CROI 2016 Abstract eBook

Abstract Listing

Poster Abstracts

577 HCV Resistance to Daclatasvir/Sofosbuvir Across Different Genotypes in the Real Life Slim Fourati 1 ; Christophe Hezode 1 ; Alexandre Soulier 1 ; Lila Poiteau 1 ; Marion Lavert 1 ; Christophe Rodriguez 2 ; Stephane Chevaliez 1 ; Jean-Michel Pawlotsky 1 1 INSERM U955, Créteil, France; 2 Inst Mondor de Recherche Biomédicale INSERM U955, Créteil, France Background: The impact of preexisting HCV NS5A and/or NS5B resistance-associated variants (RAVs) on the response to daclatasvir (DCV) in combination with sofosbuvir (SOF) is unknown in the “real life” clinical setting. We studied the impact of preexisting substitutions at RAV positions on treatment outcomes in a cohort of patients infected with HCV genotypes (GT) 1a, 1b, 3 and 4 receiving SOF plus DCV without ribavirin for 12 weeks. Methods: Population sequencing of the NS5A and NS5B genes was performed prior to SOF/DCV treatment in 177 patients (GT 1a, n=44; GT1b, n=63; GT3, n=29; GT4, n=41) and at treatment failure (relapse) in 8 patients who did not achieve SVR12. NS5A sequences were examined at 9 known RAV positions in domain I (residues 28, 29, 30, 31, 32, 58, 62, 92 and 93), while NS5B sequences were examined at 5 known RAV positions (159, 282, 316, 320, 321). Results: Prior to treatment, NS5A RAVs were detected at a frequency of 9% in GT1a-, 32% in GT1b-, 21% in GT3- and 10% in GT4-infected patients. The most frequent RAVs were Y93H in GT1b (11%) and GT3 (3.5%), and L28M in GT4 (10%). No NS5B S282T, L320F or V321A variants were detected, while L159F was found in 12% of GT1b patients at baseline. C316N and C316H were detected in GT1b patients (20% and 2%, respectively) and in GT4 patients (5% and 2%, respectively). RAVs frequency at baseline did not differ between patients with and without cirrhosis. Post-treatment relapse occurred in 8 patients, including 5 infected with GT3. Virological failure tended to be more frequent when an NS5A Y93H substitution was present at baseline (p=0.067). All relapse patients harbored dominant NS5A RAVs post-treatment, with Y93H present in all GT3-infected subjects failing treatment. In addition, a so far undescribed NS5A amino-acid substitution (H85Y) was detected in a GT3-infected patient at relapse. Conclusions: The prevalence of NS5A RAVs at baseline varied considerably across genotypes 1a, 1b, 3 and 4. Depending on the genotype, natural polymorphisms in NS5A (e.g. Y93H) differentially influence virological outcomes. A new amino acid substitution, H85Y, was selected in the NS5A region by SOF/DCV treatment. Real-life assessment will be key to better understand the mechanisms of HCV resistance and its role in interferon-free treatment failures. 578 Polymorphisms at Codon 28 of HCV NS5A Impact NS5A Inhibitor Susceptibility Alicia Newton ; Arne Frantzell; Christos Petropoulos;Wei Huang Monogram BioScis, South San Francisco, CA, USA Background: Amino acid substitutions at position 28 of NS5A in HCV genotype 1a (GT1a) represent a major resistance pathway to ledipasvir (LDV), ombitasvir (OBV) and daclatasvir (DCV). GT1a and GT1b viruses are polymorphic at position 28: methionine (M) is most common in GT1a whereas leucine (L) predominates in GT1b. To investigate the impact of amino acid 28 polymorphisms on NS5A drug susceptibility, we evaluated replicons containing M28 or L28 substitutions alone and in combination with NS5A drug resistance associated mutations (RAMs) at positions 30, 31 and 93. Methods: A panel of replicons containing M28 or L28 substitutions alone, and in combination with RAMs at positions 30, 31 and 93 was constructed using H77 (GT1a) and Con1 (GT1b) NS5A sequences. NS5A drug susceptibility (fold change in IC 50 , FC) relative to the parental reference replicon was determined using a luciferase-reporter replicon assay. Results: The introduction of an L28M substitution to the GT1b NS5A sequence conferred subtle or no reductions in susceptibility to LDV, OBV and DCV (FC=1.0 to 2.5). L28M in combination with one additional NS5A RAM notably reduced susceptibility relative to the RAM alone: L28M+L31M vs. L31M (FC: LDV=165, 4.5; OBV=2.4, 0.9; DCV=6.7, 1.5), L28M+Y93H vs. Y93H (FC: LDV=12597, 368; OBV=1371, 125; DCV=188, 8). The introduction of an M28L substitution to the GT1a NS5A sequence subtly increases susceptibility to LDV, OBV and DCV (FC=0.2 to 0.8). M28L in combination with one additional RAM notably increased susceptibility compared to the RAM alone: M28L+Q30R vs. Q30R (FC: LDV=1.4, 477; OBV=14, 4115; DCV=1.2, 298), M28L+Q30H vs. Q30H (FC: LDV=0.7, 269; OBV=0.5, 4.8; DCV=1.3, 567), M28L+L31M vs. L31M (FC: LDV=2.1, 187; OBV=0.4, 2.3; DCV=1.4, 132), M28L+Y93H vs. Y93H (FC: LDV=250, 813; OBV=1770, 47582; DCV=74, 1059). Conclusions: This study demonstrated that the M/L polymorphism at position 28 of NS5A differentially impacts the susceptibility of HCV GT1a and GT1b to NS5A inhibitors. The introduction of RAMs at positions 30, 31 or 93 conferred larger reductions in NS5A drug susceptibility in the context of the M28 substitution in GT1a compared to the L28 substitution in GT1b. These findings explain, in part, why GT1a viruses containing NS5A RAMs display larger reductions in NS5A drug susceptibility versus comparable GT1b viruses, which in turn may explain the less favorable treatment responses of GT1a HCV infected individuals treated with NS5A inhibitors. 579 Genotypic and Phenotypic Characterization of Clinical HCV NS5A Drug Resistance Wei Huang ; Alicia Newton; Arne Frantzell; Jennifer Cook; DongmeiYang; Joseph M.Volpe; Charles M.Walworth; Jeannette M.Whitcomb; Jacqueline D. Reeves; Christos Petropoulos Monogram BioScis, South San Francisco, CA, USA Background: Treatment regimens containing the NS5A inhibitors (NS5AI) ledipasvir (LDV) and ombitasvir (OBV) are approved for the treatment of HCV genotype 1 (GT1) infection. Overall, GT1b infected individuals have better treatment responses to NS5AI-containing regimens than GT1a infected individuals. Previous studies have demonstrated that the presence of NS5AI resistance-associated variants (RAVs) may impact treatment outcome. We characterized the prevalence of NS5A RAVs and NS5AI susceptibility of samples submitted for routine NS5AI resistance testing. Methods: The entire NS5A region was amplified from 564 GT1 (1a=442, 1b=122) samples submitted for routine drug resistance testing. NS5A sequencing was performed using the Illumina MiSeq platform. NS5AI RAVs at positions 28, 30, 31, 58 and 93, detected at ≥10%, were included in the analysis. Replicons containing clinical NS5A sequences, or site- directed mutations (SDMs), were evaluated for susceptibility to LDV and OBV using a luciferase-reporter assay. Results: NS5AI RAVs were identified in 113/442 (25.6%) GT1a viruses: M28A/T/V=34 (7.7%), Q30E/H/K/L/N/R/S/T/Y=53 (12.0%), L31M/V=24 (5.4%), H58D=7 (1.6%), and Y93C/F/H/N/S=38 (8.6%). Of these, 76/113 (67.2%) harbored a single RAV and 37 (32.7%) had >1 RAV. NS5A RAVs were identified in 41/122 (33.6%) GT1b viruses: L31I/M/V=18 (14.8%), Y93H=33 (27.0%). Of these, 31/41 (75.6%) harbored a single RAV and 10 (24.3%) had >1 RAV. Replicons with SDMs at position 28, 30, 31, 58 and 93, or NS5A regions from clinical samples with RAVs, exhibited large reductions in susceptibility to LDV and/or OBV. Reductions in susceptibility varied by subtype, RAV position, RAV amino acid substitution and NS5A inhibitor. GT1a viruses with RAVs generally exhibited larger reductions in NS5AI susceptibility than GT1b viruses. The GT1b Y93H mutation had a large impact on NS5AI susceptibility, conferring 367- and 125-fold reductions in LDV and OBV susceptibility, respectively. Conclusions: We observed a similar prevalence of NS5AI RAVs among GT1a and GT1b viruses submitted for routine resistance testing. GT1a viruses had a broader array of RAVs and generally exhibited larger reductions in NS5AI susceptibility than GT1b viruses. RAVs among GT1b viruses were limited to positions 31 and 93. Y93H was the most common GT1b variant observed and conferred marked resistance to NS5AI. This study illustrates the potential utility of profiling NS5AI resistance in clinical treatment settings. 580 Resistance Associated Variants: Data From the NIAID SYNERGY Trial Eleanor M. Wilson 1 ; Sarah Kattakuzhy 2 ; Hongmei Mo 3 ; Anuoluwapo Osinusi 3 ; Henry Masur 4 ; Anita Kohli 5 ; Shyam Kottilil 6 1 Inst of Human Virology, Baltimore, MD, USA; 2 Inst of Human Virology, Bethesda, MD, USA; 3 Gilead Scis, Inc, Foster City, CA, USA; 4 NIH, Bethesda, MD, USA; 5 Dignity Hlth, St. Joseph’s Hosp, Phoenix, AZ, USA; 6 Univ of Maryland Med Cntr, Baltimore, MD, USA Background: Treatment of chronic hepatitis C (HCV) has rapidly evolved frommodestly-effective interferon-based regimens to highly effective, all oral, interferon-free directly acting antiviral (DAA) therapy. While DAAs are increasingly used for treatment of HCV worldwide, the impact of emerging resistance is largely unknown. In this study, we evaluated the role of baseline, treatment-emergent and treatment-enriched resistance associated variants of HCV on sustained virologic response (SVR) at initial, short-duration, all oral DAA treatment and at re-treatment with a second DAA only regimen for patients who failed initial therapy. Methods: NIAID SYNERGY is an innovative 8 arm clinical trial that treated over 200 hepatitis C patients with varying sofosbuvir(SOF)-based DAA regimens (Figure 1). Deep sequencing was performed on plasma at baseline on all patients, at the time of relapse in patients failing therapy, and prior to retreatment, when appropriate. RAVS were classified

Poster Abstracts

230

CROI 2016