Article pubs.acs.org/jmc
Discovery of Ruzasvir (MK-8408): A Potent, Pan-Genotype HCV NS5A Inhibitor with Optimized Activity against Common ResistanceAssociated Polymorphisms Ling Tong,*,† Wensheng Yu,† Lei Chen,† Oleg Selyutin,† Michael P. Dwyer,† Anilkumar G. Nair,† Robert Mazzola,† Jae-Hun Kim,† Deyou Sha,† Jingjun Yin,⊥ Rebecca T. Ruck,⊥ Ian W. Davies,†,⊥ Bin Hu,∇ Bin Zhong,∇ Jinglai Hao,∇ Tao Ji,∇ Shuai Zan,∇ Rong Liu,‡ Sony Agrawal,‡ Ellen Xia,‡ Stephanie Curry,‡ Patricia McMonagle,‡ Karin Bystol,‡ Frederick Lahser,‡ Donna Carr,‡ Laura Rokosz,‡ Paul Ingravallo,‡ Shiying Chen,§ Kung-I Feng,# Mark Cartwright,∥ Ernest Asante-Appiah,‡ and Joseph A. Kozlowski† †
Department of Medicinal Chemistry, ‡Department of Discovery Biology, §Department of Pharmacokinetics, and ∥Department of Drug Safety, Merck & Co., Inc., 2000 Galloping Hill Road, Kenilworth, New Jersey 07033, United States ⊥ Department of Process and Analytical Chemistry, and #Department of Discovery Pharmaceutic Science, Merck & Co., Inc., 126 E. Lincoln Avenue, Rahway, New Jersey 07065, United States ∇ Department of Medicinal Chemistry, WuXi AppTec, Shanghai, 200131, China S Supporting Information *
ABSTRACT: We describe the research that led to the discovery of compound 40 (ruzasvir, MK-8408), a pan-genotypic HCV nonstructural protein 5A (NS5A) inhibitor with a “flat” GT1 mutant profile. This NS5A inhibitor contains a unique tetracyclic indole core while maintaining the imidazole− proline−valine Moc motifs of our previous NS5A inhibitors. Compound 40 is currently in early clinical trials and is under evaluation as part of an all-oral DAA regimen for the treatment of chronic HCV infection.
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INTRODUCTION Hepatitis C virus (HCV) infection has become the most common blood-born infection in developed countries.1 Recent estimates of disease burden show an increase in seroprevalence over the period of 15 years (between 1990 and 2005) to ∼2.8% of the global population, more than 185 million infections worldwide.2 HCV infection is initially asymptomatic; as the duration of chronic infection increases, symptoms become manifested as increased liver disease including fibrosis, cirrhosis (compensated and decompensated), and hepatocellular carcinoma.3 Advances in treatment regimens have resulted in significantly improved outcomes for many patients. However, not every patient becomes cured; hence, there remains a medical need in the treatment of these patient populations.3,4 The challenge in treating HCV originates from the diversity present in the virus where it is classified into seven major genotypes, with more than 50 subtypes known.5 Early treatment options such as interferon and ribavirin were poorly tolerated and showed limited efficacy due to the emergence of resistanceassociated variants (RAVs). Over time, a consensus emerged that a better tolerated, more efficacious treatment was needed © XXXX American Chemical Society
and that the diversity of HCV infection might best be addressed through administration of a combination of several direct-acting antiviral agents (DAAs), each targeting a different step in the replication cycle of the virus.3 This approach provides a chance to overcome poor treatment outcomes by allowing the combination of multiple mechanisms to suppress the emergence of RAVs escaping from any single mechanism. This approach has become the current treatment paradigm and has led to the improvement in patient outcomes. From this strategy, three major categories of drugs have been developed including NS3/4A protease inhibitors, NS5A replication complex inhibitors, and NS5B polymerase nucleotide and non-nucleotide inhibitors. The ideal profile for each component of the DAA regimen includes high potency, high barrier to resistance, pan-genotype activity, few drug−drug interactions (DDIs), minimal toxicity, and a pharmacokinetic profile that allows once-daily oral dosing.3 An area of high interest for Merck & Co., Inc. has been the identification and development of potent NS5A inhibitors to be Received: August 31, 2016 Published: November 3, 2016 A
DOI: 10.1021/acs.jmedchem.6b01310 J. Med. Chem. XXXX, XXX, XXX−XXX
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used as part of an all-oral DAA regimen.6−11 The exact mechanism of NS5A inhibition remains unclear and is poorly understood; however, NS5A plays a critical role in viral ribonucleic acid (RNA) synthesis and virion assembly.12 As a therapeutic target, NS5A has been validated clinically, where multiple inhibitors were shown to be well tolerated and to produce rapid and robust viral load declines in patients.13−15 Today NS5A inhibitors have become an integral part of DAA regimens with five NS5A inhibitors currently used in clinical practice for HCV treatment including daclatasvir (BMS-790052),13 ledipasvir (GS-5885),16 ombitasvir (ABT-267),17 elbasvir (MK-8742, 1),6 and velpatasvir (GS-5816).18
However, we also observed that molecules with improved inhibition of GT1a_Y93H often resulted in similar or better inhibition of other GT1 variants. We hypothesized that gaining the ability to better inhibit this RAV might provide a path toward the flat profile we desired, and therefore GT1a_Y93H was selected as a part of the screening set for SAR optimization. Concurrent with the selection of GT1a_Y93H, GT1a_L31V was also picked for optimization, as it offered diversity in a mutation at a different location in the protein. GT2b(31M) was also added to the screening set, as it represented a clinically important non-genotype 1 example. Inclusion of GT3a and GT4a offered verification of performance against additional genotypes. Acknowledging the risk associated with defining SAR with a focus on GT1a_Y93H, GT1a_L31V, and 2b, where the SAR may not translate to other genotypes and RAVs, we proceeded with the strategy and decided to validate it by periodically checking the full virologic profile of key compounds. The discussion herein is restricted to replicon EC90 values unless otherwise noted. The stereochemistry at the aminal position for compound 40 was determined as (S), which was more potent than the corresponding (R) configuration. Compounds reported in this paper were separated and assayed as single enantiomers with the potency for the most active isomer reported, unless otherwise noted. For the purposes of this discussion, we arbitrarily designated the aminal-phenyl group as the “Z-group”, Table 1. From our earlier results we observed sensitivity of the mutant profiles
In a continued effort to develop NS5A inhibitors, we asked the question: Is it possible to identify an inhibitor with potent activity against all genotypes and a minimal potency shift from the wild type virus to previously identified RAVs, selected by existing NS5A inhibitors, particularly in the most prevalent GT1a? We use the term “flat compound” to describe such a pan-genotype inhibitor with a minimal potency loss (∼10-fold) between the wild-type virus and clinically relevant polymorphisms. In addition, the desired flat compound should also maintain sufficiently good pharmacokinetic parameters to support once-daily dosing and suitable pharmaceutical properties in order to enable a fixed-dose combination (FDC) with partner DAAs. We now describe the structural modifications to the core structure of 1 that led to the discovery of MK-8408 (40, Figure 1),19 also known as ruzasvir, a potent NS5A inhibitor
Table 1. Potency and Shift Comparison of 1−5
Figure 1. Compound 40.
with a flat in vitro profile. Compound 40 is currently undergoing clinical testing as part of an all-oral, interferon-free DAA regimen for the treatment of chronic HCV infection.
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RESULTS AND DISCUSSION From our experience in the development of NS5A inhibitors, we observed that the GT 1a RAV 1aY93H consistently conferred a high level of resistance across a range of different molecules.6−11
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Fold-shift is relative to the WT activity, set = 1. DOI: 10.1021/acs.jmedchem.6b01310 J. Med. Chem. XXXX, XXX, XXX−XXX
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toward changes at the Z-group, as illustrated by compound 2, Table 1.7,8 Numbers in parentheses were potency shift of mutants relative to GT1a, which was used as a measurement of flatness of the profile. We also observed that improvement in the mutant profiles was achievable through addition of a fluorine atom at R1 shown by comparison of compound 39 versus compound 4.9 The combination of a chromane Z group and a fluorine atom at R1 could give a modest reduction in the shift in potency against key mutants to
