Characterization of hepatitis C virus (HCV) quasispecies dynamics upon short-term dual therapy with the HCV NS5B nucleoside polymerase inhibitor mericitabine and the NS3/4 protease inhibitor danoprevir

Abstract

In the INFORM-1 study, 73 patients with chronic hepatitis C virus infection received mericitabine plus danoprevir for up to 13 days. Seventy-two patients experienced a continuous decline in HCV RNA levels during treatment, and of these patients, 14 had viral loads that remained >1,000 IU/ml by day 13 and 1 met the definition for viral breakthrough. In-depth NS5B and NS3/4A population and clonal sequencing studies and mericitabine and danoprevir drug susceptibility testing were performed to assess the variability and quasispecies dynamics before and upon monotherapy or dual therapy. Sequence analysis of the viral quasispecies indicated that the mericitabine resistance mutation S282T was not present at baseline, nor was it selected (even at a low level) during treatment. Protease inhibitor resistance mutations, either as predominant or as minority species, were detected in 18 patients at baseline. No enrichment of minority protease inhibitor-resistant variants present at baseline was observed during treatment; viral population samples were fully susceptible to mericitabine and/or danoprevir, despite the presence within their quasispecies of minority variants confirmed to have reduced susceptibility to danoprevir or other protease inhibitors. It was also observed that certain NS3 amino acid substitutions affected protease inhibitor drug susceptibility in a compound-specific manner and varied with the genetic context. In summary, the slower kinetics of viral load decline observed in some patients was not due to the selection of danoprevir or mericitabine resistance during treatment. Over 2 weeks' therapy, mericitabine suppressed the selection of danoprevir resistance, results that could differ upon longer treatment periods.

Fig 1

(A) Viral load (VL) kinetics of patients (boxed numbers) in cohort A who received mericitabine alone on days 1 to 3 and then mericitabine plus danoprevir on days 4 to 7; (B) viral load kinetics of patients in cohort A who received danoprevir alone on days 1 to 3 and then mericitabine plus danoprevir on days 4 to 7; (C) viral load kinetics of patients in cohort D. Includes a patient with a D168E resistance mutation (patient 19) who received mericitabine and danoprevir for 13 days. Other patients from the same cohort are presented as a comparison (patients 20, 23, 24, and 31 received placebo).

Fig 2

(A) Phenotypic characterization of susceptibility to mericitabine in samples obtained at baseline and during treatment from 10 patients in cohort A with an end-of-treatment serum HCV RNA level of ≥1,000 IU/ml; (B) phenotypic characterization of susceptibility to danoprevir in samples obtained at baseline and during treatment from 10 patients in cohort A with an end-of-treatment serum HCV RNA level of ≥1,000 IU/ml.

Fig 3

Presence and viral dynamics of low-frequency PI-resistant variants in cohort A patients with an end-of-treatment serum HCV RNA level of ≥1,000 IU/ml. DNV, danoprevir; MCB, mericitabine.