Clinical relevance of detectable but not quantifiable hepatitis C virus RNA during boceprevir or telaprevir treatment

Authors

  • Patrick R. Harrington,

    Corresponding author
    1. Division of Antiviral Products, Office of Antimicrobial ProductsCenter for Drug Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, MD
    • FDA/CDER/OAP/DAVP, 10903 New Hampshire Ave., Silver Spring, MD 20993===

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    • fax: 301-796-9883

  • Wen Zeng,

    1. Division of Biometrics IV, Office of Biometrics, Office of Translational Sciences; Center for Drug Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, MD
    Search for more papers by this author
  • Lisa K. Naeger

    Corresponding author
    1. Division of Antiviral Products, Office of Antimicrobial ProductsCenter for Drug Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, MD
    • FDA/CDER/OAP/DAVP, 10903 New Hampshire Ave., Silver Spring, MD 20993===

    Search for more papers by this author
    • fax: 301-796-9883


  • Potential conflict of interest: Nothing to report.

Abstract

Boceprevir- and telaprevir-based treatments for chronic hepatitis C virus (HCV) infection use specific response-guided therapy (RGT) guidelines. Eligibility for shortened treatment duration is based on achieving undetectable HCV RNA early during treatment. It is unclear whether a detected HCV RNA level that is below the assay lower limit of quantitation (detectable/BLOQ) is comparable to an undetectable HCV RNA level for RGT decision making. We analyzed data from boceprevir and telaprevir clinical trials to obtain a comprehensive understanding of the frequency and clinical relevance of detectable/BLOQ HCV RNA measurements. In Phase 3 trials P05216 (boceprevir), C216 (telaprevir), and 108 (telaprevir), detectable/BLOQ levels were reported for approximately 10%-20% of all on-treatment HCV RNA measurements. In P05216 and C216, subjects with detectable/BLOQ HCV RNA, on average, had a reduced sustained virologic response (SVR) rate compared with subjects with undetectable HCV RNA at the same on-treatment timepoint. At key RGT timepoints (week 8 for boceprevir, week 4 for telaprevir), subjects with detectable/BLOQ HCV RNA had an approximately 20% lower SVR rate compared with subjects with undetectable HCV RNA, and this difference widened for later on-treatment timepoints. A similar trend was observed for Study 108, but the differences in SVR rates were modest, potentially explained by a higher frequency of reported detectable/BLOQ results. Analyses of Phase 2 boceprevir and telaprevir trials indicated subjects with detectable/BLOQ HCV RNA at RGT timepoints benefited from extended treatment duration. Conclusion: During boceprevir- and telaprevir-based treatment, subjects with detectable/BLOQ HCV RNA had a reduced virologic response compared with subjects with undetectable HCV RNA. Eligibility for shortened treatment duration should be based on achieving undetectable HCV RNA (i.e., HCV RNA not detected) at RGT decision timepoints. (Hepatology 2012)

Analysis of hepatitis C virus (HCV) RNA levels in plasma or serum is critical for assessing the efficacy of antiviral therapy for chronic HCV infection. The primary goal of anti-HCV therapy is achievement of a sustained virologic response (SVR), traditionally defined as undetectable serum or plasma HCV RNA 24 weeks following completion of treatment. The achievement of SVR is generally interpreted as a viral “cure” and is considered a validated surrogate of clinical efficacy because it predicts long-term clinical benefit.1, 2

The use of on-treatment HCV RNA measurements to guide treatment duration, termed response-guided therapy (RGT), has become a key component of patient management.3–5 During HCV treatment a rapid HCV RNA decline may justify a shorter treatment duration without significantly compromising efficacy. On the other hand, a slow HCV RNA decline may warrant an extended duration of treatment to maximize the chances of achieving SVR, and little or no HCV RNA decline may warrant early treatment cessation due to futility. Although a validated RGT approach may make patient management more complex, it ultimately results in a personalized treatment regimen that is optimized for safety and efficacy.

A new standard-of-care for the treatment of chronic HCV genotype 1 infection has emerged with the recent regulatory approval of the HCV NS3/4A protease inhibitors boceprevir and telaprevir. These direct-acting antiviral (DAA) drugs, each to be used in combination with pegylated interferon-alpha plus ribavirin (Peg-IFNα/RBV), significantly improve SVR rates versus Peg-IFNα/RBV alone.6–11 One of the benefits with the use of telaprevir or boceprevir is the opportunity to shorten the total duration of therapy from 48 weeks to 24-36 weeks in a large proportion of patients with HCV genotype 1 infection without compromising treatment efficacy. Treatment duration for each drug is based on multifaceted, clinically validated RGT approaches that are not only unique to each drug, but are also unique to specific patient populations using the specific drug.12, 13 Although the opportunities for greater treatment efficacy and shorter treatment duration provided by these drugs are welcomed, the complex nature of the RGT decision rules raises concerns about the potential for misinterpretation, which could result in suboptimal patient care. Therefore, providing adequate justification for RGT decision rules, and minimizing any potential sources of confusion, may help optimize the use of these drugs in clinical practice.

The boceprevir and telaprevir Phase 3 trials used the Roche COBAS TaqMan HCV 2.0 assay. For these trials the lower limit of plasma HCV RNA quantitation (LLOQ) was considered 25 international units per mL (IU/mL), and the limit of detection (LOD) was considered 9.3-10 IU/mL. Figure 1 summarizes our basic interpretation of HCV RNA results based on a hypothetical and optimally performing quantitative HCV RNA assay with an LLOQ of 25 IU/mL and LOD of 10 IU/mL. In general, there are three different qualitative levels of HCV RNA that are reported using this assay: Quantifiable, reported as a specific IU/mL number that is ≥25; detectable but below the LLOQ (detectable/BLOQ), reported as “<25 IU/mL” or “HCV RNA detected <25 IU/mL”; or undetectable, reported as “HCV RNA not detected” or “target not detected.” The LLOQ is the lowest HCV RNA level that is within the linear and analytically acceptable range of the assay, and the LOD represents the lowest HCV RNA level that is detected ≥95% of the time. The limit of blank (LOB, considered 1 IU/mL in this example) is determined by testing of blank (i.e., non-HCV) samples, and represents the minimum assay readout cutoff for calling an HCV RNA level “detected.” A common misconception is that the assay LOD is the minimum actual HCV RNA level for any result reported as “detected.” Considering that the LOD represents the lowest actual HCV RNA level with a ≥95% detection rate, HCV RNA levels that equal the LOD theoretically should not be detected <5% of the time. Therefore, detectable/BLOQ HCV RNA does not imply the actual HCV RNA concentration of the sample must be between 10 and 25 IU/mL. Actual HCV RNA levels between the LOD and LOB, and even those that fall below the LOB, can still be reported as “detected” by the assay at a given rate as a function of the HCV RNA level. In other words, undetectable and detectable HCV RNA levels are never differentiated by a single theoretical threshold, even for an optimally performing assay.

Figure 1.

Summary of different qualitative levels of HCV RNA concentrations during anti-HCV therapy based on a hypothetical, optimally performing HCV RNA assay. For the purpose of this illustration, we assume that SVR represents a virologic cure, and “Actual Viral RNA” level accurately reflects infectious virus. LLOQ, lower limit of quantitation; LOD, limit of detection; LOB, limit of blank.

In boceprevir and telaprevir trials that included RGT approaches, eligibility for shortened treatment duration was based on achieving an undetectable HCV RNA level (i.e., HCV RNA not detected) at specific treatment timepoints. The trials were not designed to assess RGT using a <LLOQ cutoff. As a result, RGT guidelines described in the boceprevir and telaprevir prescribing information are based on achieving undetectable HCV RNA at specific timepoints.12, 13 There is a general uncertainty about whether an on-treatment HCV RNA level reported as detectable/BLOQ differs clinically from an undetectable HCV RNA level. Clinicians prescribing boceprevir or telaprevir may find it confusing when confronted with detectable/BLOQ HCV RNA measurements, particularly when deciding whether a patient's virologic response meets the criteria for shortened treatment duration. Understanding the clinical relevance of on-treatment detectable/BLOQ HCV RNA measurements with respect to treatment efficacy (i.e., SVR) can provide insight regarding the potential impact of considering an on-treatment detectable/BLOQ measurement equivalent to an undetectable measurement for the purposes of RGT decisions.

This report summarizes analyses of on-treatment and follow-up HCV RNA results from selected Phase 2 and Phase 3 boceprevir and telaprevir clinical trials. These analyses were conducted to obtain a more detailed understanding of the frequency and clinical relevance of HCV RNA measures reported as detectable/BLOQ during treatment. Our analyses revealed that HCV RNA measures reported as detectable/BLOQ were common during treatment, and tended to peak in their frequency before or near key RGT decision timepoints. Furthermore, subjects with on-treatment detectable/BLOQ HCV RNA consistently had lower SVR rates compared with subjects with undetectable HCV RNA at the same timepoint. These and other analyses described in this report indicate that detectable/BLOQ HCV RNA should not be considered equivalent to undetectable HCV RNA for the purposes of making boceprevir and telaprevir RGT decisions.

Abbreviations

BLOQ, below lower limit of quantitation; DAA, direct-acting antiviral; DS, delayed start; HCV, hepatitis C virus; LOB, limit of blank; LOD, limit of detection; LLOQ, lower limit of quantitation; Peg-IFNα, pegylated interferon alpha; RBV, ribavirin; RGT, response-guided therapy; SVR, sustained virologic response.

Subjects and Methods

We analyzed HCV RNA results that were used for efficacy analysis purposes for selected Phase 2 and Phase 3 clinical trials of boceprevir and telaprevir. The data were submitted to the Food and Drug Administration (FDA) Division of Antiviral Products by Merck & Co. and Vertex Pharmaceuticals as part of the recently approved new drug applications for Victrelis and Incivek, respectively. Detailed clinical trial protocols and efficacy analyses have been described elsewhere.6, 8–11, 14–16 Brief summaries of the Phase 3 trials analyzed for this report are described below.

The Phase 3 boceprevir trial P05216 (SPRINT-2) studied RGT and non-RGT treatment regimens that included boceprevir dosed in combination with Peg-IFNα[-2b]/RBV (PR), as well as a PR control arm, in a treatment-naïve, HCV genotype 1-infected study population. The treatment protocol for both boceprevir arms was identical through week 28. Both arms included a 4-week PR lead-in period prior to addition of boceprevir to the regimen. In the BOC-RGT arm, subjects with undetectable HCV RNA at week 8 through week 24 stopped all treatment at week 28, and all others continued with an additional 20 weeks of boceprevir/PR to week 48. All subjects in the BOC-48 arm received 44 weeks of boceprevir/PR after the 4-week PR lead-in period. Subjects with detectable HCV RNA at week 24 discontinued treatment early due to futility.

The Phase 3 telaprevir Study C216 (REALIZE) included subjects who had failed prior treatment with Peg-IFNα and RBV, including prior null responders, prior partial responders, and prior relapsers. Subjects in the telaprevir arms received 12 weeks of telaprevir in combination with 48 total weeks of PR, with one arm including a 4-week PR lead-in (T12DS [delayed start]/PR48), and the other without the PR lead-in (T12/PR48), compared with a control group (PR48). Subjects were discontinued from telaprevir but continued on PR if they had greater than 100 IU/mL HCV RNA at weeks 4, 6, or 8 for the T12/PR48 arm or weeks 8, 10, or 12 for the T12(DS)/PR arm. Subjects were discontinued from all study drugs if they had less than a 2 log10 IU/mL decrease in HCV RNA from baseline at week 12 (week 16 for DS arm) or had a confirmed detectable HCV RNA measurement at week 24 (week 28 for DS arm).

The Phase 3 telaprevir Study 108 (ADVANCE), which included treatment-naïve subjects, compared telaprevir dosed in combination with PR for either the first 8 weeks (T8/PR) or the first 12 weeks (T12/PR), with a control group (PR48). Subjects who achieved an extended rapid viral response (eRVR, undetectable HCV RNA at weeks 4 and 12), received PR for a total of 24 weeks. Subjects who did not achieve eRVR were to receive PR for 48 weeks. Virologic stopping rules included the following: subjects who had greater than 1,000 IU/mL at week 4 discontinued telaprevir but continued PR; subjects discontinued all study drugs if they had a less than 2 log10 IU/mL decrease in HCV RNA by week 12; subjects with detectable HCV RNA at week 24 stopped PR.

For the analyses described here, SVR was defined as HCV RNA <LLOQ (detectable or undetectable) at follow-up week 24, with follow-up week 12 results carried forward if follow-up week 24 results were not available. The rationale for using <LLOQ for SVR determination is included in the Discussion. Subjects with no data at follow-up week 12 or follow-up week 24 were considered treatment failures. Two subjects from the boceprevir P05216 trial (both in the Peg-IFNα/RBV control arm) were censored because they had highly unusual viral RNA results during treatment and follow-up, which could not be explained biologically or interpreted simply as representing SVR or non-SVR; specifically, both subjects likely failed protocol treatment based on having ≈103 to 105 IU/mL HCV RNA levels at the end of treatment or during the early stage of follow-up, but then had one or more subsequent HCV RNA results of <LLOQ (detectable or undetectable) later during follow-up. For analyses of Phase 2 boceprevir and telaprevir trials, subjects with virologic relapse were defined as those who received the per protocol treatment duration, had undetectable HCV RNA at the end of treatment or last available on-treatment timepoint, and had quantifiable HCV RNA during follow-up. Exact (Clopper-Pearson) confidence intervals for SVR rates according to on-treatment HCV RNA status were calculated using SAS v. 9.2.

Results

We analyzed HCV RNA results from the Phase 3 boceprevir study P05216 (SPRINT-2), and the Phase 3 telaprevir studies C216 (REALIZE) and 108 (ADVANCE). Primary efficacy analyses for these clinical trials have been described in detail elsewhere.6, 8–11 Note that, whereas data from all arms studied in each of these trials are shown in this report, not all of these treatment regimens are currently recommended in the prescribing information. Please see prescribing information for VICTRELIS (boceprevir)12 and INCIVEK (telaprevir)13 for recommended treatment regimens and durations.

For P05216 and C216, HCV RNA results of detectable/BLOQ were relatively common during treatment (Fig. 2). These results tended to peak prior to, or near the key currently recommended RGT decision timepoints: week 8 for boceprevir and week 4 for telaprevir (for the non-lead-in strategy). Based on comparisons between boceprevir arms in P05216, and PR lead-in and non-lead-in telaprevir arms in C216, use of the PR 4-week lead-in period delayed the peak frequency of detectable/BLOQ results by 2 to 4 weeks. For P05216, 52% of all subjects with on-treatment HCV RNA results had at least one such result reported as detectable/BLOQ. For C216, 67% of subjects had at least one HCV RNA result reported as detectable/BLOQ.

Figure 2.

Proportion of subjects with detectable/BLOQ HCV RNA levels during treatment in (A) boceprevir clinical trial P05216, and (B) telaprevir clinical trial C216. Timepoints after week 24 are not shown because a week 24 detectable HCV RNA level futility rule was used in each trial. “DS” (Delayed Start) refers to the C216 arm in which telaprevir was initiated at week 4 following the 4-week Peg-IFNα/RBV (PR) lead-in period, similar to the boceprevir arms in P05216.

Retrospective analyses of P05216 and C216 were conducted to assess the relationship between HCV RNA qualitative results during treatment and SVR rates. For all arms in P05216, subjects with on-treatment HCV RNA results of detectable/BLOQ generally had a reduced SVR rate compared with subjects with undetectable HCV RNA at the same timepoint (Fig. 3). For the P05216 boceprevir arms, lower SVR rates for subjects with on-treatment detectable/BLOQ versus undetectable HCV RNA results began around week 6, 2 weeks prior to the RGT decision timepoint for boceprevir, and became more apparent later during treatment. Notably, most subjects with undetectable HCV RNA at week 8 in the BOC-RGT arm received the abbreviated, 28-week treatment duration, per protocol. Thus, for this arm, subjects with detectable/BLOQ HCV RNA at week 8 received a longer treatment duration compared with those with undetectable HCV RNA at week 8, yet still had an ≈24% lower SVR rate.

Figure 3.

SVR rates according to on-treatment HCV RNA status in boceprevir clinical trial P05216. Shown are %SVR rates (y-axis) for subjects with different qualitative levels of HCV RNA at specific timepoints during treatment (x-axis), for each treatment arm. WK, week. Error bars indicate the upper bound of the 95% confidence interval for the calculated SVR rates.

In C216, subjects in the T12/PR48 arm with detectable/BLOQ HCV RNA at week 4 had an ≈22% lower SVR rate compared with subjects with undetectable HCV RNA at week 4, and this difference continued to widen for subsequent timepoints (Fig. 4). Even among T12/PR48 arm subjects with undetectable HCV RNA at week 12, those with detectable/BLOQ HCV RNA at week 4 had a 14% lower SVR rate compared with those with undetectable HCV RNA at week 4 (data not shown). In the T12(DS)/PR48 arm, subjects with detectable/BLOQ HCV RNA at week 8 had an ≈40% lower SVR rate compared with subjects with undetectable HCV RNA at week 8.

Figure 4.

SVR rates according to on-treatment HCV RNA status in telaprevir clinical trial C216. Shown are %SVR rates (y-axis) for subjects with different qualitative levels of HCV RNA at specific timepoints during treatment (x-axis), for each treatment arm. WK, week. Error bars indicate the upper bound of the 95% confidence interval for the calculated SVR rates.

Taken together, these analyses of P05216 and C216 demonstrated that subjects with on-treatment HCV RNA results of detectable/BLOQ had a reduced SVR rate compared with subjects with undetectable HCV RNA at the same timepoint. These results were consistent across both trials, and also across different treatment arms within each trial. As expected, SVR rates were low for subjects with quantifiable HCV RNA during treatment, particularly at later timepoints.

Longitudinal HCV RNA analyses of P05216 confirmed the observations from the above cross-sectional analyses. A detectable/BLOQ HCV RNA level represented a viral load “transition phase” that was often followed by undetectable HCV RNA, or in some cases quantifiable HCV RNA for subjects experiencing virologic breakthrough (Fig. 5A). Across different on-treatment timepoints, subjects with declining HCV RNA levels that transitioned through the detectable/BLOQ phase prior to reaching undetectable tended to have reduced SVR rates compared with subjects whose HCV RNA levels reached undetectable just a single timepoint earlier (Fig. 5B). These results are consistent with undetectable HCV RNA reflecting a qualitatively lower HCV viral load, on average, compared with detectable/BLOQ HCV RNA.

Figure 5.

Detectable/BLOQ HCV RNA is a transition phase (boceprevir P05216 trial). (A) Subjects with detectable/BLOQ HCV RNA at one timepoint often have a different qualitative HCV RNA result at the very next timepoint (*excludes subjects without available data at next timepoint). (B) SVR rates among subjects whose HCV RNA transitioned during treatment through the detectable/BLOQ level prior to becoming undetectable, versus those whose HCV RNA became undetectable one timepoint earlier.

The effect of a shortened treatment duration based on achieving a detectable/BLOQ versus undetectable HCV RNA level at an RGT decision timepoint cannot be assessed directly using data from boceprevir and telaprevir Phase 3 clinical trials, as subjects in RGT arms were required to achieve undetectable HCV RNA to receive a shortened treatment duration. However, this question can be partially addressed using data from the Phase 2 boceprevir trial P03523 (SPRINT-1),14 and the Phase 2 telaprevir trials 104 and 104EU (PROVE 1 and PROVE 2).15, 16 In two arms in P03523, a PR 4-week lead-in period was followed by boceprevir dosed with PR for 24 additional weeks in one arm, or 44 weeks in the other arm. Each arm had a small number of subjects with detectable/BLOQ (LLOQ = 30 IU/mL) HCV RNA at week 8. Subjects with detectable/BLOQ HCV RNA at week 8 who received the 48-week duration had a higher SVR rate and lower relapse rate compared with those who received the 28-week treatment duration (Table 1). Similarly, in a pooled analysis of the telaprevir 104/104EU trials, subjects with detectable/BLOQ (LLOQ = 30 IU/mL) HCV RNA at week 4 who received telaprevir plus PR for 12 weeks, followed by PR for an additional 36 weeks, had a higher SVR rate and lower relapse rate compared with those who received telaprevir plus PR for 12 weeks, followed by PR for an additional 12 weeks. In contrast, subjects in these studies with undetectable HCV RNA at the current RGT decision timepoints had relatively high SVR rates and low relapse rates, even with shortened treatment duration. Although the numbers of subjects in these analyses are small, and the performance of the HCV RNA assays differed from the assay used in the Phase 3 trials, these results are consistent with detectable/BLOQ HCV RNA reflecting a reduced virologic response compared with undetectable HCV RNA during treatment.

Table 1. Sustained Virologic Response (SVR) and Relapse Rates According to On-treatment Viral Load Status and Treatment Duration in Boceprevir and Telaprevir Phase 2 Trials
Study, Population, and Treatment ArmSVR RateRelapse Rate
  1. Subjects with no follow-up data, or subjects who stopped treatment prematurely, were censored in calculations of relapse rates. PR, Peg-IFNα/RBV.

Boceprevir Study P03523 (SPRINT-1)
 Detectable/BLOQ HCV RNA at Week 8  
  PR for 4 Weeks, Boceprevir/PR for 24 Weeks5/13 (38%)5/10 (50%)
  PR for 4 Weeks, Boceprevir/PR for 44 Weeks9/12 (75%)0/9 (0%)
 Undetectable HCV RNA at Week 8  
  PR for 4 Weeks, Boceprevir /PR for 24 Weeks53/62 (85%)4/57 (7%)
  PR for 4 Weeks, Boceprevir /PR for 44 Weeks62/66 (94%)0/62 (0%)
Pooled Telaprevir Studies 104, 104EU (PROVE 1,2)
 Detectable/BLOQ HCV RNA at Week 4  
  Telaprevir/PR for 12 weeks, PR for 12 Weeks6/15 (40%)4/9 (44%)
  Telaprevir/PR for 12 weeks, PR for 36 Weeks4/7 (57%)2/6 (33%)
 Undetectable HCV RNA at Week 4  
  Telaprevir/PR for 12 weeks, PR for 12 Weeks89/120 (74%)6/88 (7%)
  Telaprevir/PR for 12 weeks, PR for 36 Weeks49/64 (77%)0/44 (0%)

The frequency of on-treatment detectable/BLOQ HCV RNA results and their association with SVR rates were also analyzed for the Phase 3 telaprevir Study 108. Detectable/BLOQ results in Study 108 also peaked early during treatment, but were reported more often at later timepoints throughout the 108 treatment period compared with C216 and P05216. At weeks 2 and 4, 50% and 23%, respectively, of subjects in the T12/PR arm in Study 108 had detectable/BLOQ HCV RNA results, comparable to the T12/PR48 arm in C216 (Fig. 2). However, a higher frequency of such results for the T12/PR arm in Study 108 (relative to C216) was observed later during treatment, staying close to 10% from weeks 6 to 24 (data not shown).

As in C216 and P05216, subjects in Study 108 with on-treatment HCV RNA results of detectable/BLOQ consistently had a reduced SVR rate compared with subjects with undetectable HCV RNA at the same timepoint (Fig. 6). However, these differences in SVR rates were more modest in Study 108 compared with C216 and P05216, especially at later on-treatment timepoints (Fig. 2).

Figure 6.

SVR rates according to on-treatment HCV RNA status in telaprevir clinical trial 108. Shown are %SVR rates (y-axis) for subjects with different qualitative levels of HCV RNA at specific timepoints during treatment (x-axis), for each treatment arm (PR control arm not shown). WK, week. Error bars indicate the upper bound of the 95% confidence interval for the calculated SVR rates.

During the FDA review of telaprevir, we became aware of an unexpectedly higher rate of detectable/BLOQ results reported for the treatment-free follow-up period for subjects who apparently achieved SVR in Study 108. This trend was consistent across all treatment arms. Furthermore, long-term follow-up analysis of these subjects indicated that the detectable/BLOQ results were usually transient and not reproducible, and SVR was durable. In contrast, among subjects who experienced confirmed virologic relapse, any detectable/BLOQ HCV RNA results occurred early during follow-up, before follow-up week 12, and were subsequently followed by quantifiable HCV RNA measurements often orders of magnitude above the LLOQ. The same HCV RNA assay was used for Studies P05216, C216, and 108, so we are not aware of an obvious, biologically plausible explanation for a higher rate of transient detectable/BLOQ HCV RNA levels during follow-up among SVR subjects in Study 108. However, both P05216 and C216 used the same contract laboratory for HCV RNA analyses, whereas a different contract laboratory was used for Study 108. Differences in assay performance related to the specific laboratory performing the analyses could be a possible explanation of different reporting frequencies of low level, detectable HCV RNA.

As shown in Table 2, among subjects who achieved SVR (based on <LLOQ) in C216 and P05216, reports of detectable (any level) HCV RNA measurements by Vendor A for any timepoint during follow-up were rare. Less than 2% of all SVR-achieving subjects in C216 and P05216 had a detectable HCV RNA result at any point during follow-up, and less than 1% of all follow-up results from SVR-achieving subjects were reported as detectable. All of these detectable HCV RNA measures were either below or near the assay LLOQ.

Table 2. Detectable HCV RNA Results at Any Point During Follow-up Among Subjects Who Achieved a Sustained Virologic Response (SVR) in Clinical Trial P05216, C216, or 108
 Assay Vendor AAssay Vendor B
Among SVR-achieving subjects:P05216 (Boceprevir)C216 (Telaprevir)108 (Telaprevir)
  1. In this analysis, SVR was defined as HCV RNA <LLOQ (i.e., <25 IU/mL) at follow-up week 24. Follow-up week 12 results were carried forward for any subjects missing follow-up week 24 results.

Proportion of follow-up samples with detectable HCV RNA0.6% (16/2582)0.2% (4/1946)9% (221/2456)
Proportion of subjects with detectable HCV RNA1.8% (11/610)0.6% (2/358)24% (172/730)

In contrast to the Vendor A results reported for C216 and P05216, for Study 108, Vendor B reported a 9% frequency (>15- and 45-fold higher than P05216 and C216, respectively) of detectable follow-up HCV RNA among SVR-achieving subjects, representing 24% of all SVR subjects (Table 2). As in C216 and P05216, all of these detectable HCV RNA measures were either below or near the assay LLOQ.

Reanalyses conducted by Vendor A for a subset of Study 108 samples from follow-up and various on-treatment timepoints yielded a reduced frequency of detectable/BLOQ HCV RNA results. The extent of this reduced frequency of detectable/BLOQ results varied by timepoint. For samples reported as detectable/BLOQ by Vendor B, 40% and 70% of those from week 4 and week 12 on-treatment timepoints, respectively, and 92% for follow-up timepoints, were reported by Vendor A as undetectable. Taken together, the higher frequency of follow-up detectable/BLOQ results from SVR subjects reported by Vendor B for Study 108 correlated with the higher frequency of detectable/BLOQ results reported during treatment, and was associated with less difference in SVR rates based on detectable/BLOQ versus undetectable HCV RNA during treatment.

Discussion

Our analyses of boceprevir and telaprevir clinical trials indicate that undetectable and detectable/BLOQ HCV RNA levels during treatment are qualitatively different, and this difference is clinically relevant. An on-treatment HCV RNA level that is detectable/BLOQ is, on average, indicative of a reduced virologic response compared with an HCV RNA level that is undetectable at the same timepoint. The effect of a shortened treatment duration based on achieving a detectable/BLOQ versus undetectable HCV RNA level at an RGT decision timepoint cannot be assessed directly using data from boceprevir and telaprevir Phase 3 trials. However, in Phase 2 trials subjects with detectable/BLOQ HCV RNA at the current RGT decision points (week 8 for boceprevir, week 4 for telaprevir) generally achieved a higher SVR rate and lower relapse rate with an extended duration of treatment.

Based on the totality of data presented here, clinicians prescribing boceprevir or telaprevir are cautioned not to consider detectable/BLOQ HCV RNA equivalent to undetectable HCV RNA for the purpose of shortening treatment duration. Product inserts for both boceprevir and telaprevir state that for the purpose of assessing RGT eligibility, “a confirmed ‘detectable but below limit of quantification’ HCV-RNA result should not be considered equivalent to an ‘undetectable’ HCV-RNA result.”12, 13 Using a nonvalidated, <LLOQ (detected or not detected) cutoff for on-treatment RGT decision making with boceprevir- or telaprevir-based regimens could result in suboptimal SVR rates and elevated relapse rates.

In addition, our analyses of the treatment-free follow-up period for SVR-achieving subjects demonstrated there can be variability in the rate at which detectable/BLOQ results are reported by different contract laboratories using the same HCV RNA assay. Vendor B reported an unexpectedly high rate of detectable HCV RNA results during follow-up among SVR-achieving subjects for telaprevir Study 108, which we hypothesize represents a higher rate of false-positive HCV RNA detection throughout the conduct of the trial. In fact, reanalysis of a subset of Study 108 samples by Vendor A yielded a reduced frequency of detectable/BLOQ results, particularly for follow-up samples from subjects who achieved SVR. Furthermore, the rate at which such follow-up results were reported by Vendor A for the P05216 and C216 trials is comparable to the recognized false-positive detection rate of the assay: ≈1.3%, according to the FDA-approved assay package insert.17 A higher rate of reported detectable/BLOQ results during treatment and follow-up by Vendor B could explain the decreased difference in SVR rates between subjects with on-treatment detectable/BLOQ versus undetectable HCV RNA levels relative to that observed for the P05216 and C216 trials. However, despite the unexpectedly high rate of detectable/BLOQ results during and following treatment in Study 108, SVR rates remained consistently higher for subjects with undetectable HCV RNA at any given timepoint during treatment compared with those with detectable/BLOQ at the same timepoint.

Currently, it is not clear why the detectable/BLOQ reporting frequency varied by contract laboratory, whether such variability still exists, and whether it extends across other laboratories that typically conduct HCV RNA assessments. We cannot confirm the same technical procedures were used by the different laboratories for the Roche COBAS TaqMan HCV 2.0 assay used in the clinical trials. Moreover, this assay was not FDA-approved at the time it was used for the boceprevir and telaprevir Phase 3 clinical trials. It is important that researchers and clinicians are aware of the potential for laboratory-specific variability in assay performance, and its possible impact on clinical trial efficacy analyses and patient management. The analyses described in this report warrant continued monitoring of assay performance issues, with the ultimate goal of improving the consistency of HCV RNA assay performance in clinical trials and in clinical practice.

Based on our analyses of the follow-up period after the end of treatment, we currently interpret transient detectable/BLOQ HCV RNA results during follow-up as likely representing false-positive detection of HCV RNA. There are alternative explanations for such results, such as the presence of circulating, noninfectious HCV RNA, or the presence of a small number of actively infected cells that continue to produce viral RNA while the infection remains controlled by the host immune response.18 However, given our current understanding of HCV infection biology and the effect of anti-HCV treatment,1, 19, 20 we currently do not believe transiently detectable/BLOQ HCV RNA results during follow-up are clinically significant. Therefore, to simplify analyses of SVR rates in the boceprevir and telaprevir Phase 3 trials and to limit unnecessary repeat testing in practice, the FDA used HCV RNA <LLOQ at follow-up week 24 to define SVR.

Going forward, many clinical trials of other HCV DAAs are using the assay LLOQ as the HCV RNA cutoff for RGT decision making, as it represents a more clearly defined and reproducible threshold. This will allow investigators to retrospectively validate using <LLOQ or undetectable measurements for RGT decisions by analyzing SVR and relapse rates for those subjects who met the criteria for shorter treatment duration. If validated, an LLOQ cutoff may be less subjected to sampling error and inconsistencies in assay specificity, resulting in more consistent and optimal patient management in clinical practice. Such analyses will likely be required for each specific treatment regimen that uses an RGT approach, as differences in antiviral potency and durability may influence the frequency and clinical relevance of detectable/BLOQ HCV RNA levels during treatment. Currently, however, based on the analyses presented in this report, use of an LLOQ cutoff for making RGT decisions with telaprevir- and boceprevir-containing regimens may put some subjects at risk of receiving a subtherapeutic treatment duration.

Acknowledgements

The authors thank Drs. Jules O'Rear, Jeff Murray, Debra Birnkrant, Kathleen Whitaker, Marina Kondratovich, and Uwe Scherf, as well as the FDA DAVP review teams for boceprevir and telaprevir, for providing valuable advice during the conduct of our analyses and the writing of the aritcle. The data analyzed for this report were submitted to the FDA by Merck and Vertex Pharmaceuticals as part of the new drug applications for boceprevir and telaprevir, respectively. Disclaimer: The views expressed in this report are those of the authors and do not necessarily represent official policy of the U.S. Food and Drug Administration.

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