Archives

  • 2018-07
  • 2018-10
  • 2018-11
  • 2019-04
  • 2019-05
  • 2019-06
  • 2019-07
  • 2019-08
  • 2019-09
  • 2019-10
  • 2019-11
  • 2019-12
  • 2020-01
  • 2020-02
  • 2020-03
  • 2020-04
  • 2020-05
  • 2020-06
  • 2020-07
  • 2020-08
  • 2020-09
  • 2020-10
  • 2020-11
  • 2020-12
  • 2021-01
  • 2021-02
  • 2021-03
  • 2021-04
  • 2021-05
  • 2021-06
  • 2021-07
  • 2021-08
  • 2021-09
  • 2021-10
  • br Materials and methods br Results We studied patients infe

    2021-10-13


    Materials and methods
    Results We studied 16 patients infected with HCV genotypes 1a and 1b, naïve for protease inhibitor-based therapy. All patients, chronically infected, were selected by the Hepatology Department for triple therapy. Most of them had severe liver fibrosis at the beginning of treatment. Table 1 shows the demographic, clinical, and virological characterization of the population. Six patients had received previous therapy with PEG-IFN and RBV, showing null (n=3) or relapsed response (n=3) (Table 1, Table 2). After size- and quality trimming, an average of 182,093 reads (mean size of 157bp) of the NS3 protease gene region were obtained for each sample. At 4weeks of treatment, we obtained HCV-NS3 protease variants from 6 of the 16 patients studied, while 10 had no detectable HCV RNA. NGS sequencing was able to detect even low-frequency (<20%) variants. Several synonymous and non-synonymous substitutions were identified at baseline and 4weeks of treatment (Table 2). Four of the six patients with detectable HCV-RNA levels at 4weeks were infected with HCV-1b. In those N6-Methyl-dATP patients, the total number of variants (synonymous and non-synonymous) at 4weeks was either similar to or higher than at baseline. Triple therapy was discontinued before 48weeks in three patients, either because of HCV RNA level above 1,000IU/mL (considered non-responders, NR) or death (one case). All others completed the 48weeks of treatment. Telaprevir-based therapy was effective in 11 cases: 4 patients showed sustained viral response (SVR, undetected HCV RNA level at 6months after therapy), while 7 responders (R) had undetected HCV RNA level at 48weeks and are completing the 6months follow-up to characterize SVR. Triple therapy was not effective for 2 of 3 patients with null response to previous PEG-IFN/RBV treatment, while the previously relapsed ones showed either SVR or undetected HCV RNA level at 48weeks (Table 2). Table 3, Table 4 show the frequency of variants with non-synonymous substitutions in HCV-NS3 protease region in sub-types 1a and 1b, before and at 4weeks of PI-based therapy. Low-frequency variants corresponded to 74% and 35% of non-synonymous mutations identified in HCV-1a and -1b, respectively. All patients, in both HCV genotypes, presented more than one HCV-NS3 protease non-synonymous mutation. On the other hand, most of the detected variants were identified in only one patient, while few (29% for HCV-1a and 42% for HCV-1b) showed high prevalence. Only 4.8% (out of 42 substitutions for HCV-1a) and 13.5% (out of 52 substitutions for HCV-1b) of the variants present at baseline remained at 4weeks, showing no correspondence between variant profile mutations emerged under therapy and those at baseline. In 10 out of 16 patients, we identified 9 non-synonymous substitutions in the NS3 protease variants previously associated with protease inhibitor resistance, such as V36A [associated with resistance to telaprevir (TVR), boceprevir (BOC), danoprevir (ITMN-191), paritaprevir (ABT-450)], T54S [TVR, BOC, simeprevir (TMC-435), faldaprevir (BI-201335)], V55I (BOC, ITMN-191, ABT-450), Q80K (TMC-435), Q80R (TMC-435), V107I (BOC), I132V (TVR), D168E [TVR, BOC, TMC-435, BI-201335, ITMN-191, asunaprevir (ASV)] and M175L (BOC) (Table 3, Table 4). Five of these patients presented more than one resistance mutation, most of which confer cross-resistance to more than one drug. For HCV-1a infected patients, resistance mutations had a low frequency (<4%) at baseline yet high frequency (>94%) at 4weeks. Conversely, the frequency of most resistance mutations in HCV-1b infected patients was high at both time points. It is noteworthy that, for both genotypes, virus resistance mutations identified at baseline did not persist at 4weeks, whereas those detected at the latter time point were not present at baseline. Patients with resistant mutations at baseline showed diverse outcomes—from SVR to NR—suggesting no correlation between baseline profile and outcome. On the other hand, most patients presented also non-synonymous substitutions not yet described as RAVs, both at baseline and Week 4, which could potentially impact outcome (Table 3, Table 4). The presence of high frequency telaprevir RAVs at baseline (cases 3, 14 and 19) did not predict therapy failure. Despite I132V variant high prevalence at baseline, it was not detected at week 4, while T54S, not detected at baseline, showed high frequency at week 4. Case 12, with no RAVs at baseline, presented telaprevir RAV-T54S variant at week 4, even though had SVR.