Preclinical and clinical antiviral characterization of AB-836, a potent capsid assembly modulator against hepatitis B virus.

HBV capsid assembly modulators (CAMs) target the core protein and inhibit pregenomic RNA encapsidation and viral replication. HBV CAMs also interfere with cccDNA formation during de novo infection, which in turn suppresses transcription and production of HBV antigens. In this report, we describe the antiviral activities of AB-836, a potent and highly selective HBV CAM. AB-836 inhibited viral replication (EC50 = 0.010 µM) in HepDE19 cells, and cccDNA formation (EC50 = 0.18 µM) and HBsAg production (EC50 = 0.20 µM) in HepG2-NTCP cells during de novo infection. AB-836 showed broad genotype coverage, remained active against variants resistant to nucleos(t)ide analogs, and demonstrated improved antiviral potency against core variants resistant to other CAMs. AB-836 also mediated potent inhibition of HBV replication in a hydrodynamic injection mouse model, reducing both serum and liver HBV DNA. In a Phase 1 clinical study, 28 days of once-daily AB-836 oral dosing at 50, 100, and 200 mg resulted in mean serum HBV DNA declines of 2.57, 3.04, and 3.55 log10 IU/mL from baseline, respectively. Neither on-treatment viral rebound nor the emergence of viral resistance was observed during the 28-day treatment period. Furthermore, HBV DNA sequence analysis of baseline samples from the Phase 1 study revealed that 51.4% of the chronic hepatitis B participants contained at least one core polymorphism within the CAM-binding pocket, suggesting that genetic variations exist at this site. While AB-836 was discontinued due to clinical safety findings, data from the preclinical and clinical studies could help inform future optimization of HBV CAMs.

Measuring hepatitis B pgRNA stability using an updated automated HBV pgRNA assay with increased sensitivity.

BACKGROUND: HBV pregenomic RNA (pgRNA) is a circulating biomarker for covalently closed circular DNA activity in HBV-infected individuals and has been studied for treatment efficacy, disease staging, and off-therapy outcomes; however, data on the stability are scarce. Increasing HBV pgRNA assay sensitivity may improve its predictive value and provide additional insights at low viral levels. METHODS: Modifications to a fully automated first (v1) generation HBV pgRNA assay improved sensitivity up to 15-fold over the previous assay. Flexible sample input volumes yielded lower limits of quantitation of 10 and 22 copies/mL for 0.6 and 0.2 mL assays, respectively. Results are standardized to secondary standards that are traceable to the WHO HBV DNA standard, and internal and external controls are included. RESULTS: Comparison between v1 and modified v2 assays showed increased sensitivity from 152 copies/mL with v1 to 10 (0.6 mL) and 22 (0.2 mL) copies/mL with v2, respectively. Quantitated v2 results were indistinguishable from v1, indicating that comparisons can be made to previous studies. Single timepoint treatment-naive blood donors or longitudinal draws from patients with chronic hepatitis B on AB-729, an investigational siRNA therapy, showed improved detection and quantifiable pgRNA with v2 compared with v1. Stability testing demonstrated excellent HBV pgRNA plasma stability after 3 freeze-thaw cycles, for at least 7 days at 25-37 °C and at least 30 days at 4°C, with ≤0.25 Log U/mL decrease. CONCLUSION: HBV pgRNA v2 assays with increased sensitivity and flexible input volumes demonstrated increased detection and quantitation of low viral titer samples. Highly sensitive HBV pgRNA assays may be useful in refining predictive treatment outcomes based on this marker. HBV pgRNA was stable under multiple conditions, which increases the reliability of this marker.

Safety, pharmacokinetics, and antiviral activity of the capsid inhibitor AB-506 from Phase 1 studies in healthy subjects and those with hepatitis B.

AB-506 is a potent, pan-genotypic small molecule capsid inhibitor that inhibits hepatitis B virus (HBV) pregenomic RNA encapsidation. We assessed the safety, pharmacokinetics, and antiviral activity of AB-506 in two randomized, double-blinded Phase 1 studies in healthy subjects (HS) and subjects with chronic HBV infection (CHB). Single ascending and multiple doses of AB-506 or placebo (30-1000 mg or 400 mg daily for 10 days) were assessed in HS. AB-506 or placebo was assessed at either 160 mg or 400 mg daily for 28 days in subjects with CHB. A second follow-up study examined AB-506 or placebo at 400 mg daily for 28 days in 14 Caucasian and 14 East-Asian HS. Twenty-eight days of AB-506 at 160 mg and 400 mg produced mean HBV-DNA declines from baseline of 2.1 log10 IU/ml and 2.8 log10 IU/ml, respectively. Four subjects with CHB (all Asian) had Grade 4 alanine aminotransferase (ALT) elevations (2 at each dose) as HBV DNA was declining; three events led to treatment discontinuation. In the second follow-up study, 2 Asian HS had serious transaminitis events leading to treatment and study termination. No subjects had bilirubin elevations or signs of hepatic decompensation. Conclusion: AB-506 demonstrated mean HBV-DNA declines of >2 log10 ; however, transient but severe ALT flares were observed in 4 Asian subjects with CHB. In the follow-up study in HS, 2 additional Asian HS had Grade 4 flares, suggesting that AB-506 hepatotoxicity contributed to the ALT elevations. The AB-506 development program was terminated because of these findings.

Twenty-year follow-up of an outbreak of hepatitis C in a small rural town of Argentina: The O'Brien Project.

INTRODUCTION AND OBJECTIVES: In 1999, a population-based survey showed a 5.6 % (102/1832) prevalence of HCV infection in O'Brien, a small rural town of Argentina. The aim of this study was to assess the impact of screening, clinical evaluation and antiviral therapy on elimination of HCV after 20 years of follow-up. PATIENTS AND METHODS: HCV+ subjects (n=102) underwent clinical, biochemical and histological evaluation to assess the presence and severity of liver disease. Antiviral therapy included pegylated interferon + ribavirin in 2005 and direct antiviral agents from 2017. RESULTS: All viremic subjects (n=84) had genotype 1b with 90%-97.5% sequence homology scores, suggesting the existence of a common source of infection (use of unsafe injections administered by the same health professional). Liver biopsy (n=55) showed chronic hepatitis in all patients. The prevalence of cirrhosis was 28% overall (29/102) and 34.5% among viremic patients. Sustained virological response (SVR) was obtained in 20/34 (59%) patients treated with interferon. From 2005 to 2017, when oral antivirals became available 37/50 untreated patients died. Median age of this group in 2005 was 67 years. Six interferon non-responders and five naive subjects received direct antiviral agents and all developed SVR. Only 1/31 patient (3.2%) with SVR died and none developed decompensated cirrhosis or HCC. In 2019, a new population-based study showed that the prevalence of HCV in O'Brien decreased 20-fold, from 5.6% to 0.28% (3/1070). CONCLUSIONS: Despite the high mortality rate precluding timely access to direct antiviral agents, the O'Brien Project is a good example of HCV micro-elimination studies.

Pre-existence and Persistence of Resistant Minority Hepatitis C Virus Variants in Genotype 1-Infected Patients Treated With Simeprevir/Peginterferon/Ribavirin.

Background. The pre-existence of minority hepatitis C virus (HCV) variants and their impact on treatment outcome, as well as the persistence of emerging resistant variants posttreatment in patients failing treatment with simeprevir/peginterferon/ribavirin (SMV/PR), were assessed by deep sequencing (DS). Methods. Population sequencing (PS) and Illumina DS were performed on HCV genotype 1 isolates from patients treated with SMV/PR in Phase 2b (PILLAR [NCT00882908] and ASPIRE [NCT00980330]) and Phase 3 (QUEST-1 [NCT01289782], QUEST-2 [NCT01290679], and PROMISE [NCT01281839]) trials. Results. Minority polymorphisms (ie, detected pretreatment by DS only) reducing SMV activity in vitro were uncommon (3.6%, 19 of 534 patients). These SMV-resistant minority polymorphisms were detected in similar proportions of patients achieving (3.7%) and not achieving (3.3%) sustained virologic response with SMV/PR and generally did not emerge as major variants at time of failure. SMV-resistant variants emerging at time of failure were no longer detected at end of study in 69.3% and 52.0% of the patients by PS and DS, respectively. Conclusions. Minority polymorphisms did not impact outcome of SMV/PR treatment. The majority of emerging variants that became undetectable at end of study by PS were also undetectable by DS. These results suggest no added value of DS for clinical usage of SMV.

Daclatasvir plus simeprevir with or without ribavirin for the treatment of chronic hepatitis C virus genotype 1 infection.

BACKGROUND & AIMS: We evaluated the combination of daclatasvir (pan-genotypic NS5A inhibitor) and simeprevir (NS3/4A protease inhibitor), with or without ribavirin, in hepatitis C virus genotype 1-infected patients. METHODS: This phase II, open-label study enrolled treatment-naive patients or prior null responders with genotype 1b (n=147) or 1a (n=21) infection. Genotype 1b-infected patients were randomized 1:1 to receive daclatasvir 30mg plus simeprevir 150mg once daily with or without ribavirin; those who completed the initial 12-week treatment were re-randomized 1:1 to stop treatment or continue treatment through to week 24. Genotype 1a-infected patients received daclatasvir plus simeprevir with ribavirin for 24weeks. The primary endpoint was the proportion of patients with sustained virologic response at posttreatment week 12 (SVR12). RESULTS: For genotype 1b, 84.9% (45/53) and 74.5% (38/51) of treatment-naive patients and 69.6% (16/23) and 95.0% (19/20) of prior null responders to peginterferon and ribavirin achieved SVR12 with daclatasvir plus simeprevir alone and with ribavirin, respectively. Treatment duration did not have a well-defined impact on response. For genotype 1a, daclatasvir plus simeprevir with ribavirin provided a 66.7% (8/12) response rate in treatment-naive patients and was not effective in prior null responders. Data suggest that baseline resistance polymorphisms influenced SVR12 rates. Daclatasvir plus simeprevir was well tolerated with or without ribavirin with low incidences of serious adverse events and adverse events leading to discontinuation. CONCLUSIONS: Daclatasvir plus simeprevir, with or without ribavirin, was effective with a 12- or 24-week duration in genotype 1b-infected patients and was well tolerated. ClinicalTrials.gov identifier: NCT01628692.

Deep sequencing analysis of HIV-1 reverse transcriptase at baseline and time of failure in patients receiving rilpivirine in the phase III studies ECHO and THRIVE.

Minority variants (1.0-25.0%) were evaluated by deep sequencing (DS) at baseline and virological failure (VF) in a selection of antiretroviral treatment-naïve, HIV-1-infected patients from the rilpivirine ECHO/THRIVE phase III studies. Linkage between frequently emerging resistance-associated mutations (RAMs) was determined. DS (llIumina®) and population sequencing (PS) results were available at baseline for 47 VFs and time of failure for 48 VFs; and at baseline for 49 responders matched for baseline characteristics. Minority mutations were accurately detected at frequencies down to 1.2% of the HIV-1 quasispecies. No baseline minority rilpivirine RAMs were detected in VFs; one responder carried 1.9% F227C. Baseline minority mutations associated with resistance to other non-nucleoside reverse transcriptase inhibitors (NNRTIs) were detected in 8/47 VFs (17.0%) and 7/49 responders (14.3%). Baseline minority nucleoside/nucleotide reverse transcriptase inhibitor (NRTI) RAMs M184V and L210W were each detected in one VF (none in responders). At failure, two patients without NNRTI RAMs by PS carried minority rilpivirine RAMs K101E and/or E138K; and five additional patients carried other minority NNRTI RAMs V90I, V106I, V179I, V189I, and Y188H. Overall at failure, minority NNRTI RAMs and NRTI RAMs were found in 29/48 (60.4%) and 16/48 VFs (33.3%), respectively. Linkage analysis showed that E138K and K101E were usually not observed on the same viral genome. In conclusion, baseline minority rilpivirine RAMs and other NNRTI/NRTI RAMs were uncommon in the rilpivirine arm of the ECHO and THRIVE studies. DS at failure showed emerging NNRTI resistant minority variants in seven rilpivirine VFs who had no detectable NNRTI RAMs by PS.

Comparison of three quantitative HCV RNA assays in samples from HCV genotype 1- or 4-infected patients treated with the NS3/4A protease inhibitor simeprevir.

BACKGROUND: Monitoring HCV RNA levels during treatment is an important tool for managing protease-inhibitor-based regimens, and different assays used in clinical practice can impact treatment decisions. OBJECTIVES: The concordance of three HCV RNA assays was determined, and their impact on treatment decisions assessed using samples from HCV genotype (GT) 1- and GT4-infected patients treated with the NS3/4A inhibitor simeprevir in combination with pegylated interferon-α/ribavirin. STUDY DESIGN: Plasma samples collected during the simeprevir Phase III studies QUEST-1 and QUEST-2 (GT1), and RESTORE (GT4) were analyzed with the Roche High-Pure-System COBAS(®) TaqMan(®) HCV v2.0 (HPS), the Roche AmpliPrep COBAS(®) TaqMan(®) HCV v2.0 (CAP), and the Abbott RealTime HCV (ART) assay. RESULTS: In GT1, of the 440 samples, 81% were undetectable (rapid virological response; RVR) by HPS at Week 4, 76% by CAP and 44% by ART. In GT4 (103 samples), RVR rates were 67% by HPS and 24% by ART. HCV RNA <25IU/mL at Week 4 was observed for 95-96% and 92% GT1 samples and 86% and 74% GT4 samples by HPS/CAP and ART, respectively. At Week 12, assay concordance for undetectability was high in GT1 and GT4, (95-98% and 93%, respectively). CONCLUSIONS: While different HCV RNA assays can lead to substantially different RVR rates, a good concordance was observed with a cut-off of 25IU/mL. Sustained virologic response rates among GT1 patients achieving RVR or <25IU/mL at Week 4 were high and similar between assays used. At later time points, when viremia is low, assay concordance was high.

An OPTIMIZE study retrospective analysis for management of telaprevir-treated hepatitis C virus (HCV)-infected patients by use of the Abbott RealTime HCV RNA assay.

Protease inhibitor (PI)-based response-guided triple therapies for hepatitis C virus (HCV) infection are still widely used. Noncirrhotic treatment-naive and prior relapser patients receiving telaprevir-based treatment are eligible for shorter, 24-week total therapy if HCV RNA is undetectable at both weeks 4 and 12. In this study, the concordance in HCV RNA assessments between the Roche High Pure System/Cobas TaqMan and Abbott RealTime HCV RNA assays and the impacts of different HCV RNA cutoffs on treatment outcome were evaluated. A total of 2,629 samples from 663 HCV genotype 1 patients receiving telaprevir/pegylated interferon/ribavirin in OPTIMIZE were analyzed using the High Pure System and reanalyzed using Abbott RealTime (limits of detection, 15.1 IU/ml versus 8.3 IU/ml; limits of quantification, 25 IU/ml versus 12 IU/ml, respectively). Overall, good concordance was observed between the assays. Using undetectable HCV RNA at week 4, 34% of the patients would be eligible for shorter treatment duration with Abbott RealTime versus 72% with the High Pure System. However, using <12 IU/ml for Abbott RealTime, a similar proportion (74%) would be eligible. Of the patients receiving 24-week total therapy, 87% achieved a sustained virologic response with undetectable HCV RNA by the High Pure System or <12 IU/ml by Abbott RealTime; however, 92% of the patients with undetectable HCV RNA by Abbott RealTime achieved a sustained virologic response. Using undetectable HCV RNA as the cutoff, the more sensitive Abbott RealTime assay would identify fewer patients eligible for shorter treatment than the High Pure System. Our data confirm the <12-IU/ml cutoff, as previously established in other studies of the Abbott RealTime assay, to determine eligibility for shortened PI-based HCV treatment. (The study was registered with ClinicalTrials.gov under registration no. NCT01241760.).

Virology analyses of HCV isolates from genotype 1-infected patients treated with simeprevir plus peginterferon/ribavirin in Phase IIb/III studies.

BACKGROUND & AIMS: Simeprevir is an oral hepatitis C virus (HCV) NS3/4A protease inhibitor approved for treatment of chronic HCV infection. Baseline NS3 polymorphisms in all patients and emerging mutations in patients who failed to achieve sustained virologic response (SVR) with simeprevir plus peginterferon/ribavirin (PR) in Phase IIb/III studies are described. METHODS: Baseline sequencing data were available for 2007 genotype 1 (GT1)-infected patients. Post-baseline data were available for 197/245 simeprevir-treated patients who did not achieve SVR. In vitro simeprevir susceptibility was assessed in a transient replicon assay as site-directed mutants or in chimeric replicons with patient-derived NS3 protease sequences. RESULTS: Baseline NS3 polymorphisms at positions associated with reduced in vitro susceptibility to simeprevir (43, 80, 122, 155, 156, and/or 168; EC50 fold change >2.0) were uncommon (1.3% [26/2007]), with the exception of Q80K, which confers ∼10-fold reduction in simeprevir activity in vitro (13.7% [274/2007]; GT1a 29.5% [269/911], GT1b 0.5% [5/1096]). Baseline Q80K had minor effect on initial response to simeprevir/PR, but resulted in lower SVR rates. Overall, 91.4% of simeprevir-treated patients [180/197] without SVR had emerging mutations at NS3 positions 80, 122, 155, and/or 168 at failure (mainly R155K in GT1a with and without Q80K, and D168V in GT1b), conferring high-level resistance in vitro (EC50 fold change >50). Emerging mutations were no longer detectable by population sequencing at study end in 50% [90/180] of patients (median follow-up 28.4weeks). CONCLUSIONS: Simeprevir treatment failure was usually associated with emerging high-level resistance mutations, which became undetectable over time in half of the patients.

Low-density lipoprotein and other predictors of response with telaprevir-based therapy in treatment-experienced HCV genotype 1 patients: REALIZE study.

BACKGROUND & AIMS: Predictors of response to treatment with peginterferon plus ribavirin are well established. In these post-hoc analyses of the REALIZE study, we sought to identify predictors of response for telaprevir-based triple therapy. METHODS: Patients from the REALIZE study with baseline data for all predictors evaluated (including baseline disease characteristics and demographics, prior treatment response and baseline laboratory assessments) were included in the post-hoc analyses (n = 465). Univariate and multivariate analyses were used to evaluate factors predicting treatment outcomes. RESULTS: Sustained viral response (SVR) rates were 86% in prior relapsers, 63% in prior partial responders and 32% in prior null-responders. In the final multivariate analysis, baseline factors predicting SVR were prior response to treatment [Odds ratio (OR) = 2.80; 95% confidence interval (CI), 2.13-3.69], low-density lipoprotein (LDL) (≥2.6 mmol/L) (OR = 2.11; 95% CI, 1.52-2.93), HCV genotype (OR = 0.58; 95% CI, 0.36-0.93), and maximum alanine amino transferase and aspartate amino transferase (OR = 0.62; 95% CI, 0.40-0.97). CONCLUSIONS: Prior response to peginterferon plus ribavirin treatment and LDL levels are the main independent predictive markers of response with telaprevir-based triple therapy.

Simeprevir plus sofosbuvir, with or without ribavirin, to treat chronic infection with hepatitis C virus genotype 1 in non-responders to pegylated interferon and ribavirin and treatment-naive patients: the COSMOS randomised study.

BACKGROUND: Interferon-free regimens are needed to treat hepatitis C virus (HCV) infections. We investigated the efficacy of combined simeprevir and sofosbuvir. METHODS: We enrolled patients with chronic HCV genotype 1 infections who had previously not responded to pegylated interferon (peginterferon) and ribavirin or were treatment naive. Patients were randomly assigned in a 2:1:2:1 ratio to receive 150 mg simeprevir and 400 mg sofosbuvir daily for 24 weeks with (group 1) or without (group 2) ribavirin or for 12 weeks with (group 3) or without (group 4) ribavirin, in two cohorts: previous non-responders with METAVIR scores F0-F2 (cohort 1) and previous non-responders and treatment-naive patients with METAVIR scores F3-F4 (cohort 2). The primary endpoint was sustained virological response 12 weeks after stopping treatment (SVR12). Analysis was done by intention to treat. Safety data from cohorts 1 and 2 were pooled for analysis. This study is registered with ClinicalTrials.gov, number NCT01466790. FINDINGS: 168 patients were enrolled and randomised, and 167 started treatment (n=80 in cohort 1 and n=87 in cohort 2). SVR12 was achieved in 154 (92%) patients (n=72 [90%, 95% CI 81-96] in cohort 1 and n=82 [94%, 87-98] in cohort 2). The most common adverse events in the pooled groups were fatigue (n=52 [31%]), headache (n=33 [20%]), and nausea (n=26 [16%]). Grade 4 adverse events were seen in one (2%) of 54 patients in each of groups 1 and 3 and in three (10%) of 31 patients in group 2, whereas grade 3-4 events were reported in less than 5% of all patients, except increased blood amylase concentration. Serious adverse events were seen in four (2%) patients, all in groups 1 and 2. Four (2%) patients discontinued all study treatment because of adverse events, three before week 12. INTERPRETATION: Combined simeprevir and sofosbuvir was efficacious and well tolerated. FUNDING: Janssen.

Modeling viral evolutionary dynamics after telaprevir-based treatment.

For patients infected with hepatitis C virus (HCV), the combination of the direct-acting antiviral agent telaprevir, pegylated-interferon alfa (Peg-IFN), and ribavirin (RBV) significantly increases the chances of sustained virologic response (SVR) over treatment with Peg-IFN and RBV alone. If patients do not achieve SVR with telaprevir-based treatment, their viral population is often significantly enriched with telaprevir-resistant variants at the end of treatment. We sought to quantify the evolutionary dynamics of these post-treatment resistant variant populations. Previous estimates of these dynamics were limited by analyzing only population sequence data (20% sensitivity, qualitative resistance information) from 388 patients enrolled in Phase 3 clinical studies. Here we add clonal sequence analysis (5% sensitivity, quantitative) for a subset of these patients. We developed a computational model which integrates both the qualitative and quantitative sequence data, and which forms a framework for future analyses of drug resistance. The model was qualified by showing that deep-sequence data (1% sensitivity) from a subset of these patients are consistent with model predictions. When determining the median time for viral populations to revert to 20% resistance in these patients, the model predicts 8.3 (95% CI: 7.6, 8.4) months versus 10.7 (9.9, 12.8) months estimated using solely population sequence data for genotype 1a, and 1.0 (0.0, 1.4) months versus 0.9 (0.0, 2.7) months for genotype 1b. For each individual patient, the time to revert to 20% resistance predicted by the model was typically comparable to or faster than that estimated using solely population sequence data. Furthermore, the model predicts a median of 11.0 and 2.1 months after treatment failure for viral populations to revert to 99% wild-type in patients with HCV genotypes 1a or 1b, respectively. Our modeling approach provides a framework for projecting accurate, quantitative assessment of HCV resistance dynamics from a data set consisting of largely qualitative information.

Virologic characterization of genotype 4 hepatitis C virus variants in patients treated with telaprevir.

BACKGROUND: Study C210 was a Phase IIa, exploratory trial to assess the activity of telaprevir on hepatitis C virus (HCV) early viral kinetics in treatment-naïve patients infected with genotype 4 (G4) HCV. METHODS: Patients were randomized to receive peginterferon and ribavirin alone, telaprevir monotherapy (T arm), or telaprevir in combination with peginterferon/ribavirin (TPR arm) for 15 days, followed by a 46- or 48-week standard treatment phase. The current analysis aimed to characterize the genotype and phenotype of HCV G4 variants emerging during telaprevir treatment. RESULTS: Five of the 8 (62.5%) patients in the telaprevir (T) arm had viral breakthrough (vBT) during the investigational treatment phase (between baseline and Day 15), compared to no patients in the TPR arm. HCV G4 viral variants with a T54A/T mutation were detected in two of these patients, as well as two other patients with detectable HCV RNA at the end of telaprevir treatment. Emergence of the T54A/T mutation was associated with a 2- to 4-fold decreased susceptibility to telaprevir. All patients with vBT during the investigational treatment phase or with a T54A/T mutation achieved undetectable HCV RNA 12 or 24 weeks after end of treatment with subsequent peginterferon/ribavirin treatment. CONCLUSIONS: In this analysis in G4 HCV-infected patients, more patients in the telaprevir monotherapy arm experienced vBT with resistant variants compared to none with telaprevir combination therapy. The most commonly selected mutation T54A in telaprevir-treated G4 HCV patients was previously described in the context of G1 infection. TRIAL REGISTRATION: The trial was registered with ClinicalTrials.gov (NCT00580801).

Deep-sequencing analysis of the gene encoding the hepatitis C virus nonstructural 3-4A protease confirms a low prevalence of telaprevir-resistant variants at baseline and the end of the REALIZE study.

BACKGROUND: Population sequencing (PS) has shown that telaprevir-resistant variants are not typically detectable at baseline (prevalence, ≤5% of patients), and most variants present at the time of treatment failure are no longer detectable at the end of the study. METHODS: To gain insight into the evolution of telaprevir-resistant variants, their baseline prevalence and persistence after treatment was investigated using a more sensitive, deep-sequencing (DS) technique in a large number of treatment-experienced patients from the REALIZE study who were infected with hepatitis C virus genotype 1. RESULTS: Before treatment initiation, telaprevir-resistant variants (T54A, T54S, or R155K in 1%-2% of the viral population) were detected by DS in a fraction (2%) of patients for whom PS failed to detect resistance; these variants were not necessarily detected at the time of treatment failure. Of 49 patients in whom telaprevir-resistant variants were detected by PS at the time of treatment failure but not at the end of the study, DS revealed the presence of variants (V36A/L/M, T54S, or R155K in 1%-36% of the viral population) in 16 patients (33%) at the end of the study. CONCLUSIONS: Similar to PS findings, DS analysis revealed that the frequency of telaprevir-resistant variants before treatment was also low, and variants detected at the time of treatment failure were no longer detectable in the majority of patients during follow-up.

Prevalence in the USA of rilpivirine resistance-associated mutations in clinical samples and effects on phenotypic susceptibility to rilpivirine and etravirine.

BACKGROUND: The prevalence of rilpivirine resistance-associated mutations (RAMs) in the USA, and their effect on phenotypic susceptibility to rilpivirine and etravirine, was evaluated in clinical samples from HIV-1-infected patients. METHODS: In total, 15,991 samples submitted to Monogram Biosciences (South San Francisco, CA, USA) for routine resistance testing between January 2010 and June 2011 were assessed for the presence of known rilpivirine RAMs K101E/P, E138A/G/K/Q/R, V179L, Y181C/I/V, Y188L, H221Y, F227C and M230I/L; non-nucleoside reverse transcriptase inhibitor (NNRTI) RAMs K103N, L100I and L100I+K103N; and the nucleoside reverse transcriptase inhibitor (NRTI) RAMs M184I/V and their combinations with rilpivirine RAMs. Phenotypic susceptibility (PhenoSenseGT(®) assay; Monogram Biosciences) was evaluated, with reduced susceptibility defined as fold change (FC) in 50% inhibitory concentration (IC50)>2.0 for rilpivirine and FC>2.9 for etravirine. RESULTS: Of the 15,991 samples, 17% harboured ≥1 rilpivirine RAMs. The prevalence of most rilpivirine RAMs and combinations of NNRTI RAMs of interest was low (≤3%), except for Y181C (7%). Rilpivirine RAMs were often associated with reduced rilpivirine phenotypic susceptibility. Median FC values >2.0 were observed for clinical isolates with rilpivirine RAMs K101P, E138Q/R, Y181C/I/V, Y188L or M230L, and for the combination of E138K with M184I/V, and K101E with M184I. Most rilpivirine FC values >2.0 were associated with etravirine FC values >2.9 for individual rilpivirine RAMs and those combined with M184I/V. There was no relationship between the presence of K103N and rilpivirine FC. However, the L100I+K103N combination (without rilpivirine RAMs), at <2% prevalence, was associated with a rilpivirine FC>2.0. CONCLUSIONS: Based on 15,991 US clinical samples from HIV-1-infected patients, the frequency of most known rilpivirine RAMs apart from Y181C was low.

HCV RNA quantification with different assays: implications for protease-inhibitor-based response-guided therapy.

BACKGROUND: Response-guided therapy (RGT) for HCV treatment, whereby therapy duration is shortened according to on-treatment virological response, requires patient HCV RNA concentrations below the lower limit of quantification (LLOQ) or limit of detection (LOD) of the viral load assay at weeks 4 and 12. Concordance of two assays and impact on treatment decisions were investigated. METHODS: Plasma samples (n=1,411; baseline to week 12) from HCV genotype-1-infected patients (n=290) receiving simeprevir (TMC435) plus pegylated interferon-α2a/ribavirin in the PILLAR study (NCT00882908) were analysed using Roche High-Pure-System/COBAS(®) TaqMan(®) v2.0 assay (HPS; LLOQ 25 IU/ml and LOD 15 IU/ml; Roche Diagnostics, Indianapolis, IN, USA) and reanalysed using Abbott realtime assay (ART; LLOQ and LOD 12 IU/ml; Abbott Molecular Inc., Des Plaines, IL, USA). RESULTS: Overall, 217/766 (28.3%) samples from different time points with HCV RNA undetectable by HPS had HCV RNA detectable by ART. Conversely, 35/584 (6.0%) samples undetectable by ART were detectable by HPS. For both assays, most discrepant samples (96-100%) had HCV RNA<25 IU/ml. At week 4, 75.5% of samples were undetectable by HPS, whereas 49.4% were undetectable by ART, resulting in different RGT assessment in 26.1% (P<0.0001). At week 12, 95.4% and 91.9% of samples were undetectable with HPS and ART, respectively. CONCLUSIONS: Lower rates of undetectable HCV RNA with ART at week 4 suggest that if RGT criteria are determined with ART, the proportion of patients qualifying for shorter treatment duration may be significantly lower (26%). Therefore, different RGT criteria may be necessary for ART to maximize numbers benefiting from shortened treatment. Further testing and validation are required.

Patterns of viral load decline with telaprevir-based therapy in patients with genotype 1 chronic HCV infection.

BACKGROUND: Telaprevir-based therapy is associated with rapid decline in HCV RNA, enabling the application of early futility rules. OBJECTIVES: To familiarize physicians with this paradigm, a comprehensive analysis of the most frequent HCV viral load profiles observed during treatment with telaprevir/Peg-IFN/RBV in Phase III trials is provided. DESIGN: HCV RNA profiles were analyzed from 320 HCV genotype 1 treatment-naïve patients enrolled in the ADVANCE study, and 225 prior Peg-IFN/RBV treatment-experienced patients enrolled in the REALIZE study. Patients received 12 weeks of telaprevir with either 24 or 48 weeks of Peg-IFN alfa-2a/RBV. Patients with missing SVR assessments during follow-up, detectable HCV RNA at end of treatment but who did not have viral breakthrough (vBT), or with early vBT who discontinued telaprevir before time of failure were excluded. RESULTS: All analyzed patients experienced a rapid decline in HCV RNA (>2.0 log(10)) by Day 14, irrespective of baseline characteristics and/or prior response to Peg-IFN/RBV (relapse, partial response and null response). Subsequently, HCV RNA continued to decline to undetectable levels in most patients. These patients went on to have one of the following outcomes: sustained virologic response, late vBT (after Week 12, i.e. during the Peg-IFN/RBV phase), or relapse. In the small subset of patients with early vBT or meeting a futility rule before Week 12, HCV RNA usually never became undetectable and/or increased rapidly after reaching the nadir. CONCLUSIONS: HCV RNA profiles with telaprevir/Peg-IFN/RBV are different from those with Peg-IFN/RBV alone. It is important that clinicians understand these HCV RNA profiles and monitor patient viral load in order to apply futility rules correctly.

Telaprevir activity in treatment-naive patients infected hepatitis C virus genotype 4: a randomized trial.

BACKGROUND: This partially blinded, randomized, phase 2a C210 study evaluated the antiviral activity of telaprevir-based regimens in treatment-naive patients with chronic hepatitis C virus (HCV) genotype 4 infection. METHODS: Twenty-four patients received telaprevir 750 mg every 8 hours for 15 days (T; n = 8), telaprevir in combination with pegylated interferon alfa-2a and ribavirin (Peg-IFN/RBV) for 15 days (TPR; n = 8), or Peg-IFN/RBV plus placebo for 15 days (PR; n = 8), followed by Peg-IFN/RBV for 46 or 48 weeks. The primary objective was to assess the effect of telaprevir on HCV RNA levels. RESULTS: HCV RNA levels decreased slightly with T and PR; TPR produced substantial, rapid declines. On day 15, median reductions in the HCV RNA load from baseline were -0.77, -4.32, and -1.58 log10 IU/mL for T, TPR, and PR, respectively, and 0 patients in the T group, 1 in the TPR group, and 0 in the PR group had undetectable HCV RNA. Five of 8 patients who received telaprevir monotherapy had viral breakthrough within 15 days of treatment. Adverse event incidence was similar across treatments and comparable with the incidences from previous clinical trials. One patient (in T group) had a serious adverse event (considered unrelated to telaprevir) that led to treatment discontinuation. CONCLUSIONS: Telaprevir with Peg-IFN/RBV had greater activity than Peg-IFN/RBV treatment or telaprevir monotherapy against HCV genotype 4. Telaprevir was generally safe and well tolerated. Further investigation of telaprevir combination therapy in patients with HCV genotype 4 infection is warranted.

96-Week resistance analyses of rilpivirine in treatment-naive, HIV-1-infected adults from the ECHO and THRIVE Phase III trials.

BACKGROUND: In the ECHO/THRIVE 96-week efficacy analysis, the response rate was 78% with rilpivirine (RPV) and efavirenz (EFV) plus two nucleoside/nucleotide reverse transcriptase inhibitors. METHODS: For resistance analyses, virological failures (VFs) were genotyped and/or phenotyped at baseline and failure. RESULTS: In the overall 96-week resistance analyses, the proportion of VFs was higher with RPV (96/686, 14%) versus EFV (52/682, 8%), but similar within weeks 48-96 (22/686, 3% versus 16/682, 2%). In genotyped VFs, treatment-emergent non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance-associated mutations (RAMs) were as common with RPV (46/86, 53%) versus EFV (20/42, 48%), but nucleoside/nucleotide reverse transcriptase inhibitor RAMs were more common with RPV (48/86, 56%) than EFV (11/42, 26%). In RPV VFs, E138K+M184I remained the most frequent combination. Among RPV VFs with phenotypic RPV resistance, cross-resistance was observed with nevirapine (16/35, 46%), EFV (30/35, 86%) and etravirine (32/35, 91%). Among patients with baseline viral load (VL)≤100,000 copies/ml, there were fewer VFs (RPV: 28/368, 8%; EFV: 20/329, 6%), fewer VFs with treatment-emergent NNRTI RAMs (RPV: 10/27, 37%; EFV: 6/17, 35%), and less phenotypic resistance to RPV and other NNRTIs, than in patients with baseline VL>100,000 copies/ml (VFs: 68/318, 21% [RPV], 32/353, 9% [EFV]; NNRTI RAMs: 36/59, 61% [RPV], 14/32, 56% [EFV]). Among RPV VFs with baseline VL≤100,000 copies/ml observed within weeks 48-96, only 1/7 had phenotypic resistance to RPV. CONCLUSIONS: During the second year of treatment in ECHO/THRIVE, few VFs with emerging NNRTI RAMs (no new RPV RAMs) occurred.