Reduction In HCV RNA Levels Research Articles

Silencing HCV Replication in Its Reservoir.

BACKGROUND:HCV infection and its complications are among the leading public health challenges; the emergence of drug-resistant variants are expected to be a major problem. A novel combinatorial small interfering RNA (siRNA) could be a novel triple therapy that could be suitable for genotype 4. Although HCV is a hepatotropic virus, there is reliable evidence about its replication in peripheral blood mononuclear cells (PBMC) of chronically infected patients; these cells act as an extra-hepatic reservoir for viral recurrence and persistence. The patients with HCV-RNA in PBMC showed a significantly lower response to therapy that supports to be one of the factors influencing therapeutic response. Almost all regions of HCV show potential for siRNA target with relative efficiencies of individual siRNA sequences.AIM:This study aims to test the efficacy of siRNA against HCV-4 replication in PBMC in vitro, to introduce an alternative therapeutic option for HCV-4 suitable to eradicate it from both hepatic and extra-hepatic reservoirs.METHODS:Efficacy of synthesised siRNA molecule that targets 5/UTR of domain IIIC within IRES of HCV RNA to eradicate HCV intra-PBMC in vitro was tested and compared with IFN/RBV in vitro, by using both qRT-PCR and western blot. Sixty genotype-4 chronic HCV patients who are naïve for any HCV treatment were enrolled and tested for the presence of HCV intra-PBMC using qRT-PCR before and after siRNA treatment in vitro.RESULTS:Real-time PCR analysis showed a significant reduction of HCV RNA levels after 24hr post-HCV-positive-PBMCs treatment by siRNA with cell vitality reached up to 98%. Besides a complete inhibition of NS5A viral protein expression, that is functionally essential for viral assembly, replication and egress.CONCLUSION:So, Targeting HCV infection using RNA interference technology might be a reliable therapeutic option for chronic HCV patients with HCV minus strand within PBMCs.

Potent Immune Activation in Chronic Hepatitis C Patients upon Administration of An Oral Inducer of Endogenous Interferons that Acts via Toll-Like Receptor 7

ANA773, an oral prodrug of a small-molecule Toll-like receptor (TLR)7 agonist, induces a dose-related decrease in serum HCV RNA levels in chronic hepatitis C patients. The prodrug ANA773 was administered to healthy individuals and chronic hepatitis C patients. At different time points during the course of treatment, modulation of the phenotype and function of peripheral leukocytes were evaluated to determine the role of distinct immune cells on the clinical outcome of therapy. Early after administration of the TLR7 agonist, a mild transient reduction of the number of lymphocytes was observed in both healthy individuals and chronic hepatitis C patients. Moreover, repeated administration of ANA773 resulted in transiently reduced numbers of myeloid and plasmacytoid dendritic cells (DC) in blood. Interestingly, reduced plasmacytoid DC numbers as well as increased serum interferon (IFN)-α and IFN-γ inducible protein (IP)-10 levels were observed only in virological responders (≥1 log(10) IU/ml reduction of HCV RNA levels upon ANA773 treatment), but were absent in virological non-responders. In vitro stimulation of peripheral blood mononuclear cells from virological responders showed a high frequency of IFN-α-producing plasmacytoid DC upon stimulation in vitro with ANA773, whereas no IFN-α was induced in non-responders. These findings indicate that the viral load decline in chronic hepatitis C patients treated with the TLR7 agonist ANA773 is likely due to intrinsic differences in the induction of endogenous IFNs and IFN-stimulated gene products (IFN-α and IP-10) upon TLR7 ligation.

Overview of the PROVE studies evaluating the use of telaprevir in chronic hepatitis C genotype 1 patients

Current treatment for genotype 1 HCV infection with pegylated interferon (PEG IFN) and ribavirin (RBV) is effective in less than 50% of patients. The advent of direct-acting antiviral agents that target replication of HCV promises to improve therapy for the disease. Telaprevir is a new peptidomimetic serine protease inhibitor that specifically targets the NS3/4a HCV serine protease to cause rapid reduction in HCV RNA levels. Three Phase II Protease Inhibition for Viral Evaluation (PROVE) studies have assessed the efficacy and safety of telaprevir in genotype 1 patients. The studies examined sustained virological response (SVR) rates and also the adverse events related to the use of this drug in different groups. The results of these studies suggested that the addition of this specific protease inhibitor to PEG IFN alfa-2a and RBV can significantly improve the results of treatment in patients affected with chronic HCV infection with genotype 1, when compared with the standard treatment, PEG IFN alfa-2a and RBV alone. The key observations in these Phase II trials of telaprevir were higher rate of SVR above current standard of care (61–69% for T12PR24 treatment-naive patients compared with 46–48% for standard of care in naive patients). Low rates of relapse were observed in T12PR24-treated patients (2–14% vs 22–23%). The studies suggest that the duration of treatment could be reduced for rapidly responsive naive patients from 48 to 24 weeks while maintaining improved SVR rates. RBV remains an essential component of treatment with protease inhibitors combined with PEG IFN. The main adverse reactions of note with its use were rashes, anemia and nausea.

Dynamic Hepatitis C Virus Genotypic and Phenotypic Changes in Patients Treated With the Protease Inhibitor Telaprevir

Telaprevir (VX-950), a hepatitis C virus (HCV) NS3.4A protease inhibitor, has shown strong antiviral activity in phase 1 clinical studies. Because of high levels of HCV replication and the low fidelity of HCV polymerase, selection of resistant isolates during therapy may occur. A highly sensitive sequencing method was developed in which approximately 80 clones/sample were analyzed to identify mutations in the NS3 protease catalytic domain in HCV genotype-1-infected patients dosed with 450 mg every 8 hours, 750 mg every 8 hours, or 1250 mg every 12 hours of telaprevir for 14 days. Mutations that confer low-level resistance (V36A/M, T54A, R155K/T, and A156S) and high-level resistance (A156V/T, 36+155, 36+156) to telaprevir were detected and correlated with telaprevir exposure and virologic response. Changes in the frequency of mutations after the end of dosing showed an inverse relationship between in vivo viral fitness and resistance. In the absence of telaprevir selective pressure the majority of resistant variants were replaced by wild-type virus within 3-7 months. Resistant HCV isolates are selected rapidly during therapy with the highly active protease inhibitor telaprevir. Combination therapy with pegylated interferon-alfa or other direct antiviral drugs seem mandatory to avoid developing resistance.

Clinical evaluation (Phase I) of a human monoclonal antibody against hepatitis C virus: Safety and antiviral activity

Background/Aims: HCV-AB68, a human monoclonal antibody against the envelope protein of hepatitis C virus (HCV), neutralizes HCV in cell-culture and in the HCV-Trimera mouse model. A Phase 1 clinical trial was designed to test safety, tolerability, and antiviral activity of HCV-AB68 in patients with chronic HCV-infection. Methods/Results: Single doses of HCV-AB68, 0.25–40 mg, administered to 15 patients were well tolerated with no moderate or serious adverse events (SAEs) reported. In six patients, HCV-RNA levels transiently decreased by 2- to 100-fold immediately following infusion and rebound to baseline in 24–48 h. Multiple doses of HCV-AB68, 10–120 mg, were administered to 25 patients. Doses were given weekly for 3 weeks, then 3× a week during the fourth week, after which patients were followed for 3 months. No drug-related SAEs were reported and no specific pattern of adverse events was evident. Eight out of 25 patients had at least a 1-log reduction and 17 had at least a 0.75-log reduction in HCV-RNA levels from baseline at one or more time points following HCV-AB68 infusion. Conclusions: These data support the investigation of HCV-AB68 in the prevention of recurrent HCV-infection in patients who had received hepatic allografts for end-stage liver disease.

First phase hepatitis c viral kinetics in previous nonresponders patients

A large proportion of patients fails to respond to treatment for hepatitis C. Initiation of interferon therapy is associated with a rapid first phase decline in viremia, reflecting inhibition of viral production or release from infected cells. We characterized first phase viral kinetics in previous nonresponder patients and compared the antiviral efficacy of interferon in nonresponders to that observed in naive patients. Twenty nonresponders with genotype 1 infection were evaluated. Ten received a single 15 mcg dose of interferon alfacon-1 and ten were given a 30 mcg dose. Viral kinetic data from previously untreated historical control patients with genotype 1 infection who received 9 mcg (n = 12) or 15 mcg (n = 13) of interferon alfacon-1 provided a basis for comparison. Antiviral efficacy was evaluated by calculating the reduction in HCV RNA levels during the first 24 h after interferon administration (log effectiveness). Hepatitis C virus levels decreased exponentially in previous nonresponder patients. Non-responders treated with 30 mcg of interferon alfacon-1 exhibited a greater log drop than non-responders receiving 15 mcg (P = 0.01). The log effectiveness of 15 mcg of interferon alfacon-1 in nonresponders was similar to 9 mcg in naives and was significantly < 15 mcg (P = 0.04) in naïve patients. The 30 mcg dose provided similar log effectiveness in nonresponders compared with 15 mcg in naive patients and exceeded the log effectiveness of 9 mcg in previously untreated patients (P = 0.035). Nonresponders who had greater than a 50% decrease in HCV RNA level from baseline at the end of previous treatment had a larger reduction in viral load at 24 h compared with those who had not achieved that level of response with prior therapy (P = 0.04). In conclusion, the log effectiveness of interferon was lower in nonresponders compared with treatment naive patients. The difference in antiviral effectiveness in previous nonresponders was overcome by higher interferon doses.

The impact of interferon plus ribavirin on response to therapy in black patients with chronic hepatitis C

Black patients with chronic hepatitis C have lower response rates than white patients to interferon monotherapy. The factors responsible for these differences are unknown, as is the impact of combination antiviral therapy on responsiveness among ethnic groups. We evaluated the impact of race on response to therapy in these patients. A total of 1744 patients with chronic hepatitis C were randomized in 2 recent clinical trials to receive 24 or 48 weeks of interferon monotherapy or interferon-ribavirin combination therapy. Sustained virologic responses occurred in 27% of 1600 whites, 11% of 53 blacks (P = 0.01 vs. white), 44% of 32 Asians, and 16% of 27 Hispanics. No black patient had a sustained virologic response to interferon monotherapy, but 20% and 23% had sustained responses to 24 and 48 weeks, respectively, of combination therapy. Among black patients, 96% had hepatitis C genotype 1 compared with 65% of white subjects (P < 0.0001). Sustained response rates were similar for black and white patients with genotype 1 infection (23% vs. 22%, respectively). Compared with whites, black patients were older, weighed more, and had higher median Histologic Activity Index scores but did not differ in sex, baseline alanine aminotransferase or hepatitis C virus (HCV)-RNA levels, degree of fibrosis or percentage with cirrhosis, or other demographic variables. White subjects had a significantly greater reduction in HCV-RNA levels than blacks at weeks 4, 12, 24, and 48 of therapy, but only for black patients treated with interferon monotherapy. The decreased reduction of HCV-RNA reduction among blacks was eliminated by combination therapy. These observations suggest that the impaired responsiveness of black patients to interferon monotherapy can be overcome partially by combination interferon-ribavirin therapy.

Chronic Hepatitis C Treatment: A Review

Chronic Hepatitis C Treatment: A Review

Quantification of serum hepatitis C virus RNA with daily or standard interferon doses plus ribavirin in nonresponder patients with chronic hepatitis C.

Less than 10% of patients with chronic hepatitis C infected with genotype 1 achieve a sustained response with 12 months of interferon therapy. Viral kinetics studies have shown that HCV may replicate in less than 24 hr, generating over 10(12) copies per day and suggesting the need for more aggressive therapy. The aim of the study was to determine the effect of a higher and daily dosage of IFN-alpha2b plus ribavirin on the viral load and on the response rate in patients infected by genotype 1 and previous nonresponders to interferon. Ten patients with chronic hepatitis C infected with genotype 1 were allocated to receive IFN-alpha2b, 5 MU daily or three times a week for four weeks followed by 5 MU three times a week until week 24 plus ribavirin for the entire period. At week 4 of therapy, a 2 log reduction in HCV RNA levels was achieved in three (60%) patients in the daily group and in one (20%) patient in the three times a week group. At week 24, HCV RNA was negative in four of the five patients in the three times a week group, and three of the four patients in the daily group had a virological response. However, in follow-up, none of these patients experienced a sustained response. The safety of and ability to tolerate the combination therapy was good, anemia being the most common adverse effect. In conclusion, patients previously not responding to interferon achieved a greater virological reduction early in combination therapy compared to a three times a week interferon schedule. However, the virological response at the end of therapy was similar between the two regimens, and no sustained response was observed in any of the treatment groups.

HEPATITIS C VIRAL DYNAMICS

It is apparent that the sooner the virus is cleared from the serum following IFN monotherapy, the better the sustained virologic response rates. It is also clear that in patients infected with HCV genotype 1a and 1b, standard dosages of IFN-alpha 2b (3 MU) administered three times a week are inadequate for a substantial and sustained lowering of HCV RNA serum levels. Understanding the kinetics and dynamics of HIV and HBV has greatly improved the understanding of the life cycle of these viruses and their response to therapy. Studies of the kinetics of HCV following initiation of IFN monotherapy have revealed that IFN-alpha 2b causes a rapid dose-dependent (3 < 5 < 10 = 15 MU) reduction in HCV RNA levels within 24 to 48 hours. This rapid exponential decline in RNA levels is best explained by an effect of IFN on viral production or release. The dose of other IFN products that maximally suppresses viral levels needs to be determined. Mathematical calculations reveal that HCV has a serum half-life of 3 hours and a viral production rate of 1.0 x 10(12) virions/d. After this rapid decline, there is a slower phase of viral decline that varies widely among patients and is attributed to the death rate of infected liver cells. The rate of decline of the second phase, which is probably mediated by immune clearance of infected liver cells, is the best viral kinetic predictor of early viral clearance. This kinetic information indicates that in patients infected with HCV genotype 1a or 1b, initial therapy with IFN should be daily and initial doses should be sufficient to reduce viral levels by more than 95% within 48 hours. Whether higher doses of IFN will alter or enhance the second phase of viral decline needs to be investigated. Also, the effect of ribavirin on IFN-mediated changes in HCV RNA levels needs to be investigated in carefully performed kinetics studies to better determine its mechanism of action. Defining the viral kinetics in patients infected with HCV genotype 2 or 3 and in patients who do not respond to IFN therapy will also improve the approach to therapy.

Effect of obesity on pharmacokinetics and biologic effect of interferon-alpha in hepatitis C.

To examine potential adverse effects of obesity in reducing the response to interferon-alpha (IFN-alpha) in chronic hepatitis C (HCV), IFN-alpha and HCV RNA levels in serum and the 2',5'-oligoadenylate synthetase (2-5 OAS) levels in peripheral blood mononuclear cells (PBMC) were compared between six obese and five nonobese patients before and after a single, 10 mIU dose of IFN-alpha2b. There were no differences in the mean histologic activity index between the two groups. The maximal IFN concentration and the area under the serum IFN concentration-time curve were higher in nonobese patients. These two parameters were inversely correlated with body weight and body surface area. No differences were found in the mean reduction in HCV RNA levels between the two groups following IFN-alpha. The maximal 2-5 OAS level after treatment divided by the pretreatment 2-5 OAS level (2-5 OAS response ratio) was greater in the nonobese patients, suggesting stronger biologic response upon exposure to exogenous IFN-alpha in nonobese patients.