3-(1,1-Dioxo-2<i>H</i>-(1,2,4)-benzothiadiazin-3-yl)-4-hydroxy-2(1<i>H</i>)-quinolinones, Potent Inhibitors of Hepatitis C Virus RNA-Dependent RNA Polymerase

Tedesco, Rosanna; Shaw, Antony N.; Bambal, Ramesh; Chai, Deping; Concha, N.O.; Darcy, Michael G.; Dhanak, Dashyant; Fitch, Duke M.; Gates, Adam; GERHARDt, W.; Halegoua, Dina L.; Han, Chao; Hofmann, Glenn A.; Johnston, Victor K.; Kaura, Arun C.; Liu, Nannan; Keenan, Richard M.; Lin-Goerke, Juili; Sarisky, Robert T.; Wiggall, Kenneth J.; Zimmerman, Michael N.; Duffy, Kevin J.

doi:10.1021/jm050855s

Cited by 152 publications

(101 citation statements)

References 23 publications

(76 reference statements)

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“…Selection of replicons dually resistant to combination treatment with thumb and palm site inhibitors. We have solved the cocrystal structure of the HCV NS5B polymerase bound to NNI-2 ( Table 4) and show that the compound binds to HCV polymerase in the palm domain at a site located at the base of the relatively large internal polymerase cavity, in agreement with a recent report (38). The bound conformation of NNI-2 shows an approximately 22°bend between the thiadiazine and quinolinedione ring systems.…”

Section: Resultssupporting

confidence: 87%

“…Several chemical classes of nonnucleoside inhibitors that inhibit the isolated enzyme and replication in the replicon system have been shown to bind at distinct sites on HCV polymerase. These polymerase inhibitors include benzothiadiazines, binding to the palm domain near the active site (38,40), thiophene carboxylic acids which bind at the outer surface of the thumb domain (thumb I site), and benzimidazoles and indoles which bind to the thumb domain near the fingertips (thumb II site) (12,20,39).…”

Section: Multiple Nonnucleoside Inhibitor Binding Sites Have Been Idementioning

confidence: 99%

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Selection and Characterization of Replicon Variants Dually Resistant to Thumb- and Palm-Binding Nonnucleoside Polymerase Inhibitors of the Hepatitis C Virus

Pogam

Kang

Harris

et al. 2006

J Virol

129

117

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Multiple nonnucleoside inhibitor binding sites have been identified within the hepatitis C virus (HCV)polymerase, including in the palm and thumb domains. After a single treatment with a thumb site inhibitor (thiophene-2-carboxylic acid NNI-1), resistant HCV replicon variants emerged that contained mutations at residues Leu419, Met423, and Ile482 in the polymerase thumb domain. Binding studies using wild-type (WT) and mutant enzymes and structure-based modeling showed that the mechanism of resistance is through the reduced binding of the inhibitor to the mutant enzymes. Combined treatment with a thumb-and a palmbinding polymerase inhibitor had a dramatic impact on the number of replicon colonies able to replicate in the presence of both inhibitors. A more exact characterization through molecular cloning showed that 97.7% of replicons contained amino acid substitutions that conferred resistance to either of the inhibitors. Of those, 65% contained simultaneously multiple amino acid substitutions that conferred resistance to both inhibitors. Double-mutant replicons Met414Leu and Met423Thr were predominantly selected, which showed reduced replication capacity compared to the WT replicon. These findings demonstrate the selection of replicon variants dually resistant to two NS5B polymerase inhibitors binding to different sites of the enzyme. Additionally, these findings provide initial insights into the in vitro mutational threshold of the HCV NS5B polymerase and the potential impact of viral fitness on the selection of multiple-resistant mutants.Hepatitis C virus (HCV), a positive-strand RNA virus, is a member of the genus Hepacivirus in the Flaviviridae family and is the leading cause of liver disease worldwide. It is estimated that over 170 million individuals are infected with HCV (43). The current standard of care provides good clinical efficacy for patients infected with genotype 2 and 3 but is less efficacious for patients infected with the most prevalent genotype, genotype 1, thereby emphasizing the urgent need for more effective HCV-specific antiviral therapies (15,27).The HCV RNA-dependent RNA polymerase is an essential enzyme for viral RNA replication and represents an attractive therapeutic target. HCV polymerase has the "right-hand" polymerase fold with finger, thumb, and palm domains (22). As with other RNA-dependent RNA polymerases, the extended "fingertips" contact a thicker thumb domain to create an encircled active site constituting the closed, active conformation of the enzyme (7,16,22,32). With the advent of the HCV replicon system there have been extensive developments supporting the discovery of new HCV polymerase nonnucleoside inhibitors (1-3, 5, 6, 11, 36). Several chemical classes of nonnucleoside inhibitors that inhibit the isolated enzyme and replication in the replicon system have been shown to bind at distinct sites on HCV polymerase. These polymerase inhibitors include benzothiadiazines, binding to the palm domain near the active site (38, 40), thiophene carboxylic acids which bind at the...

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Section: Resultssupporting

confidence: 87%

Section: Multiple Nonnucleoside Inhibitor Binding Sites Have Been Idementioning

confidence: 99%

Selection and Characterization of Replicon Variants Dually Resistant to Thumb- and Palm-Binding Nonnucleoside Polymerase Inhibitors of the Hepatitis C Virus

Pogam

Kang

Harris

et al. 2006

J Virol

129

117

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“…The NNI-3 site is located adjacent to the active site. Reported NNI-3 ligands include benzothiadiazine (11,47), proline sulfonamide (18), benzylidene (24,42), and acrylic acid (40,41) derivatives. In drug discovery, knowledge of the inhibitor site of action is crucial to guiding medicinal chemistry efforts.…”

mentioning

confidence: 99%

“…Structural activity relationships are further complicated by the variation observed for each of the NNI binding sites between genotype and subtypes. These issues can be addressed using X-ray crystallography, as demonstrated by others (2,13,18,24,29,40,41,42,47,55). However, this is a considerable undertaking and requires both time and abundant purified enzyme.…”

mentioning

confidence: 99%

Binding-Site Identification and Genotypic Profiling of Hepatitis C Virus Polymerase Inhibitors

Pauwels¹,

Mostmans²,

Quirynen³

et al. 2007

J Virol

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The search for hepatitis C virus polymerase inhibitors has resulted in the identification of several nonnucleoside binding pockets. The shape and nature of these binding sites differ across and even within diverse hepatitis C virus genotypes. These differences confront antiviral drug discovery with the challenge of finding compounds that are capable of inhibition in variable binding pockets. To address this, we have established a hepatitis C virus mutant and genotypic recombinant polymerase panel as a means of guiding medicinal chemistry through the elucidation of the site of action of novel inhibitors and profiling against genotypes. Using a genotype 1b backbone, we demonstrate that the recombinant P495L, M423T, M414T, and S282T mutant enzymes can be used to identify the binding site of an acyl pyrrolidine analog. We assess the inhibitory activity of this analog and other nonnucleoside inhibitors with our panel of enzyme isolates generated from clinical sera representing genotypes 1a, 1b, 2a, 2b, 3a, 4a, 5a, and 6a.Hepatitis C is estimated to affect 3% of the global population. In a number of individuals, it can lead to liver fibrosis, cirrhosis, and death. Although virus can be cleared by a combination of pegylated interferon and ribavirin, the treatment is successful in only around 50% of treated patients and has considerable liabilities. These weaknesses highlight the need for new drugs to treat hepatitis C virus (HCV) in patients who have failed current therapy, as well as in untreated patients (12,56).HCV is an enveloped virus with an RNA genome of ϳ9.6 kb. Its single-stranded RNA has a positive polarity and encodes a polyprotein of ϳ3,300 amino acids comprising 4 structural proteins (Core, E1, E2, and p7) and 6 nonstructural proteins (NS2, -3, -4A, -4B, -5A, and -5B) (43). These proteins, as well as the viral translation process using the internal ribosomal entry site and a range of host factors, are candidate targets for therapeutic intervention (3, 46). The remarkable clinical success of human immunodeficiency virus reverse transcriptase and protease inhibitors, as well as the availability of several crystal structures, has motivated HCV drug discovery efforts to focus mainly on the development of protease and polymerase inhibitors. HCV NS5B is an RNA-dependent RNA polymerase that is responsible for the replication of the viral genome, which is thought to occur through a primer-independent de novo mechanism (6, 31). Due to the lack of proofreading capacity, this replication process is subject to a high error rate (36). As a result, the virus has evolved into multiple variant strains, classified into six different genotypes (1 to 6) and several subtypes (a, b, c, etc.) (45). To add to this complexity, HCV-infected individuals also harbor different variants or quasispecies of the virus, together representing a pool of genomes on which selective pressure can act (16). It has been speculated that drug resistance will rapidly emerge upon administration of specific HCV antivirals and that together with viral ...

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“…Among the hydrophobic groups present in the molecule, Quinoline ring is positively correlated with activity. Substitution of fluorine at C6 position of the quinolone ring has been shown to improve the cellular activity [20]. In the less active compound, nitrogen atoms of 5-hydroxy pyridazine ring (hydrogen bond donor) and benzyl moiety of benzothiadiazine ring (hydrophobic group) are negatively correlated with activity ( fig.…”

Section: Fig 2: Pharmacophore Site Distances (A) Pharmacophore Site mentioning

confidence: 99%

Combinatorial Pharmacophore Modeling and Atom Based 3d Qsar Studies of Benzothiadiazines as HCV-Ns5b Inhibitors

Polamreddy

Vishwakarma

Mahto

2018

Int J Pharm Pharm Sci

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Objective: The objective of the current study was to elucidate the 3D pharmacophoric features of benzothiadiazine derivatives that are crucial for inhibiting Hepatitis C virus (HCV) Non-structural protein 5B (NS5B) and quantifying the features by building an atom based 3D quantitative structure-activity relationship (3D QSAR) model. Methods:Generation of QSAR model was carried out using PHASE 3.3. Results:A five-point pharmacophore model with two hydrogen bond acceptors, one negative ionization potential and two aromatic rings (AANRR) was found to be common among a maximum number of benzothiadiazine based NS5B inhibitors. A statistically significant 3D QSAR model was obtained from AANRR.6 which had correlation-coefficient (R 2 ) value of 0.924, cross-validated correlation-coefficient (Q 2 ) of 0.774, high Fisher ratio of 138 and low root mean square standard error (RMSE=0.29). There is another parameter, Pearson's R, its value emphasizes correlation between predicted and observed activities of the test set. For the current model, Pearson's R-value is 0.90, hence underlining the good quality of the model. The present study suggests that nitrogen atom of benzothiadiazine sulfamide ring, oxyacetamide group attached to C7 carbon of benzothiadiazine and sulfonamide oxygens are crucial for NS5B inhibitory activity. Prediction of activities of hit drugs generated in earlier research suggests that Aprepitant (Phase predicted activity: 6.9) could be a potential NS5B inhibitor.Conclusion: This 3D QSAR model developed was statistically good and can be used to predict the activities of newly designed NS5B inhibitors and virtual screening as well. Predict the activities of newly designed NS5B inhibitors and virtual screening as well.

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3-(1,1-Dioxo-2H-(1,2,4)-benzothiadiazin-3-yl)-4-hydroxy-2(1H)-quinolinones, Potent Inhibitors of Hepatitis C Virus RNA-Dependent RNA Polymerase

Cited by 152 publications

References 23 publications

Selection and Characterization of Replicon Variants Dually Resistant to Thumb- and Palm-Binding Nonnucleoside Polymerase Inhibitors of the Hepatitis C Virus

Selection and Characterization of Replicon Variants Dually Resistant to Thumb- and Palm-Binding Nonnucleoside Polymerase Inhibitors of the Hepatitis C Virus

Binding-Site Identification and Genotypic Profiling of Hepatitis C Virus Polymerase Inhibitors

Combinatorial Pharmacophore Modeling and Atom Based 3d Qsar Studies of Benzothiadiazines as HCV-Ns5b Inhibitors

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