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The Immune Response to HCV: implications for HCV/HIV co-infection
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Reported by Jules Levin
Several interesting papers were presented at AASLD on the immune response to HCV. These papers provide insight to understanding to why some individuals have a better immune response. The study by Golia immediately below contrasts a difference between the response to therapy and an expectation of response based on the pharmacokinetics of drugs. These papers, particularly the paper on dendritic cells and the immune response, provide insights into why HIV/HICV co-infected patients may not respond as well to HCV therapy or mount as good a response to HCV. Results from preliminary data of studies for peginterferon plus ribavirin in HCV/HIV co-infected patients suggest lower viral response rates than we see in studies of HCV monoinfected patients. An impaired immune response due to HIV, partcularly at the level of dendrtitic cells, may contribute to reduced viral response rates.
Host Factors Responsible for a Viral Response May Be Different in Responders Compared with Nonresponders, Independent of serum PEG-IFN alfa-2b Concentrations
PHARMACODYNAMICS AND PHARMACOKINETICS OF HEPATITIS C VIRUS (HCV) AND PEGYLATED INTERFERON-ALFA2B IN HUMAN IMMUNODEFICIENCY VIRUS (HIV)/HCV CO-INFECTED PATIENTS
Reported in poster at AASLD by Preeti Golia and colleagues (Weill Medical College of Cornell University, New York, NY; Los Alamos National Laboratory, Los Alamos, NM; New York Blood Center, New York, NY).
Approximately one half of hepatitis C virus (HCV) infected patients will fail to respond to current treatment regimens, pegylated interferon alfa and ribavirin. Modeling of HCV RNA decay in response to antiviral therapy can provide insight into viral and host determinants of viral response.
Pharmacokinetic, pharmacodynamic, and immunologic outcomes were evaluated on 24 co-infected patients: 1) To evaluate the relationship between HCV RNA decay and PEG-IFN alfa2b serum concentrations in HIV/HCV co-infected patients and 2) to evaluate association between HCV-specific immune responses and virologic outcome.
24 co-infected, therapy-naïve patients were treated with PEG-IFN alfa2b 1.5ug/kg/wk and RBV 13 ± 2 mg/kg/day. Serum was obtained for HCV RNA, HIV-1 RNA and serum PEG-IFN alfa2b levels at baseline, after the first dose (6, 12, 24 hours) and on days 2, 3, 5, 6; after the second dose (6, 12, 24 hours) and on days 8, 9; and after the third dose on days 14, 15, and 16. Peripheral blood mononuclear cells were obtained at baseline, weekly for the first month, and monthly thereafter.
HCV and HIV-1 RNA levels were determined by reverse-transcriptase polymerase chain reaction and PEG-IFN alfa2b levels were measured by ELISA. CD4+ cell proliferative responses to HCV antigens (core, NS3, NS4, NS5) and HIV-1 gag were performed at baseline, day 7 and 14, and weeks 4, 8, 12, and 24. Among 24 co-infected patients, 21 are males, the mean age was 46 ± 5.8 years, 8 are Caucasian, 12 are African-American and 4 are Hispanic. 20/24 patients were infected with genotype 1, 3/24 patients with genotype 2, and 1/24 with genotype 3a.
RESULTS
Pharmacokinetics:
To date, 21 patients have been evaluated. 10/21 patients were end of treatment responders, but 2 of these were "late responders" (i.e. HCV RNA declines after week 8 and is undetectable by week 24). Data fitting during the first seven days revealed a free virion clearance rate c of 13.3 (day-1) and an infected cell clearance rate of 0.21 (day-1). The half-lives were 2.7 hours and 4.0 days, respectively.
Pharmacodynamics:
Viron production was blocked after the first dose of PEG-IFN alfa with an average maximum effectiveness (max) of 89%. We estimated that the fifty percent effective concentration (EC50) of PEG-IFN alfa2b, the theoretical in vivo drug concentration at which the drug efficacy is 50%, was five-fold lower in responders compared with non-responders while the maximum (Cmax) and minimum (Cmin) concentrations did not differ significantly (Table I). The calculated minimum (min) and maximum (max) efficacy were also significantly increased in responders.
Immunologic:
The mean number of CD4+, CD8+ and CD56+ cells did not differ significantly at baseline or during treatment between responders and nonresponders. Peripheral blood CD4+ HCV-specific proliferative responses did not differ significantly between responders and nonresponders except that the response to core antigen was increased in non-responders compared with responders at weeks 4 and 12. HIV gag CD4+ proliferative responses were stronger than HCV-specific responses at all time points evaluated.
The authors concluded that the serum PEG-IFN alfa2b concentration necessary for an end of treatment response is five-fold lower in responders compared with non-responders while there are no significant differences in the maximum and minimum PEG-IFN alfa2b concentrations in responders and nonresponders. These data suggest that host factors responsible for a viral response may be different in responders compared with nonresponders, independent of serum PEG-IFN alfa2b concentrations.
PEGINTERFERON alfa-2A (40KD) ENHANCES HCV SPECIFIC T-CELL RESPONSES BY RESTORATION OF ALLOSTIMULATORY FUNCTION OF DENDRITIC CELLS IN CHRONIC HEPATITIS C.
Sanaa M Kamal and colleagues reported on the immune system and its response to HCV. (Harvard Institutes of Medicine, Boston, MA; University of Freiburg, Freiburg, Germany; University, Cairo, Egypt; University of Freiburg, Freiburg, Germany; Ain Shams University, Cairo, Egypt; University of Freiburg, Freiburg, Germany)
Peginterferon alfa - 2a (40KD)/ribavirin combination therapy markedly improves the sustained virological response in chronic hepatitis C compared to conventional interferon alfa-2 therapy. We have previously demonstrated that Peginterferon a- 2a (40 KD) alone or in combination with ribavirin enhances HCV specific CD4+ T helper 1 responses in patients with chronic hepatitis C but the underlying mechanism by which the HCV-specific responses are restored is not yet fully understood.
Dendritic cells (DCs) are the most efficient type of cells involved in antigen presentation (APC), however HCV proteins affect DC function resulting in abnormal priming of anti-HCV-specific T cells and defective antiviral immunity. We hypothesized that peginterferon activates CD4+ T cells through restoration of the DC antigen-presenting function.
Monocyte-derived dendritic cells (DCs) generation (magnatic sorting and positive selection), phenotypic analysis and allogeneic stimulatory capacity of DC (S.I.) as well as HCV-specific CD4+ T-cell proliferative responses and cytokine production to HCV proteins (ELISPOT assay using autologous DCs as APCs) were prospectively assessed in 64 patients with chronic hepatitis C (genotypes 1 and 4) before, during and after treatment with either conventional IFN alfa-2a/ribavirin therapy, or Peginterferon alfa - 2a (40 KD)/ribavirin combination therapy and the results were correlated to the therapy outcome.
The SVR in genotype 1 was 49% with Peginterferon alfa - 2a (40KD)/ribavirin, and 26% with conventional IFN a-2/ ribavirin while in genotype 4 the SVR was 54% in subjects treated with Peginterferon alfa - 2a (40KD) /ribavirin combination therapy vs 28% of HCV in subjects treated with conventional IFN a-2a/ ribavirin.
Before induction of therapy, DCs from HCV infected subjects exhibited a pattern of incomplete activation and the stimulatory capacity of HCV-DCs was significantly lower than that of the normal-DCs (7.8± 3.9 vs. 67.2± 19.7, respectively; P < 0.001).
The same pattern of incomplete activation was observed in CD4+ T cells in the form of absent or weak pre-treatment HCV specific CD4+ responses. Initiation of Peginterferon 2a (40KD)/ ribavirin therapy markedly ameliorated the allostimulatory capacity in DCs of HCV patients and induced significant increase in the frequency, strength and breadth of HCV-specific CD4+T-h1 responses; compared to conventional IFN- based regimen.
The stimulatory capacity of HCV-DC was restored to normal in subjects who achieved SVR (62.05 ± 17.7 in SR versus 11.88 ± 2.55 in NR; P = 0.007). Sustained responders developed early restoration of DC functions and strong multi-specific persistent HCV specific CD4+T- cell responses with preferential IFN-; production and IL-10 suppression.
Interestingly patients who had breakthrough or relapse had transient increase in DC stimulatory capacity, which coincided with the absence of HCV viremia; however DC abnormalities were detected with recurrence of viremia. Viral clearance and HAI improvement but not HCV genotype correlated with the DC stimulatory capacity and HCV-specific CD4+ responses.
Peginterferon alfa - 2a (40 KD) in combination with ribavirin enhances HCV specific CD4+ T responses through restoration of the antigen presenting functions of dendritic cells. This finding has important implications for development of novel immunotherapeutic strategies.
EFFECT OF EARLY ANTIVIRAL THERAPY ON THE CELLULAR IMMUNE RESPONSE IN ACUTE HEPATITIS C
Fareed Rahman and colleagues from the NIH reported on this study exploring the immune response to HCV.
Spontaneous recovery occurs in a minority of patients with acute hepatitis C, but is associated with vigorous and long-lasting cellular immune responses (J. Exp. Med. 2001; 194:1395, Nat. Med. 2000; 6: 578). Treatment-induced recovery from hepatitis C can be achieved in the majority of patients who are treated during the acute phase (NEJM 2001; 20: 1452), but the mechanisms of viral clearance and its effect on cellular immune responsiveness are not known. Both direct effects, such as immune modulation of Th2 to Th1 responses, and indirect effects, such as prevention of downregulation and exhaustion of cellular responses by rapid and early reduction of viral load, have been proposed.
To investigate how early antiviral therapy and successful reduction of HCV load affect HCV-specific T cell responses, we performed detailed, prospective clinical, immunological and virological studies on 7 patients with acute hepatitis C who received antiviral therapy and were followed over a period of 1-2 years.
The peripheral blood CD4+ and CD8+ T cell response against all potential HCV epitopes in the context of all given HLA class I and class II alleles of each patient was assessed in ex vivo IFN- ELISpot and in vitro proliferation assays using 600 overlapping HCV peptides and a panel of recombinant HCV proteins. Results:
This comprehensive analysis demonstrated that none of patients suffered from a primary failure of the HCV-specific cellular immune response in the acute phase prior to treatment. Even in the absence of detectable T cell proliferation, multispecific IFN- + T cell responses were found in all patients. In individual patients, phases of apparent immune control and decreased viral load were followed by phases of weaker immune responses and increased viral load.
After initiation of peginterferon/ribavirin or interferon/ribavirin combination therapy (mean of 20 weeks after defined exposure to HCV), all patients with a sustained response (n=6) demonstrated a rapid decrease of HCV RNA levels followed by a more gradual decrease of the HCV-specific T cell responses to undetectable level. In contrast, the sole patient who maintained HCV-specific T cell responses through the course of therapy did not clear HCV completely as shown by viral breakthrough at week 20 of peginterferon monotherapy.
In summary, this study demonstrates active cellular immune responses and periods of transient HCV control in all patients with acute hepatitis C, regardless of the potential outcome of the acute infection. The significant decrease of HCV-specific T cell responses seen in association with a sustained response to therapy suggests that HCV virus and antigens are cleared as a result of the antiviral rather than immunomodulatory actions of interferon and ribavirin. The T cell responses in these patients differ from those reported to occur in spontaneously recovered cohorts, where HCV-specific T cell remain readily detectable and persist long-term. Because maintenance of cellular immune responses has been associated with immune protection upon rechallenge (J. Virol., 2003; 77: 4781), patients who resolve acute hepatitis C as a result of antiviral therapy may have less protective immunity than those who recover spontaneously. The first two authors contributed equally to this work.
HCV EXPOSURE IN HUMANS: STIMULATION OF CELLULAR, BUT NOT HUMORAL IMMUNE RESPONSES IN THE ABSENCE OF DETECTABLE VIREMIA
Theo Heller and colleagues reported on the immune system and HCV. (Washington Hospital Center, Washington, DC; Inova Fairfax Hospital, Falls Church, VA; Thomas Jefferson University, Philadelphia, PA; National Institutes of Health, Bethesda, MD)
Most studies of HCV-specific cellular and humoral immune responses have been performed on patients who were exposed to a large volume of HCV-infected blood and developed HCV viremia. Here, we prospectively analyzed the humoral and cellular immune response of 25 subjects with a documented exposure to HCV that did not result in any detectable viremia.
Cellular and humoral immune responses of the exposed subjects were compared to those of a control group of 29 healthy blood donors.
The cohort consisted of 13 female and 12 male subjects with percutaneous (needlestick, cut or bite; n=19), cutaneous (blood on skin; n=3) or mucosal (splash of blood into the eye, n=3) exposure to HCV.
Virological and immunological analyses were performed on the day of exposure and at weeks 2, 4, 6, 12, 24 and 48 after exposure. Peripheral blood mononuclear cells were tested for HCV-specific proliferation using recombinant HCV core, NS3, NS4, NS5A and NS5B proteins. Direct, ex vivo effector functions of HCV-specific CD4+ and CD8+ T cells were assessed by IFN- ELISpot analysis using 228 overlapping 15-mer peptides spanning the HCV core, NS3, NS4A and NS4B proteins.
No subject tested positive for HCV-RNA (Amplicor Assay, Roche Diagnostics) or HCV antibodies (2nd generation EIA, Abbott Laborat.) at any study time point .
Eight of 25 subjects (32%) had a positive proliferative T cell response at baseline (day 0 of the study) which was defined as higher than the mean response of 29 healthy blood donors plus 3 standard deviations. Three of 8 subjects (38%) with a baseline response had a further increase of their HCV-specific proliferative T cell response and 7/17 subjects (41%) without a baseline response developed a primary HCV-specific proliferative T cell response within 6 weeks after HCV exposure.
The HCV-specific proliferative response was targeted against structural and nonstructural HCV proteins and in 9 of 10 (90%) cases, associated with IFN- production, thus indicating direct, ex vivo effector functions of the responding cells. HCV-specific T cell responses returned to baseline levels in all but 2 subjects within 12 weeks after exposure.
Collectively, these results demonstrate that low level exposure to HCV can induce and/or boost HCV-specific T cell responses in the absence of antibody production and detectable viremia. Thus, cellular immune responses might be a more sensitive indicator of HCV exposure than humoral responses. The data also suggest that repeated exposure to HCV is sufficient to immunize the exposed individuals without causing clinically evident infection.
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