronic hepatitis C viral (HCV) infection is the number one causative agent of cirrhosis necessitating liver transplantation and is the major cause of the recent doubling of hepatocellular carcinoma in the United States. We have identified heat shock proteins (HSP)s 40 and 70 as cellular proteins that are important for HCV infection. We propose to study HSP40, HSP70 and HSP synthesis inhibitors effect on HCV infection in this bench to beside proposal that could lead to treatment of chronic HCV infection thereby reducing the incidence of cirrhosis and hepatocellular carcinoma.