Preclinical Characterization of GS-9669, a Thumb Site II Inhibitor of the Hepatitis C Virus NS5B Polymerase

Abstract

GS-9669 is a highly optimized thumb site II nonnucleoside inhibitor of the hepatitis C virus (HCV) RNA polymerase, with a binding affinity of 1.35 nM for the genotype (GT) 1b protein. It is a selective inhibitor of HCV RNA replication, with a mean 50% effective concentration (EC 50 ) of <11 nM in genotype 1 and 5 replicon assays, but lacks useful activity against genotypes 2 to 4. The M423T mutation is readily generated clinically upon monotherapy with the thumb site II inhibitors filibuvir and lomibuvir, and it is notable that GS-9669 exhibited only a 3-fold loss in potency against this variant in the genotype 1b replicon. Rather than M423T, resistance predominantly tracks to residues R422K and L419M and residue I482L in GT 1b and 1a replicons, respectively. GS-9669 exhibited at least additive activity in combination with agents encompassing four other direct modes of action (NS3 protease, NS5A, NS5B via an alternative allosteric binding site, and NS5B nucleotide) as well as with alpha interferon or ribavirin in replicon assays. It exhibited high metabolic stability in in vitro human liver microsomal assays, which, in combination with its pharmacokinetic profiles in rat, dog, and two monkey species, is predictive of good human pharmacokinetics. GS-9669 is well suited for combination with other orally active, direct-acting antiviral agents in the treatment of genotype 1 chronic HCV infection. (This study has been registered at ClinicalTrials.gov under registration number NCT01431898.)

PTC725, an NS4B-Targeting Compound, Inhibits a Hepatitis C Virus Genotype 3 Replicon, as Predicted by Genome Sequence Analysis and Determined Experimentally

“…Danoprevir, an HCV NS3/4A protease inhibitor, had EC 50 s of 3.6 and 2.6 μM, as determined by luciferase assay and qRT-PCR, respectively, an observation consistent with prior reports indicating low susceptibility of HCV gt3 replicons to this compound (36). VX-222 (lomibuvir), a nonnucleoside HCV NS5B polymerase inhibitor, was similarly a poor inhibitor of gt3, as has been previously demonstrated for polymerase thumb site II inhibitors as a class (39), with EC 50 s of 5.8 and 4.8 μM, as determined by luciferase assay and qRT-PCR, respectively. Therefore, a lack of preexisting resistance mutations in the HCV gt3 NS4B coding sequence predicted the susceptibility of this genotype to inhibition by PTC725.…”

“…Clinical experience demonstrates that different genotypes can have differential responses to therapeutic intervention. In some cases, as with some narrow-spectrum HCV NS3/4A protease inhibitors, drugs may be active against one genotype but inactive against others due to critical sequence divergence in the drug binding site (39). PTC725 was previously shown to be an effective inhibitor of HCV gt1 replicon replication but with greatly decreased potency against HCV gt2 (18).…”

Biophysical Mode-of-Action and Selectivity Analysis of Allosteric Inhibitors of Hepatitis C Virus (HCV) Polymerase

“…However there are some small variations in DSF data, which can be explained by different experimental set ups, and SPR data especially with lomibuvir, where the interactions were described by a simple 1:1 interaction model [29,40,41]. …”

Antiviral response and resistance analysis of treatment‐naïve HCV‐infected patients receiving single and multiple doses of GS‐9190

“…The barrier to resistance of NNIs is lower compared to NIs in general. Cross‐resistance between NNIs targeting the same NS5B polymerase binding site has been observed, such as GS‐9669 and filibuvir and between GS‐9190 and dasabuvir . Clinically relevant RAVs to DAAs are important to define for management of HCV infection.…”