Hepatitis C Virus GT1a Research Articles

Inhibition of Hepatitis C Virus in Chimeric Mice by Short Synthetic Hairpin RNAs: Sequence Analysis of Surviving Virus Shows Added Selective Pressure of Combination Therapy

We have recently shown that a cocktail of two short synthetic hairpin RNAs (sshRNAs), targeting the internal ribosome entry site of hepatitis C virus (HCV) formulated with lipid nanoparticles, was able to suppress viral replication in chimeric mice infected with HCV GT1a by up to 2.5 log10 (H. Ma et al., Gastroenterology 146:63-66.e5, http://dx.doi.org/10.1053/j.gastro.2013.09.049) Viral load remained about 1 log10 below pretreatment levels 21 days after the end of dosing. We have now sequenced the HCV viral RNA amplified from serum of treated mice after the 21-day follow-up period. Viral RNA from the HCV sshRNA-treated groups was altered in sequences complementary to the sshRNAs and nowhere else in the 500-nucleotide sequenced region, while the viruses from the control group that received an irrelevant sshRNA had no mutations in that region. The ability of the most commonly selected mutations to confer resistance to the sshRNAs was confirmed in vitro by introducing those mutations into HCV-luciferase reporters. The mutations most frequently selected by sshRNA treatment within the sshRNA target sequence occurred at the most polymorphic residues, as identified from an analysis of available clinical isolates. These results demonstrate a direct antiviral activity with effective HCV suppression, demonstrate the added selective pressure of combination therapy, and confirm an RNA interference (RNAi) mechanism of action. This study presents a detailed analysis of the impact of treating a hepatitis C virus (HCV)-infected animal with synthetic hairpin-shaped RNAs that can degrade the virus's RNA genome. These RNAs can reduce the viral load in these animals by over 99% after 1 to 2 injections. The study results confirm that the viral rebound that often occurred a few weeks after treatment is due to emergence of a virus whose genome is mutated in the sequences targeted by the RNAs. The use of two RNA inhibitors, which is more effective than use of either one by itself, requires that any resistant virus have mutations in the targets sites of both agents, a higher hurdle, if the virus is to retain the ability to replicate efficiently. These results demonstrate a direct antiviral activity with effective HCV suppression, demonstrate the added selective pressure of combination therapy, and confirm an RNAi mechanism of action.

Resistance Analysis of Hepatitis C Virus Genotype 1 Prior Treatment Null Responders Receiving Daclatasvir And Asunaprevir

In a sentinel cohort, hepatitis C virus (HCV) patients (primarily genotype [GT] 1a) were treated with daclatasvir (NS5A inhibitor) and asunaprevir (NS3 protease inhibitor). Preexistence, emergence, and persistence of resistance variants in patients who failed this treatment are described. HCV-infected null responders received daclatasvir (60 mg once daily) and asunaprevir (600 mg twice daily) alone (Group A, 11 patients) or with peginterferon alfa-2a and ribavirin (Group B, 10 patients) for 24 weeks. Resistance testing was performed on baseline samples and samples with HCV RNA ≥1,000 IU/mL at Week 1 through posttreatment Week 48. Resistance substitution susceptibility to inhibition by asunaprevir and daclatasvir was assessed using HCV replicon assays. In Group A, six GT1a patients experiencing viral breakthrough and one GT1a patient who relapsed had detectable NS5A (Q30E/R, L31V/M, Y93C/N) and NS3 (R155K, D168A/E/V/Y) resistance-associated variants at failure. Two of six viral breakthrough patients achieved SVR48 after treatment intensification with peginterferon alfa-2a and ribavirin. For 2/4 viral breakthrough patients not responding to treatment intensification, NS3 resistance variants changed (D168Y to D168T; R155K to V36M-R155K). At posttreatment Week 48, daclatasvir-resistant variants persisted while asunaprevir-resistant variants were generally replaced by wild-type sequences. The NS3 sequence remained unchanged in the one patient with NS3-R155K at baseline, relapse, and posttreatment Week 48. In Group B, no viral breakthrough was observed. The treatment failure of daclatasvir and asunaprevir in HCV GT1a patients was associated with both NS5A and NS3 resistance variants in prior null responders. NS5A resistance variants persisted while NS3 resistance variants generally decayed, suggesting a higher relative fitness of NS5A variants.

Together … to Take Care: Multidisciplinary Management of Hepatitis C Virus Treatment in Randomly Selected Drug Users With Chronic Hepatitis

Hepatitis C Virus (HCV) infection is treated with peg-interferon α2a or α2b and ribavirin. International studies show that drug user adherence to treatment is 40% to 60% and increases if the patient is in addiction treatment. The aim of the Together To Take Care (TTTC) study was to achieve better adherence to HCV therapy in randomly selected drug users, who are considered "difficult to treat." The secondary aim of the TTTC Study Group was to standardize a method for a multidisciplinary management of the liver disease in drug users. The TTTC group data were matched with a control group. Adherence: The 93.7% of patients followed therapy prescribed; of the patients infected by HCV genotype (gt) 3, all completed therapy as scheduled. For the 48-week treatment group, 66.7% of patients completed therapy (2 of 9 patients stopped treatment for breakthrough). Toxicological results: 10 (62.5%) patients were negative in the toxicological tests (opiates, cocaine, and alcohol). Virological results: 8 of 16 patients were infected by HCV gt 1, and 8 were infected by gt 3; 2 of 16 (12.5%) patients were human immunodeficiency virus (HIV) coinfected (1 HCV gt 1a and 1 HCV gt 3). All patients: 11 of 16 (68.75%) patients were HCV ribonucleic acid undetectable 24 weeks after completing therapy (sustained virological response, SVR). Gt 1: 4 of 8 (50.0%) showed SVR. Gt 3: 7 of 8 (87.5%) showed SVR. Overall, the HCV gt 3 patients had 87.5% probability of SVR, whereas gt 1 patients had 50% probability of SVR (gt 3/gt 1 patients odds ratio = 7). The results were analyzed by Fisher exact test. Our results show that good healthcare management plays an important role in increasing patients' adherence to therapy. In the project "TTTC," the patients work with the physicians to take responsibility for their health and acquire self-efficacy and self-awareness, thanks to the special care.