In Vivo Emergence of a Novel Mutant L159F/L320F in the NS5B Polymerase Confers Low-Level Resistance to the HCV Polymerase Inhibitors Mericitabine and Sofosbuvir

Tong, Xiao; Pogam, Sophie Le; Li, Lewyn; Haines, Kristin; Piso, Katherine; Baronas, Victoria A.; Yan, Junmei; So, Sung‐Sau; Klumpp, Klaus; Nájera, Isabel

doi:10.1093/infdis/jit562

Cited by 113 publications

(98 citation statements)

References 16 publications

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“…The absence of detectable viremia, even with the very sensitive TMA assay, for a month following infection of 4x0193 with H77S.2 virus (Fig. 1C) is relevant to recent clinical trials of DAAs where some subjects were found to relapse after completing treatment with mericitabine or sofosbuvir, potent nucleosides or nucleotide inhibitors of the NS5B polymerase that effectively suppressed replication (56,57). S282T is a key mutation that confers resistance to these compounds, and its absence following the reappearance of viremia has been interpreted as indicative of the NS5A residues involved in mutations are highlighted in yellow on the surface of domain I of the protein from the genotype 1b Con1 strain of HCV (PDB entry 3FQQ) (left).…”

Section: Discussionmentioning

confidence: 86%

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Evolution of a Cell Culture-Derived Genotype 1a Hepatitis C Virus (H77S.2) during Persistent Infection with Chronic Hepatitis in a Chimpanzee

Joyce

et al. 2014

J Virol

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Persistent infection is a key feature of hepatitis C virus (HCV). IMPORTANCEThis study shows that mutations promoting the production of infectious genotype 1a HCV in cell culture have the opposite effect and attenuate replication in the liver of the only fully permissive animal species other than humans. It provides the only example to date of persistent infection in a chimpanzee challenged with cell culture-produced virus and provides novel insight into the forces shaping molecular evolution of that virus during 5 years of persistent infection. It demonstrates that a poorly fit virus can replicate for weeks within the liver in the absence of detectable viremia, an observation that expands current concepts of HCV pathogenesis and that is relevant to relapses observed with direct-acting antiviral therapies.

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Section: Discussionmentioning

confidence: 86%

“…Residues are labeled and colored as described in the legend to Fig. 8. absence of virologic resistance (56,57). Importantly, because the S282T mutation impacts the active site of the polymerase, S282T viruses are very impaired in replication.…”

Section: Discussionmentioning

confidence: 99%

Evolution of a Cell Culture-Derived Genotype 1a Hepatitis C Virus (H77S.2) during Persistent Infection with Chronic Hepatitis in a Chimpanzee

Joyce

et al. 2014

J Virol

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“…In an analysis of SOF and LDV/SOF studies, the presence of baseline L159F was shown to be associated with virologic failure in a subset of GT1b patients with advanced liver disease treated for a shorter duration with SOF plus RBV before liver transplantation but not for patients receiving LDV/SOF 17. Moreover, the substitution L320F, which is associated with low‐level resistance to mericitabine,18 has been observed in a few patients experiencing virologic failure in SOF studies, but in no patients in LDV/SOF studies. In vitro analyses of L320F and the combination of L320F with L159F showed a low EC 50 fold reduction in SOF susceptibility (1.7‐ to 2.2‐fold for L320F and L320F+L159F, respectively) 17…”

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confidence: 99%

The emergence of NS5B resistance associated substitution S282T after sofosbuvir‐based treatment

Gane

Métivier

Nahass³

et al. 2017

Hepatology Communications

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S282T in NS5B is the primary amino acid substitution associated with resistance to sofosbuvir (SOF) but has rarely been detected in patients treated with a SOF‐based regimen. Here, the emergence and fitness of the S282T substitution in virologic failure patients administered SOF‐based regimens across the SOF and ledipasvir (LDV)/SOF phase 2 and 3 programs was evaluated. Plasma samples collected at baseline and at virologic failure were amplified and deep sequenced (1% cutoff). To date, over 12,000 patients have been treated in SOF or LDV/SOF phase 2 and 3 studies. Of these, deep sequencing was available at baseline in 8598 patients (62.4% genotype [GT] 1, 10.7% GT2, 20.9% GT3, and 6.0% GT4‐6) and at virologic failure in 901 patients. In the 8598 patients, no S282T substitution was detected at baseline; at virologic failure, 10 of the 901 (1%) patients had S282T detected. The SOF‐based regimen associated with treatment‐emergent S282T was SOF monotherapy in two patients, retreatment with LDV/SOF in prior LDV/SOF failures in three patients, LDV/SOF for 8 weeks in 1 GT1 patient, LDV/SOF for 12 weeks in 1 patient each with GT3, GT4, and GT5, and LDV/SOF + ribavirin for 12 weeks in 1 GT6 patient. Nine of 10 patients with emergent S282T received an SOF‐based retreatment regimen, eight of whom achieved sustained virologic response 12 weeks after treatment and one of whom failed retreatment. Conclusion: The emergence of S282T substitution was rare in patients who fail SOF‐based regimens. Successful retreatment of prior SOF failure patients is possible in the presence of S282T substitution with SOF in combination with various direct‐acting antiviral agents. (Hepatology Communications 2017;1:538–549)

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“…Although several resistance-associated substitutions and baseline polymorphisms recently have been associated with treatment, their phenotypic impact on conferring resistance to rNAI remains to be determined (17)(18)(19). For example, the infrequent emergence of resistance-conferring mutation S282T (20)(21)(22) is probably due to the unfit nature of the mutated virus (23).…”

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confidence: 99%

Biochemical Characterization of the Active Anti-Hepatitis C Virus Metabolites of 2,6-Diaminopurine Ribonucleoside Prodrug Compared to Sofosbuvir and BMS-986094

Ehteshami

Tao

Öztürk

et al. 2016

Antimicrob Agents Chemother

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bRibonucleoside analog inhibitors (rNAI) target the hepatitis C virus (HCV) RNA-dependent RNA polymerase nonstructural protein 5B (NS5B) and cause RNA chain termination. Here, we expand our studies on ␤-D-2=-C-methyl-2,6-diaminopurine-ribonucleotide (DAPN) phosphoramidate prodrug 1 (PD1) as a novel investigational inhibitor of HCV. DAPN-PD1 is metabolized intracellularly into two distinct bioactive nucleoside triphosphate (TP) analogs. The first metabolite, 2=-C-methyl-GTP, is a wellcharacterized inhibitor of NS5B polymerase, whereas the second metabolite, 2=-C-methyl-DAPN-TP, behaves as an adenosine base analog. In vitro assays suggest that both metabolites are inhibitors of NS5B-mediated RNA polymerization. Additional factors, such as rNAI-TP incorporation efficiencies, intracellular rNAI-TP levels, and competition with natural ribonucleotides, were examined in order to further characterize the potential role of each nucleotide metabolite in vivo. Finally, we found that although both 2=-C-methyl-GTP and 2=-C-methyl-DAPN-TP were weak substrates for human mitochondrial RNA (mtRNA) polymerase (POLRMT) in vitro, DAPN-PD1 did not cause off-target inhibition of mtRNA transcription in Huh-7 cells. In contrast, administration of BMS-986094, which also generates 2=-C-methyl-GTP and previously has been associated with toxicity in humans, caused detectable inhibition of mtRNA transcription. Metabolism of BMS-986094 in Huh-7 cells leads to 87-fold higher levels of intracellular 2=-C-methyl-GTP than DAPN-PD1. Collectively, our data characterize DAPN-PD1 as a novel and potent antiviral agent that combines the delivery of two active metabolites. Chronic infection with hepatitis C virus (HCV) represents a significant disease burden on global health. It is estimated that 3% of the human population carries a chronic HCV infection, which translates to roughly 170 million infections worldwide and 3.2 million infections in the United States (1). Although up to 25% of acutely infected individuals clear the virus spontaneously, chronic HCV infection develops in the remaining 75% of those infected and can lead to the development of liver cirrhosis and hepatocellular carcinoma (2-4).HCV contains a 9.6-kb positive-strand RNA genome that codes for three structural and seven nonstructural proteins. The nonstructural protein 5B (NS5B) is the viral RNA-dependent RNA polymerase and is responsible for genome replication. Because of the high mutation rate of this enzyme, as well as the high virus replication rate, HCV propagation results in the generation of thousands of viral quasispecies (5, 6). Therefore, drug combination therapy is required in order to overcome development of resistance and to achieve successful viral clearance. HCV treatment comprised of ribavirin and pegylated interferon alpha, with or without protease inhibitors, can achieve sustained virologic responses in 40 to 80% of treated patients (7-9). However, curative outcomes historically have been suboptimal due to host polymorphisms, viral genotypic variability, and onset ...

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In Vivo Emergence of a Novel Mutant L159F/L320F in the NS5B Polymerase Confers Low-Level Resistance to the HCV Polymerase Inhibitors Mericitabine and Sofosbuvir

Abstract: NCT00869661, NCT01057667.

Cited by 113 publications

References 16 publications

Evolution of a Cell Culture-Derived Genotype 1a Hepatitis C Virus (H77S.2) during Persistent Infection with Chronic Hepatitis in a Chimpanzee

Evolution of a Cell Culture-Derived Genotype 1a Hepatitis C Virus (H77S.2) during Persistent Infection with Chronic Hepatitis in a Chimpanzee

The emergence of NS5B resistance associated substitution S282T after sofosbuvir‐based treatment

Biochemical Characterization of the Active Anti-Hepatitis C Virus Metabolites of 2,6-Diaminopurine Ribonucleoside Prodrug Compared to Sofosbuvir and BMS-986094

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