• Humans
• Hepacivirus/genetics
• Cohort Studies
• Virus Internalization
• Hepatitis C/drug therapy

• Antiviral
• Fused bicyclic derivative of pyrrolidine and imidazolidinone
• Hepatitis C virus (HCV)

• Humans
• Antiviral Agents/pharmacology
• Antiviral Agents/therapeutic use
• Hepatitis C, Chronic
• Hepatitis C
• Hepacivirus/genetics
• Virus Replication
• Pyrrolidines/pharmacology

• compartment formation dynamics
• positive sense RNA virus
• stochastic fate of infection
• synergy in virus inhibition
• viral intracellular life cycle model

• Animals
• Humans
• Life Cycle Stages
• Positive-Strand RNA Viruses
• RNA Viruses
• RNA, Viral/genetics
• Virus Replication

• The Wellcome Trust DBT India Alliance
• Council of Scientific and Industrial Research, India
• Prime Minister Research Fellowship

• A549 cells
• COVID-19
• Huh7.5 cells
• coronavirus
• galidesivir
• molnupiravir
• nucleotide analogs
• remdesivir
• sofosbuvir
• virus evolution

• lipopolysaccharides
• macrophages
• scavenger receptor class B type 1
• vitamin D3

• bistability
• direct-acting antiviral agents
• interferon signaling
• multiscale modeling
• sustained virological response
• viral kinetics

• adaptive mutation
• hepatitis C virus
• infectivity
• thermostability

Hepatitis C virus (HCV) continues to spread worldwide with an annual increase of 1.75 million new infections. The number of HCV cases in the U.S. is now greater than the number of HIV cases and is increasing in young adults because of the opioid epidemic sweeping the country. HCV-related liver disease is the leading indication of liver transplantation. An effective vaccine is of paramount importance to control and prevent HCV infection. While this vaccine will need to induce both cellular and humoral immunity, this review is focused on the required antibody responses. For highly variable viruses, such as HCV, isolation and characterization of monoclonal antibodies mediating broad virus neutralization are an important guide for vaccine design. The viral envelope glycoproteins, E1 and E2, are the main targets of these antibodies. Epitopes on the E2 protein have been studied more extensively than epitopes on E1, due to higher antibody targeting that reflects these epitopes having higher degrees of immunogenicity. E2 epitopes are overall organized in discrete clusters of overlapping epitopes that ranged from high conservation to high variability. Other epitopes on E1 and E1E2 also are targets of neutralizing antibodies. Taken together, these regions are important for vaccine design. Another element in vaccine design is based on information on how the virus escapes from broadly neutralizing antibodies. Escape mutations can occur within the epitopes that are involved in antibody binding and in regions that are not involved in their epitopes, but nonetheless reduce the efficiency of neutralizing antibodies. An understanding on the specificities of a protective B cell response, the molecular locations of these epitopes on E1, E2, and E1E2, and the mechanisms, which enable the virus to negatively modulate neutralizing antibody responses to these regions will provide the necessary guidance for vaccine design.

• antigenic domains
• epitopes
• hepatitis C virus
• human monoclonal antibodies
• vaccine design
• virus neutralization

• clinical trial
• direct-acting antivirals
• hepatitis C virus
• host-targeting agents
• treatment

To investigate the effect of a single amino acid mutation in human class B scavenger receptor I (SR-BI) on the infectivity of cell culture-derived hepatitis C virus (HCVcc) in SR-BI knock-down Huh7-siSR-BI cells.

Site-directed mutagenesis was used to construct the SR-BI S112F mutation, and the mutation was confirmed by nucleotide sequencing. SR-BI knock-down Huh7-siSR-BI cells were transfected with SR-BI S112F, SR-BI wild type (WT) and control plasmids, and then infected with HCVpp (HCV pseudoparticles) and hepatitis C virus derived from cell culture (HCVcc). A fluorescence assay was performed to analyze the effect of the S112F mutation on HCV entry; quantitative real-time PCR, immunofluorescence, and Western blot assays were used to analyze the effect of the S112F mutation on HCV infectivity. CHO cells expressing WT and SR-BI S112F were incubated with the HCV E2 protein expressed in HEK 293T cells, and flow cytometry was performed to examine the ability of SR-BI S112F to bind to the HCV E2 protein. Huh7-siSR-BI cells were transfected with SR-BI WT and the S112F mutant, and then DiI-HDL was added and images captured under the microscope to assess the ability of SR-BI S112F to take up HDL.

The SR-BI S112F mutation was successfully constructed. The S112F mutation decreased the expression of the SR-BI mRNA and protein. SR-BI S112F decreased HCV entry and HCVcc infectivity in Huh7-siSR-BI cells. The S112F mutation impaired the binding of SR-BI to HCV E2 protein and decreased the HDL uptake of SR-BI.

The S112F single amino acid mutation in SR-BI decreased the levels of the SR-BI mRNA and protein, as well as the ability of SR-BI to bind to the HCV E2 protein. Amino acid 112 in SR-BI plays important roles in HCV entry and the infectivity of HCVcc in vitro.

• Hepatitis C virus
• Class B scavenger receptor I
• Mutant
• Infectivity

• Animals
• CHO Cells
• Cell Line, Tumor
• Cricetinae
• Cricetulus
• Down-Regulation
• Flow Cytometry
• Gene Knockdown Techniques
• HEK293 Cells
• Hepacivirus/metabolism
• Hepacivirus/pathogenicity
• Hepatitis C/genetics
• Hepatitis C/virology
• Humans
• Mutagenesis, Site-Directed
• Point Mutation
• RNA Interference
• RNA, Messenger/metabolism
• RNA, Small Interfering/metabolism
• Real-Time Polymerase Chain Reaction
• Scavenger Receptors, Class B/genetics
• Scavenger Receptors, Class B/metabolism
• Transfection
• Viral Envelope Proteins/metabolism
• Virus Internalization

• CD5
• Cancer immunotherapy
• Danger signal
• Immune-checkpoint
• Scavenger receptor
• Scavenger receptor class B type 1
• Toll-like receptors

One of the most exciting developments in antiviral research has been the discovery of the direct-acting antivirals (DAAs) that effectively cure chronic hepatitis C virus (HCV) infections. Based on more than 100 clinical trials and real-world studies, we provide a comprehensive overview of FDA-approved therapies and newly discovered anti-HCV agents with a special focus on drug efficacy, mechanisms of action, and safety. We show that HCV drug development has advanced in multiple aspects: (i) interferon-based regimens were replaced by interferon-free regimens; (ii) genotype-specific drugs evolved to drugs for all HCV genotypes; (iii) therapies based upon multiple pills per day were simplified to a single pill per day; (iv) drug potency increased from moderate (∼60%) to high (>90%) levels of sustained virologic responses; (v) treatment durations were shortened from 48 to 12 or 8 weeks; and (vi) therapies could be administered orally regardless of prior treatment history and cirrhotic status. However, despite these remarkable achievements made in HCV drug discovery, challenges remain in the management of difficult-to-treat patients.

•

HCV genotype-specific drugs evolve to pan-genotypic drugs.

• Direct-acting antivirals
• NS3/4A drugs
• NS5A drugs
• NS5B drugs

• ANTIVIRAL THERAPY
• CHRONIC VIRAL HEPATITIS
• HCV
• HEPATITIS C
• LIVER

• Antiviral Agents/therapeutic use
• Disease Progression
• Hepacivirus/drug effects
• Hepatitis C/drug therapy
• Hepatitis C, Chronic/complications
• Hepatitis C, Chronic/drug therapy
• Hepatitis C, Chronic/epidemiology
• Humans
• Incidence
• Liver Cirrhosis/drug therapy
• Liver Cirrhosis/etiology
• Liver Transplantation
• Virus Activation/drug effects

• K24 DK078772 NIDDK NIH HHS
• U19 AI123862 NIAID NIH HHS

• Antivirals
• HCV
• Lipidoid
• Nanoparticles
• PRK2
• siRNA

• Animals
• Antiviral Agents/administration & dosage
• Hepacivirus
• Mice
• Nanoparticles
• RNA, Small Interfering/administration & dosage
• Viral Nonstructural Proteins
• Virus Replication

• Intestinal SR-BI
• Nonabsorbed SR-BI inhibitor
• SR-BI inhibitor
• Scavenger receptor class B type I

• allograft
• antiviral
• entry
• inhibitor
• recurrence

MEAN (6‐methoxyethylamino‐numonafide) is a small molecule compound, and here, we report that it effectively inhibits hepatitis C virus (HCV) infection in an HCV cell culture system using a JC1‐Luc chimeric virus, with a 50% effective concentration (EC50) of 2.36 ± 0.29 μM. Drug combination usage analyses demonstrated that MEAN was synergistic with interferon α, ITX5061 and ribavirin. In addition, MEAN effectively inhibits N415D mutant virus and G451R mutant viral infections. Mechanistic studies show that the treatment of HCV‐infected hepatocytes with MEAN inhibits HCV replication but not translation. Furthermore, treatment with MEAN significantly reduces polypyrimidine tract‐binding protein (PTB) levels and blocks the cytoplasmic redistribution of PTB upon infection. In the host cytoplasm, PTB is directly associated with HCV replication, and the inhibition of HCV replication by MEAN can result in the sequestration of PTB in treated nuclei. Taken together, these results indicate that MEAN is a potential therapeutic candidate for HCV infection, and the targeting of the nucleo‐cytoplasmic translocation of the host PTB protein could be a novel strategy to interrupt HCV replication.

• anti-HCV drug
• novel strategy
• nucleo-cytoplasmic translocation
• polypyrimidine tract-binding protein

• Animals
• Antiviral Agents/chemistry
• Cell Survival
• Computational Biology
• Disease Models, Animal
• Genotype
• HEK293 Cells
• Hepacivirus/drug effects
• Hepacivirus/physiology
• Hepatitis C/drug therapy
• Humans
• Internal Ribosome Entry Sites/genetics
• Mice
• Nanoparticles/chemistry
• Nucleotides/chemistry
• Protein Structure, Tertiary
• RNA, Small Interfering/chemistry
• RNA, Viral/chemistry
• Transfection
• Virus Replication

Hepatitis C virus (HCV) entry inhibitors (EIs) act synergistically with drugs targeting other stages of the HCV lifecycle. The origin of this synergy remains unknown. Here, we argue that the synergy may arise from the complementary activities of the drugs across cell subpopulations expressing different levels of HCV entry receptors. We employ mathematical modeling of viral kinetics in vitro, where cells with a distribution of entry receptor expression levels are exposed to HCV with or without drugs. The drugs act independently in each cell, as expected in the absence of underlying interactions. Yet, at the cell population level our model predicts that the drugs exhibit synergy. EIs effectively block infection of cells with low receptor levels. With high receptor levels, where EIs are compromised, other drugs are potent. This novel mechanism of synergy, arising at the cell population level may facilitate interpretation of drug activity and treatment optimization.

A potent class of indolinyl-thiazole based inhibitors of cellular lipid uptake mediated by scavenger receptor, class B, type I (SR-BI) was identified via a high-throughput screen of the National Institutes of Health Molecular Libraries Small Molecule Repository (NIH MLSMR) in an assay measuring the uptake of the fluorescent lipid DiI from HDL particles. This class of compounds is represented by ML278 (17–11), a potent (average IC₅₀ = 6 nM) and reversible inhibitor of lipid uptake via SR-BI. ML278 is a plasma-stable, noncytotoxic probe that exhibits moderate metabolic stability, thus displaying improved properties for in vitro and in vivo studies. Strikingly, ML278 and previously described inhibitors of lipid transport share the property of increasing the binding of HDL to SR-BI, rather than blocking it, suggesting there may be similarities in their mechanisms of action.

• HCV
• HDL receptor
• HTS
• ML278
• MLP
• SR-BI inhibitor
• indoline
• reverse cholesterol transport
• thiazole

• Cholesterol transport
• DOS macrocycle
• HDL receptor
• ML312
• SR-BI inhibitor

• Epitopes
• Genotype
• Hepatitis C Virus
• Peptides

• Animals
• Antiviral Agents/pharmacology
• Antiviral Agents/therapeutic use
• Cell Line
• Cell Line, Tumor
• Curcumin/pharmacology
• Curcumin/therapeutic use
• Hepacivirus/drug effects
• Hepacivirus/physiology
• Hepatitis C/prevention & control
• Hepatitis C/virology
• Hepatocytes/virology
• Humans
• Mice
• Virus Assembly/drug effects
• Virus Internalization/drug effects
• Virus Release/drug effects
• Virus Replication/drug effects

• R01 AI072613 NIAID NIH HHS
• R01 AI099284 NIAID NIH HHS
• R01AI099284 NIAID NIH HHS
• R01AI072613 NIAID NIH HHS

Hepatitis C virus (HCV) is transmitted between hepatocytes via classical cell entry but also uses direct cell-cell transfer to infect neighboring hepatocytes. Viral cell-cell transmission has been shown to play an important role in viral persistence allowing evasion from neutralizing antibodies. In contrast, the role of HCV cell-cell transmission for antiviral resistance is unknown. Aiming to address this question we investigated the phenotype of HCV strains exhibiting resistance to direct-acting antivirals (DAAs) in state-of-the-art model systems for cell-cell transmission and spread. Using HCV genotype 2 as a model virus, we show that cell-cell transmission is the main route of viral spread of DAA-resistant HCV. Cell-cell transmission of DAA-resistant viruses results in viral persistence and thus hampers viral eradication. We also show that blocking cell-cell transmission using host-targeting entry inhibitors (HTEIs) was highly effective in inhibiting viral dissemination of resistant genotype 2 viruses. Combining HTEIs with DAAs prevented antiviral resistance and led to rapid elimination of the virus in cell culture model. In conclusion, our work provides evidence that cell-cell transmission plays an important role in dissemination and maintenance of resistant variants in cell culture models. Blocking virus cell-cell transmission prevents emergence of drug resistance in persistent viral infection including resistance to HCV DAAs.

In spite of the rapid development of antiviral agents, antiviral resistance remains a challenge for the treatment of viral infections including hepatitis B and C virus (HBV, HCV), human immunodeficiency virus (HIV) and influenza. Virus spreads from infected cells to surrounding uninfected host cells to develop infection through cell-free and cell-cell transmission routes. Understanding the spread of resistant virus is important for the development of novel antiviral strategies to prevent and treat antiviral resistance. Here, we characterize the spread of resistant viruses and its impact for emergence and prevention of resistance using HCV as a model system. Our results show that cell-cell transmission is the main transmission route for antiviral resistant HCV strains and is crucial for the maintenance of infection. Monoclonal antibodies or small molecules targeting HCV entry factors are effective in inhibiting the spread of resistant HCV in cell culture models and thus should be evaluated clinically for prevention and treatment of HCV resistance. Combination of inhibitors targeting viral entry and clinically used direct-acting antivirals (DAAs) prevents antiviral resistance and leads to viral eradication in cell culture models. Collectively, the investigation provides a new strategy for prevention of viral resistance to antiviral agents.

• Antibodies, Neutralizing/metabolism
• Antiviral Agents/pharmacology
• Carbamates
• Cell Communication/immunology
• Cells, Cultured
• Drug Resistance, Viral/immunology
• Hepacivirus/drug effects
• Hepacivirus/growth & development
• Hepacivirus/physiology
• Hepatitis C/immunology
• Hepatitis C/pathology
• Hepatitis C/virology
• Humans
• Imidazoles/pharmacology
• Oligopeptides/pharmacology
• Proline/analogs & derivatives
• Proline/pharmacology
• Pyrrolidines
• Valine/analogs & derivatives
• Viral Load/immunology
• Virus Internalization/drug effects

• MC_U130184144 Medical Research Council
• MC_UU_12014/2 Medical Research Council

The current therapeutic regimen to combat chronic hepatitis C is not optimal due to substantial side effects and the failure of a significant proportion of patients to achieve a sustained virological response. Recently developed direct-acting antivirals targeting hepatitis C virus (HCV) enzymes reportedly increase the virologic response to therapy but may lead to a selection of drug-resistant variants. Besides direct-acting antivirals, another promising class of HCV drugs in development include host targeting agents that are responsible for interfering with the host factors crucial for the viral life cycle. A family of host proteins known as DEAD-box RNA helicases, characterized by nine conserved motifs, is known to play an important role in RNA metabolism. Several members of this family such as DDX3, DDX5 and DDX6 have been shown to play a role in HCV replication and this review will summarize our current knowledge on their interaction with HCV. As chronic hepatitis C is one of the leading causes of hepatocellular carcinoma, the involvement of DEAD-box RNA helicases in the development of HCC will also be highlighted. Continuing research on the interaction of host DEAD-box proteins with HCV and the contribution to viral replication and pathogenesis could be the panacea for the development of novel therapeutics against HCV.

• Hepatitis C virus
• Chronic hepatitis C
• Hepatitis C virus therapy
• DEAD-box helicases
• Host factors
• Hepatocellular carcinoma

• Animals
• Antiviral Agents/therapeutic use
• Carcinoma, Hepatocellular/etiology
• DEAD-box RNA Helicases/metabolism
• Genome, Viral
• Hepacivirus/physiology
• Hepatitis C/drug therapy
• Hepatitis C/enzymology
• Host-Pathogen Interactions
• Humans
• Liver Neoplasms/etiology
• Viral Proteins/metabolism

• Difficult-to-treat patients
• Direct-acting antivirals
• Entry inhibitors
• Liver transplantation
• Resistance

• Cyclophilin A
• Hepatitis C
• Host-targeting antivirals
• MicroRNA-122
• Phosphatidylinositol 4-kinase III alpha
• Scavenger receptor class B type I

• Alisporivir
• Cyclophilin A
• HCV
• Host-targeting antivirals

• HCV
• SRB1
• entry inhibitors

• Animals
• Antiviral Agents/therapeutic use
• Genotype
• Hepacivirus/drug effects
• Hepacivirus/genetics
• Hepacivirus/physiology
• Hepatitis C/drug therapy
• Hepatitis C/virology
• Host-Pathogen Interactions/genetics
• Host-Pathogen Interactions/immunology
• Humans
• Models, Biological
• Treatment Outcome
• Virus Replication/physiology

• R01 AI072613 NIAID NIH HHS
• R01 AI091707 NIAID NIH HHS
• AI090055 NIAID NIH HHS
• R01 AI075099 NIAID NIH HHS
• R01 AI090055 NIAID NIH HHS
• AI075099 NIAID NIH HHS
• AI072613 NIAID NIH HHS
• DK085713 NIDDK NIH HHS
• R01 AI099284 NIAID NIH HHS
• AI099284 NIAID NIH HHS
• AI091707 NIAID NIH HHS
• R01 CA057973 NCI NIH HHS
• UL1RR024143 NCRR NIH HHS
• R01 DK085713 NIDDK NIH HHS
• CA057973 NCI NIH HHS
• UL1 RR024143 NCRR NIH HHS

Efforts to treat HCV patients are focused on developing antiviral combinations that lead to the eradication of infection. Thus, it is important to identify optimal combinations from the various viral inhibitor classes. Based on viral dynamic models, HCV entry inhibitors are predicted to reduce viral load in a monophasic manner reflecting the slow death rate of infected hepatocytes (t_1/2 = 2–70 days) and the protection of naïve, un-infected cells from HCV infection. In contrast, replication inhibitors are predicted to reduce viral load in a biphasic manner. The initial rapid reduction phase is due to the inhibition of virus production and elimination of plasma virus (t_1/2∼3 hours). The second, slower reduction phase results from the elimination of infected hepatocytes. Here we sought to compare the ability of HCV entry and replication inhibitors as well as combinations thereof to reduce HCV infection in persistently-infected Huh7 cells. Treatment with 5×EC₅₀ of entry inhibitors anti-CD81 Ab or EI-1 resulted in modest (≤1 log₁₀ RNA copies/ml), monophasic declines in viral levels during 3 weeks of treatment. In contrast, treatment with 5×EC₅₀ of the replication inhibitors BILN-2016 or BMS-790052 reduced extracellular virus levels more potently (∼2 log₁₀ RNA copies/ml) over time in a biphasic manner. However, this was followed by a slow rise to steady-state virus levels due to the emergence of resistance mutations. Combining an entry inhibitor with a replication inhibitor did not substantially enhance the rate of virus reduction. However, entry/replication inhibitor and replication/replication inhibitor combinations reduced viral levels further than monotherapies (up to 3 log₁₀ RNA copies/ml) and prolonged this reduction relative to monotherapies. Our results demonstrated that HCV entry inhibitors combined with replication inhibitors can prolong antiviral suppression, likely due to the delay of viral resistance emergence.

• Animals
• Antibodies, Monoclonal/immunology
• Antibodies, Neutralizing/immunology
• Cell Membrane/immunology
• Hepacivirus/immunology
• Hepacivirus/physiology
• Hepatitis C/immunology
• Hepatitis C/virology
• Hepatitis C Antibodies/immunology
• Humans
• Immunization
• Rats
• Rats, Wistar
• Tetraspanin 28/immunology
• Viral Envelope Proteins/immunology
• Virus Internalization

Only 20 years after the discovery of the Hepatitis C Virus (HCV), a cure is now likely for most people affected by this chronic infection, which carries a substantial disease burden, not only in the United States but also worldwide.1 The recent approval of two direct-acting antiviral agents that specifically inhibit viral replication has dramatically increased the viral clearance rate, from less than 10% with the initial regimen of interferon monotherapy to more than 70% with current therapy. Moreover, many other drugs targeting viral or host factors are in development, and some will almost certainly be approved in the coming years. The questions of who should be treated and with what regimen will be increasingly complex to address and will require careful consideration. As therapy improves, systemwide identification and care of patients who need treatment will be the next challenge. Because most infected persons are unaware of their diagnosis, the Centers for Disease Control and Prevention recently recommended screening for HCV all persons born between 1945 and 1965.2 ,3 It is anticipated that in the course of such a screening process, a large number of persons will be found to be infected with the virus; whether it will be possible to treat all these people is unclear. This article reviews the current therapy for HCV infection and the landscape of drug development.

• Antiviral Agents/therapeutic use
• Drug Therapy, Combination
• Genotype
• Hepacivirus/genetics
• Hepacivirus/growth & development
• Hepatitis C/drug therapy
• Humans
• Interferon-alpha/therapeutic use
• Standard of Care

• ZIA DK054505-17 Intramural NIH HHS
• ZIA DK054504-17 Intramural NIH HHS
• Z99 OD999999 Intramural NIH HHS
• ZIA DK054504 Intramural NIH HHS
• Z99 DK999999 Intramural NIH HHS
• ZIA DK054505-16 Intramural NIH HHS
• ZIA DK054511-06 Intramural NIH HHS
• ZIA DK054511-07 Intramural NIH HHS
• ZIA DK054505 Intramural NIH HHS
• Z99 DA999999 Intramural NIH HHS
• ZIA DK054504-16 Intramural NIH HHS
• ZIA DK054511 Intramural NIH HHS

• Hepatitis C virus
• Life cycle
• Virology

• Genome, Viral
• Hepacivirus/genetics
• Hepacivirus/physiology
• Humans
• RNA, Viral/metabolism
• Scavenger Receptors, Class B/metabolism
• Tetraspanin 28/metabolism
• Viral Envelope Proteins/chemistry
• Viral Envelope Proteins/metabolism
• Viral Nonstructural Proteins/chemistry
• Viral Nonstructural Proteins/metabolism
• Virus Assembly
• Virus Internalization
• Virus Replication