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About this book

​Hepatitis C is a liver disease caused by the hepatitis C virus (HCV) and infects approximately 75 million individuals worldwide. It is also one of the major causes of liver cancer and liver transplants. The elucidation of the HCV genome, and the development of a whole cell system to study the virus spurred the search for novel direct acting antiviral drugs to cure this disease. This global effort culminated in the development of direct acting antiviral drugs that led to cure rates approaching 100% in all patient populations after only 8-12 weeks of therapy. These efforts resulted in one of the greatest achievements in public health and provides the potential for eliminating HCV as a major disease worldwide.

This volume is aimed at a broad audience of academic and industrial scientists interested in the discovery and development of drugs to treat viral diseases and those interested in reading about one of the most unique accomplishments in biomedical research. The volume will provide a one of a kind reference work that highlights the many efforts, from the discovery of the HCV virus, to the invention of breakthrough medicines and their use in the real world to cure patients. It is the companion book to the volume "HCV: The Journey from Discovery to a Cure - Volume I".

Table of Contents


HCV NS5A Inhibitors


NS5A as a Target for HCV Drug Discovery

Discovery and development of HCV inhibitors is one of the most successful stories in the history of antiviral research. After more than 30 years of effort by academic and pharmaceutical researchers, HCV infection is a curable disease. In fact, HCV is the first chronic infectious disease to be cured with combinations of direct antiviral agents. Among these antiviral agents, NS5A inhibitors are the most potent. The unprecedented low pM potency, pan-genotype coverage, and well-tolerated clinical profile have made NS5A inhibitors an essential component of all interferon-/ribavirin-free regimens in currently approved HCV therapies. Since NS5A has no known enzymatic activity and is not a traditional antiviral target, this review focuses on the challenges and concerns that arose during the discovery of this class of inhibitor, the mode of action/inhibition, and the value of NS5A inhibitors in the treatment of HCV infection.
Donald R. O’Boyle, Min Gao

The Discovery and Development of Daclatasvir: An Inhibitor of the Hepatitis C Virus NS5A Replication Complex

The discovery of the hepatitis C virus (HCV) NS5A replication inhibitor daclatasvir (1) had its origins in a phenotypic screening lead. However, during the optimization campaign, it was observed that some members of the chemotype underwent a radical dimerization under the assay conditions. This redirected the effort to focus on palindromic molecules, a species subsequently shown to complement the dimeric nature of the NS5A protein. The most challenging aspect of the discovery program was extending antiviral activity to encompass GT-1a virus which was accomplished only after the development of extensive structure-activity relationships. In clinical trials, oral administration of daclatasvir (1) produced a profound effect on viral load with onset that was more rapid than had been seen previously with either NS3 protease or NS5B polymerase inhibitors. A groundbreaking clinical trial that combined daclatasvir (1) with the protease inhibitor asunaprevir (52) established that a chronic HCV infection could be cured with small molecule therapy in the absence of immune stimulation, setting the stage for approval of this regimen for the treatment of GT-1b-infected subjects by the Japanese health authorities on July 4, 2014.
Nicholas A. Meanwell, Makonen Belema

The Discovery of Ledipasvir (GS-5885): The Potent Once-Daily Oral HCV NS5A Inhibitor in the Single-Tablet Regimen Harvoni®

The advent of direct-acting antiviral agents (DAAs) has revolutionized the treatment and cure of chronic hepatitis C virus (HCV) infection. Herein is described the discovery of ledipasvir (LDV), an orally available HCV nonstructural protein 5A inhibitor with picomolar antiviral potency and a long pharmacokinetic half-life. The combination of LDV with the nonstructural protein 5B inhibitor sofosbuvir (SOF) is Harvoni® and represents the first approved single-tablet regimen for the treatment of HCV infection. This safe simple and efficacious regimen affords clinical trial cure rates over 95% and comparable effectiveness in real-world studies and has treatment durations as short as 8 weeks. The approval of Harvoni® heralded a new era for the treatment of HCV infection.
John O. Link

The Discovery of Velpatasvir (GS-5816): The Potent Pan-Genotypic Once-Daily Oral HCV NS5A Inhibitor in the Single-Tablet Regimens Epclusa and Vosevi®

The initial approval of the single-tablet regimen (STR) Harvoni®, containing the hepatitis C virus (HCV) nonstructural 5A protein (NS5A) inhibitor ledipasvir and the nonstructural 5B protein (NS5B) nucleotide inhibitor sofosbuvir (SOF), provided a major advancement in the treatment of individuals with chronic genotype 1 (GT1) HCV infection. Herein is described the discovery of velpatasvir (VEL, GS-5816), a pan-genotypic NS5A inhibitor with low picomolar activity against GT1–6 HCV and a high resistance barrier. The combinations of SOF/VEL as Epclusa® and SOF/VEL/voxilaprevir (VOX, NS3/4a protease inhibitor) as Vosevi® are the only pan-genotypic STRs for the treatment and cure of HCV infection. Epclusa® is the first approved pan-genotypic STR and affords high cure rates with a single 12-week treatment duration regardless of genotype, cirrhosis status, or the presence of baseline resistance variants. Vosevi® provides high cure rates for GT1–6-infected individuals who have previously failed therapy (96% cure rates for GT1–6 patients who had failed regimens with an NS5A inhibitor or 98% for those who had failed regimens without an NS5A inhibitor). With pan-genotype activity, no need for on-treatment monitoring, and real-world effectiveness comparable to that observed in clinical trials, the safe, simple, and effective STR Epclusa® is an important agent for eradication of HCV infection worldwide.
John O. Link

Discovery of Elbasvir

On January 28, 2016, the US Food and Drug Administration approved Zepatier™ (a fixed-dose two-drug combination containing the NS5A inhibitor elbasvir and the NS3/4A protease inhibitor grazoprevir) for the treatment of adult patients with chronic hepatitis C virus genotype 1 or genotype 4 infection. The discovery of elbasvir (EBR) was the result of a concerted research effort within Merck’s newly formed External Basic Research (also, EBR) group and a team of scientists from WuXi AppTec. Lead ID efforts combined components from both internal and literature compounds to generate a first-generation benzofuran-based NS5A inhibitor (MK-4882) that demonstrated preclinical proof of concept before entering phase 1 clinical trials. Lead optimization efforts and refinement of the core structure ultimately led to the identification of elbasvir, a ring-constrained tetracyclic indole-based analogue of MK-4882 which showed increased potency against clinical resistance variants and an improved resistance profile.
Craig Coburn

HCV NS5A as an Antiviral Therapeutic Target: From Validation to the Discovery and Development of Ombitasvir and Pibrentasvir as Components of IFN-Sparing HCV Curative Treatments

While IFN-based hepatitis C virus (HCV) treatment regimens required long treatment duration, they only achieved a limited cure rate in HCV-infected patients and were accompanied by significant therapy-based side effects. The first curative IFN-sparing therapies revolutionized HCV treatment by utilizing a cocktail of mechanistically orthogonal direct-acting antiviral (DAA) agents to achieve much higher cure rates in a shorter period of time and with fewer side effects. One of the drug targets that these therapies usually engaged was the HCV NS5A protein. This chapter reviews the Abbott/AbbVie HCV NS5A program, which discovered inhibitors of this protein using an in vitro phenotypic screen, validated the mechanism in vivo, and ultimately discovered two FDA-approved NS5A inhibitors ombitasvir (OMB) and pibrentasvir (PIB). OMB, a first-generation NS5A inhibitor, is a component of two FDA-approved IFN-sparing DAA therapies (Viekira Pak™ and Technivie™) with approval to treat genotypes 1 and 4, respectively. PIB, a next-generation NS5A inhibitor included in AbbVie’s next-generation therapy Mavyret™ (or Maviret™), prevents replication of HCV genotypes 1–6 and exhibits an improved resistance profile relative to other FDA-approved first-generation NS5A inhibitors.
Rolf Wagner, David A. DeGoey, John T. Randolph, Allan C. Krueger, Mark A. Matulenko, Warren M. Kati

HCV NS4B Inhibitors


Evolution of HCV NS4B Inhibitors

NS4B has remained for a long time an undisclosed target within the HCV drug discovery programs. However, impressive drug discovery efforts from 2005 to 2016 led to the identification of different chemical classes targeting NS4B as effective anti-HCV agents, and some of them act by impairing AH2-mediated membranous web formation or RNA-binding property. This book chapter aims to discuss research published on NS4B inhibitors focusing on hit identification and hit-to-lead optimization, also with respect to pharmacokinetic properties and structure-activity relationships raised for the different chemical classes taken into account. To date, the only clinical trial conducted with molecules targeting NS4B was focused on clemizole hydrochloride. However, even if NS4B ligands are not currently used in therapy, they can serve in the near future as new weapons to combat resistance to the current therapy.
Giuseppe Manfroni, Rolando Cannalire

Clinical Trials and Combination Therapy


The Evolution of Clinical Trials for Hepatitis C

The development of well-tolerated treatments that attain nearly universal cure of hepatitis C virus (HCV) infection, less than 30 years after the long-sought discovery of the causative agent, ranks as a landmark achievement of modern medicine. In the broadest sense, the international effort to address this global public health problem can be divided into an era of nonspecifically targeted therapy centering on interferon, a relatively brief “hybrid period” combining interferon and ribavirin with direct-acting antiviral agents (DAAs), and the latest era of DAA combination regimens. One of the most notable features of this story is the quantum leap in efficacy for DAA therapy to extraordinarily high levels instead of the years-long incremental steps that might have been anticipated. Similarly gratifying is the foundation on which the concept of curability, unique to HCV thus far in human virology, has been solidified based on the combination of our understanding of the molecular biology of the virus and the rarity, dating back to the interferon era, of virologic relapse after attainment of sustained virologic response. Although, at least until recently, the number of therapeutic agents was very limited, the combination of viral and host diversity ensured the development of a rich literature reflecting hundreds of treatment studies which dominated the scientific programs of the international liver meetings for many years. Viewed panoramically through a retrospective lens, the field developed in a logical sequence by first making the most out of the limited tools which were available and later by building on the remarkable elucidation of HCV biology by the scientific community and the paradigm of combination therapy for viral infection established in the HIV field to get us where we are today.
Viviana Figueroa Diaz, Mary Olson, Ira M. Jacobson

The Clinical Development of Ledipasvir/Sofosbuvir (LDV/SOF, Harvoni®)

The fixed-dose combination tablet of ledipasvir (LDV), an HCV NS5A inhibitor, and sofosbuvir (SOF), an HCV nucleotide analog NS5B polymerase inhibitor, was the first all-oral (one-pill once daily), interferon-free and ribavirin-free regimen approved for the treatment of patients with chronic hepatitis C. With over 5,900 HCV-infected patients enrolled in LDV/SOF clinical trials through late 2017, the accelerated clinical development program was able to generate safety and efficacy data across a broad range of patient populations. The initial registration trials demonstrated that 12 weeks of treatment with LDV/SOF resulted in high cure rates of over 95% in HCV genotype 1 patients regardless of historical negative treatment predictors including cirrhosis or prior treatment history. The program subsequently expanded to include other HCV genotypes and special populations with significant unmet medical need including but not limited to decompensated liver disease, HIV/HCV coinfection, posttransplantation, and children. With favorable pharmacokinetic properties, good safety profile, and high efficacy rates, the approval of LDV/SOF (Harvoni®) ushered in a new era of treatment and management for the millions of HCV-infected patients globally.
Anu Osinusi, John G. McHutchison

The Clinical Development of Sofosbuvir/Velpatasvir (SOF/VEL, Epclusa®)

The single-tablet regimen of sofosbuvir (SOF), an HCV nucleotide analog NS5B polymerase inhibitor, and velpatasvir (VEL), a second-generation HCV NS5A inhibitor, provides a highly efficacious, safe, and simple treatment regimen for patients with genotype 1–6 HCV infection. The clinical development program for SOF/VEL focused on generating safety and efficacy data across a broad range of patient populations to support a single treatment duration for all patients and therapeutic options for patients with compensated and decompensated liver disease. Three Phase 2 studies defined the optimal dose of VEL as 100 mg for a fixed-dose combination tablet with 400 mg of SOF and demonstrated that the treatment duration of 12 weeks provided high SVR rates across all genotypes irrespective of cirrhosis status, prior treatment history, or the presence of baseline resistance-associated substitutions (RASs). The Phase 3 studies enrolled and treated over 1,000 genotype 1–6 HCV-infected patients with 12 weeks of SOF/VEL. In patients with compensated cirrhosis, the overall SVR rate was 98%, and with SOF/VEL + RBV in patients with decompensated cirrhosis, the SVR rate was 94%. With minimal drug-drug interactions and no need for on-treatment safety monitoring, SOF/VEL for 12 weeks provides an important treatment option for patients of all genotypes and is ideally suited to address the global epidemic of chronic HCV infection.
Diana M. Brainard, John G. McHutchison

The Clinical Development of Sofosbuvir/Velpatasvir/Voxilaprevir (SOF/VEL/VOX, Vosevi®)

The single-tablet regimen of sofosbuvir (SOF), an HCV nucleotide analog NS5B polymerase inhibitor; velpatasvir (VEL), an HCV NS5A inhibitor; and voxilaprevir (VOX), an NS3/4 protease inhibitor, provides a highly efficacious, safe, and salvage regimen for patients with genotype 1 to 6 HCV infection with and without compensated cirrhosis who were previously unsuccessfully treated with direct-acting antivirals (DAAs). The clinical development program for SOF/VEL/VOX focused on generating safety and efficacy data in DAA-experienced patients without retreatment options as well as assessing the possibility of shortening treatment duration for DAA-naive patients. The Phase 3 studies enrolled and treated over 1,000 genotype 1–6 HCV-infected patients with SOF/VEL/VOX. In DAA-experienced patients treated with 12 weeks of SOF/VEL/VOX, the overall SVR rate was 97%, and high SVR rates were observed across all genotypes irrespective of prior DAA regimen, cirrhosis status, or the presence of baseline resistance-associated substitutions (RASs), supporting its use as a retreatment regimen. In DAA-naive patients treated with 8 weeks of SOF/VEL/VOX, the overall SVR rate was 95% making it an alternative treatment option for regions in which a shorter duration is of particular interest.
Luisa M. Stamm, John G. McHutchison

Clinical Development of Viekira Pak to Mavyret

Viekira Pak (ombitasvir/paritaprevir/ritonavir and dasabuvir) was one of the first interferon-free direct-acting antiviral (DAA) regimens to be approved for the treatment of genotype 1 HCV infection. The research and development team at Abbott/AbbVie based their approach to HCV cure on the use of three DAAs to avoid emergence of resistance, anchored by a potent protease inhibitor and NS5A inhibitor. Clinical trials were designed to answer multiple questions within a single study, in order to advance the regimen as quickly as possible in a highly competitive environment. The global phase 2 and 3 development program allowed for rapid identification of optimal treatment regimens and durations for populations defined by HCV subtype, prior treatment experience, and the presence of specific comorbidities such as cirrhosis and orthotopic liver transplant. The clinical trial program also clarified the limitations of this first-generation DAA regimen, including activity that was limited to genotypes 1 and 4 and the need for ribavirin for some patients, which defined a target product profile for a next-generation regimen. This continued research and development activity ultimately led to the approval of the pangenotypic regimen glecaprevir/pibrentasvir (Mavyret).
Daniel E. Cohen

Development of ZEPATIER®

ZEPATIER® (MK-5172A; elbasvir and grazoprevir, Merck & Co., Inc.) is a fixed-dose combination treatment for individuals with chronic hepatitis C virus (HCV) infection. This novel direct-acting antiviral (DAA) regimen combines elbasvir, a selective inhibitor of the HCV nonstructural protein 5A, and grazoprevir, a reversible competitive inhibitor of the HCV nonstructural protein 3/4A protease. After extensive preclinical testing and evaluation of safety and pharmacokinetics (PK) in healthy volunteers, the efficacy of these agents was evaluated in a systematic and comprehensive clinical development program culminating in phase 3 clinical trials in a broad population of participants with HCV infection, including treatment-naive and treatment-experienced participants, those with chronic kidney disease or inherited blood disorders, and those receiving opioid agonist therapy. These studies led to the approval of the elbasvir/grazoprevir combination therapy for the treatment of people with HCV genotype 1 or genotype 4 infection in the United States, Europe, Canada, and many other countries worldwide.
Michael N. Robertson, Eliav Barr

Impact of Curative Therapies


Real-World Evidence and Hepatitis C

Direct-acting antiviral agents (DAAs) are the treatment of choice for patients with chronic hepatitis C. Their efficacy across diverse patient populations and safety among those with all stages of liver disease, including cirrhosis, have been repeatedly demonstrated in studies encompassing all classes of DAAs. Real-world evidence has confirmed that DAA therapies used in usual clinical practice achieved similar rates of sustained virological response when compared to those reported in rigorously controlled clinical trials. These data, developed from large cohort studies performed around the world, have instilled greater confidence in the management of patients with chronic hepatitis C using DAAs. Furthermore, real-world evidence contributed to better understanding the strengths and limitations of DAA treatment among unique populations of patients with chronic hepatitis C who were underrepresented in the original registration trials of these agents.
Michael W. Fried, David R. Nelson

The Benefit of Direct-Acting Antiviral HCV Cure Therapies

Although more than 600,000 patients in the USA have now been cured of HCV, the clinical benefits of cure have been seen primarily in patients with more advanced liver disease. This has resulted in a reduced incidence of liver cancers and reduced liver-related mortality. Benefits of cure in patients without cirrhosis have not been seen yet in systemic review, but are likely to be seen in the future years. There remain unresolved issues regarding patients with advanced fibrosis or cirrhosis who are cured of their HCV. It is not clear if fibrosis reverses after cure in everyone, if so by how much, and for how long patients with cirrhosis need to be monitored after cure. We have shown that 62% had improved liver stiffness measured by transient elastography (TE) that was consistent with regression of at least one stage of fibrosis over 1 year. Fifteen patients with matched liver biopsies prior to SVR underwent a biopsy after SVR. However, the post-SVR liver biopsies of only 4 patients showed F1–F2, while 11 patients still showed F3–F4, indicating that TE improvements are overstated when compared to histologic staging and that patients with cirrhosis before DAA therapy need to be monitored for hepatocellular carcinoma after cure.
Paul J. Pockros

Cure and Control: What Will It Take to Eliminate HCV?

In May 2016, the World Health Organization adopted the first global hepatitis strategy, setting the ambitious goal of “elimination of viral hepatitis as a public health threat” by 2030. HCV-specific targets included a 65% reduction in HCV-related mortality and an 80% reduction in HCV incidence. Globally, an estimated 71 million people were living with chronic HCV infection in 2015. Approximately two million new HCV infections occur annually, with injecting drug use and unsafe health-care practices (including unsterile health-care injection), the predominant modes of HCV transmission.
The development and availability of highly effective direct-acting antiviral agents (DAAs) have revolutionised HCV management and provide the therapeutic tools required to strive for elimination. For HCV treatment as prevention to have greatest impact, HCV testing and treatment coverage must be high, with new diagnoses linked expediently to care and treatment. In 2015, of the 71 million people living with HCV infection, only 20% (14 million) were diagnosed, and of those diagnosed, 13% (1.1 million in 2015, 1.76 million in 2016) had initiated DAA treatment.
This chapter outlines the United Nations and World Health Organization elimination targets; defines the epidemiological concepts of control, elimination and eradication; and discusses the important lessons learnt from control and elimination efforts in other infectious diseases epidemics. The biological and technical feasibility of HCV control and elimination is discussed, followed by related financial, political and social considerations. Examples of national HCV strategies are presented, highlighting the facilitators and barriers to successful implementation of HCV elimination strategies. Control and elimination of HCV infection will require an enormous public health, political and economic commitment. The costs and risks may be high, but so too are the potential benefits.
Marianne Martinello, Behzad Hajarizadeh, Jason Grebely, Gail V. Matthews, Gregory J. Dore

Perspectives on HCV Cure

The development of interferon-free cures for hepatitis C has revolutionized the treatment of patients chronically infected with the hepatitis C virus. Since 2010, ten new curative regimens have been introduced into clinical practice. These new regimens have delivered cure rates in excess of 95% in as little as 8–10 weeks on therapy. Never before has there been an absolute cure for a chronic viral disease. This medical breakthrough has been made possible by the commitment of scientists and clinicians from both academia and industry working toward a common goal. Because of the availability of these curative regimens, it is now possible to contemplate eliminating HCV as a global public health problem as outlined by the World Health Organization. This perspective will give a brief commentary of the achievements and future possibilities provided by direct-acting antiviral interferon-free HCV cure therapies.
Michael J. Sofia


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