Hepatocarcinogenesis Secondary to Hepatitis C

描述（由申请人提供）：丙型肝炎病毒（HCV）是一个主要的公共卫生问题，目前有超过1.7亿人感染。大多数感染会变成慢性，通常会导致肝硬化和癌症。在美国，每年有10，000至20，000人死于慢性HCV感染。这是肝移植最常见的原因。目前认为，HCV感染继发的长期慢性炎症是肝细胞癌的主要原因。长期慢性炎症和癌症发展的潜在机制尚未得到很好的理解。目前治疗慢性HCV感染的方法是IFN-α和利巴韦林的联合治疗，但只有一半的患者可以通过这种方案摆脱病毒感染。丙型肝炎病毒感染导致人类慢性感染率高，并经常对干扰素治疗产生耐药性的原因尚不清楚。本提案的总体目标是了解慢性HCV感染中干扰素作用和耐药性的机制，并开发抑制HCV复制的替代抗病毒策略。在过去的几年里，我们的研究产生了以下证据：（一）。我们已经开发了干扰素抗性复制子细胞系，并确定Jak-Stat信号通路中的缺陷可导致ISRE启动子（IFN-启动子）和干扰素抗性表型的低水平活化。（ii）。我们已经发表的数据表明，IFN-α，IFN-γ和IFN-？每种都能抑制HCV复制。干扰素的这种抑制作用是在核糖体加载到病毒通过内部核糖体进入位点（IRES）依赖性机制翻译其基因组所使用的5 'UTR序列的水平上。（三）.最近，我们报道了针对IRES区域的小干扰RNA（siRNA）可以抑制六种不同HCV基因型的翻译。基于这些初步研究，我们的假设是，控制干扰素诱导基因转录的Jak-Stat信号分子的表达在肝脏中受感染的肝细胞中存在差异。具有缺陷的Jak-Stat信号传导的肝细胞在IRES翻译水平逃避干扰素作用，导致慢性持续性病毒复制。我们建议，通过使用可生物降解的聚合物将siRNA-74封装到纳米颗粒中将有效地将siRNA递送到肝细胞，并可能为对干扰素无应答的慢性HCV患者提供新的治疗策略。为了验证我们的假设，我们制定了三个具体目标。在具体目标1中，我们将研究HCV慢性感染者对IFN-α的肝耐药性。在具体目标2中，我们将研究IFN-α对丙型肝炎病毒的抗病毒机制。在具体目标3中，我们将配制可生物降解的纳米胶囊作为一种非病毒方法来递送siRNA以抑制干扰素的病毒靶点，以克服耐药机制。如果这些实验是成功的，那么它将增加我们对干扰素的作用机制和对慢性HCV的抵抗力的理解。这项研究可能会为干扰素治疗无效的慢性丙型肝炎患者带来一种创新的治疗策略。 公共卫生相关性：慢性丙型肝炎病毒感染是美国肝癌的主要原因。本研究拟发展细胞内免疫策略以抑制HCV。如果这些实验成功，它可能导致慢性HCV感染的治疗和预防肝癌。

DESCRIPTION (provided by applicant): Hepatitis C virus (HCV) is a major public health problem with more than 170 millions people are currently infected. Most infections become chronic often leading to liver cirrhosis and cancer. In the United States 10,000 to 20,000 deaths a year are caused by chronic HCV infection. It is the most common cause of liver transplantation. It is now believed that long-standing chronic inflammation secondary to HCV infection is the main cause of hepatocellular carcinoma. The mechanisms underlying the development of long-lasting chronic inflammation and cancer are not well understood. Current therapy for chronic HCV infection, a combination of IFN-a and ribavirin, but only half of the patients can get rid of the virus infection by this regimen. The reasons why HCV infection leads to a high rate of chronic infection in human and often develops resistance to interferon therapy are not clear. The overall goals of this proposal are to understand the mechanisms of interferon action and resistance in chronic HCV infection, and develop alternative antiviral strategies to inhibit HCV replication. During the last couple of years our research has generated the following evidences: (i). We have developed interferon-resistant replicon cell lines and determined that a defect in the Jak-Stat signaling pathway can lead to low-level activation of ISRE promoter (IFN-promoter) and interferon resistance phenotypes. (ii). We have published data showing that IFN-a, IFN-¿ and IFN-? each inhibits HCV replication. This inhibitory effect of interferon is at the level of ribosome loading to the 5'UTR sequences used by the virus to translate its genome by an internal ribosome entry site (IRES) dependent mechanism. (iii). Recently, we have reported that small interfering RNA (siRNA) targeted to the IRES region that can inhibit translation of six different HCV genotypes. Based on these preliminary studies our hypothesis is that the expression of the Jak-Stat signaling molecules that control the transcription of interferon-inducible genes varies among infected hepatocytes in the liver. Hepatocytes with a defective Jak-Stat signaling escape interferon action at the level of IRES translation leading to chronic persistent virus replication. We propose that encapsulation of siRNA-74 into nanoparticles by the use of biodegradable polymers will efficiently deliver siRNA to the hepatocytes and may provide a novel therapeutic strategy for chronic HCV patients who are non-responders to interferon. To test our hypothesis we have developed three Specific Aims. In Specific Aim 1, we will investigate hepatic resistance to IFN-alpha in HCV chronically infected humans. In Specific Aim 2, we will investigate the antiviral mechanisms of IFN-alpha against hepatitis C virus. In Specific Aim 3, we will formulate biodegradable nanocapsules as a non-viral method to deliver siRNA to inhibit viral target of interferon to overcome mechanisms of resistance. If these experiments are successful then it will increase our understanding on the mechanisms of interferon action and resistance against chronic HCV. This research will potentially leads to an innovative therapeutic strategy for chronic hepatitis C patients not responding to interferon therapy. Public Health Relevance: Chronic hepatitis C virus infection is the major cause of liver cancer in the United States. This research proposal intends to develop intracellular immunization strategy to inhibit HCV. If these experiments are successful it can potentially lead to a therapy for chronic HCV infection and prevent liver cancer.