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Treatment failure with DAA therapy: Importance of resistance

  • Christoph Sarrazin
    Correspondence
    Corresponding author. Address: Goethe-University Hospital, Medizinische Klinik 1, Theodor-Stern-Kai 7, 60590 Frankfurt am Main, Germany; Tel.: +49-611-177-1251, fax: +49-0611-177-1252.
    Affiliations
    St. Josefs-Hospital, Beethovenstr. 20, 65189 Wiesbaden, Germany

    Goethe-University Hospital, Medizinische Klinik 1, Theodor-Stern-Kai 7, 60590 Frankfurt am Main, Germany
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Published:March 11, 2021DOI:https://doi.org/10.1016/j.jhep.2021.03.004

      Summary

      Viral resistance is a major reason for virological failure in patients being treated with direct-acting antivirals (DAAs) for chronic HCV infection. However, the importance of viral resistance mainly depends on the DAA regimen and HCV genotype. For first-line therapy with glecaprevir/pibrentasvir (G/P) or velpatasvir/sofosbuvir (VEL/SOF) no general baseline resistance analysis is required because of the high antiviral activity and high barrier to resistance. If available, resistance testing may help to optimise therapy in certain subgroups of patients with HCV genotype 3 and other rare HCV geno/subtypes. Voxilaprevir/velpatasvir/sofosbuvir (VOX/VEL/SOF) is the first choice for the second-line treatment of patients following a previous DAA failure, with rates of viral eradication above 90% irrespective of the presence of resistance-associated substitutions (RASs). However, in resource-limited settings, only first-generation DAAs may be available for second-line therapy. Here, RASs selected during initial antiviral therapy should be considered if testing is available and rescue treatment should include a switch to a regimen with a new DAA class to optimise treatment response. Patients with HCV genotype 3 are overrepresented in the group who experience DAA treatment failure. Limited data are available for third-line therapies, but promising results have been achieved with G/P plus SOF or VOX/VEL/SOF with or without ribavirin for 12 to 24 weeks; these regimens should be administered irrespective of a patient’s RAS profile.

      Keywords

      Introduction

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      Figure thumbnail gr1
      Fig. 1Important aspects of HCV resistance analysis.
      Potential parameters which influence importance/detection of RASs for virologic response to DAA therapies. DAA, direct-acting antiviral; RASs, resistance-associated substitutions.

      Baseline resistance analysis for pangenotypic DAA regimens

      While no general sequencing is required in subgroups of patients infected with HCV genotype 3, baseline resistance analysis may be useful to optimise the selection of DAA regimens for treatment-naïve patients.

      VEL/SOF and genotype 3

      In phase III approval studies for VEL/SOF, baseline NS5A RASs were not associated with virologic treatment outcome in patients infected with HCV genotypes 1, 2, 4-6. For HCV genotype 3, SVR rates with and without VEL-specific RASs were 93% and 98%, respectively. The frequency of NS5A RASs (15% cut-off by deep sequencing) specific for VEL was 12% overall and 5% for Y93H, respectively.
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      Resistance analysis in patients with genotype 1-6 HCV infection treated with sofosbuvir/velpatasvir in the phase III studies.
      In addition, in the approval study, SVR rates in genotype 3-infected patients with and without compensated cirrhosis were 91% and 97%, respectively.
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      For certain HCV genotype 3 subtypes, RASs are associated with reduced SVR rates on VEL/SOF or G/P.
      Figure thumbnail gr2
      Fig. 2VEL/SOF±RBV in HCV genotype 3-infected patients with compensated cirrhosis.
      Comparison of SVR rates following treatment with VEL/SOF vs. VEL/SOF+RBV for 12 weeks in all patients, patients without NS5A RASs, any NS5A RASs and Y93H as the most relevant NS5A RAS for VEL. For VEL/SOF, SVR rates in HCV genotype 3-infected patients with compensated cirrhosis were obtained from an integrated analysis of phase II/III approval studies
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      and the VEL/SOF arm of the controlled randomised Spanish study for comparison with VEL/SOF+RBV.
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      Efficacy of sofosbuvir and velpatasvir, with and without ribavirin, in patients with hepatitis C virus genotype 3 infection and cirrhosis.
      RASs, resistance-associated substitutions; RBV, ribavirin; SOF, sofosbuvir; SVR, sustained virologic response; VEL, velpatasvir.
      Table 1Importance of RASs for pangenotypic regimens in DAA-naïve HCV genotype 3-infected patients.
      HCV genotype 3-infected patients
      RegimenSubgroupRAS with reduced SVRNatural RAS prevalenceAlternative regimen
      VEL/SOF 12 weeksCompensated cirrhosisY93HApprox. 5%VEL/SOF+RBV 12 weeks

      G/P 8/16 weeks
      G/P 8 weeks.Treatment-naïve, without (with?) cirrhosisA30KApprox. 6%VEL/SOF 12 weeks

      G/P and genotype 3

      Early studies for G/P in HCV genotype 3 revealed higher relapse and lower SVR rates in comparison to other HCV genotypes, especially for treatment-experienced patients.
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      Glecaprevir and pibrentasvir yield high response rates in patients with HCV genotype 1-6 without cirrhosis.
      Therefore, initially only 8 weeks of G/P was approved for treatment-naïve patients with HCV genotype 3 without cirrhosis.
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      Glecaprevir-pibrentasvir for 8 or 12 Weeks in HCV genotype 1 or 3 infection.
      Baseline RASs within NS3 or NS5A in HCV genotype 1, 2, 4-6 were not significantly associated with virologic treatment failure.
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      Pooled resistance analysis in patients with hepatitis C virus genotype 1 to 6 infection treated with glecaprevir-pibrentasvir in phase 2 and 3 clinical trials.
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      Systematic review with meta-analysis: impact of baseline resistance-associated substitutions on the efficacy of glecaprevir/pibrentasvir among chronic hepatitis C patients.
      However, for HCV genotype 3, A30K was associated with differences in treatment responses in treatment-naïve patients without cirrhosis treated for 8 and 12 weeks, although this variant does not seem to confer resistance to pibrentasvir in vitro (Fig. 3).
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      Pooled resistance analysis in patients with hepatitis C virus genotype 1 to 6 infection treated with glecaprevir-pibrentasvir in phase 2 and 3 clinical trials.
      In a pooled resistance analysis of phase II/III studies, baseline A30K was associated with differences in SVR rates in treatment-naïve patients receiving 8-week therapy (78% vs. 99%, respectively) and treatment-experienced (interferon-alfa) patients receiving 12-week therapy (25% vs. 96%, respectively).
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      Pooled resistance analysis in patients with hepatitis C virus genotype 1 to 6 infection treated with glecaprevir-pibrentasvir in phase 2 and 3 clinical trials.
      ,
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      These differences disappeared with longer treatment durations of 12 weeks in treatment-naïve and 16 weeks in treatment-experienced patients. However, the number of patients treated for 16 weeks was low and real-world experiences are very limited. As for treatment-experienced patients with HCV genotype 3 the approved treatment duration is 16 weeks, though alternative treatment options may be preferred and no resistance analysis is recommended in this situation. However, for treatment-naïve patients with HCV genotype 3 without cirrhosis baseline resistance analysis may be considered prior to the standard 8-week regimen. A30K naturally occurs at a frequency of approximately 6%.
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      • Pilot-Matias T.
      • Schnell G.
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      • Ng T.I.
      • Reisch T.
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      Pooled resistance analysis in patients with hepatitis C virus genotype 1 to 6 infection treated with glecaprevir-pibrentasvir in phase 2 and 3 clinical trials.
      If A30K is detected, slightly lower SVR rates are to be expected and G/P may not be the first choice. No data are available on the potential efficacy of G/P plus RBV for 8 weeks in patients with A30K and the longer duration of 12 weeks is not approved and may lead to reimbursement issues. Thus, baseline resistance analysis in treatment-naïve patients with HCV genotype 3 without cirrhosis may only be used in settings with common access to HCV sequencing to select an optimal alternative treatment regimen and to avoid rescue therapy, which is also associated with higher failure rates in HCV genotype 3.
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      While baseline Y93H is the major RAS associated with treatment failure on VEL/SOF, A30K is for G/P. A combination of a dual A30K/Y93H mutation is associated with failure on both therapies but is an exceedingly rare event (Table 1).
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      Resistance analysis in patients with genotype 1-6 HCV infection treated with sofosbuvir/velpatasvir in the phase III studies.
      ,
      • Krishnan P.
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      • Tripathi R.
      • Ng T.I.
      • Reisch T.
      • et al.
      Pooled resistance analysis in patients with hepatitis C virus genotype 1 to 6 infection treated with glecaprevir-pibrentasvir in phase 2 and 3 clinical trials.
      Figure thumbnail gr3
      Fig. 3G/P in treatment-naïve HCV genotype 3-infected patients without cirrhosis.
      Comparison of SVR rates for treatment with G/P for 8 vs. 12 weeks in all patients, patients without NS5A RASs, any NS5A RASs and A30K as the most relevant NS5A RAS for pibrentasvir. Data are obtained from supplementary material to the publication of the approval study.
      • Zeuzem S.
      • Foster G.R.
      • Wang S.
      • Asatryan A.
      • Gane E.
      • Feld J.J.
      • et al.
      Glecaprevir-pibrentasvir for 8 or 12 Weeks in HCV genotype 1 or 3 infection.
      G/P, glecaprevir/pibrentasvir; RASs, resistance-associated substitutions; SVR, sustained virologic response.
      More recently, 8 weeks treatment with G/P was also approved for treatment-naïve patients with compensated cirrhosis.
      • Brown Jr., R.S.
      • Buti M.
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      Glecaprevir/pibrentasvir for 8 weeks in treatment-naive patients with chronic HCV genotypes 1-6 and compensated cirrhosis: the EXPEDITION-8 trial.
      In 63 patients with HCV genotype 3 only 1 virologic treatment failure was observed, and this patient had no baseline RASs within NS3 or NS5A. Thus, data are currently insufficient to assess the importance of A30K or other RASs in treatment-naïve HCV genotype 3-infected patients with compensated cirrhosis treated for 8 weeks.

      Selection of RASs after failure on first-line DAA treatment

      Pattern of RASs in DAA failures

      In most patients with virologic failure on DAA regimens, RASs are detectable within the target regions of the respective DAAs. Diverse patterns of resistance to NS3, NS5A and NS5B inhibitors have been characterised based on data from approval studies and real-world studies with larger cohorts of patients with virologic treatment failure (Table 2).
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      • et al.
      Treatment outcomes in hepatitis C virus genotype 1a infected patients with and without baseline NS5A resistance-associated substitutions.
      ,
      • Hezode C.
      • Reau N.
      • Svarovskaia E.S.
      • Doehle B.P.
      • Shanmugam R.
      • Dvory-Sobol H.
      • et al.
      Resistance analysis in patients with genotype 1-6 HCV infection treated with sofosbuvir/velpatasvir in the phase III studies.
      ,
      • Kwo P.
      • Gane E.J.
      • Peng C.Y.
      • Pearlman B.
      • Vierling J.M.
      • Serfaty L.
      • et al.
      Effectiveness of elbasvir and grazoprevir combination, with or without ribavirin, for treatment-experienced patients with chronic hepatitis C infection.
      ,
      • Krishnan P.
      • Pilot-Matias T.
      • Schnell G.
      • Tripathi R.
      • Ng T.I.
      • Reisch T.
      • et al.
      Pooled resistance analysis in patients with hepatitis C virus genotype 1 to 6 infection treated with glecaprevir-pibrentasvir in phase 2 and 3 clinical trials.
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      • Zhao Y.
      • et al.
      Grazoprevir-elbasvir combination therapy for treatment-naive cirrhotic and noncirrhotic patients with chronic hepatitis C virus genotype 1, 4, or 6 infection: a randomized trial.
      • Wyles D.
      • Thompson A.
      • Lawitz E.
      • Willems B.
      • Gane E.
      • Svarosvskaia E.S.
      • et al.
      No impact of RASs on the high efficacy of SOF/VEL/VOX for 8 weeks in DAA-naïve patients: an integrated resistance analysis of the POLARIS-2 and POLARIS-3 studies.
      • Dietz J.
      • Di Maio V.C.
      • de Salazar A.
      • Merino D.
      • Vermehren J.
      • Paolucci S.
      • et al.
      Failure to voxilaprevir, velpatasvir, sofosbuvir and efficacy of rescue therapy.
      • de Salazar A.
      • Dietz J.
      • di Maio V.C.
      • Vermehren J.
      • Paolucci S.
      • Mullhaupt B.
      • et al.
      Prevalence of resistance-associated substitutions and retreatment of patients failing a glecaprevir/pibrentasvir regimen.
      Rescue treatment with multiple targeted therapies (VOX/VEL/SOF) is the first choice for failures on first-generation DAAs or G/P.
      Table 2Typical resistance pattern in patients with virologic failure on DAAs.
      Failure onGT1aGT1bGT2GT3GT4GT5GT6
      NS3 inhibitors
       SMVR155K

      D168E
      D168Vn.a.n.a.Q80R, D168En.a.n.a.
       PTVrR155K

      D168V
      Y56H

      D168V
      n.a.n.a.Y56H

      D168V
      n.a.n.a.
       GZRQ80K

      A156V/T

      D168A/E
      A156G/T

      D168A/E/G/V
      n.a.n.a.A156Tn.a.n.a.
       GLEA156VY56F

      Q80L

      S122G
      no RASsY56H

      Q80R/K

      A156G

      Q168L/R
      no datano datano data
       VOXQ80Kno RASsno RASsno RASsA156Sno RASsno data
      NS5A inhibitors
       DACQ30H/R

      L31M
      L31M

      Y93H
      no dataY93HL28M

      L30R

      Y93C/H
      no datano data
       LDVQ30H/R

      L31M
      L31M

      Y93H
      n.a.no RASsL28M

      Y93C/H
      n.a.n.a.
       OMVM28T/V

      Q30R
      Y93Hn.a.n.a.L28V

      L30R

      M31L/V

      Y93H
      n.a.n.a.
       EBVM28T

      Q30H/R

      L31M

      Y93C/H
      R30H

      L31F/M/V

      Y93H
      n.a.n.a.L30R

      M31L/V
      n.a.n.a.
       VELQ30R/H

      L31M

      Y93N/H
      L31M

      Y93H
      L31MA30K

      L31M

      Y93H
      L28M/V

      L30R

      M31L/V

      Y93C/H
      no dataT58S
       PIBQ30R

      L31M

      H58D

      Y93N
      L31M/F

      P32del

      Y93H
      F28C

      L31M
      M28G

      A30K/G

      L31F/M

      Y93H
      no datano datano data
      NonNUC inhibitors
       DSVS556GS556Gn.a.n.a.n.a.n.a.n.a.
      NUC inhibitor
       SOFS282TL159F C316N S282Tno dataL159F

      S282T
      S282Tno datano data
      Data from DAA combination therapies (SMV/SOF, PTVr/OMV±DSV, GZR/EBR, GLE/PIB, DAC/SOF, LDV/SOF, VEL/SOF, VOX/VEL/SOF with and without RBV. Only variants selected in the majority of patients with virologic treatment failure are shown as presented in different studies and from own unpublished data of the European Resistance Data bank. Other RASs also have been detected in individual patients. For HCV genotypes 2-6 RASs selected depend substantially on HCV subtypes.
      • Dietz J.
      • Vermehren J.
      • Matschenz K.
      • Buggisch P.
      • Klinker H.
      • Schulze zur Wiesch J.
      • et al.
      Treatment outcomes in hepatitis C virus genotype 1a infected patients with and without baseline NS5A resistance-associated substitutions.
      ,
      • Hezode C.
      • Reau N.
      • Svarovskaia E.S.
      • Doehle B.P.
      • Shanmugam R.
      • Dvory-Sobol H.
      • et al.
      Resistance analysis in patients with genotype 1-6 HCV infection treated with sofosbuvir/velpatasvir in the phase III studies.
      ,
      • Kwo P.
      • Gane E.J.
      • Peng C.Y.
      • Pearlman B.
      • Vierling J.M.
      • Serfaty L.
      • et al.
      Effectiveness of elbasvir and grazoprevir combination, with or without ribavirin, for treatment-experienced patients with chronic hepatitis C infection.
      ,
      • Krishnan P.
      • Pilot-Matias T.
      • Schnell G.
      • Tripathi R.
      • Ng T.I.
      • Reisch T.
      • et al.
      Pooled resistance analysis in patients with hepatitis C virus genotype 1 to 6 infection treated with glecaprevir-pibrentasvir in phase 2 and 3 clinical trials.
      ,
      • Dietz J.
      • Susser S.
      • Vermehren J.
      • Peiffer K.H.
      • Grammatikos G.
      • Berger A.
      • et al.
      Patterns of resistance-associated substitutions in patients with chronic HCV infection following treatment with direct-acting antivirals.
      • Dietz J.
      • Kalinina O.V.
      • Vermehren J.
      • Peiffer K.H.
      • Matschenz K.
      • Buggisch P.
      • et al.
      Resistance-associated substitutions in patients with chronic hepatitis C virus genotype 4 infection.
      • Zeuzem S.
      • Ghalib R.
      • Reddy K.R.
      • Pockros P.J.
      • Ben Ari Z.
      • Zhao Y.
      • et al.
      Grazoprevir-elbasvir combination therapy for treatment-naive cirrhotic and noncirrhotic patients with chronic hepatitis C virus genotype 1, 4, or 6 infection: a randomized trial.
      • Wyles D.
      • Thompson A.
      • Lawitz E.
      • Willems B.
      • Gane E.
      • Svarosvskaia E.S.
      • et al.
      No impact of RASs on the high efficacy of SOF/VEL/VOX for 8 weeks in DAA-naïve patients: an integrated resistance analysis of the POLARIS-2 and POLARIS-3 studies.
      • Dietz J.
      • Di Maio V.C.
      • de Salazar A.
      • Merino D.
      • Vermehren J.
      • Paolucci S.
      • et al.
      Failure to voxilaprevir, velpatasvir, sofosbuvir and efficacy of rescue therapy.
      • de Salazar A.
      • Dietz J.
      • di Maio V.C.
      • Vermehren J.
      • Paolucci S.
      • Mullhaupt B.
      • et al.
      Prevalence of resistance-associated substitutions and retreatment of patients failing a glecaprevir/pibrentasvir regimen.
      DAA, direct-acting antiviral; DAC, daclatasvir; DSV, dasabuvir; EBV, elbasvir; GLE, glecaprevir; GT, genotype; GZR, grazoprevir; LDV, ledipasvir; Non-NUC, non-nucleosidic NS5B polymerase inhibitor; NUC, nucleotide NS5B polymerase inhibitor; OMV, ombitasvir; PIB, pibrentasvir; PTVr, paritaprevir/ritonavir; RASs, resistance-associated substitutions; RBV, ribavirin; SMV, simeprevir; SOF, sofosbuvir; VEL, velpatasvir; VOX, voxilaprevir. n.a., not applicable as no (general) use of respective DAAs in these HCV genotypes.
      Overall, RAS patterns are partially overlapping for the different NS3 and NS5A inhibitors. However, there are also differences with non-overlapping resistance profiles which may be important for selection of rescue DAA regimens. For example, the first- and second-generation NS3 protease inhibitors (PIs) simeprevir, paritaprevir/ritonavir, grazoprevir vs. glecaprevir, voxilaprevir showed different RAS profiles (Table 2). For NS5A inhibitors, RAS profiles are remarkably similar for HCV genotype 1b, but some differences are observed for other HCV geno-/subtypes (Table 2). For dasabuvir, as the only non-nucleoside NS5B polymerase inhibitor, a single variant (S556G) was most commonly selected in HCV genotype 1a/b (Table 2). For sofosbuvir, as a nucleotide NS5A polymerase inhibitor, S282T was selected very rarely, mainly in HCV genotypes 1, 3 and 4. The importance of selection of other variants so far is unclear (Table 2). Moreover, for rare HCV genotypes/subtypes, for HCV genotypes 5 and 6, and for pangenotypic regimens and HCV genotype 2, no or only few valid data are available as no larger cohorts of virologic failure patients have been reported to date (Table 2). Finally, for voxilaprevir/velpatasvir/sofosbuvir (VOX/VEL/SOF), as a regimen targeting 3 HCV proteins with a high genetic barrier to resistance in the majority of virologic failure patients, no RASs have been selected (Table 2). Resistance levels of single RASs are highly variable for the different NS3 and NS5A inhibitors. Pangenotypic DAA regimens generally display higher barriers to resistance. However, several combined NS5A RASs and the P32del variant which was observed mainly in HCV genotype 1b-infected DAA failure patients from Asia also confer high levels of resistance to G/P and VEL/SOF.
      • Gottwein J.M.
      • Pham L.V.
      • Mikkelsen L.S.
      • Ghanem L.
      • Ramirez S.
      • Scheel T.K.H.
      • et al.
      Efficacy of NS5A inhibitors against hepatitis C virus genotypes 1-7 and escape variants.
      • Jensen S.B.
      • Fahnoe U.
      • Pham L.V.
      • Serre S.B.N.
      • Tang Q.
      • Ghanem L.
      • et al.
      Evolutionary pathways to persistence of highly fit and resistant hepatitis C virus protease inhibitor escape variants.
      • Hikita H.
      • Takehara T.
      NS5A-P32 deletion in hepatitis C genotype 1b infection is the most refractory treatment-mediated amino acid change exhibiting resistance to all NS5A inhibitors.

      Persistence of RASs

      RASs selected during administration of NS3 PIs and the NS5B inhibitor sofosbuvir will disappear relative rapidly (within weeks to months) during follow-up depending on HCV geno-/subtype and based on population sequencing.
      • Sarrazin C.
      The importance of resistance to direct antiviral drugs in HCV infection in clinical practice.
      ,
      • Pawlotsky J.M.
      Hepatitis C virus resistance to direct-acting antiviral drugs in interferon-free regimens.
      During long-term follow-up for several years no or only minor NS3 RASs for different PIs were observed.
      • Zoulim F.
      • Moreno C.
      • Lee S.S.
      • Buggisch P.
      • Horban A.
      • Lawitz E.
      • et al.
      A 3-year follow-up study after treatment with simeprevir in combination with pegylated interferon-alpha and ribavirin for chronic hepatitis C virus infection.
      • Susser S.
      • Vermehren J.
      • Forestier N.
      • Welker M.W.
      • Grigorian N.
      • Fuller C.
      • et al.
      Analysis of long-term persistence of resistance mutations within the hepatitis C virus NS3 protease after treatment with telaprevir or boceprevir.
      • Lahser F.
      • Galloway A.
      • Hwang P.
      • Palcza J.
      • Brunhofer J.
      • Wahl J.
      • et al.
      Interim analysis of a 3-year follow-up study of NS5A and NS3 resistance-associated substitutions after treatment with grazoprevir-containing regimens in participants with chronic HCV infection.
      An exception is the NS3 variant Q80K in HCV subtype 1a-infected patients, which typically persists during long-term follow-up.
      • Dietz J.
      • Vermehren J.
      • Welzel T.
      • Müllhaupt B.
      • Buggisch P.
      • Matschenz K.
      • et al.
      Analysis of long-term persistence of HCV resistance-associated substitutions within NS3, NS5A and NS5B in genotype 1 and 3 after DAA treatment failure.
      In contrast, RASs selected on treatment with NS5A inhibitors seem to persist for more than 2 years in >80% of patients.
      • Wyles D.
      • Mangia A.
      • Cheng W.
      • Shafran S.
      • Schwabe C.
      • Ouyang W.
      • et al.
      Long-term persistence of HCV NS5A resistance-associated substitutions after treatment with the HCV NS5A inhibitor, ledipasvir, without sofosbuvir.
      However, some changes within the HCV quasispecies have been observed by deep sequencing. Also, during long-term follow-up (for up to 4 years) a decline in several NS5A RASs has been reported, depending on the NS5A inhibitor and HCV genotype.
      • Sarrazin C.
      The importance of resistance to direct antiviral drugs in HCV infection in clinical practice.
      ,
      • Pawlotsky J.M.
      Hepatitis C virus resistance to direct-acting antiviral drugs in interferon-free regimens.
      ,
      • Dietz J.
      • Vermehren J.
      • Welzel T.
      • Müllhaupt B.
      • Buggisch P.
      • Matschenz K.
      • et al.
      Analysis of long-term persistence of HCV resistance-associated substitutions within NS3, NS5A and NS5B in genotype 1 and 3 after DAA treatment failure.

      Reselection of RASs during repeated DAA therapy

      For HIV infection it is well known that RASs selected during direct-antiviral therapy can remain in the quasispecies below the limit of detection of sequencing assays. During retreatment, these variants then reappear very rapidly leading to immediate development of resistance. For HCV, few studies – with contradictory results – have investigated the persistence and reselection of RASs after DAA treatment. In several patients, persistence and reselection of NS3 RASs was observed during sequential treatment with a PI at short-term intervals.
      • Vermehren J.
      • Susser S.
      • Lange C.M.
      • Forestier N.
      • Karey U.
      • Hughes E.
      • et al.
      Mutations selected in the hepatitis C virus NS3 protease domain during sequential treatment with boceprevir with and without pegylated interferon alfa-2b.
      However, in patients who were re-exposed to a NS3 PI after more than 1 year, there was no clear evidence of persistence but de novo generation of resistance was observed (by deep sequencing) in most patients.
      • Susser S.
      • Flinders M.
      • Reesink H.W.
      • Zeuzem S.
      • Lawyer G.
      • Ghys A.
      • et al.
      Evolution of hepatitis C virus quasispecies during repeated treatment with the NS3/4A protease inhibitor telaprevir.
      ,
      • Lenz O.
      • de Bruijne J.
      • Vijgen L.
      • Verbinnen T.
      • Weegink C.
      • Van Marck H.
      • et al.
      Efficacy of re-treatment with TMC435 as combination therapy in hepatitis C virus-infected patients following TMC435 monotherapy.
      As NS5A RASs typically persist at high frequencies and selection of RASs on treatment with sofosbuvir is rare, no comparable data are available for NS5A or NS5B RASs.
      There is no evidence that known RASs impact on rescue treatment with multiple targeted therapies (VOX/VEL/SOF).

      Importance of RASs for rescue treatment

      A limited number of studies on retreatment after DAA failure have been published. Therefore, evidence for recommendations on suitable retreatment options is limited (Fig. 4).
      Figure thumbnail gr4
      Fig. 4General concepts and recommendations for retreatment in patients with virologic failure on interferon-free DAA combination therapies.
      Recommendations according to pretreatment with first- or second-generation DAA regimens and according to availability of first- or second-generation DAA regimens for first- and second-line retreatment. Regarding combinations of a PI together with SOF as rescue treatment, data are only available for simeprevir (which is no longer available) while no data exist for other PIs like Narlaprevir or asunaprevirDAA, direct-acting antiviral; G/P, glecaprevir/pibrentasvir; RASs, resistance-associated substitutions; RBV, ribavirin; SOF, sofosbuvir; SVR, sustained virologic response; VEL, velpatasvir; VOX, voxilaprevir.

      Retreatment with first-generation DAAs

      Retreatment with first-generation DAAs is the only option in many healthcare settings globally; thus, data on the importance of RASs and other predictors for the selection of DAA regimens and the duration of rescue treatment is of importance. Moreover, ribavirin was shown to increase SVR rates in many DAA studies and thus should generally be considered for rescue treatment with first-generation DAAs. However, epidemiological, clinical and virologic characteristics of published cohorts are highly heterogeneous, and no prospective evaluation of retreatment strategies is available. After failure on a combination regimen with first-generation DAAs, RASs selected during the previous DAA therapy are observed in most patients and these RASs seem to influence virologic treatment outcome. For example, retreatment with an identical regimen is generally not recommended, as high rates of virologic failure have been reported.
      • Lawitz E.
      • Flamm S.
      • Yang J.
      • Pang P.S.
      • Zhu Y.
      • Svarosvskaia E.S.
      • et al.
      Retreatment of patients who failed 8 or 12 weeks of ledipasvir/sofosbuvir-based regimens with ledipasvir/sofosbuvir for 24 weeks.
      Thus, RAS testing may be helpful to tailor salvage therapy. However, RAS testing is often not available or affordable. In this setting, as a general rule, a new DAA class (NS3-, NS5A-, NS5B-inhibitor) should be introduced for rescue treatment to optimise treatment response if possible.

      Failure on PI plus NS5A inhibitor

      In patients with virologic failure on a first-generation PI (simeprevir, asunaprevir, grazoprevir, paritaprevir) in combination with a first-generation NS5A inhibitor (daclatasvir, ombitasvir, elbasvir) typically drug-specific NS3 and NS5A RAS patterns are detectable.
      • Dietz J.
      • Susser S.
      • Vermehren J.
      • Peiffer K.H.
      • Grammatikos G.
      • Berger A.
      • et al.
      Patterns of resistance-associated substitutions in patients with chronic HCV infection following treatment with direct-acting antivirals.
      These regimens have mainly been administered to patients with HCV genotype 1 and less frequently to those with genotype 4 infections. In patients with HCV genotype 1b infection and previous failure on asunaprevir/daclatasvir, retreatment with sofosbuvir plus ledipasvir led to SVR rates of 64–70% in a study from Japan.
      • Kanda T.
      • Nirei K.
      • Matsumoto N.
      • Higuchi T.
      • Nakamura H.
      • Yamagami H.
      • et al.
      Retreatment of patients with treatment failure of direct-acting antivirals: focus on hepatitis C virus genotype 1b.
      Rescue therapy with sofosbuvir plus velpatasvir was shown to be associated with equally high SVR rates in a small cohort; the presence of the P32del NS5A variant seems to be associated with reduced efficacy.
      • Hikita H.
      • Takehara T.
      NS5A-P32 deletion in hepatitis C genotype 1b infection is the most refractory treatment-mediated amino acid change exhibiting resistance to all NS5A inhibitors.
      ,
      • Takaki S.
      • Imamura M.
      • Yamaguchi S.
      • Fukuhara T.
      • Mori N.
      • Tsuji K.
      • et al.
      Real-word efficacy of sofosbuvir, velpatasvir plus ribavirin therapy for chronic hepatitis patients who failed to prior DAA therapy with NS5A-P32 deletion mutated HCV infection.
      Similarly, sofosbuvir plus an NS5A inhibitor was reported to be highly effective for the retreatment of patients who failed on paritaprevir/ombitasvir plus dasabuvir (3D) with or without ribavirin.
      • Vermehren J.
      • Dietz J.
      • Susser S.
      • Von Hahn T.
      • Petersen J.
      • Hinrichsen H.
      • et al.
      Retreatment of patients who failed direct-acting antiviral (DAA) therapies: real world experience from a large European hepatitis C resistance database.
      Therefore, although no systematic study data are available, switching to a nucleotide inhibitor (sofosbuvir) in combination with a NS5A inhibitor (ideally pangenotypic) seems to be highly effective for the retreatment of patients after virologic failure on a PI plus NS5A inhibitor (with or without a non-nucleosidic NS5B polymerase inhibitor [dasabuvir]). Ribavirin may be administered in addition and treatment duration may be extended from 12 to 24 weeks in difficult to treat patients. A high range of SVR rates between 60% and 100% were reported with this approach; the presence of NS5A RASs may be associated with reduced SVR rates (Fig. 4).

      Failure on PI plus sofosbuvir

      After virologic treatment failure on a PI (mainly simeprevir) in combination with sofosbuvir as a first-line DAA therapy, NS3 RASs are typically detectable in most patients. Data on rescue treatment for these mainly HCV genotype 1-infected patients are scarce, but high SVR rates (75–100%) have been reported, irrespective of the existence of RASs, following a switch to a NS5A inhibitor (ledipasvir, daclatasvir, velpatasvir) in combination with sofosbuvir. The addition of ribavirin does not seem to be required for further efficacy in this setting.
      • Vermehren J.
      • Dietz J.
      • Susser S.
      • Von Hahn T.
      • Petersen J.
      • Hinrichsen H.
      • et al.
      Retreatment of patients who failed direct-acting antiviral (DAA) therapies: real world experience from a large European hepatitis C resistance database.
      ,
      • Kondili L.A.
      • Gaeta G.B.
      • Brunetto M.R.
      • Di Leo A.
      • Iannone A.
      • Santantonio T.A.
      • et al.
      Incidence of DAA failure and the clinical impact of retreatment in real-life patients treated in the advanced stage of liver disease: interim evaluations from the PITER network.
      In addition, especially in HCV genotype 1b-infected patients, the 3D regimen (paritaprevir/ombitasvir plus dasabuvir) was highly effective in a small number of patients.
      • Vermehren J.
      • Dietz J.
      • Susser S.
      • Von Hahn T.
      • Petersen J.
      • Hinrichsen H.
      • et al.
      Retreatment of patients who failed direct-acting antiviral (DAA) therapies: real world experience from a large European hepatitis C resistance database.
      Thus, in patients not exposed to NS5A inhibitors, retreatment together with sofosbuvir seems to be highly effective and can be recommended for rescue treatment in resource-limited settings.
      If multiple targeted therapies (VOX/VEL/SOF) are not available or affordable rescue treatment with first-generation DAAs is possible with reduced efficacy. In such cases, resistance analysis should be performed if available to tailor salvage therapy. As general rule a switch to a regimen including a new DAA class is of importance to optimise the efficacy of retreatment.

      Failure on NS5A inhibitor plus sofosbuvir

      Retreatment of patients with virologic failure on a NS5A inhibitor (ledipasvir, daclatasvir, velpatasvir) in combination with sofosbuvir is more challenging. Data are mainly available for HCV genotype 1- and 3-infected patients. For HCV genotype 1, retreatment with the same or another NS5A inhibitor plus sofosbuvir was associated with SVR rates ranging from 25% to 100%. Significant differences between patients with (33-67%) and without NS5A RASs (88%) have been observed.
      • Dietz J.
      • Spengler U.
      • Mullhaupt B.
      • Schulze zur Wiesch J.
      • Piecha F.
      • Mauss S.
      • et al.
      Efficacy of retreatment after failed direct-acting antiviral therapy in patients with HCV genotype 1-3 infections.
      A switch to other regimens led to variable SVR rates of 89–92% for simeprevir/sofosbuvir with or without ribavirin, 67% for grazoprevir/elbasvir and 60–88% for the 3D regimen. Generally, SVR rates were higher in HCV genotype 1b vs. 1a infected patients and the addition of ribavirin seems to increase efficacy further.
      • Dietz J.
      • Spengler U.
      • Mullhaupt B.
      • Schulze zur Wiesch J.
      • Piecha F.
      • Mauss S.
      • et al.
      Efficacy of retreatment after failed direct-acting antiviral therapy in patients with HCV genotype 1-3 infections.
      For patients infected with HCV genotype 3, no switch to a new DAA class is possible if only first-generation DAAs are available. In such cases, retreatment of daclatasvir/sofosbuvir failures with the same or another NS5A inhibitor for durations between 12 and 36 weeks led to an overall SVR rate of only 60%. The combination of velpatasvir/sofosbuvir was most effective (50–86% SVR). However, the presence of cirrhosis and Y93H as a major NS5A RAS was associated with reduced efficacy (50% and 56%, respectively).
      • Dietz J.
      • Spengler U.
      • Mullhaupt B.
      • Schulze zur Wiesch J.
      • Piecha F.
      • Mauss S.
      • et al.
      Efficacy of retreatment after failed direct-acting antiviral therapy in patients with HCV genotype 1-3 infections.
      Based on data from a large cohort, resistance analysis was recommended as an efficacious way to guide rescue therapy in patients with NS5A inhibitor/sofosbuvir failures. In patients without NS5A RASs, combination therapy with a NS5A inhibitor plus sofosbuvir with ribavirin led to SVR rates of approximately 90%. For HCV genotype 3, velpatasvir/sofosbuvir plus ribavirin for 24 weeks can be recommended, with similar efficacy if only Y93H is observed.
      • Perez A.B.
      • Chueca N.
      • Garcia-Deltoro M.
      • Martinez-Sapina A.M.
      • Lara-Perez M.M.
      • Garcia-Bujalance S.
      • et al.
      High efficacy of resistance-guided retreatment of HCV patients failing NS5A inhibitors in the real world.
      Generally, the addition of ribavirin is also recommended for rescue treatment in genotype 3-infected patients as it seems to enhance efficacy.
      However, because patients without NS5A resistance are rare after virologic failure on NS5A inhibitors, and the efficacy of NS5A inhibitor-free regimens is limited, multiple targeted DAA regimens are recommended for rescue treatment in most patients (see below).

      Retreatment with multiple targeted therapies

      Inhibition of the NS3 protease, the NS5A replicase and the NS5B polymerase with DAAs following virologic failure represents a promising retreatment approach, as viral escape may become impossible – despite the selection of highly resistant RASs during initial antiviral therapy – because of the broad antiviral targeting potential. Studies have tested the combinations of different first- and second-generation PIs, NS5A inhibitors and sofosbuvir.
      • de Ledinghen V.
      • Laforest C.
      • Hezode C.
      • Pol S.
      • Renault A.
      • Alric L.
      • et al.
      Retreatment with sofosbuvir plus grazoprevir/elbasvir plus ribavirin of patients with hepatitis C virus genotype 1 or 4 who previously failed an NS5A- or NS3-containing regimen: the ANRS HC34 REVENGE study.
      ,
      • Lok A.S.
      • Sulkowski M.S.
      • Kort J.J.
      • Willner I.
      • Reddy K.R.
      • Shiffman M.L.
      • et al.
      Efficacy of glecaprevir and pibrentasvir in patients with genotype 1 hepatitis C virus infection with treatment failure after NS5A inhibitor plus sofosbuvir therapy.
      So far, only the combination of the second-generation pangenotypic PI voxilaprevir, with the pangenotypic NS5A inhibitor velpatasvir and sofosbuvir (VOX/VEL/SOF) has been approved for retreatment of patients following DAA failure.

      Retreatment with VOX/VEL/SOF

      In the approval studies, patients with and without previous exposure to a NS5A inhibitor were enrolled.
      • Bourliere M.
      • Gordon S.C.
      • Flamm S.L.
      • Cooper C.L.
      • Ramji A.
      • Tong M.
      • et al.
      Sofosbuvir, velpatasvir, and voxilaprevir for previously treated HCV infection.
      Retreatment of patients without previous administration of NS5A inhibitors led to SVR rates of 90% and 98% after VEL/SOF or VOX/VEL/SOF. The approval study for VEL/SOF, also included HCV genotype 1-infected patients with previous exposure to NS3 PIs in combination with peg-interferon-alfa and ribavirin.
      • Feld J.J.
      • Jacobson I.M.
      • Hezode C.
      • Asselah T.
      • Ruane P.J.
      • Gruener N.
      • et al.
      Sofosbuvir and velpatasvir for HCV genotype 1, 2, 4, 5, and 6 infection.
      Interestingly, for this group, the SVR rate after 12 weeks VEL/SOF in the VEL/SOF approval study was 100% in contrast to 91% after VEL/SOF in the DAA retreatment study.
      • Bourliere M.
      • Gordon S.C.
      • Flamm S.L.
      • Cooper C.L.
      • Ramji A.
      • Tong M.
      • et al.
      Sofosbuvir, velpatasvir, and voxilaprevir for previously treated HCV infection.
      ,
      • Feld J.J.
      • Jacobson I.M.
      • Hezode C.
      • Asselah T.
      • Ruane P.J.
      • Gruener N.
      • et al.
      Sofosbuvir and velpatasvir for HCV genotype 1, 2, 4, 5, and 6 infection.
      A potential explanation for this difference may be the previous exposure to sofosbuvir in all patients in the DAA retreatment study, while in the VEL/SOF approval studies neither nucleotide inhibitors nor NS5A inhibitors were allowed as previous therapy.
      G/P plus SOF or VOX/VEL/SOF with or without RBV for 12 to 24 weeks are options for the retreatment of VOX/VEL/SOF failures.
      In patients who were previously exposed to a NS5A inhibitor, rescue treatment with VOX/VEL/SOF yielded an overall SVR rate of 96%.
      • Bourliere M.
      • Gordon S.C.
      • Flamm S.L.
      • Cooper C.L.
      • Ramji A.
      • Tong M.
      • et al.
      Sofosbuvir, velpatasvir, and voxilaprevir for previously treated HCV infection.
      Virologic treatment failure was observed in 7 patients, of whom 4 were infected with HCV genotype 3a. HCV resistance patterns assessed by deep sequencing were typically identical before and after VOX/VEL/SOF rescue treatment. The frequency of RASs in the HCV quasispecies increased in 2 patients and new NS5A RASs were selected after VOX/VEL/SOF failure in only 2 patients.
      • Sarrazin C.
      • Cooper C.L.
      • Manns M.P.
      • Reddy K.R.
      • Kowdley K.V.
      • Roberts S.K.
      • et al.
      No impact of resistance-associated substitutions on the efficacy of sofosbuvir, velpatasvir, and voxilaprevir for 12 weeks in HCV DAA-experienced patients.
      In the real-world experience with VOX/VEL/SOF, with and without ribavirin, SVR rates of between 91–96% were reported following rescue treatment in cohorts from different countries.
      • Degasperi E.
      • Spinetti A.
      • Lombardi A.
      • Landonio S.
      • Rossi M.C.
      • Pasulo L.
      • et al.
      Real-life effectiveness and safety of sofosbuvir/velpatasvir/voxilaprevir in hepatitis C patients with previous DAA failure.
      • Llaneras J.
      • Riveiro-Barciela M.
      • Lens S.
      • Diago M.
      • Cachero A.
      • Garcia-Samaniego J.
      • et al.
      Effectiveness and safety of sofosbuvir/velpatasvir/voxilaprevir in patients with chronic hepatitis C previously treated with DAAs.
      • Belperio P.S.
      • Shahoumian T.A.
      • Loomis T.P.
      • Backus L.I.
      Real-world effectiveness of sofosbuvir/velpatasvir/voxilaprevir in 573 direct-acting antiviral experienced hepatitis C patients.
      • Papaluca T.
      • Roberts S.K.
      • Strasser S.I.
      • Stuart K.A.
      • Farrell G.
      • MacQuillan G.
      • et al.
      Efficacy and safety of sofosbuvir/velpatasvir/voxilaprevir for HCV NS5A-inhibitor experienced patients with difficult to cure characteristics.
      • Vermehren J.
      • Serfert Y.
      • Cornberg M.
      • Stoehr A.
      • Klinker H.
      • Simon K.G.
      • et al.
      Sofosbuvir, velpatasvir, and voxilaprevir for patients with failure of previous direct-acting antiviral therapy for chronic hepatitis C: results from the German Hepatitis C-Registry (DHC-R).
      • Smith D.A.
      • Bradshaw D.
      • Mbisa J.
      • Manso C.
      • Bibby D.
      • Singer J.
      • et al.
      Real-world retreatment of HCV-infected patients with prior failure to direct antiviral therapy using sofosbuvir, velpatasvir and voxilaprevir.
      • Janjua N.
      • Wilton J.C.
      • Cook D.
      • Wong S.
      • Butt Z.
      • Barlett S.
      • et al.
      Real-world effectiveness of sofosbuvir/velpatasvir/voxilaprevir as a hepatitis C virus infection salvage treatment.
      • Flamm S.
      • Tsai N.
      • Bacon B.
      • Curry M.
      • Milligan S.
      • Wick N.
      • et al.
      Pangenotypic therapies glecaprevir-pibrentasvir (G-P) and sofosbuvir-velpatasvir-voxilaprevir (S-V-V) after failure with interferon (IFN)-frei direct acting antiviral (DAA) treatment for hepatitis C.
      Although the different studies are highly heterogeneous with respect to distribution of HCV genotypes and rate of patients with cirrhosis, based on larger numbers of patients it seems that HCV genotype 3a and cirrhosis are weak negative predictors of response, with SVR rates of 81–92% and 81–90%, respectively. Also, pretreatment with VEL/SOF was associated with lower SVR rates in some studies, while this was not observed in others (Table 3). Furthermore, SVR rates in patients with active or previous liver cancer were highly variable in the different studies. Finally, it was also shown that in patients with end-stage renal disease, as well as those with decompensated cirrhosis, administration of sofosbuvir and voxilaprevir, respectively, was safe.
      • Papaluca T.
      • Roberts S.K.
      • Strasser S.I.
      • Stuart K.A.
      • Farrell G.
      • MacQuillan G.
      • et al.
      Efficacy and safety of sofosbuvir/velpatasvir/voxilaprevir for HCV NS5A-inhibitor experienced patients with difficult to cure characteristics.
      ,
      • Borgia S.M.
      • Dearden J.
      • Yoshida E.M.
      • Shafran S.D.
      • Brown A.
      • Ben-Ari Z.
      • et al.
      Sofosbuvir/velpatasvir for 12 weeks in hepatitis C virus-infected patients with end-stage renal disease undergoing dialysis.
      Due to the potential liver toxicity of PIs in patients with decompensated cirrhosis, a lead-in phase with SOF plus a NS5A inhibitor may be considered to downstage patients to Child-Pugh class A cirrhosis before starting VOX/VEL/SOF.
      • Papaluca T.
      • Roberts S.K.
      • Strasser S.I.
      • Stuart K.A.
      • Farrell G.
      • MacQuillan G.
      • et al.
      Efficacy and safety of sofosbuvir/velpatasvir/voxilaprevir for HCV NS5A-inhibitor experienced patients with difficult to cure characteristics.
      Table 3SVR rates after VOX/VEL/SOF with and without ribavirin as rescue treatment after DAA failure.
      StudyRescue treatmentPatients, nCirrhosis (%)SVR allSVR cirrhosisSVR HCV GT3aSVR VEL/SOF failure
      Bourliere et al.VOX/VEL/SOF26346%96%93%95%no data
      Degasperi et al.VOX/VEL/SOF±R16944%96%91%92%94%
      Llaneras et al.VOX/VEL/SOF13534%95%89%80%no data
      Belperio et al.VOX/VEL/SOF55135%91%90%91%83%
      Papaluca et al.VOX/VEL/SOF9178%90%90%89%90%
      Vermehren et al.VOX/VEL/SOF±R10227%98%100%100%100%
      Smith et al.VOX/VEL/SOF14440%91%81%81%74%
      Janjua et al.VOX/VEL/SOF±R19115%95%88%92%89%
      Flamm et al.VOX/VEL/SOF17679%94%no datano data95%
      DAA, direct-acting antiviral; R, ribavirin; SVR, sustained virologic response; VOX/VEL/SOF, voxilaprevir/velpatasvir/sofosbuvir. SVR data from Bourliere et al. based on intention to treat analysis.
      • Bourliere M.
      • Gordon S.C.
      • Flamm S.L.
      • Cooper C.L.
      • Ramji A.
      • Tong M.
      • et al.
      Sofosbuvir, velpatasvir, and voxilaprevir for previously treated HCV infection.
      ,
      • Degasperi E.
      • Spinetti A.
      • Lombardi A.
      • Landonio S.
      • Rossi M.C.
      • Pasulo L.
      • et al.
      Real-life effectiveness and safety of sofosbuvir/velpatasvir/voxilaprevir in hepatitis C patients with previous DAA failure.
      • Llaneras J.
      • Riveiro-Barciela M.
      • Lens S.
      • Diago M.
      • Cachero A.
      • Garcia-Samaniego J.
      • et al.
      Effectiveness and safety of sofosbuvir/velpatasvir/voxilaprevir in patients with chronic hepatitis C previously treated with DAAs.
      • Belperio P.S.
      • Shahoumian T.A.
      • Loomis T.P.
      • Backus L.I.
      Real-world effectiveness of sofosbuvir/velpatasvir/voxilaprevir in 573 direct-acting antiviral experienced hepatitis C patients.
      • Papaluca T.
      • Roberts S.K.
      • Strasser S.I.
      • Stuart K.A.
      • Farrell G.
      • MacQuillan G.
      • et al.
      Efficacy and safety of sofosbuvir/velpatasvir/voxilaprevir for HCV NS5A-inhibitor experienced patients with difficult to cure characteristics.
      • Vermehren J.
      • Serfert Y.
      • Cornberg M.
      • Stoehr A.
      • Klinker H.
      • Simon K.G.
      • et al.
      Sofosbuvir, velpatasvir, and voxilaprevir for patients with failure of previous direct-acting antiviral therapy for chronic hepatitis C: results from the German Hepatitis C-Registry (DHC-R).
      • Smith D.A.
      • Bradshaw D.
      • Mbisa J.
      • Manso C.
      • Bibby D.
      • Singer J.
      • et al.
      Real-world retreatment of HCV-infected patients with prior failure to direct antiviral therapy using sofosbuvir, velpatasvir and voxilaprevir.
      • Janjua N.
      • Wilton J.C.
      • Cook D.
      • Wong S.
      • Butt Z.
      • Barlett S.
      • et al.
      Real-world effectiveness of sofosbuvir/velpatasvir/voxilaprevir as a hepatitis C virus infection salvage treatment.
      • Flamm S.
      • Tsai N.
      • Bacon B.
      • Curry M.
      • Milligan S.
      • Wick N.
      • et al.
      Pangenotypic therapies glecaprevir-pibrentasvir (G-P) and sofosbuvir-velpatasvir-voxilaprevir (S-V-V) after failure with interferon (IFN)-frei direct acting antiviral (DAA) treatment for hepatitis C.
      Patients with failure on G/P were not enrolled in the VOX/VEL/SOF approval study. However, rescue treatment with VOX/VEL/SOF for 12 weeks seems to be highly effective, as shown in a small study in 31 HCV genotype 1- or 3-infected patients, which reported an overall SVR rate of 94% (29/31).
      • Pearlman B.
      • Perrys M.
      • Hinds A.
      Sofosbuvir/velpatasvir/voxilaprevir for previous treatment failures with glecaprevir/pibrentasvir in chronic hepatitis C infection.
      Alternatively, G/P plus sofosbuvir and ribavirin may be an effective retreatment option; an SVR rate of 96% (22/23) was reported in a small clinical study with treatment durations of 12 or 16 weeks.
      • Wyles D.
      • Weiland O.
      • Yao B.
      • Weilert F.
      • Dufour J.F.
      • Gordon S.C.
      • et al.
      Retreatment of patients who failed glecaprevir/pibrentasvir treatment for hepatitis C virus infection.
      Similar to data from the approval study, no single or combined RASs, including the NS5A P32del variant which confers high level resistance to all NS5A inhibitors, were predictive of treatment response to VOX/VEL/SOF.
      • Gottwein J.M.
      • Pham L.V.
      • Mikkelsen L.S.
      • Ghanem L.
      • Ramirez S.
      • Scheel T.K.H.
      • et al.
      Efficacy of NS5A inhibitors against hepatitis C virus genotypes 1-7 and escape variants.
      ,
      • Hikita H.
      • Takehara T.
      NS5A-P32 deletion in hepatitis C genotype 1b infection is the most refractory treatment-mediated amino acid change exhibiting resistance to all NS5A inhibitors.
      Taken together, VOX/VEL/SOF for 12 weeks represents a standard treatment for patients with failure on previous DAA regimens, including G/P. SVR rates of around 90% can be expected. The addition of ribavirin may be considered in difficult to treat patients with HCV genotype 3a, cirrhosis or liver cancer.

      Retreatment after VOX/VEL/SOF failure as rescue treatment

      So far only case reports are published on retreatment in patients with failure on VOX/VEL/SOF rescue therapy, with the exception of 1 larger analysis. In this study, a cohort of 40 VOX/VEL/SOF failure patients were collected in 3 European countries for resistance analysis. Retrospective data on retreatment efficacy was included in a subgroup of patients.
      • Dietz J.
      • Di Maio V.C.
      • de Salazar A.
      • Merino D.
      • Vermehren J.
      • Paolucci S.
      • et al.
      Failure to voxilaprevir, velpatasvir, sofosbuvir and efficacy of rescue therapy.
      As in previous studies, resistance analysis within NS3, NS5A and NS5B genes before and after VOX/VEL/SOF failure showed only minor changes in RAS frequencies and profiles.
      • Dietz J.
      • Di Maio V.C.
      • de Salazar A.
      • Merino D.
      • Vermehren J.
      • Paolucci S.
      • et al.
      Failure to voxilaprevir, velpatasvir, sofosbuvir and efficacy of rescue therapy.
      ,
      • Sarrazin C.
      • Cooper C.L.
      • Manns M.P.
      • Reddy K.R.
      • Kowdley K.V.
      • Roberts S.K.
      • et al.
      No impact of resistance-associated substitutions on the efficacy of sofosbuvir, velpatasvir, and voxilaprevir for 12 weeks in HCV DAA-experienced patients.
      In line with data on characteristics of VOX/VEL/SOF failures, most patients were infected with HCV genotype 3 (45%) and had cirrhosis (70%). Rescue treatments included G/P+SOF with or without ribavirin (n = 15), G/P alone (n = 2), VOX/VEL/SOF with or without ribavirin (n = 4) and VEL/SOF plus ribavirin (n = 1), all for 12 to 24 weeks. SVR was reported in 17/21 patients (81%). Two patients who died of decompensated liver disease and 2 virologic failures (1 each after VEL/SOF plus ribavirin and G/P plus SOF and ribavirin) were reported. No clear association of NS5A RASs with failure on retreatment after VOX/VEL/SOF was observed in this small cohort of patients. Taken together, repetition of multiple targeted therapies with second-generation DAAs like VOX/VEL/SOF or G/P+SOF with or without ribavirin for 12 to 24 weeks is effective after virologic treatment failure on VOX/VEL/SOF rescue treatment. However, more data on baseline characteristics and RAS profiles are required before more specific recommendations can be made for this difficult to treat subgroup of patients (Fig. 4).

      Conclusions

      Virologic resistance is a major cause of failure on direct antiviral therapy. Viral variants, particularly in the NS5A protein, occur naturally and are found in most patients with failure on DAA therapy. For pangenotypic regimens in first-line therapy, RASs play only a minor role. Only in HCV genotype 3a does NS5A resistance lead to lower virologic response rates. This is particularly true for Y93H and VEL/SOF in patients with cirrhosis, and A30K and G/P. For patients after virologic failure on any DAA therapy, a pangenotypic, multiple targeted regimen (VOX/VEL/SOF) is recommended as second-line treatment. In resource limited settings where only first-generation DAA regimens are available, a new antiviral target should be introduced together with sofosbuvir and eventually ribavirin. In addition, treatment duration should be extended. Few data are available for third-line therapy after failure of VOX/VEL/SOF. However, rescue treatment with VOX/VEL/SOF or G/P/SOF with ribavirin seems to be associated with high SVR rates even in this situation.

      Abbreviations

      DAA, direct-acting antiviral; G/P, glecaprevir/pibrentasvir; PI, protease inhibitor; RASs, resistance-associated substitutions; RAVs, resistance-associated viral variants; SOF, sofosbuvir; VEL, velpatasvir; VOX, voxilaprevir.

      Financial support

      CS was supported by a research grant “Deutsches Zentrum für Infektionsforschung (DZIF), TTU Hepatitis”.

      Conflict of interest

      Advisory Committees: Abbvie, Gilead, Merck/MSD,. Grant/Research Support: Abbvie, Gilead, MSD. Speaking and Teaching: Abbvie, Gilead, Merck/MSD.
      Please refer to the accompanying ICMJE disclosure forms for further details.

      Acknowledgements

      The author thanks Dr. Julia Dietz for search of literature and collection of data.

      Supplementary data

      The following is the supplementary data to this article:

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