Hepatitis C virus (HCV) is an important cause of chronic liver disease, cirrhosis and cancer. Its limited host range (humans and chimpanzees) and poor replication in cell culture have made it difficult to study. In this program, four groups of investigators join forces in a multidisciplinary effort that connects clinically based investigation with fundamental studies on virus-host interactions. The four groups (C. Rice, H. Greenberg, M. Kay and T. Wright) have a proven history of productive collaboration, and bring a broad range of talents and interests to this program. By combining resources, reagents and ideas, the program team hopes to advance our knowledge of HCV immunity, pathogenesis and treatment. Projects address important questions relevant to vaccine design, improvement of current therapy, and testing of cutting-edge new therapeutic approaches, all in the context of improving our understanding of the natural history of HCV-associated liver disease. Dr. Rice, who is the overall program director, will study the role of neutralizing antibodies (nAbs) in control of HCV. He and co-Pi Jane McKeating will refine and employ new technologies (retrovirus pseudotypes and high titer stocks of cell culture infectious HCV) to study and characterize nAbs in patients who resolve acute infection. These studies will be done in close collaboration with Dr. Wright and the acute IDU cohort of Dr. Kim Page-Shafer. Dr. Greenberg will continue his ex vivo microarray studies to better understand the racial differences and mechanisms underlying the variable responses to IFN therapy. These studies will set the stage for future genetic admixture mapping experiments in search of specific genes underlying distinct treatment outcomes. Dr. Greenberg will benefit from Dr. Wright's large cohort of chronic HCV patients for these studies. Dr. Kay will explore RNA interference for HCV therapy. He has developed new vectors for efficient in vivo gene delivery that will be engineered to express short hairpin shRNAs that target and destroy the HCV genome. Prior to testing in mouse and chimpanzee models of HCV infection, he will work with Dr. Rice to evaluate their anti-HCV efficacy in cell culture. Dr. Wright, in addition to providing clinical resources for all projects, will continue to investigate the clinical consequences of HCV disease and the factors that influence disease outcomes. She will take advantage of a cohort of well characterized patients with HCV disease, with complete information regarding demographics, alcohol intake, duration of infection and histological stage of liver disease, to examine the importance of these baseline characteristics in determining clinical and histological outcomes with medium-term follow-up. ? ? PROJECT 1: HCV Infection Systems and Role of Neutralizing Antibodies in Hepatitis C (Rice, C.) ? ? PROJECT 1 DESCRIPTION (provided by applicant): Hepatitis C virus (HCV) is a major etiological agent of liver disease in most regions of the world, with approximately 170 million individuals infected. Cellular and humoral immune responses are mounted but fail to clear HCV in most individuals. Although more difficult to study, the -30% of patients who do resolve infection are a valuable resource for identifying immune correlates that lead to virus control and clearance. Recent studies of re-challenge after acute resolving infection indicate that protective immunity exists, mediated largely by memory CD4+ and CD8+ T cell responses. While early and strong intrahepatic CD4+ and CD8+ T-cell responses are also believed to be important for control of primary infections, the role of neutralizing antibodies (nAbs) is less clear. This gap in our knowledge has stemmed largely from technical limitations in our ability to measure HCV neutralization. The development of retrovirus pseudotype particles (HCVpp) and cell culture infectious virus (HCVcc) now allows these studies to move forward.
The aims outlined in this proposal will refine and generalize the HCVcc approach, create convenient reporter viruses for high throughput and quantitative measurements of infectivity and neutralization, and compare the cell tropism, receptor usage and neutralization characteristics of HCVcc to the well-established HCVpp system. This will result in a standardized panel of virus stocks, representing all HCV genotypes, which can be used for basic and applied studies (see CRC Project 3). These tools will be used to expand longitudinal studies of nAbs in acute HCV infection in the unique cohorts described in Translational Research Core A. The underlying premise of this work, supported by preliminary data, is that nAb responses during acute infection may play a role in controlling and resolving virus infection. The nAb responses and the HCV epitopes they recognize will be defined and used to establish a database of conserved neutralization determinants that can be integrated into strategies for vaccination or passive immunotherapy. In addition, this panel of recombinant HCVcc expressing convenient reporters will facilitate screening efforts to identify pan-HCV small molecule inhibitors of virus assembly and entry. Tools and reagents developed in the course of these studies will be made available to other HCV CRCs and the broader HCV research community in an effort to speed progress on combating this important public health problem. ? ?
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