Virology and Pathogenesis of Hepatitis C

Introduction

Hepatitis C virus (HCV) infection is a prominent cause of chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma (HCC), with up to 185 million chronically infected individuals worldwide, most of whom are undiagnosed and untreated. A protective vaccine is not available, and those who clear infection spontaneously or with treatment are susceptible to reinfection. The number of untreated patients presenting with long-term seqeula of chronic hepatitis C infection, including hepatocellular carcinoma, is expected to increase during the next decade. Recent advances in HCV therapeutics with the development and clinical application of directly acting antiviral agents (DAAs) have transformed HCV management in the developed world, although DAAs are not yet widely available worldwide. This chapter will provide an overview of the principles of HCV virology and the immunopathogenic mechanisms of disease.

Overview of the Virion

HCV was identified in 1989 as the most common etiologic agent of posttransfusion and sporadic non-A, non-B hepatitis, a clinical entity first reported in 1975. Despite difficulties propagating HCV infection in vitro , marked scientific progress has been made during the past 26 years with use of heterologous expression systems, functional infectious complementary DNA clones in chimpanzees, the replicon system, retroviral pseudoparticles displaying functional HCV glycoproteins, and complete cell culture systems.

HCV is classified in the genus Hepacivirus within the family Flaviviridae, which also includes yellow fever virus, dengue virus, and West Nile virus. The genetic organization and polyprotein processing of HCV are illustrated in Fig. 28-1 . HCV contains a 9.6-kb positive-strand RNA genome composed of a 5′ noncoding region with an internal ribosome entry site, an open reading frame that encodes the structural and nonstructural proteins, and a 3′ noncoding region. The structural proteins include the core and envelope glycoproteins E1 and E2, and the p7 viroporin. The nonstructural proteins include the NS2-3 and NS3-4A proteases, the NS3 RNA helicase, the NS4B and NS5A proteins, and the NS5B RNA-dependent RNA polymerase (RdRp). Infectious virus is associated with low-density and very low density lipoproteins, as well as host apolipoproteins B, C1, and E. HCV infection is a highly dynamic process, with an estimated serum HCV half-life of less than 1 hour, an estimated infected hepatocyte half-life of approximately 1 week, and production and clearance of up to 10 virions per day in an infected individual. High replicative activity and the lack of a proofreading function of the viral RdRp is the basis of the high genetic variability of HCV. HCV isolates are classified into genotypes and subtypes. The seven HCV genotypes differ in their nucleotide sequence by 30% to 35%, and within an HCV genotype, subtypes (designated a, b, etc.) can be defined, and differ in nucleotide sequence by approximately 20%. Genotype distribution varies globally, and HCV genotype is associated with variable responses to interferon (IFN)-based and IFN-free treatment with DAAs.

Model Systems

The development of selectable subgenomic, dicistronic HCV RNA replicons, capable of autonomous replication in Huh-7 cells (a human hepatocyte-derived cellular carcinoma cell line) transformed opportunities to conduct HCV research ( Fig. 28-2 ). This model allows in vitro study of HCV RNA, genetic dissection of HCV RNA elements and proteins, and biochemical and ultrastructural characterization of the viral replication complex, and has facilitated the discovery and development of DAAs and the investigation of antiviral resistance in vitro . Furthermore, it has been used to study interactions between viral and host cell components. A major breakthrough came with the discovery that a genotype 2a strain of virus, derived from a Japanese patient with a rare case of fulminant hepatitis and designated JFH-1 (for Japanese fulminant hepatitis 1), was able to replicate and release infectious particles in cell culture, which was subsequently also achieved with the H77 genotype 1a isolate, facilitating studies to explore the viral life cycle. Cell culture–derived HCV particles were found to be infectious in vivo in chimpanzees and immunodeficient mice with chimeric human livers, and viral inocula derived from these animal models were infectious for naïve Huh-7 cells in vitro . This enabled the study of the life cycle steps, including viral entry, genome packaging, virion assembly, maturation, and release. The challenge remains to develop in vivo and ex vivo models that mimic the natural environment of the liver, as primary HCV isolates show a poor ability to replicate in tissue culture.

Progress in the development of a small animal model of HCV replication has been achieved with the successful infection of immunodeficient mice reconstituted with human hepatocytes. Inoculation with serum from patients with hepatitis C results in persistent HCV viremia in mice with high-level human hepatocyte engraftment, with viral titers similar to those found in infected humans ( Fig. 28-3 ). This model has been used successfully to investigate the in vivo phenotype of specific mutations introduced into the HCV genome, selected aspects of viral pathogenesis, and neutralization of viral entry, and to evaluate novel antiviral strategies. Table 28-1 summarizes the current model systems and the advantages and limitations of each of them.

Structure and Function of the Viral Proteins

The structures and membrane association of HCV proteins are illustrated in Figs. 28-4 and 28-5 .