Novel substituted pyrimidines as HCV replication (replicase) inhibitors.

Compound 1 was identified as a HCV replication inhibitor from screening/early SAR triage. Potency improvement was achieved via modulation of substituent on the 5-azo linkage. Due to potential toxicological concern, the 5-azo linkage was replaced with 5-alkenyl or 5-alkynyl moiety. Analogs containing the 5-alkynyl linkage were found to be potent inhibitors of HCV replication. Further evaluation identified compounds 53 and 63 with good overall profile, in terms of replicon potency, selectivity and in vivo characteristics. Initial target engagement studies suggest that these novel carbanucleoside-like derivatives may inhibit the HCV replication complex (replicase).

Pyridofuran substituted pyrimidine derivatives as HCV replication (replicase) inhibitors.

Introduction of nitrogen atom into the benzene ring of a previously identified HCV replication (replicase) benzofuran inhibitor 2, resulted in the discovery of the more potent pyridofuran analogue 5. Subsequent introduction of small alkyl and alkoxy ligands into the pyridine ring resulted in further improvements in replicon potency. Replacement of the 4-chloro moiety on the pyrimidine core with a methyl group, and concomitant monoalkylation of the C-2 amino moiety resulted in the identification of several inhibitors with desirable characteristics. Inhibitor 41, from the monosubstituted pyridofuran and inhibitor 50 from the disubstituted series displayed excellent potency, selectivity (GAPDH/MTS CC(50)) and PK parameters in all species studied, while the selectivity in the thymidine incorporation assay (DNA·CC(50)) was low.

Synthesis and SAR of geminal substitutions at the C5' carbosugar position of pyrimidine-derived HCV inhibitors.

The installation of geminal substitution at the C5' position of the carbosugar in our pyrimidine-derived hepatitis C inhibitor series is reported. SAR studies around the C5' position led to the installation of the dimethyl group as the optimal functionality. An improved route was subsequently designed to access these substitutions. Expanded SAR at the C2 amino position led to the utilization of C2 ethers. These compounds exhibited good potency, high selectivity, and excellent plasma exposure and bioavailability in rodent as well as in higher species.

Synthesis and SAR of pyridothiazole substituted pyrimidine derived HCV replication inhibitors.

Introduction of a nitrogen atom into the benzene ring of a previously identified HCV replication (replicase) benzothiazole inhibitor 1, resulted in the discovery of the more potent pyridothiazole analogues 3. The potency and PK properties of the compounds were attenuated by the introductions of various functionalities at the R(1), R(2) or R(3) positions of the molecule (compound 3). Inhibitors 38 and 44 displayed excellent potency, selectivity (GAPDH/MTS CC(50)), PK parameters in all species studied, and cross genotype activity.

5-Benzothiazole substituted pyrimidine derivatives as HCV replication (replicase) inhibitors.

Based on a previously identified HCV replication (replicase) inhibitor 1, SAR efforts were conducted around the pyrimidine core to improve the potency and pharmacokinetic profile of the inhibitors. A benzothiazole moiety was found to be the optimal substituent at the pyrimidine 5-position. Due to potential reactivity concern, the 4-chloro residue was replaced by a methyl group with some loss in potency and enhanced rat in vivo profile. Extensive investigations at the C-2 position resulted in identification of compound 16 that demonstrated very good replicon potency, selectivity and rodent plasma/target organ concentration. Inhibitor 16 also demonstrated good plasma levels and oral bioavailability in dogs, while monkey exposure was rather low. Chemistry optimization towards a practical route to install the benzothiazole moiety resulted in an efficient direct C-H arylation protocol.

Building a Culture of Medicinal Chemistry Knowledge Sharing.

Increasing the efficiency of the drug discovery process is a challenge faced by drug hunters everywhere. One strategy medicinal chemists employ to meet this challenge is learning from knowledge sources within and beyond their organization. In this Perspective, we discuss the evolution of mechanisms for medicinal chemistry knowledge capture and sharing at Merck & Co. over the past 15 years. We describe our approach to knowledge management and report on the multiple enduring and complementary teams and initiatives we have created to capture and share knowledge within a geographically diverse medicinal chemistry community. In addition, this Perspective will share the benefits we have observed and also reflect on what has allowed our efforts to be both successful and sustainable.

The introduction of P4 substituted 1-methylcyclohexyl groups into Boceprevir: a change in direction in the search for a second generation HCV NS3 protease inhibitor.

In the search for a second generation HCV protease inhibitor, molecular modeling studies of the X-ray crystal structure of Boceprevir1 bound to the NS3 protein suggest that expansion into the S4 pocket could provide additional hydrophobic Van der Waals interactions. Effective replacement of the P4 tert-butyl with a cyclohexylmethyl ligand led to inhibitor 2 with improved enzyme and replicon activities. Subsequent modeling and SAR studies led to the pyridine 38 and sulfone analogues 52 and 53 with vastly improved PK parameters in monkeys, forming a new foundation for further exploration.

Discovery of potent sulfonamide P4-capped ketoamide second generation inhibitors of hepatitis C virus NS3 serine protease with favorable pharmacokinetic profiles in preclinical species.

Hepatitis is a disease characterized by inflammation of the liver, usually producing swelling and, in many cases, permanent damage to liver tissues. Viral hepatitis C (HCV), a small (+)-RNA virus, infects chronically 3% of the world's population. Boceprevir, SCH 503034, (1) our first generation HCV inhibitor, has already established proof-of- concept and is currently in late stage (phase III) clinical trials. In view of the positive data from our first generation compound, further work aimed at optimizing its overall profile was undertaken. Herein, we report that extension of our earlier inhibitor to the P(4) pocket by introducing a new sulfonamide moiety and optimization of the P1/P(1)' capping led to the discovery of a novel series of inhibitors of the HCV NS3 serine protease. Optimization of the P(1) residue significantly improved potency and selectivity. The combination of optimal moieties led to the discovery of compound 47 which, in addition to being a potent inhibitor of HCV subgenomic RNA replication, was also found to have good PK profile in rat, dog and monkey.

Discovery of novel P3 sulfonamide-capped inhibitors of HCV NS3 protease. Inhibitors with improved cellular potencies.

Hepatitis C Virus (HCV) infection is the major cause of chronic liver disease, leading to cirrhosis and hepatocellular carcinoma, which affects more than 200 million people worldwide. Currently the only therapeutic regimens are subcutaneous interferon-alpha or PEG-interferon alone or in combination with oral ribavirin. Although combination therapy is reasonably successful with the majority of genotypes, its efficacy against the predominant genotype (genotype 1) is moderate at best, with only approximately 50% of the patients showing sustained virological response. We recently disclosed the discovery of Boceprevir, SCH 503034 (1), which is a novel, potent, selective, orally bioavailable NS3 protease inhibitor that has been shown to be efficacious in humans and is currently undergoing clinical trials. As second generation compounds, we have further explored various novel structures with the aim of improving enzyme and cellular binding activities of 1. Herein, we disclose our efforts toward the identification of a novel P(3) sulfonamide-capped inhibitor that demonstrated improved binding and cellular activity compared to 1. X-ray structure of one of these inhibitors bound to the enzyme revealed a hydrogen bond of the P(3) sulfonamide group to Cys-159 which resulted in improved binding and cellular potency.

Discovery of the HCV NS3/4A protease inhibitor (1R,5S)-N-[3-amino-1-(cyclobutylmethyl)-2,3-dioxopropyl]-3- [2(S)-[[[(1,1-dimethylethyl)amino]carbonyl]amino]-3,3-dimethyl-1-oxobutyl]- 6,6-dimethyl-3-azabicyclo[3.1.0]hexan-2(S)-carboxamide (Sch 503034) II. Key steps in structure-based optimization.

The structures of both the native holo-HCV NS3/4A protease domain and the protease domain with a serine 139 to alanine (S139A) mutation were solved to high resolution. Subsequently, structures were determined for a series of ketoamide inhibitors in complex with the protease. The changes in the inhibitor potency were correlated with changes in the buried surface area upon binding the inhibitor to the active site. The largest contribution to the binding energy arises from the hydrophobic interactions of the P1 and P2 groups as they bind to the S1 and S2 pockets [the numbering of the subsites is as defined in Berger, A.; Schechter, I. Philos. Trans. R. Soc. London, Ser. B 1970, 257, 249-264]. This correlation of the changes in potency with increased buried surface area contributed directly to the design of a potent tripeptide inhibitor of the HCV NS3/4A protease that is currently in clinical trials.

Novel potent hepatitis C virus NS3 serine protease inhibitors derived from proline-based macrocycles.

The hepatitis C virus (HCV) NS3 protease is essential for viral replication. It has been a target of choice for intensive drug discovery research. On the basis of an active pentapeptide inhibitor, 1, we envisioned that macrocyclization from the P2 proline to P3 capping could enhance binding to the backbone Ala156 residue and the S4 pocket. Thus, a number of P2 proline-based macrocyclic alpha-ketoamide inhibitors were prepared and investigated in an HCV NS3 serine protease continuous assay (K(i*)). The biological activity varied substantially depending on factors such as the ring size, number of amino acid residues, number of methyl substituents, type of heteroatom in the linker, P3 residue, and configuration at the proline C-4 center. The pentapeptide inhibitors were very potent, with the C-terminal acids and amides being the most active ones (24, K(i*) = 8 nM). The tetrapeptides and tripeptides were less potent. Sixteen- and seventeen-membered macrocyclic compounds were equally potent, while fifteen-membered analogues were slightly less active. gem-Dimethyl substituents at the linker improved the potency of all inhibitors (the best compound was 45, K(i*) = 6 nM). The combination of tert-leucine at P3 and dimethyl substituents at the linker in compound 47 realized a selectivity of 307 against human neutrophil elastase. Compound 45 had an IC(50) of 130 nM in a cellular replicon assay, while IC(50) for 24 was 400 nM. Several compounds had excellent subcutaneous AUC and bioavailability in rats. Although tripeptide compound 40 was 97% orally bioavailable, larger pentapeptides generally had low oral bioavailability. The X-ray crystal structure of compounds 24 and 45 bound to the protease demonstrated the close interaction of the macrocycle with the Ala156 methyl group and S4 pocket. The strategy of macrocyclization has been proved to be successful in improving potency (>20-fold greater than that of 1) and in structural depeptization.

Discovery of SCH446211 (SCH6): a new ketoamide inhibitor of the HCV NS3 serine protease and HCV subgenomic RNA replication.

Introduction of various modified prolines at P(2) and optimization of the P(1) side chain led to the discovery of SCH6 (24, Table 2), a potent ketoamide inhibitor of the HCV NS3 serine protease. In addition to excellent enzyme potency (K(i)*= 3.8 nM), 24 was also found to be a potent inhibitor of HCV subgenomic RNA replication with IC(50) and IC(90) of 40 and 100 nM, respectively. Recently, antiviral activity of 24 was demonstrated with inhibition of the full-length genotype 2a HCV genome. In addition, 24 was found to restore the responsiveness of the interferon regulatory factor 3 (IRF-3) in cells containing HCV RNA replicons.

Discovery of (1R,5S)-N-[3-amino-1-(cyclobutylmethyl)-2,3-dioxopropyl]- 3-[2(S)-[[[(1,1-dimethylethyl)amino]carbonyl]amino]-3,3-dimethyl-1-oxobutyl]- 6,6-dimethyl-3-azabicyclo[3.1.0]hexan-2(S)-carboxamide (SCH 503034), a selective, potent, orally bioavailable hepatitis C virus NS3 protease inhibitor: a potential therapeutic agent for the treatment of hepatitis C infection.

Hepatitis C virus (HCV) infection is the major cause of chronic liver disease, leading to cirrhosis and hepatocellular carcinoma, which affects more than 170 million people worldwide. Currently the only therapeutic regimens are subcutaneous interferon-alpha or polyethylene glycol (PEG)-interferon-alpha alone or in combination with oral ribavirin. Although combination therapy is reasonably successful with the majority of genotypes, its efficacy against the predominant genotype (genotype 1) is moderate at best, with only about 40% of the patients showing sustained virological response. Herein, the SAR leading to the discovery of 70 (SCH 503034), a novel, potent, selective, orally bioavailable NS3 protease inhibitor that has been advanced to clinical trials in human beings for the treatment of hepatitis C viral infections is described. X-ray structure of inhibitor 70 complexed with the NS3 protease and biological data are also discussed.

Stereoselective synthesis of beta-substituted beta-amino sulfones and sulfonamides via addition of sulfonyl anions to chiral N-sulfinyl imines.

[reaction: see text] A highly stereoselective synthesis of beta-amino sulfones and sulfonamides via addition of sulfonyl anions to chiral N-sulfinyl imines is described. The addition reaction proceeds in good yield (75-99%) and stereoselectivity.

2'-Modified Guanosine Analogs for the Treatment of HCV.

Novel 2'-modified guanosine nucleosides were synthesized from inexpensive starting materials in 7-10 steps via hydroazidation or hydrocyanation reactions of the corresponding 2'-olefin. The antiviral effectiveness of the guanosine nucleosides was evaluated by converting them to the corresponding 5'-O-triphosphates (compounds 38-44) and testing their biochemical inhibitory activity against the wild-type NS5B polymerase.

Non-peptidic small-molecule inhibitors of the single-chain hepatitis C virus NS3 protease/NS4A cofactor complex discovered by structure-based NMR screening.

NMR-based screening of a customized fragment library identified 16 small-molecule hits that bind weakly (K(D) approximately 100 microM to 10 mM) to substrate binding sites of the NS4A-bound NS3 protease of the hepatitis C virus (HCV). Analogues for five classes of NMR hits were evaluated by a combination of NMR and biochemical data yielding SAR and, in most cases, optimized hits with improved potencies (K(D) approximately K(I) approximately 40 microM to 1 mM). NMR chemical shift perturbation data were used to establish the binding location and orientation of the active site directed scaffolds in these five analogue series. Two of these scaffolds, which bind the enzyme at the proximal S1-S3 and S2' substrate binding sites, were linked together producing competitive inhibitors of the HCV NS3 protease with potencies in the micromolar range. This example illustrates that the low molecular weight scaffolds discovered from structure-based NMR screening can be optimized with focused structure-guided chemistry to produce potent nonpeptidic small-molecule inhibitors of the HCV NS3 protease.

Cyclic sulfones as novel P3-caps for hepatitis C virus NS3/4A (HCV NS3/4A) protease inhibitors: synthesis and evaluation of inhibitors with improved potency and pharmacokinetic profiles.

HCV infection affects more than 170 million people worldwide and many of those patients will reach the end stage complications of the disease which include hepatocarcinoma and liver failure. The success rate for treatment of patients infected with genotype-1 is about 40%. Therefore, novel treatments are needed to combat the infection. The HCV NS3 protease inhibitor Boceprevir (1) was reported by our research group and efforts continue for the discovery of more potent compounds with improved pharmacokinetic profiles. A new series of HCV NS3 protease inhibitors having a cyclic sulfone P3-cap have been discovered. Compounds 43 and 44 showed K(i)* values in the single-digit nM range and their cellular potency was improved by 10-fold compared to 1. The pharmacokinetic profiles of 43 and 44 in rats and monkeys were also improved to achieve higher plasma levels after oral administration.

Discovery of Narlaprevir (SCH 900518): A Potent, Second Generation HCV NS3 Serine Protease Inhibitor.

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.

Second-generation highly potent and selective inhibitors of the hepatitis C virus NS3 serine protease.

The hepatitis C virus (HCV) infection is a leading cause of chronic liver disease. The moderate efficacy along with side effects of the current pegylated interferon and ribavirin combination therapy underscores the need for more effective and safer new treatment. In an effort to improve upon our current clinical candidate, Boceprevir (SCH 503034), extensive SAR studies were performed on the P3 capping moieties. This led to the discovery of tert-leucinol derived cyclic imides as a potent series of novel P3 capping groups. Thus, the introduction of these imide caps improved the cell-based replicon EC(90) by more than 10-fold. A number of imides with various substitutions, ring sizes, bicyclic systems, and heterocyclic rings were explored. The 4,4-dimethyl substituted glutarimide emerged as the best cap as exemplified in compound 21 (K(i)* = 4 nM, EC(90) = 40 nM). Systematic optimization of different positions (P', P3, and P1) of the inhibitor resulted in the identification of the lead compound 46, which had an excellent potency (K(i)* = 4 nM, EC(90) = 30 nM) and good pharmacokinetic profile (22% and 35% bioavailability in rats and dogs, respectively). X-ray structure of inhibitor 46 bound to the enzyme revealed that there was an additional hydrogen bonding interaction between one of the imide carbonyls and Cys159.

Toward the back-up of boceprevir (SCH 503034): discovery of new extended P4-capped ketoamide inhibitors of hepatitis C virus NS3 serine protease with improved potency and pharmacokinetic profiles.

Hepatitis C is the most prevalent liver disease. Viral hepatitis C (HCV), a small (+)-RNA virus, infects chronically an estimated 300 million people worldwide. Results of Phase I clinical studies with our first generation HCV inhibitor Boceprevir, SCH 503034 (1), presented at the 56th Annual Meeting of the American Association for the Study of Liver Diseases (AASLD) were encouraging, and thus, additional human clinical studies are underway. In view of the positive data from our first generation compound, further work aimed at optimizing its overall profile was undertaken. Herein, we report that extension of our earlier inhibitor to the P(4) pocket and optimization of the P(1)' capping led to the discovery of new ketoamide inhibitors of the HCV NS3 serine protease with improved in vitro potency. In addition to being potent inhibitors of HCV subgenomic RNA replication, some of the new P(4)-capped inhibitors were also found to have improved PK profile.