Initiation of Hepatitis B Virus Replication

描述（申请人提供）：乙型肝炎病毒（HBV）感染是一个全球性的健康问题。据估计，世界上有2亿至5亿HBV慢性携带者，迄今尚无可靠的治疗方法。HBV可引起急性和慢性肝病，慢性HBV携带者患原发性肝细胞癌（PHC）的相对危险度估计约为未感染个体的100倍。因此，需要对慢性HBV感染进行有效治疗。在这些研究中，将研究调节乙型肝炎病毒（HBV） DNA合成初始步骤的机制。HBV DNA的合成是通过病毒聚合酶与位于HBV基因组前RNA 5'端茎环结构epsilon结合而启动的。最初，HBV负链DNA的前三个核苷酸是利用聚合酶的氨基末端结构域作为引物和epsilon的凸起区域作为模板合成的。带有共价三核苷酸序列的HBV聚合酶随后被转移到基因组前RNA 3'端的DR1序列上。HBV负链DNA合成随后通过基因组前RNA的逆转录进行。调控易位步骤的机制尚不清楚。最近，一个调控序列元件phi被发现，它位于DR1序列的上游，位于妊娠RNA的3‘端，对有效的病毒复制很重要，并且与epsilon的5’-一半互补。这一发现表明，负链引物从epsilon到DR1的易位可能是由基因组前RNA的构象变化介导的，该构象变化使引物靠近基因组前RNA的3'端DR1序列。土拨鼠肝炎病毒（WHV）和鸭乙肝病毒（DHBV）基因组中epsilon和phi之间的互补性守恒也支持了这一论点。因此，epsilon和phi之间的互补性在调节HBV复制中的作用将通过对这些序列元件的突变分析直接进行检验。该方法旨在确定慢性HBV感染治疗干预的可能靶点。

DESCRIPTION (provided by applicant): Hepatitis B virus (HBV) infection is a worldwide health problem. It is estimated that there are 200 to 500 million HBV chronic carriers in the world for whom, to date, there is no reliable treatment. HBV causes both acute and chronic liver disease and the estimated relative risk of primary hepatocellular carcinoma (PHC) in chronic HBV carriers is approximately 100 times greater than in uninfected individuals. Therefore, effective treatments for chronic HBV infection are required. In these studies, the mechanism(s) regulating the initial steps in the synthesis of hepatitis B virus (HBV) DNA will be investigated. HBV DNA synthesis is initiated by binding of the viral polymerase to a stem-loop structure, epsilon, located at the 5'-end of the HBV pregenomic RNA. Initially, the first three nucleotides of HBV minus-strand DNA are synthesized utilizing the amino-terminal domain of the polymerase as a primer and the bulge region of epsilon as a template. The HBV polymerase with the covalently attached trinucleotide sequence is subsequently translocated to the DR1 sequence at the 3'-end of the pregenomic RNA. HBV minus-strand DNA synthesis then proceeds by the reverse transcription of the pregenomic RNA. The mechanism(s) regulating the translocation step are unknown. Recently, a regulatory sequence element, phi, located immediately upstream of the DR1 sequence at the 3'-end of the pregnomic RNA that is important for efficient viral replication and is complementary to the 5'-half of epsilon was identified. This finding suggests that the translocation of the minus-strand primer from epsilon to DR1 might be mediated by a conformational change in the pregenomic RNA that brings the primer into proximity with the DR1 sequence at the 3'-end of the pregenomic RNA. The conservation of the complementarity between epsilon and phi in the woodchuck hepatitis virus (WHV) and the duck hepatitis B virus (DHBV) genomes also supports this contention. Therefore, the role of the complementarity between epsilon and phi in regulating HBV replication will be examined directly by mutational analysis of these sequence elements. This approach is aimed at identifying possible targets for therapeutic intervention in chronic HBV infection.