NS5A: A new target for antiviral drugs in the treatment of hepatitis C virus infection
Authors: Jose D. Debes, Coleman I. Smith
Journal: Hepatology (2012)

DAA, direct-acting antiviral agents; HCV, hepatitis C virus; ISDR, interferon-alpha sensitivity determining region. Hepatitis C virus (HCV) is a major cause of liver disease and mortality affecting 170 million people worldwide.1 Chronic infection with HCV can lead to liver fibrosis, cirrhosis, and hepatocellular carcinoma2 and is the most common reason for liver transplantation in the United States.3 HCV is an enveloped single-strand RNA virus that belongs to the flaviviridae family. It has a 9.6-kb RNA genome encoding a single polyprotein of 3,010 amino acids. This polyprotein is composed of structural and nonstructural proteins. HCV entry into hepatocytes is mediated by the structural proteins, including glycoproteins E1 and E2, which interact with cellular receptors. Once HCV enters the host cell, replication of its genome is orchestrated by the nonstructural proteins, including proteases (NS2-3, NS3-4A), helicases (NS3), polymerases (NS5B), and NS5A, a protein with no known enzymatic activity.4-6 NS5A is essential to the replication machinery of the virus and critical in the assembly of infectious viral particles. However, its specific role remains unclear.6 A number of properties of this viral protein have been described. NS5A interacts with a variety of host proteins, including some which modulate host cellular signaling pathways.7, 8 Studies have demonstrated that NS5A interacts with p53 and p21, affecting cell cycle control.7 More recently, NS5A was found to interact with proteins related to focal adhesions, gap-junction, and host signaling pathways.7, 8 NS5A also interacts with other nonstructural viral proteins. During HCV viral replication, NS5A interacts with the RNA- dependent RNA-polymerase NS5B, which is essential to maintain HCV replication in cell culture. Through its domain 1 NS5A has RNA-binding capacity, a property that can be critical for RNA replication.9 In addition to being involved in HCV replication, NS5A is involved in the assembly of the virus by forming stable complexes with NS2, the main protein involved in HCV assembly.10 NS5A attracted attention due to its interferon-alpha sensitivity determining region (ISDR), which can confer resistance to interferon treatment. Moreover, NS5A has an effect on interferon activity by up-regulating interleukin (IL)-8, which has been reported to attenuate the antiviral properties of interferon.11 These observations suggest that NS5A plays a critical role in many aspects of the life cycle of HCV and is therefore an attractive target for antiviral therapy (Fig. 1). The potential effects of targeting NS5A. Increase in interferon sensitivity by direct effect of blocking NS5A ISDR or by down-regulation of IL-8. Alteration of NS5A interaction with NS5B and NS2 affecting HCV replication and viral assembly. Downregulation of host signaling pathways such as those involving p21, p53, and Cdk2. Until recently, the standard treatment for HCV infection has been pegylated interferon-alpha and ribavirin. These drugs have significant and often serious side effects. Moreover, in patients with genotype 1 (the most prevalent HCV genotype), sustained viral clearance is achieved in less than 50% of cases.12 In recent years, several viral inhibitors targeting nonstructural proteins of HCV have been developed (direct-acting antiviral agents [DAA]). Most of the research has been focused on inhibitors of the NS3-NS4A and NS5B proteins, mainly because of a better understanding of the role of these proteins. A variety of NS3-NS4A and NS5B inhibitors are currently in various stages of development in preclinical and clinical studies.13 Telaprevir and boceprevir, which are both inhibitors of the NS3-NS4A serine protease, have recently been approved for use in combination with pegylated interferon and ribavirin in patients with genotype 1 HCV infection.14, 15 Treatment with pegylated interferon, ribavirin, and either of these protease inhibitors results in improved rates of viral eradication (both in previously untreated and in those who have failed prior treatment). However, the recommended treatment regimens are complex and are associated with significant side effects from interferon and ribavirin, as well as adverse events from the protease inhibitors. Furthermore, these protease inhibitors need to be administered every 8 hours with food and there is also a very real problem of interaction with other drugs. In addition, the rapid replication of HCV makes the appearance of viral resistance a potential problem, requiring the need for combination therapies with drugs directed against a variety of viral targets BMS-790052 is an agent with activity against the NS5A protein. This agent was identified in 2010 using a screening strategy that was designed to identify new agents that inhibit HCV replication with no effect on NS3-4A or N5B (which already had known inhibitors). The compound was found to inhibit all HCV genotypes at low concentrations. Through coprecipitation it was demonstrated that BMS-790052 interacted with NS5A.16 BMS-790052 inhibits a number of aspects of viral replication17, 18 and has been shown in the replicon system to be additive or synergistic with other DAAs.16 Early clinical testing of BMS-790052 showed that a single dose of the NS5A inhibitor was able to markedly decrease HCV viral levels in patients with chronic HCV infection.16 Nettles et al.19 showed a marked decline in HCV RNA after administration of BMS-790052 in a double-blind, placebo-controlled, multiple ascending dose study in 30 patients infected with HCV genotype 1. The study demonstrated a dose-dependent effect on HCV RNA. There were no significant side effects observed in this study. Pharmacokinetic studies showed that this drug is effective in a once-a-day dose. Most patients had viral rebound by day 7. This viral rebound was associated with the emergence of mutations associated with viral resistance, suggesting that treatment with this drug alone will be unlikely to result in a sustained suppression of HCV RNA. Preliminary studies with BMS-790052 combined with another antiviral, with and without pegylated interferon and ribavirin, have been shown to successfully lower HCV levels for a longer period of time.20 BMS-790052 is a new, promising agent in the treatment of chronic HCV infection. It is the first drug with activity against NS5A. Further studies with longer courses of therapy of BMS-790052 in combination with other DAAs and pegylated interferon and ribavirin are required to establish the long-term efficacy and safety of BMS-790052. BMS 790052 is one of over 50 new DAAs that are in different stages of development from preclinical to phase 3 clinical trials. These agents act on one of three targets on the HCV. They inhibit NS3-NS4A protease, NS5B polymerase (nucleoside or nonnucleoside), or the NS5A protein. Many of these agents result in viral suppression in over 80% of cases. These classes of agents exhibit different properties, with respect to development of resistance mutations, genotypic coverage, potency, and toxicity (Table 1). Most of these molecules appear to be well tolerated and exhibit pharmacokinetics that allow for daily or twice-daily administration. The pattern of resistance mutations induced by these agents varies among the different classes of agents. This suggests that a combination of agents may be effective in treating hepatitis C and minimizing the development of resistance mutations. Studies are under way with single DAAs and combinations of these agents with interferon and ribavirin. Preliminary results appear promising. It has now been shown that treatment of hepatitis C with a combination of DAAs (including BMS 790052) alone, without interferon, may result in sustained viral clearance.20 There are a number of ongoing studies utilizing combinations of DAAs without interferon that are showing promise. It still remains to be seen whether ribavirin will be needed as part of these regimens to produce a sustained viral response. Oral, interferon-free treatment of hepatitis C appears a possibility. It will require a combination of agents aimed at different targets on the HCV. We are approaching the dawn of a new era in the treatment of HCV with the possibility of oral, effective, and well-tolerated combination therapy. These regimens have the possibility of eradicating HCV in the majority of patients. Much work is still needed to define the most clinically effective and least toxic regimens.