Review Article: Specifically Targeted Anti-viral Therapy for Hepatitis C – A New Era in Therapy

C. M. Lange; C. Sarrazin; S. Zeuzem


Aliment Pharmacol Ther. 2010;32(1):14-28. 

In This Article

Compounds Targeting HCV Polyprotein Procession

NS3/4A Protease Inhibitors

The design of NS3/4A inhibitors is relatively difficult because the active site of the NS3/4A protease is located in a shallow groove between two β-barrels of the protease.[6,7] Nevertheless, many NS3/4A protease inhibitors are under development, which in general provide a high anti-viral efficacy but a low genetic barrier to resistance. Protease inhibitors can be divided into two chemical classes, macrocyclic inhibitors and linear tetra-peptide α-ketoamid derivatives. NS3/4A protease inhibitors of both classes strongly inhibit HCV replication during monotherapy, but also frequently cause the selection of resistant mutants which may be followed by viral breakthrough.[13,22] However, it was shown that the frequency of resistance development against protease inhibitors can be vastly reduced by the additional administration of pegylated interferon and ribavirin. The most advanced compounds are telaprevir and boceprevir that are currently under phase III evaluation and are expected to be approved in 2011/2012.

Ciluprevir (BILN 2061)

The first NS3/4A inhibitor applied in clinical studies was ciluprevir (BILN 2061), an orally bioavailable, peptidomimetic, macrocyclic drug binding noncovalently to the active centre of the enzyme.[23] Ciluprevir monotherapy was evaluated in a double-blind, placebo-controlled pilot study in treatment-naïve genotype 1 patients with compensated liver disease.[24] In this study, ciluprevir, administered twice daily for 2 days at doses ranging from 25 to 500 mg, led to a mean 2–3 log10 decrease of HCV RNA serum levels in most patients. Another study with equivalent design assessed the influence of the HCV genotype on treatment with this protease inhibitor. Compared with genotype 1 patients, the anti-viral activity of ciluprevir was less pronounced and more variable in patients infected with genotypes 2 or 3.[25] Although the development of ciluprevir was stopped because of serious cardiotoxicity observed in an animal model, these studies provided the proof-of-principle for successful suppression of HCV replication by NS3/4A inhibitors in patients with chronic hepatitis C.

Viral Resistance to Ciluprevir. As a result of the high replication rate of HCV and the poor fidelity of its RNA-dependent RNA polymerase, numerous variants (quasispecies) are continuously produced during HCV replication. Amongst them, variants carrying mutations altering the conformation of the binding sites of STAT-C compounds can develop. During treatment with specific anti-virals, these pre-existing drug-resistant variants have a fitness advantage and can be selected to become the dominant viral quasispecies. Many of these resistant mutants exhibit an attenuated replication fitness with the consequence that, after termination of exposure to specific anti-virals, the wild-type may again replace the resistant variants.[22,26] Nevertheless, HCV quasispecies resistant to NS3/4A protease inhibitors or non-nucleoside polymerase inhibitors can be detected at very low levels in some patients who were never treated with specific anti-virals before.[27–29] The clinical relevance of these pre-existing mutants is not completely understood, although there is evidence that they may reduce the chance to achieve a SVR by therapies based on HCV protease or non-nucleoside polymerase inhibitors.

Exposure of genotype 1 replicon cells to ciluprevir and subsequent sequence analyses of the NS3 region have led to the identification of several mutations conferring ciluprevir-resistance: A156T, R155Q and D168V/A. These mutations result in a 357-fold, 24-fold and 144-fold reduced susceptibility to ciluprevir, respectively, compared with wild-type.[30–32] The A156T mutant confers varying levels of cross-resistance to ciluprevir, telaprevir and boceprevir. The A156T mutation causes a significantly reduced enzymatic function attenuating the HCV life cycle, which, however, can be overcome by additional mutations at P89L, Q86R or G162R.[30–32] No data are available on clinically selected resistance mutations after administration of ciluprevir in patients with chronic hepatitis C.

Telaprevir (VX-950)

Telaprevir is an orally bioavailable NS3 protease inhibitor which belongs to the α-ketoamids and binds the enzyme covalently but reversibly, with a half-life of 58 min of the enzyme-inhibitor complex. Currently, telaprevir is under phase III evaluation (ADVANCE- and ILLUMINATE-Study for treatment-naïve patients, REALIZE study for nonresponders).

Phase I Studies. Telaprevir Monotherapy Study A double-blind, randomized placebo-controlled phase Ib clinical trial evaluating telaprevir monotherapy over 14 days was performed in patients with chronic HCV genotype 1 infection.[33] In this study, anti-viral activity, safety, optimal dosage and pharmacokinetics were assessed in treatment-naïve patients, relapsers or nonresponders to standard treatment. Doses of telaprevir were 450 mg or 750 mg every 8 h or 1250 mg every 12 h. Telaprevir was well tolerated and led to a rapid decline of HCV RNA serum levels in all groups. The best results were obtained in the 750 mg telaprevir q8h dose group with a median reduction of HCV RNA of 4.4 log10 after 14 days of treatment, which is the basis for telaprevir-dosage in most of the following clinical trials. However, viral rebound because of selected mutants occurred in all patients after treatment completion and in some patients even during therapy. The selection of resistant mutants was more frequent in patients who received suboptimal doses.[22]

Telaprevir/pegylated (peg) Interferon α-2a/ribavirin Combination Studies A second phase I study investigated the safety, viral kinetics and the development of telaprevir-resistant mutants of telaprevir monotherapy and in combination with pegIFN-α 2a in treatment-naïve genotype 1 patients.[34] Telaprevir dosage was 750 mg every 8 h after an initial loading dose of 1250 mg and it was administered either alone or in combination with pegIFN-α 2a in comparison to pegIFN-α 2a monotherapy. Treatment was given for 14 days and caused a median reduction of HCV RNA of 1.09 log10 in the pegIFN-α 2a/placebo group, of 3.99 log10 in the telaprevir/placebo group and of 5.49 log10 in the telaprevir/pegIFN-α 2a group at the end of therapy. As observed before, selection of telaprevir-resistant mutants occurred during telaprevir monotherapy. However, their frequency was significantly lower during combination therapy with pegIFN-α 2a and no viral breakthrough was seen during the combination therapy within 14 days.[34]

A parallel study evaluated the safety and efficacy of telaprevir (750 mg every 8 h) in combination with pegIFN-α 2a and weight-based ribavirin in treatment-naive genotype 1 patients for 28 days.[35] At the end of the 28-day treatment period, all patients had undetectable HCV RNA serum levels.

Phase II Studies. Telaprevir and peg Interferon with and without Ribavirin

Studies in treatment naïve patients (PROVE 1 and 2, C208, C209, C210) Larger phase II clinical trials (PROVE 1 and 2) in treatment naïve genotype 1 patients assessed whether with additional telaprevir to pegIFN-α 2a and ribavirin, overall treatment duration can be reduced and/or SVR rates be improved (Figures 2 and 3). PROVE 1 was conducted in the USA whereas PROVE 2 was conducted in Europe. In addition, a study comparing two vs. three times daily administration of telaprevir in combination with either pegylated interferon alfa 2a or 2b (C208) and studies in genotype 2, 3 and 4 infected patients were performed (C209, C210).

Figure 2.

Results of PROVE 1 (USA). Combination therapy of telaprevir (TVR) and pegIFN-α 2a + ribavirin in treatment-naive genotype 1 patients.

Figure 3.

Results of PROVE 2 (Europe). Combination therapy of telaprevir (TVR) and pegIFN-α 2a ± ribavirin in treatment-naive genotype 1 patients.

In PROVE 1, telaprevir, pegIFN-α 2a and ribavirin were administered for 12 weeks in combination, followed by pegIFN-α 2a and ribavirin alone for 0 (n = 17), 12 (n = 79) or 36 (n = 79) weeks in comparison to standard treatment. SVR rates were 35%, 61% and 67%, respectively, compared to 41% with standard treatment. According to the study protocol, treatment was only stopped after 12 or 24 weeks when a rapid virological response (RVR) was achieved. Serious adverse effects led to premature treatment termination in 18% of all subjects treated with telaprevir in contrast to 4% of patients with standard-therapy. Most common adverse events were skin rash, anaemia and gastrointestinal disorders.[15]

The study design of PROVE 2 was similar to PROVE 1 with the main difference being that treatment termination after 12 or 24 weeks was independent of achieving an RVR and one treatment arm was ribavirin-free. The recently published final results showed SVR rates of 36%, 60% and 69% for patients treated with telaprevir plus pegIFN alone for 12 weeks (n = 78), telaprevir and pegIFN and ribavirin for 12 weeks (n = 82), and with telaprevir, pegIFN and ribavirin for 12 weeks followed by 12 weeks of pegIFN plus ribavirin alone (n = 81) respectively. The SVR rate achieved by standard treatment was 46%. However, the rate of relapse in the groups treated for 12 weeks was relatively high with 30% and 48% of all patients who were treated with and without ribavirin respectively. Two patients who discontinued treatment at day 60 and 65 experienced a late relapse 36 and 48 weeks after the end of treatment respectively.[14]

The results of PROVE 1 and 2 indicate that 12 weeks of triple therapy was too short because of the high rate of relapse after treatment completion. Moreover, ribavirin is necessary in therapies with telaprevir to achieve high SVR rates. However, 24 to 48 weeks of total therapy including 12 weeks of triple therapy with telaprevir in addition to standard treatment greatly improved SVR rates in treatment-naïve genotype 1 patients compared with the standard of care. The RVR during triple therapy is an important predictor for treatment success and can be applied for defining individualized treatment durations.

The most important side effects of telaprevir are rash, gastrointestinal disorders and anaemia. Although severe rash may require treatment discontinuation, moderate forms can be treated successfully with topical steroids. The median decline of blood haemoglobin concentration related to telaprevir was approximately 1 g/dL. As telaprevir was administered in most trials for only 12 weeks, the use of erythropoietin-analogues was rarely necessary.

C208 was a small study (n = 161) comparing three times daily 750 mg with two times daily 1125mg telaprevir combined with pegylated interferon alfa 2a or 2b, respectively, and ribavirin.[36] In all four treatment arms comparable SVR rates were observed (81–85%). These high overall SVR rates underline the potential of the triple therapy approach. They are explained in part by experienced study centres with very low discontinuation rates (5%) in comparison with the PROVE studies. In addition, in this study the response-guided therapy approach was investigated. Treatment duration was shortened to 24 weeks in patients who achieved a RVR, while the remaining patients received 48 weeks therapy. Between 80–83% of all patients treated with pegIFN-α 2a, and 67–69% of all patients treated with pegIFN-α 2b achieved an RVR and could therefore be treated for 24 weeks.

As the amino acid sequence of the NS3 protease domain varies significantly between HCV genotypes, protease inhibitors may have a different anti-viral efficacy in patients infected with different genotypes. Like ciluprevir, telaprevir alone or in combination with pegIFN and ribavirin was less effective in treatment-naïve patients infected with other genotypes than genotype 1. For HCV genotype 2, a somewhat weaker anti-viral activity in comparison with HCV genotype 1 was observed with a mean viral decline of 3.9 log10 IU/mL during 14 days of monotherapy with telaprevir. In genotype 3 and 4 infected patients, no significant anti-viral activity was detectable (0.5–0.9 log10 decline).[37,38]

Studies in Nonresponders and Relapsers (PROVE 3) The PROVE 3 trial was conducted to determine SVR rates of treatment with telaprevir in combination with pegIFN-α and ribavirin in treatment-experienced patients (Figure 4). Telaprevir was administered in combination with pegIFN-α 2a with and without ribavirin for 12 to 24 weeks followed by pegIFN-α 2a and ribavirin alone for up to 24 weeks. Retreatment of previous nonresponders with 12 weeks of triple therapy followed by 12 weeks of standard treatment led to a SVR rate of 51% (69% relapser, 39% nonresponder), which is significantly higher compared with SVR rates achieved with the standard of care (14%). Retreatment of nonresponders with 24 weeks of triple therapy followed by 24 weeks of standard treatment led to a SVR rate of 53% (76% relapser, 38% nonresponder) and retreatment of nonresponders with 24 weeks of telaprevir and pegIFN-α 2a without ribavirin followed by 24 weeks of pegIFN-α 2a alone led to a SVR rate of only 24% (42% relapser, 11% nonresponder). The latter result indicates that ribavirin is required for a successful treatment of nonresponders with telaprevir. As in the PROVE 1 and 2 studies viral breakthrough was observed more frequently in patients infected with genotype 1a than in patients infected with genotype 1b.[39]

Figure 4.

Results of PROVE 3. Combination therapy of telaprevir (TVR) and pegIFN-α 2a ± ribavirin in HCV genotype 1 patients with prior nonresponse or relapse to standard treatment.

Phase III Studies. Design of Phase III Clinical Trials: Telaprevir with Pegylated Interferon-alfa and Ribavirin Phase III clinical trials evaluating telaprevir in combination with pegIFN-α and ribavirin have been initiated. The ADVANCE trial enrolled more than 1000 treatment-naïve HCV genotype 1 patients to evaluate 24 weeks of telaprevir-based therapy. Telaprevir was dosed at 750 mg every 8 h and given for 8 or 12 weeks in combination with pegIFN-α 2a and ribavirin followed by pegIFN-α 2a and ribavirin alone until treatment week 24. Patients who did not achieve an RVR were treated with pegIFN-α 2a and ribavirin until week 48. In the ILLUMINATE trial, telaprevir was given for 12 weeks in combination with pegIFN-α 2a and ribavirin followed by pegIFN-α 2a and ribavirin alone until treatment week 24 or 28. The aim of the ILLUMINATE trial is to assess whether treatment extension beyond 24 weeks of total therapy improves SVR rates in patients with RVR or EVR. The REALIZE study enrolled more than 650 patients with prior failure to standard treatment. PegIFN-α 2a and ribavirin were given for 48 weeks including 12 weeks of telaprevir at a dose of 750 mg every 8 h. In one treatment arm, telaprevir treatment was initiated after a 4 week lead-in phase of pegIFN-α 2a and ribavirin alone. SVR data of the ADVANCE, ILLUMINATE and REALIZE study are expected to be published in 2010.

Viral Resistance to Telaprevir To date, mutations conferring telaprevir-resistance have been identified at four positions, V36A/M/L, T54A, R155K/M/S/T and A156S//T,[22,30,31,40] see Table 2 and Table 3 . The A156 mutation was shown by in vitro analyses in the replicon assay while the other mutations could only be detected in vivo by a clonal sequencing approach during telaprevir administration in patients with chronic hepatitis C. A detailed kinetic analysis of telaprevir-resistant variants was performed in genotype 1 patients during 14 days of telaprevir monotherapy and combination therapy with pegIFN-α 2a.[41] Telaprevir monotherapy initially led to a rapid HCV RNA decline in all patients as a result of a strong reduction in wild-type virus. In patients who developed a viral rebound during telaprevir monotherapy, mainly the single mutation variants R155K/T and A156/T were uncovered by wild-type reduction and became dominant after day 8. These single mutation variants were selected from pre-existing quasispecies. During the viral rebound phase these variants typically were replaced by highly resistant double-mutation variants (e.g., V36M/A +R155K/T). The combination of telaprevir and pegIFN-α 2a was sufficient to inhibit the breakthrough of resistant mutations in a 14-day study. It is important to note that after up to 3 years after telaprevir treatment low to medium levels of V36 and R155 variants were still observed in single patients.[42] Another study modelling the dynamics of wild type HCV genotype 1 in patients treated with telaprevir with and without pegylated interferon-alfa and ribavirin showed a first and second phase reduction in virus decline which was up to 10-fold stronger than reported for the standard of care.[43]

As shown for other NS3/4A protease inhibitors as well (e.g. ITMN-191), the genetic barrier to telaprevir resistance differs significantly between HCV subtypes. In all clinical studies of telaprevir alone or in combination with pegIFN-α and ribavirin, viral resistance and breakthrough occurred much more frequently in patients infected with HCV genotype 1a compared with genotype 1b. This difference was shown to result from nucleotide differences at position 155 in HCV subtype 1a (aga, encodes R) vs. 1b (cga, also encodes R). The mutation most frequently associated with resistance to telaprevir is R155K; changing R to K at position 155 requires 1 nucleotide change in HCV subtype 1a and 2 nucleotide changes in subtype 1b isolates.[44]

Boceprevir (SCH 503034)

Boceprevir is another novel peptidomimetic orally bioavailable α-ketoamid HCV protease inhibitor that forms a covalent but reversible complex with the NS3 protein.[45] Like telaprevir, boceprevir is currently in phase III evaluation.

Phase I Studies. Boceprevir Monotherapy Study An initial phase I trial evaluated safety, tolerability and anti-viral efficacy of boceprevir monotherapy (100 to 400 mg daily) in HCV genotype 1 patients with prior failure to standard therapy.[46] After the 14-day treatment period, a mean log10 reduction in HCV RNA load of 2.06 was achieved in patients treated with 400 mg boceprevir daily. Boceprevir was well tolerated at all doses without significant adverse effects. However, viral breakthrough with selection of resistant variants occurred in some patients with a frequency depending on boceprevir dosage.[47]

Boceprevir/peg Interferon α-2b Combination Study A subsequent phase Ib study evaluated the combination of boceprevir and pegIFN-α 2b in HCV genotype 1-infected nonresponders to standard therapy.[48] In this randomized, double-blind crossover study, boceprevir was administered at doses of 200 or 400 mg every 8 h either alone for 7 days or in combination with pegIFN-α 2b for 14 days in comparison to 14 days of pegIFN-α 2b monotherapy. As HCV genotype 1 nonresponders to standard treatment are heterogeneous, the study design intended each patient to receive boceprevir alone, in combination with pegIFN-α 2b and pegIFN-α 2b alone with washout-periods in between in a randomized crossover sequence. Mean maximum reductions in HCV RNA load were 2.45 and 2.88 log10 for boceprevir 200 mg and 400 mg plus pegIFN-α 2b, 1.08 and 1.61 log10 for boceprevir monotherapy and 1.08 and 1.26 log10 for pegIFN-α 2b monotherapy. Boceprevir was well-tolerated alone and in combination with pegIFN-α 2b. Viral breakthrough resulting from selection of pre-existing resistant mutants was observed in some patients, in particular during boceprevir monotherapy.[49]

Phase II Studies. Boceprevir and peg Interferon with and without Ribavirin

Treatment Naïve Phase II Study (SPRINT-1) The aim of the SPRINT 1 trial was to investigate safety, tolerability and anti-viral efficacy of boceprevir (800 mg three times a day) in combination with pegIFN-α 2b and ribavirin in treatment-naïve HCV genotype 1 patients.[16] Treatment with boceprevir in combination with pegIFN-α 2b and ribavirin was either performed continuously for 28 or 48 weeks or for 24 or 44 weeks after a previous 4-week lead-in phase of pegIFN-α 2b and ribavirin alone. The lead-in design was chosen to determine a potential benefit of pre-treatment with pegIFN-α 2b and ribavirin on avoiding resistance development. The control group was treated with pegIFN-α 2b and ribavirin for 48 weeks. SVR rates after 28 weeks of triple treatment were 54% and 56% after 24 weeks with an additional 4 weeks of pre-treatment lead in with pegIFN-α2 and ribavirin (Figure 5). SVR rates after 48 weeks of triple treatment were 67% and 75% after 44 weeks with an additional 4 weeks of pre-treatment lead in with pegIFN-α 2b and ribavirin. After 4 weeks triple therapy with boceprevir, pegIFN and ribavirin 38% of patients achieved an RVR. The most common side-effects related to boceprevir were anaemia, nausea, vomiting and dysgeusia. In general, SPRINT-1 has proven a higher anti-viral efficacy of combination therapy with boceprevir in comparison to the standard of care with slightly better results after a 4 week lead-in phase. However, RVR rates of only 38% during boceprevir triple therapy indicate that boceprevir is potentially less potent than telaprevir which, during triple therapy with pegIFN-α 2b, lead to an RVR rate of approximately 70%.

Figure 5.

Results of SPRINT-1. Combination therapy of boceprevir and pegIFN-α 2b + ribavirin (RBV) in treatment-naive genotype 1 patients.

Studies in Nonresponders and Relapsers In a complex study of HCV genotype 1 nonresponders, the addition of boceprevir to pegIFN-α 2b and ribavirin resulted in only slightly increased SVR rates compared with standard treatment (14% vs. 2%).[50]

Design of Phase III Studies. A phase III clinical trial (SPRINT-2) evaluating boceprevir in treatment-naïve patients was initiated recently and has enrolled more than 1000 patients. Equivalent to the SPRINT-1 study design, patients receive 800 mg boceprevir three times daily in combination with pegIFN-α 2b and weight based ribavirin for 28 or 48 weeks. RESPOND-2 evaluates boceprevir in combination with pegIFN-α 2b and ribavirin at the same doses but for 36 and 48 weeks in relapsers and nonresponders. In all investigational arms a lead-in strategy with pegIFN-α 2b and ribavirin is followed.

Viral Resistance to Boceprevir. In the replicon system, mutations at three positions conferring boceprevir resistance were discovered (Table 3). T54A, A156S and V170A confer low level resistance to boceprevir whereas A156T that also confers telaprevir and ciluprevir resistance exhibited greater levels of resistance.[26] In patients with chronic hepatitis C three additional mutations were detected during boceprevir monotherapy (V36G/M/A, V55A, R155K).[47] In a number of these patients one year and in single patients even 4 years after the end of boceprevir treatment still resistant variants were detected in the HCV quasispecies by clonal sequence analysis.[51] However, an additional study revealed that the anti-viral activity of boceprevir was not impaired in patients who were treated with boceprevir with and without pegIFN-α before.[49]

Other NS3 Protease Inhibitors

Other NS3 protease inhibitors are currently in phase 1–2 development (R7227/ITMN191, MK7009, BI201335, TMC435350, SCH900518, BMS-650032, PHX1766, ACH-1625).[13,52,53] In general, they exhibit a high anti-viral activity in HCV genotype 1 patients, comparable with telaprevir and boceprevir (Figure 6). Triple therapy studies for a number of compounds have been initiated and confirm that resistance development is significantly reduced by combination with pegylated interferon and ribavirin. Whereas linear tetrapeptide and macrocyclic inhibitors do not differ in general with respect to their anti-viral activity, their resistance profile differs significantly. However, R155 is an overlapping position for resistance and different mutations at this amino acid site within the NS3 protease confer resistance to all protease inhibitors which are currently in advanced clinical development.[13]

Figure 6.

Anti-viral activity of NS3/4A protease inhibitors during monotherapy for 3–14 days (modified from [13]).

NS4A Inhibitors

ACH-806. NS4A is a crucial cofactor of NS3, mandatory for proper folding of the protease and capable to enhance the enzymatic activity of NS3 manifold. ACH-806 targets NS4A and therefore inhibits the NS3/4A protease by a different mechanism than peptidomimetic NS3 inhibitors. ACH-806 binds to newly synthesized NS4A molecules, which leads to the blockade of their assembly with NS3 proteins. A phase Ib trial in HCV genotype 1-infected patients demonstrated that ACH-806 has a significant inhibitory impact on HCV replication.[54] Although the development of ACH-806 was halted as a result of reversible serum creatinine elevations, the concept of NS4A inhibition was proven. Importantly, no cross-resistance between ACH-806 and peptidomimetic NS3/4A protease inhibitors was observed in vitro.[55,56] Novel NS4A inhibitors (e.g. ACH-1095) are currently under preclinical development.


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