“
“See Covering the Cover synopsis on page 946; see editorial on page 959. Infection with the hepatitis C virus (HCV) is a major cause of chronic liver disease and accounts

for a large proportion of liver cirrhosis cases and hepatocellular carcinomas.1 Given an estimated 130 to 170 million infected individuals worldwide and the high prevalence in industrialized countries, intensive efforts are being undertaken to improve antiviral therapy.2 Very recently, HCV-specific direct-acting antiviral drugs Apoptosis Compound Library mw have become available that allow virus elimination in the majority of treated patients, however, drug resistance, incomplete genotype coverage, and high costs are important limitations.3 HCV is a plus-strand RNA virus encoding a single polyprotein that is proteolytically cleaved into 10 different products (reviewed in Moradpour and Penin4). Of these, nonstructural protein (NS) 3, NS4A, NS4B, NS5A, and NS5B form a multiprotein complex mediating viral replication. Like all plus-strand RNA viruses, HCV replication occurs in cytoplasmic membranous factories. These are composed primarily of double- and multimembrane vesicles forming a heterogeneous Atezolizumab meshwork designated “membranous web” (MW).5 and 6 It is induced by a concerted action

of HCV replicase proteins6 together with host cell factors, most notably phosphatidylinositol-4 kinase IIIα (PI4KIIIα).7 and 8 This lipid kinase is recruited to viral replication sites by interaction with NS5A, leading to the accumulation of high amounts of PI4-phosphate (PI4P) at intracellular membranes. NS5A is a multifunctional zinc-binding protein (reviewed in Moradpour and Penin4). It is phosphorylated at several sites by cellular kinases giving rise to basal (p56) and hyperphosphorylated (p58) NS5A. Phosphorylation is thought to regulate the multitude

of NS5A functions, including RNA binding and self-interaction. NS5A is composed of an N-terminal amphipathic α-helix (AH) tethering the protein to membranes,9 D-malate dehydrogenase a highly structured domain I (DI)10 and 11 and 2 intrinsically unfolded “domains” with limited sequence conservation between genotypes (reviewed in Moradpour and Penin4). Four x-ray crystal structures of NS5A DI of genotype 1 revealed virtually identical monomer conformations, but distinct dimer organizations that have been proposed to form multimeric complexes.10, 11 and 12 High-throughput screening with HCV replicons and optimization of lead compounds led to the development of highly potent direct-acting antiviral drugs targeting NS5A and efficiently inhibiting viral RNA synthesis and virus assembly.13, 14 and 15 As illustrated with daclatasvir, the first inhibitor of this class, these drugs are characterized by a symmetric structure with a rigid central core and unparalleled antiviral activity.