More than 170 million people worldwide are affected by the hepatitis C virus (HCV). The disease has been described as a "silent epidemic" and "a serious global health crisis". HCV infection is a leading cause of chronic liver disease such as cirrhosis, carcinoma, or liver failure. The current pegylated interferon and ribavirin combination therapy is effective in only 50% of patients. Its moderate efficacy and apparent side effects underscore the need for safer and more effective treatments. The nonstructural NS3 protease of the virus plays a vital role in the replication of the HCV virus. The development of small molecule inhibitors of NS3 protease as antiviral agents has been intensively pursued as a viable strategy to eradicate HCV infection. However, it is a daunting task. The protease has a shallow and solvent-exposed substrate binding region, and the inhibitor binding energy is mainly derived from weak lipophilic and electrostatic interactions. Moreover, lack of a robust in vitro cell culture system and the absence of a convenient small animal model have hampered the assessment of both in vitro and in vivo efficacy of any antiviral compounds. Despite the tremendous challenges, with access to a recently developed cell-based replicon system, major progress has been made toward a more effective small molecule HCV drug. In our HCV program, facing no leads from our screening effort, a structure-based drug design approach was carried out. An alpha-ketoamide-type electrofile was designed to trap the serine hydroxyl of the protease. Early ketoamide inhibitors mimicked the structures of the peptide substrates. With the aid of X-ray structures, we successfully truncated the undecapeptide lead that had a molecular weight of 1265 Da stepwise to a tripeptide with a molecular weight of 500 Da. In an attempt to depeptidize the inhibitors, various strategies such as hydrazine urea replacement of amide bonds and P2 to P4 and P1 to P3 macrocyclizations were examined. Further optimization of the tripeptide inhibitors led to the identification of the best moieties for each site: primary ketoamide at P', cyclobutylalanine at P1, gem-dimethylcyclopropylproline at P2, tert-leucine at P3, and tert-butyl urea as capping agent. The combination of these led to the discovery of compound 8 (SCH 503034, boceprevir), our clinical candidate. It is a potent inhibitor in both enzyme assay (Ki* = 14 nM) and cell-based replicon assay (EC 90 = 0.35 microM). It is highly selective (2200x) against human neutrophil elastase (HNE). Boceprevir is well tolerated in humans and demonstrated antiviral activity in phase I clinical trials. It is currently in phase II trials. This Account details the complexity and challenges encountered in the drug discovery process.

Boceprevir (SCH 503034), 1, a novel HCV NS3 serine protease inhibitor discovered in our laboratories, is currently undergoing phase III clinical trials. Detailed investigations toward a second generation protease inhibitor culminated in the discovery of narlaprevir (SCH 900518), 37, with improved potency (∼10-fold over 1), pharmacokinetic profile and physicochemical characteristics, currently in phase II human trials. Exploration of synthetic sequence for preparation of 37 resulted in a route that required no silica gel purification for the entire synthesis.KEYWORDS Hepatitis C virus NS3 serine protease inhibitor, R-ketoamide, narlaprevir, SCH 900518 H epatitis C virus (HCV) infection is a global health crisis leading to liver cirrhosis, hepatocellular carcinoma and liver failure in humans. 1 An estimated 3% of the human population worldwide is chronically infected with HCV. 2 Currently the only available treatment regimens are subcutaneous R-interferon or long-acting pegylated-interferon, alone or in combination with oral antiviral agent ribavirin. 3 The approved therapy is still far from ideal for the hard to treat genotype-1 patients 4 and is frequently accompanied by adverse side effects. There is an unmet medical need to discover new, more effective and tolerable regimens for the treatment of HCV infection. Advances in the understanding of molecular pathways of HCV replication have resulted in several small molecule direct-acting antivirals entering clinical trials in the past few years.Since identification of this virus, the NS3 serine protease contained within the N-terminal region of the NS3 protein has been studied extensively. 5 This chymotrypsin-like serine protease plays a pivotal role in viral replication and, therefore, is an attractive target for HCV antiviral therapeutics. 6,7 Intense efforts were focused in the past decade to discover novel small molecule agents that inhibit NS3 serine protease. 8 Proof of concept studies in humans with BILN 2061, a noncovalent P1-P3 macrocyclic inhibitor, validated this hypothesis. 9 Since then, several NS3 protease inhibitors have progressed to human clinical trials. Currently the most advanced among those are boceprevir (SCH 503034), 1, 10,11 and telaprevir (VX950), 12,13 from the slow-binding reversible R-ketoamide class, in phase III human evaluation. Inhibitors in phase II studies, from the structurally distinct noncovalent macrocyclic class, include 14 15 and MK-7009 (P2-P4 macrocycle). 16 Other NS3 protease inhibitors currently in clinical evaluation (structure not yet disclosed) include BI-201335, ABT-450, PHX-1766, ACH-1625 and VX-813. 8 Inhibitor 1 exhibited K i * = 14 nM in the enzyme binding assay, 17 EC 90 = 350 nM in the cell-based replicon assay, 18 and acceptable pharmacokinetic profile in rats and dogs (Figure 1). In our efforts to discover a second generation HCV protease inhibitor, we focused mainly on improving the in vitro potency and preclinical pharmacokinetic profile of the inhibitor, specifically exposure in monkeys. Further...

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