MOLECULAR BIOLOGY OF HEPATITIS C VIRUS

丙型肝炎病毒的分子生物学

• 批准号: 6431596
• 负责人: Robert H. Purcell
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6431596
• 关键词: Pan; enzyme linked immunosorbent assay; genotype; hepatitis C virus; hepatitis vaccine; human tissue; immunization; microorganism culture; microorganism immunology; molecular cloning; neutralizing antibody; nonhuman therapy evaluation; nucleic acid sequence; nucleocapsid; virus RNA; virus genetics; virus replication; Pan; enzyme linked immunosorbent assay; genotype; hepatitis C virus; hepatitis vaccine; human tissue; immunization; microorganism culture; microorganism immunology; molecular cloning; neutralizing antibody; nonhuman therapy evaluation; nucleic acid sequence; nucleocapsid; virus RNA; virus genetics; virus replication

Hepatitis C virus (HCV) is a major cause of community-acquired viral hepatitis. Prototype strains of the various genotypes of HCV, including some of those discovered in this laboratory, are being biologically amplified in chimpanzees, packaged and distributed for use as challenge inocula in studies of passive and active immunoprophylaxis, etc. Full-length cDNA clones of HCV (genotypes 1a, 1b and 2a) have been constructed and transcribed RNA used to transmit hepatitis C to chimpanzees by in vivo hepatic transfection. Chimpanzees, transfected with infectious cDNA clones of HCV, are being followed to determine the natural history of infection. Infectivity pools have been prepared from chimpanzees infected with monoclonal HCV (derived by in vivo transfection with RNA transcripts of infectious cDNA); these have been titered for infectivity in other chimpanzees. In addition, the availability of infectious cDNA clones of HCV has permitted for the first time a mutational analysis of genomic regions. For example, individual portions of the 3' NCR have been deleted from the full-length clone and the resultant deletion mutant clones inoculated into chimpanzees by intrahapatic transfection. Certain regions of the NCR have been identified as critical for in vivo replication of HCV. We have constructed an infectious cDNA clone of GB virus-B (GBV-B), a monkey virus that is the closest relative to HCV. In addition, we have prepared challenge pools of GBV-B and have determined the infectivity titer of these in tamarins. We plan to use the GBV-B tamarin system to study characteristics of the virus that it shares with HCV, which must be studied in chimpanzees. In other studies, we have constructed chimeric genomes from infectious cDNA clones of HCV and bovine viral diarrhea virus. These genomes can replicate in transfected cells but the resultant viral products cannot assemble into infectious virus in the absence of helper virus.We have determined the genetic heterogeneity of HCV isolates that were recovered from patients who were infected following transfusion. The sequence of the hypervariable region and adjacent portions of envelope proteins 1 and 2 were determined for multiple clones obtained from patients who had fulminant hepatitis, from patients who convalesced following acute hepatitis and from patients who progressed to chronic hepatitis C. Distinctive patterns of dynamic change in sequence of clones during the first several weeks of infection were observed. Patients with fulminant or resolving hepatitis had few changes in the sequences of clones, whereas there were many changes in the sequences of clones from patients who progressed to chronic hepatitis. Thus, the outcome of an HCV infection could be predicted in the first few weeks of the infection

丙型肝炎病毒（HCV）是社区获得性病毒肝炎的主要原因。 HCV各种基因型的原型菌株（包括在该实验室中发现的一些基因型）在黑猩猩中在生物学上放大，包装和分布在被动和主动免疫预防的研究中，以用作挑战接种。黑猩猩通过体内肝转染。正在遵循用感染性cDNA克隆转染的黑猩猩，以确定感染的自然病史。 感染性池是由感染单克隆HCV的黑猩猩制备的（通过体内转染，用感染性cDNA的RNA转录得出）； 这些已被替代其他黑猩猩的感染性。此外，HCV的传染性cDNA克隆的可用性首次允许对基因组区域进行突变分析。 例如，3'NCR的各个部分已从全长克隆中删除，所得的缺失突变克隆通过缩写内转染接种到黑猩猩中。 NCR的某些区域已被确定为HCV体内复制至关重要。 我们已经构建了GB病毒-B（GBV-B）的传染性cDNA克隆，这是一种猴子病毒，是最接近HCV的猴子病毒。 此外，我们已经准备了GBV-B的挑战池，并确定了菜他蛋白中的感染性滴度。 我们计划使用GBV-B tamarin系统研究与HCV共享的病毒特征，必须在黑猩猩中研究。在其他研究中，我们从HCV和牛病毒腹泻病毒的传染性cDNA克隆中构建了嵌合基因组。 这些基因组可以在转染的细胞中复制，但是在没有辅助病毒的情况下，由此产生的病毒产物不能将其组装成传染性病毒。我们确定了从被感染后感染的患者中回收的HCV分离株的遗传异质性。 确定了从患有暴发性肝炎的患者获得的多个克隆，从急性肝炎后康复的患者以及从第一次感染序列的动态序列中获得独特的动态变化的患者，从患有暴发的肝炎的患者，从患有暴发的肝炎的患者中获得多个克隆的序列，并从患有肝炎的患者中获得了多个克隆的序列。 暴发或解决肝炎的患者的克隆序列几乎没有变化，而从发展为慢性肝炎的患者的克隆序列序列发生了许多变化。 因此，可以在感染的前几周预测HCV感染的结果