Search

Hepatitis C virus infection is a major cause of chronic liver disease. CD4(+) T cells play a key role in disease outcome. However, the critical functions and associated phenotypes of intrahepatic CD4(+) T cells are not well defined. We have previously shown that CD8(+) T cells expressing the C type lectin CD161 are highly enriched in the human liver, especially during chronic hepatitis. These cells are associated with a type 17 differentiation pattern and express cytokines including IL-17A, IL-22, and IFN-γ. We therefore analyzed expression of CD161 on CD4(+) T cells in blood and liver and addressed the relevant phenotype and functional capacity of these populations. We observed marked enrichment of CD161(+)CD4(+) T cells in the liver during chronic hepatitis such that they are the dominant subtype (mean 55% of CD4(+) T cells). IL-22 and IL-17 secreting CD4(+) T cells were readily found in the livers of HCV(+) and NASH donors, although not enriched compared to blood. There was, however, specific enrichment of a novel subset of IL-22/IFN-γ dual secretors (p = 0.02) compared to blood, a result reconfirmed with direct ex vivo analyses. These data indicate the dominance of CD161(+) expressing lymphocyte populations within the hepatic infiltrate, associated with a distinct cytokine profile. Given their documented roles as antiviral and hepatoprotective cytokines respectively, the impact of co-secretion of IFN-γ and IL-22 in the liver may be particularly significant.

Over recent years, it has become increasingly accepted that virus-specific CD4+ and CD8+ T cell responses play a major role in outcome and pathogenesis of hepatitis C virus (HCV) infection. Indeed, while the emergence of strong and multispecific T cell responses may correlate with spontaneous viral clearance, the virus has developed several mechanisms to avoid T cell control in the majority of acutely HCV-infected patients that subsequently develop persistent HCV infection. In this review, we will discuss the current knowledge about the role of cellular immune responses in HCV infection. Specifically, we will emphasise recent new insights into the effector functions of T cells, possible mechanisms of their failure and the host-virus interactions occurring at the site of the disease, the liver.

Background & Aims: Perforin plays a central role in the immunopathogenesis of different viral infections. However, its role in hepatitis C virus (HCV) infection has not been fully understood. Here, we analyzed two closely related questions: first, is CD8+ T cell-mediated killing of HCV-replicating human hepatoma cells mediated by perforin? Second, if so, do HCV-specific CD8+ T cells obtained from chronically HCV infected patients express and upregulate perforin?, Methods: Susceptibility of HCV-replicating human hepatoma cells to the cytotoxic pathway was tested in vitro by addition of perforin substitute streptolysin O and granzyme B and by co-culture experiments with a perforin-expressing HCV-specific CD8+ T cell clone in the presence of perforin or caspase inhibitors. HCV-specific CD8+ T cells were obtained and analyzed for perforin expression and differentiation markers ex vivo from 12 chronically infected patients and 12 patients with resolved HCV infection., Results: HCV-replicating human hepatoma cells were susceptible to cytotoxic killing in vitro and a dominant role of perforin in HCV-specific CD8+ T cell-mediated cytolysis was observed. However, HCV-specific CD8+ T cells obtained ex vivo from chronically HCV infected patients expressed only low levels of perforin and showed an impaired ability to upregulate perforin. This was tightly linked to the distinct differentiation stage of HCV-specific CD8+ T cell differentiation ex vivo since early and intermediate differentiated HCV-specific CD8+ T cells only showed weak perforin expression in contrast to late differentiated CD8+ T cells that displayed strong perforin expression., Conclusions: Our results suggest that perforin plays a dominant role in CD8+ T cell-mediated lysis of HCV-replicating human hepatoma cells but that lysis may be limited in human chronic viral infection by the low perforin expression of early/intermediate differentiated HCV-specific CD8+ T cells., (Copyright © 2012 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background & Aims: BI 207127 is a potent non-nucleoside hepatitis C virus (HCV) NS5B polymerase inhibitor in vitro., Methods: In this double-blind, placebo-controlled study, 57 HCV genotype (GT)-1 patients (n=27 treatment-naïve [TN]; n=30 treatment-experienced [TE]) with compensated liver disease were randomised for 28-day treatment with 400, 600, or 800 mg BI 207127 three times daily (TID) or placebo (only TN) in combination with peginterferon alfa 2a and ribavirin (PegIFN/RBV). Plasma HCV RNA was measured by Roche COBAS TaqMan assay., Results: HCV RNA decreased in a dose-dependent manner with little difference between 600 mg (TN 5.6 log(10), TE 4.2 log(10)) and 800 mg (TN 5.4 log(10), TE 4.5 log(10)). Rapid virological response (RVR; HCV RNA <15 IU/ml) at day 28 occurred in 11/19 TN and 4/30 TE patients treated with BI 207127. GT-1b patients had stronger reductions in HCV RNA than GT-1a (RVR: TN 64% vs. 43%; TE 33% vs. 5%). There were no breakthroughs (HCV RNA rebound >1 log(10) from nadir) in the TN groups, whereas 3/30 TE patients experienced breakthrough due to P495-mutations. Gastrointestinal adverse events (AEs) and rash were the major AEs and most frequent at higher doses. One and four patients discontinued due to AEs in the 600 and 800 mg groups, respectively. Overall, tolerability was good and better at 600 mg than 800 mg., Conclusions: BI 207127 in combination with PegIFN/RBV demonstrated strong antiviral activity with a favourable safety and tolerability profile. The best benefit/risk ratio was observed at 600 mg., (Copyright © 2012 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Introduction: Hepatocellular carcinoma (HCC) is a tumor of increasing incidence and high mortality worldwide. Diagnosis of HCC is often difficult, especially at early stages of disease. Additionally, current treatment options are limited. HCC usually develops in an environment of chronic liver disease. The immune system has an important role in shaping this environment, especially in chronic viral hepatitis, the leading cause of HCC. However, the immune system also plays a role in natural immunity against HCC although this is apparently not sufficient to control the majority of tumors. This failure in tumor control is due to multiple immunomodulatory mechanisms employed by HCC to subvert the immune system. In this review, we will summarize the current knowledge about the role of the immune system in hepatocarcinogenesis. Additionally, we will describe the mechanisms used by the immune system to control established lesions and the reasons why these immune responses apparently fail so often. Finally, possible implications for the design of novel immunotherapeutic strategies will be discussed.

Dipeptidylpeptidase IV (CD26) is a multifunctional ectoenzyme involved in T cell activation that has been implicated in autoimmune pathophysiology. Because IL-17-producing CD4(+) T cells (Th17 cells) are important mediators of autoimmune disease, we analyzed the expression of CD26 and its enzymatic function on human Th17 cells. Analysis of CD26 expression on different CD4(+) T helper subsets showed that CD26 expression is highest on CD4(+) T cells producing type 17 cytokines (e.g., IL-22, IL-17, GM-CSF, or TNF) compared with Th1, Th2, and regulatory T cells. Phenotypic analysis revealed that CD26(++)CD4(+) T cells express the type 17 differentiation molecules CD161, CCR6, lL-23R, and retinoic acid-related orphan receptor-γt. Furthermore, sorted CD26(++)CD4(+) T cells contain >90-98% of Th17 cells, indicating that CD26(++) T cells harbor the Th17 lineage. A comparison with CD161 and CCR6 indicated that analysis of CD26 coexpression may improve the phenotypic characterization of Th17 cells. Of note, CD26(++) Th17 cells are enriched in the inflamed tissue of patients with hepatitis and inflammatory bowel disease. Functional analysis in migration assays revealed that CD26 expressed on Th17 cells is enzymatically active. Indeed, CD26 negatively regulates the chemotactic CD4(+) T cell response to the inflammatory chemokines CXCL9-12 that can be restored by pharmacological blockade of the enzymatic center of CD26. In summary, these results strongly suggest that CD26 may contribute to the orchestration of the immune response by Th17 cells in human inflammatory diseases. They also suggest that the phenotypic analysis of Th17 cells may be facilitated by determination of CD26 expression.

The standard antiviral therapy for chronic hepatitis C is pegylated interferon-alfa (PegIFN) and ribavirin since about 10 years. This treatment regimen leads to a sustained virological response (SVR) in 40-50 % of patients infected with HCV genotype 1 and in approx. 80 % of those infected with HCV genotype 2 or 3. In recent years, many direct antiviral agents (DAA) have been developed and are being explored in clinical studies. These antiviral agents target different viral proteins that are central for HCV replication, incl. the NS3/4A protease, NS5B polymerase, and the NS5A protein. The protease inhibitors telaprevir and boceprevir have recently been approved for the treatment of chronic HCV genotype 1 infection in combination with PegIFN and ribavirin. These triple therapies increase the SVR rates in HCV genotype 1 patients from 40-50 % to approx. 70 %. Other DAAs will likely be approved in the near future and may result in an IFN-free antiviral therapy., (© Georg Thieme Verlag KG Stuttgart · New York.)

Effective innate and adaptive immune responses are essential for the control of hepatitis C virus (HCV) infection. Indeed, elimination of HCV during acute infection correlates with an early induction of innate and a delayed induction of adaptive immune responses. However, in the majority of acutely HCV-infected individuals, these responses are insufficient to clear the virus and persistence develops. In recent years, different mechanisms responsible for the failure of innate and adaptive immune responses have been identified. These include the proteolytic cleavage of molecules playing key roles in the induction of the interferon response, manipulation of interferon-induced effector proteins, interference with CD8+ T-cell function or immune escape in T- and B-cell epitopes. In this review, we discuss the possible roles of innate and adaptive immune responses in HCV clearance and the different evasion strategies used by the virus to escape these immune responses., (© 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.)

A subset of CD8(+) T cells can secrete interleukin 17 (IL-17). However, very little information is currently available about their antigen specificity, tissue distribution, and biological relevance in chronic human viral infection. To address these issues, we comprehensively analyzed peripheral and intrahepatic CD8(+) T-cell responses in a cohort of patients with chronic hepatitis C virus (HCV) infection for the antigen-specific production of IL-17 and interferon (IFN) γ. We found that HCV-specific IL-17-producing and retinoic acid receptor related orphan receptorγt-expressing CD8(+) T cells are detectable in blood and liver and target different epitopes, compared with IFN-γ-producing CD8(+) T cells. Their highest frequency was found in patients with low inflammatory activity, suggesting a protective role in chronic HCV infection.

Stromal interaction molecule 1 (STIM1) deficiency is a rare genetic disorder of store-operated calcium entry, associated with a complex syndrome including immunodeficiency and immune dysregulation. The link from the molecular defect to these clinical manifestations is incompletely understood. We report two patients with a homozygous R429C point mutation in STIM1 completely abolishing store-operated calcium entry in T cells. Immunological analysis of one patient revealed that despite the expected defect of T cell proliferation and cytokine production in vitro, significant antiviral T cell populations were generated in vivo. These T cells proliferated in response to viral Ags and showed normal antiviral cytotoxicity. However, antiviral immunity was insufficient to prevent chronic CMV and EBV infections with a possible contribution of impaired NK cell function and a lack of NKT cells. Furthermore, autoimmune cytopenia, eczema, and intermittent diarrhea suggested impaired immune regulation. FOXP3-positive regulatory T (Treg) cells were present but showed an abnormal phenotype. The suppressive function of STIM1-deficient Treg cells in vitro, however, was normal. Given these partial defects in cytotoxic and Treg cell function, impairment of other immune cell populations probably contributes more to the pathogenesis of immunodeficiency and autoimmunity in STIM1 deficiency than previously appreciated.

Approximately 550 million people worldwide are chronically infected with hepatitis B (HBV) or hepatitis C (HCV) virus. The clinical course of HBV and HCV infection is variable and can lead to a chronic but mild hepatitis or to liver cirrhosis and hepatocellular carcinoma (HCC)1. Therefore, antiviral therapy should be considered in patients with chronic hepatitis to prevent progression of disease. Here, we review the current indications and guidelines for antiviral therapy in chronic HBV and HCV infection.

Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide, with a continuously high mortality. Thus, the development of new therapeutic strategies is crucial to decrease recurrence rates and to improve the overall survival rates of HCC patients. The rationale for immunotherapy is based on the findings of several studies showing specific CD8(+) T-cell responses against various tumor-associated antigens (TAAs) in HCC patients and a clinical benefit of T-cell infiltration in the tumor tissue. However, the impact of TAA-specific CD8(+) T-cell responses on tumor control seems to be rather weak. Several different mechanisms contribute to the failure of the cellular immune response and will be summarized in this review. The aim of immune-based therapies is to overcome these mechanisms of T-cell failure and to induce or boost TAA-specific CD8(+) and CD4(+) T-cell responses. Several preclinical and clinical studies of immune-based therapeutic approaches show encouraging results and will be discussed in this review., (Copyright © 2012 S. Karger AG, Basel.)

HLA-B*27 exerts protective effects in hepatitis C virus (HCV) and human immunodeficiency virus (HIV) infections. While the immunological and virological features of HLA-B*27-mediated protection are not fully understood, there is growing evidence that the presentation of specific immunodominant HLA-B*27-restricted CD8+ T-cell epitopes contributes to this phenomenon in both infections. Indeed, protection can be linked to single immunodominant CD8+ T-cell epitopes and functional constraints on escape mutations within these epitopes. To better define the immunological mechanisms underlying HLA-B*27-mediated protection in HCV infection, we analyzed the functional avidity, functional profile, antiviral efficacy and naïve precursor frequency of CD8+ T cells targeting the immunodominant HLA-B*27-restricted HCV-specific epitope as well as its antigen processing and presentation. For comparison, HLA-A*02-restricted HCV-specific epitopes were analyzed. The HLA-B*27-restricted CD8+ T-cell epitope was not superior to epitopes restricted by HLA-A*02 when considering the functional avidity, functional profile, antiviral efficacy or naïve precursor frequency. However, the peptide region containing the HLA-B*27-restricted epitope was degraded extremely fast by both the constitutive proteasome and the immunoproteasome. This efficient proteasomal processing that could be blocked by proteasome inhibitors was highly dependent on the hydrophobic regions flanking the epitope and led to rapid and abundant presentation of the epitope on the cell surface of antigen presenting cells. Our data suggest that rapid antigen processing may be a key immunological feature of this protective and immunodominant HLA-B*27-restricted HCV-specific epitope.

Hepatocellular carcinoma (HCC) is the fifth most common malignancy worldwide with an increasing incidence. The clinical outcome is influenced by the underlying liver cirrhosis, the size of the tumour at the time of diagnosis and the few therapeutic options currently available. In recent years there has been a lot of progress in the understanding of HCC immunobiology. This review summarizes our current knowledge of HCC biology, the role of chronic inflammation in carcinogenesis and the role of tumour-specific immune responses. Furthermore, we will present potentially new, immune-based therapies that might open up new avenues for the treatment of HCC., (© Georg Thieme Verlag KG Stuttgart · New York.)

Background & Aims: Guidelines recommend that patients with chronic hepatitis C virus (HCV) infection be treated with pegylated interferon and ribavirin for 24, 48, or 72 weeks, based on their virologic response to treatment. We investigated the effects of treating patients for individualized durations., Methods: We treated 398 treatment-naïve patients who had HCV genotype 1 infections with pegylated interferon alfa-2b and ribavirin for 24, 30, 36, 42, 48, 60, or 72 weeks (mean of 39 weeks, termed individualized therapy); the duration of therapy was determined based on baseline viral load and the time point at which HCV RNA levels became undetectable (measured at weeks 4, 6, 8, 12, 24, and 30). Results were compared with those of 225 patients who received standard treatment for 48 weeks (mean of 38 weeks)., Results: Rates of sustained virologic response (SVR) were 55% among patients who received individualized treatment and 48% among those who received standard treatment (P < .0001 for noninferiority). SVR rates, according to the time point at which HCV RNA levels became undetectable, did not differ significantly between groups. Patients with a rapid virologic response (undetectable levels of HCV RNA at week 4) who were treated for 24 to 30 weeks achieved high rates of SVR (86%-88%). Rates of SVR increased among slow responders who first tested negative for HCV RNA at week 24 and were treated for 60 to 72 weeks compared with those treated for 48 weeks (60%-68% vs 43%-44%). The CC polymorphism at IL28B rs129797860 was associated with an increased rate of SVR compared with the CT/TT polymorphism (P < .0001) at baseline but not among patients who had undetectable levels of HCV RNA following treatment., Conclusions: Individualizing treatment of patients with chronic HCV genotype 1 infections for 24 to 72 weeks results in high rates of SVR among rapid responders and increases SVR among slow responders., (Copyright © 2011 AGA Institute. Published by Elsevier Inc. All rights reserved.)

While human leukocyte antigen B57 (HLA-B57) is associated with the spontaneous clearance of hepatitis C virus (HCV), the mechanisms behind this control remain unclear. Immunodominant CD8(+) T cell responses against the B57-restricted epitopes comprised of residues 2629 to 2637 of nonstructural protein 5B (NS5B(2629-2637)) (KSKKTPMGF) and E2(541-549) (NTRPPLGNW) were recently shown to be crucial in the control of HCV infection. Here, we investigated whether the selection of deleterious cytotoxic T lymphocyte (CTL) escape mutations in the NS5B KSKKTPMGF epitope might impair viral replication and contribute to the B57-mediated control of HCV. Common CTL escape mutations in this epitope were identified from a cohort of 374 HCV genotype 1a-infected subjects, and their impact on HCV replication assessed using a transient HCV replicon system. We demonstrate that while escape mutations at residue 2633 (position 5) of the epitope had little or no impact on HCV replication in vitro, mutations at residue 2629 (position 1) substantially impaired replication. Notably, the deleterious mutations at position 2629 were tightly linked in vivo to upstream mutations at residue 2626, which functioned to restore the replicative defects imparted by the deleterious escape mutations. These data suggest that the selection of costly escape mutations within the immunodominant NS5B KSKKTPMGF epitope may contribute in part to the control of HCV replication in B57-positive individuals and that persistence of HCV in B57-positive individuals may involve the development of specific secondary compensatory mutations. These findings are reminiscent of the selection of deleterious CTL escape and compensatory mutations by HLA-B57 in HIV-1 infection and, thus, may suggest a common mechanism by which alleles like HLA-B57 mediate protection against these highly variable pathogens.

Common variable immunodeficiency (CVID) is a heterogeneous antibody deficiency syndrome with alterations in T cell regulation and function in a subgroup of patients. We assessed phenotype and function of bulk and virus-specific CD8+ T cells of a cohort of 34 HLA-A2+ CVID patients by pentamer technology and flow cytometry in relationship to their immunological and clinical phenotypes. Bulk CD8+ T cells displayed a shift toward a more antigen experienced and activated differentiation state. The advanced differentiation pattern was mainly found in a subgroup of CVID patients with lymphadenopathy and granulomatous disease. This effect existed independently of the patients' CMV status even so CMV-associated immunosenescence was more evident in CVID patients than in CMV-positive immunocompetent controls. As the phenotype and function of virus-specific CD8+ T cells were normal in CVID the induction of antiviral immunity by prophylactic immunization appears to be a logical and desirable aim for this group of patients., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Hepatocellular carcinoma (HCC) is one of the most common malignancies with a poor prognosis and limited therapeutic options that is often characterized by the expression of the tumor-associated antigen α-fetoprotein (AFP). CD4+ helper T cells are important in generating potent anticancer immunity as they prime and expand CD8+ T-cell memory and may also have direct antitumor activity. However, very little information is currently available about the relative frequency, immunodominance and peripheral versus intratumoral distribution of AFP-specific CD4+ T-cell responses in patients with HCC. We, therefore, analyzed AFP-specific CD4+ responses in blood and tumor tissue of patients with HCC by using overlapping peptides spanning the entire AFP protein and novel sensitive approaches such as antigen-specific upregulation of CD154. We found that AFP-specific CD4+ T-cell responses were not detectable in the peripheral blood ex vivo. However, after in vitro stimulation, AFP-specific CD4+ T-cell responses were detectable in a large fraction of patients targeting different previously unreported epitopes with no clear immunodominance. These results indicate that AFP-specific CD4+ T-cell responses are not completely deleted but only present at very low frequencies. Importantly, AFP-specific CD4+ T-cell responses were also rarely detectable in tumor tissue, suggesting that the relative absence of these cells in the circulation ex vivo is not due to a rapid accumulation to the tumor side. Taken together, these results suggest that the lack of sufficient CD4+ T-cell help, especially within the tumor tissue, may be one central mechanism responsible for the failure of AFP-specific immune responses to control HCC progression., (Copyright © 2010 UICC.)

Unlabelled: Human leukocyte antigen B27 is associated with spontaneous viral clearance in hepatitis C virus (HCV) infection. Viral escape within the immunodominant, HLA-B27-restricted, HCV-specific, cluster of differentiation (CD)8(+) T-cell epitope, nonstructural protein (NS)5B(2841-2849) (ARMILMTHF), has been shown to be limited by viral fitness costs as well as broad T-cell cross-recognition, suggesting a potential mechanism of protection by HLA-B27. Here, we studied the subdominant HLA-B27-restricted epitope, NS5B(2936-2944) (GRAAICGKY), to further define the mechanisms of protection by HLA-B27. We identified a unique pattern of escape mutations within this epitope in a large cohort of HCV genotype 1a-infected patients. The predominant escape mutations represented conservative substitutions at the main HLA-B27 anchor residue or a T-cell receptor contact site, neither of which impaired viral replication capacity, as assessed in a subgenomic HCV replicon system. In contrast, however, in a subset of HLA-B27(+) subjects, rare escape mutations arose at the HLA-B27 anchor residue, R(2937) , which nearly abolished viral replication. Notably, these rare mutations only occurred in conjunction with the selection of two equally rare, and structurally proximal, upstream mutations. Coexpression of these upstream mutations with the rare escape mutations dramatically restored viral replication capacity from <5% to ≥ 70% of wild-type levels., Conclusion: The selection of rare CTL escape mutations in this HLA-B27-restricted epitope dramatically impairs viral replicative fitness, unless properly compensated. These data support a role for the targeting of highly constrained regions by HLA-B27 in its ability to assert immune control of HCV and other highly variable pathogens., (Copyright © 2011 American Association for the Study of Liver Diseases.)

Background & Aims: Inhibitory receptors such as programmed death 1 (PD-1) and cytotoxic T lymphocyte-associated antigen (CTLA)-4 mediate CD8+ T-cell exhaustion during chronic viral infection, but little is known about roles in dysfunction of CD4+ T cells., Methods: We investigated the functions of inhibitory molecules on hepatitis C virus (HCV)-, influenza-, and Epstein-Barr virus (EBV)-specific CD4+ T cells in patients with chronic infections compared with patients with resolved HCV infection and healthy donors. Expression of PD-1, CTLA-4, CD305, and CD200R were analyzed on HCV-specific CD4+ T cells, isolated from peripheral blood using major histocompatibility complex class II tetramers. We investigated the effects of in vitro inhibition of various inhibitory pathways on proliferation and cytokine production by CD4+ T cells, and we compared these effects with those from inhibition of interleukin (IL)-10 and transforming growth factor (TGF)-β1., Results: PD-1 and CTLA-4 were up-regulated on virus-specific CD4+ T cells from patients with chronic HCV infections. PD-1 expression was lower on influenza- than on HCV-specific CD4+ T cells from subjects with chronic HCV infection, whereas CTLA-4 was expressed at similar levels, independent of their specificity. CD305 and CD200R were up-regulated in HCV resolvers. Blockade of PD-L1/2, IL-10, and TGF-β1 increased expansion of CD4+ T cells in patients with chronic HCV, whereas inhibition of IL-10 and TGF-β1 was most effective in restoring HCV-specific production of interferon gamma, IL-2, and tumor necrosis factor α., Conclusions: We characterized expression of inhibitory molecules on HCV-, influenza-, and EBV-specific CD4+ T cells and the effects of in vitro blockade on CD4+ T-cell expansion and cytokine production. Inhibition of PD-1, IL-10, and TGF-β1 is most efficient in restoration of HCV-specific CD4+ T cells., (Copyright © 2011 AGA Institute. Published by Elsevier Inc. All rights reserved.)

With up to 400 million affected people worldwide, chronic hepatitis B virus (HBV) infection is still a major health care problem. During the last decade, several novel therapeutic approaches have been developed and evaluated. In most regions of the world, interferon-α, and nucleos(t)ide analogues (NUCs) are currently approved. Despite major improvements, none of the existing therapies is optimal since viral clearance is rarely achieved. Recently, a better understanding of the HBV life cycle and the development of novel model systems of HBV infection have led to the development of novel antiviral strategies and drug targets. This review will focus on current and potential future drug targets in the HBV life cycle and strategies to modulate the virus-host interaction.

The impact of naïve-precursor frequency on human virus-specific CD8(+) T cell immunodominance is not well understood. Using a recently developed major histocompatibility complex (MHC) class I tetramer enrichment protocol, we found a conserved hierarchy and a >10-fold difference in naïve-precursor frequencies across three HLA-A2-restricted hepatitis C virus (HCV)-specific epitopes. Importantly, the NS3(1406) epitope with the highest naïve-precursor frequency in healthy donors was also the most frequently targeted epitope in a large cohort of chronically HCV-infected patients, both ex vivo and after in vitro stimulation. These results indicate for the first time that immunodominance in a human viral infection is linked to naïve-precursor frequency.

Upon infection with hepatitis C virus (HCV), only few patients spontaneously clear the virus, while most patients develop chronic HCV infection. The host innate and adaptive immune response is believed to be the key determinant of viral clearance or persistence. Several host factors have been demonstrated to influence the efficiency of the antiviral immune response, including IL-28B polymorphisms, inhibitory natural killer cell receptors, as well as HLA class I and II alleles presenting viral antigens to CD8(+) and CD4(+) T cells. The understanding of the respective mechanisms is essential for the development of successful vaccination strategies against HCV.

Primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC) are important causes of cholestatic liver disease. IgG4-associated cholangitis (IAC) also belongs to the same entity. Overlap syndromes combine characteristics of cholestatic liver diseases and autoimmune hepatitis. The diagnosis of PBC is based on the detection of anti-mitochondrial antibodies. PBC is frequently associated with other autoimmune disorders. The treatment of choice is ursodeoxycholic acid. PSC is frequently associated with inflammatory bowel disease (IBD). The cholangiography shows characteristic bile duct lesions. Bile duct strictures and bacterial cholangitis should be treated by dilatation and antibiotics, respectively. Cirrhosis may ultimately develop in PBC and PSC. In advanced PBC or PSC, liver transplantation might be indicated. The clinical course of IAC is similar to PSC. In contrast to PSC, however, there is no association with IBD.

Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide with limited therapeutic options. Since HCC has been shown to be immunogenic, T cell-based immunotherapy is considered a promising treatment. In this review, we summarize current knowledge of T cell responses against tumour-associated antigens, as well as the mechanisms underlying the poor quality of these responses in patients with HCC. Insights into these important aspects of HCC immunology are crucial for the further development of novel immunotherapies., (Copyright © 2010 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Effective host immune responses are essential for the control of hepatitis C virus (HCV) infection and persistence of HCV has indeed been attributed to their failure. In recent years, several in vitro and in vivo experimental models have allowed studies of host immune responses against HCV. Numerous observations derived from these models have improved our understanding of the mechanisms responsible for the host's ability to clear the virus as well as of the mechanisms responsible for the host's failure to control HCV replication. Importantly, several findings obtained with these model systems have been confirmed in studies of acutely or chronically HCV-infected individuals. Collectively, several mechanisms are used by HCV to escape host immune responses, such as poor induction of the innate immune response and escaping/impairing adaptive immunity. In this review, we summarize current findings from experimental models available for studies of the immune response targeting HCV and discuss the relevance of these findings for the in vivo situation in HCV-infected humans.

Hepatitis C virus (HCV) infection is only cleared in a minority of infected individuals, the majority of patients develop chronic infection. Chronic HCV infection potentially leads to liver fibrosis, cirrhosis and finally hepatocellular carcinoma. The host immune response is an important determinant in the outcome of HCV infection. Innate as well as adaptive cellular and humoral immune responses mediate important antiviral actions; however, virus-specific T cell responses appear to be most critical. Indeed, strong and multispecific CD4+ as well as CD8+ T cell responses are required for viral clearance. Interestingly, individuals who express certain HLA alleles (which are important for antigen presentation to CD4+ and CD8+ T cells) have a higher chance to clear the virus. The mechanisms of protection by HLA class I alleles such as HLA-B27 have been characterized recently. In most individuals, however, the HCV-specific immune response fails to clear the virus. Several mechanisms underlying this HCV-specific T cell failure have been identified. These include viral factors such as viral escape mutations and immunological factors such as the expression of inhibitory receptors, which lead to CD8+ T cell dysfunction. An in-depth understanding of the determinants of success or failure of the HCV-specific T cell response is critical for the development of prophylactic as well as therapeutic vaccination regimes against HCV. Here, we will discuss the virological and immunological determinants of HCV clearance and persistence., (Copyright © 2011 S. Karger AG, Basel.)

Genome-wide association studies (GWAS) are aimed to identify genetic markers of complex human diseases and individual traits. In this context more than 150 gene loci have been found to be associated with about 60 different diseases and personal characteristics. A recent example is the relevance of a polymorphism in the interleukin 28B gene for the natural course of hepatitis C virus infection and the efficacy of antiviral therapy. It is to be expected that GWAS will increasingly contribute to the understanding of the pathogenesis and consquently improve prediction, prevention, diagnosis and therapy of human diseases and eventually will become part of clinical practice., (© Georg Thieme Verlag KG Stuttgart · New York.)

Whether the recently identified innate lymphocyte population coexpressing natural killer cell receptors (NKRs) and the nuclear receptor RORγt is part of the NK or lymphoid tissue inducer (LTi) cell lineage remains unclear. By using adoptive transfer of genetically tagged LTi-like cells, we demonstrate that NKR⁻RORγt(+) innate lymphocytes but not NK cells were direct progenitors to NKR(+)RORγt(+) cells in vivo. Genetic lineage tracing revealed that the differentiation of LTi-like cells was characterized by the stable upregulation of NKRs and a progressive loss of RORγt expression. Whereas interleukin-7 (IL-7) and intestinal microbiota stabilized RORγt expression within such NKR-LTi cells, IL-12 and IL-15 accelerated RORγt loss. RORγt(+) NKR-LTi cells produced IL-22, whereas RORγt⁻ NKR-LTi cells released IFN-γ and were potent inducers of colitis. Thus, the RORγt gradient in NKR-LTi cells serves as a tunable rheostat for their functional program. Our data also define a previously unappreciated role of RORγt⁻ NKR-LTi cells for the onset or maintenance of inflammatory bowel diseases., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Exhausted CD8+ T cell responses during chronic viral infections are defined by a complex expression pattern of inhibitory receptors. However, very little information is currently available about the coexpression patterns of these receptors on human virus-specific CD8+ T cells and their correlation with antiviral functions, T cell differentiation and antigen recognition. We addressed these important aspects in a cohort of 38 chronically HCV infected patients and found a coexpression of inhibitory receptors such as 2B4, CD160 and KLRG1 in association with PD-1 in about half of the HCV-specific CD8+ T cell responses. Importantly, this exhaustive phenotype was associated with low and intermediate levels of CD127 expression, an impaired proliferative capacity, an intermediate T cell differentiation stage and absence of sequence variations within the corresponding epitopes, indicating ongoing antigen triggering. In contrast, a low expression of inhibitory receptors by the remaining HCV-specific CD8+ T cells occurred in concert with a CD127hi phenotype, an early T cell differentiation stage and presence of viral sequence variations within the corresponding epitopes. In sum, these results suggest that T cell exhaustion contributes to the failure of about half of HCV-specific CD8+ T cell responses and that it is determined by a complex interplay of immunological (e.g. T cell differentiation) and virological (e.g. ongoing antigen triggering) factors.

Background & Aims: Virus-specific CD4+ T cells play a major role in hepatitis C virus (HCV) infection. Viral clearance is associated with vigorous and multispecific CD4+ T cell responses, while chronic infection has been shown to be associated with weak or absent T cell responses. Most of these studies, however, have used functional assays to analyse virus-specific CD4+ T cell responses. Therefore, the important question, of whether virus-specific CD4+ T cells are completely absent or primarily impaired in specific effector functions during chronic infection, has yet to be analysed in detail., Methods: To address this issue, a novel assay, where CD4+ T cell frequencies can be determined by de novo CD154 (CD40 ligand) expression in response to HCV antigens, was used in a cohort of chronically infected HCV patients and patients who spontaneously resolved HCV infection. These responses were compared to functional assays, such as the IFN-gamma ELISpot and flow cytometry-based proliferative assays., Results: Our results reveal that using the CD154 assay, virus-specific CD4+ T cells are readily detectable during chronic HCV infection albeit at a lower frequency when compared to patients who spontaneously resolved the infection. Importantly, no CD4+ T cell responses were detectable from these patients when using functional assays. Finally, these cell populations were enriched in the intrahepatic compartment., Conclusions: Our findings suggest that HCV-specific CD4+ T cell responses are readily detectable in chronic HCV infection and enriched in the infected liver., (Copyright 2010 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

CD8(+) T lymphocytes play a key role in host defense, in particular against important persistent viruses, although the critical functional properties of such cells in tissue are not fully defined. We have previously observed that CD8(+) T cells specific for tissue-localized viruses such as hepatitis C virus express high levels of the C-type lectin CD161. To explore the significance of this, we examined CD8(+)CD161(+) T cells in healthy donors and those with hepatitis C virus and defined a population of CD8(+) T cells with distinct homing and functional properties. These cells express high levels of CD161 and a pattern of molecules consistent with type 17 differentiation, including cytokines (e.g., IL-17, IL-22), transcription factors (e.g., retinoic acid-related orphan receptor gamma-t, P = 6 x 10(-9); RUNX2, P = 0.004), cytokine receptors (e.g., IL-23R, P = 2 x 10(-7); IL-18 receptor, P = 4 x 10(-6)), and chemokine receptors (e.g., CCR6, P = 3 x 10(-8); CXCR6, P = 3 x 10(-7); CCR2, P = 4 x 10(-7)). CD161(+)CD8(+) T cells were markedly enriched in tissue samples and coexpressed IL-17 with high levels of IFN-gamma and/or IL-22. The levels of polyfunctional cells in tissue was most marked in those with mild disease (P = 0.0006). These data define a T cell lineage that is present already in cord blood and represents as many as one in six circulating CD8(+) T cells in normal humans and a substantial fraction of tissue-infiltrating CD8(+) T cells in chronic inflammation. Such cells play a role in the pathogenesis of chronic hepatitis and arthritis and potentially in other infectious and inflammatory diseases of man.

Systematic studies of the circulation of hepatitis C virus (HCV) recombinants in different parts of the world have been initiated only recently, and no detailed information on this subject is available. The aim of the current investigation was to determine the frequency of HCV recombinants in intravenous drug users (IVDU) from two European countries. HCV RNA from serum samples was tested by RT-PCR with primers derived from the core and NS5B regions with subsequent sequencing and genotype assignment. The 118 samples from Germany (100%) and 45 out of 47 (96%) sera from Russia demonstrated concordant genotyping results. In the two genotype discrepant sera from Russia 2k/1b recombinants were identified. In order to test the hypothesis that the individuals from the IVDU group might be multiply exposed to various genotypes, 145 out of 165 genotyped serum samples, which were found to be positive for anti-NS4 antibodies, were serotyped with the Murex HCV serotyping kit that is based on detection of antibodies to type-specific peptides derived from the NS4 proteins of different HCV genotypes. Discrepancy in genotype and serotype attributions was observed in 11% cases. Retesting of 99 type 1a or 3a samples with a set of type- and subtype-specific primers revealed the presence of a mixed infection only in one case (1a/3a). Thus, the cases of the mixed infection with different HCV genotypes as well as the recombinant forms of HCV are very rare even in such a highly exposed group as IVDU., ((c) 2009 Wiley-Liss, Inc.)

Hepatitis C virus (HCV)-specific CD8(+) T cells in persistent HCV infection are low in frequency and paradoxically show a phenotype associated with controlled infections, expressing the memory marker CD127. We addressed to what extent this phenotype is dependent on the presence of cognate antigen. We analyzed virus-specific responses in acute and chronic HCV infections and sequenced autologous virus. We show that CD127 expression is associated with decreased antigenic stimulation after either viral clearance or viral variation. Our data indicate that most CD8 T-cell responses in chronic HCV infection do not target the circulating virus and that the appearance of HCV-specific CD127(+) T cells is driven by viral variation.

Unlabelled: Human leukocyte antigen B27 (HLA-B27) is associated with protection in human immunodeficiency virus (HIV) and hepatitis C virus (HCV) infection. This protective role is linked to single immunodominant HLA-B27-restricted CD8+ T-cell epitopes in both infections. In order to define the relative contribution of a specific HLA-B27-restricted epitope to the natural course of HCV infection, we compared the biological impact of the highly conserved HCV genotype 1 epitope, for which the protective role has been described, with the corresponding region in genotype 3 that differs in its sequence by three amino acid residues. The genotype 3a peptide was not recognized by CD8+ T cells specific for the genotype 1 peptide. Furthermore, patients with acute or chronic infection with HCV genotype 3a did not mount T-cell responses to this epitope region, and their autologous viral sequences showed no evidence of T-cell pressure. Finally, we found a significantly higher frequency of HLA-B27 positivity in patients with chronic HCV genotype 3a infection compared to genotype 1 infection, indicating that there is no protection by HLA-B27 in HCV genotype 3 infection., Conclusion: Our data indicate that the protective effect of HLA-B27 is limited to HCV genotype 1 infection and does not expand to other genotypes such as genotype 3a. This can most likely be explained by intergenotype sequence diversity leading to the loss of the immunodominant HLA-B27 epitope in viral strains other than genotype 1. Our results underline the central role of a single HLA-B27-restricted epitope-specific CD8+ T-cell response in mediating protection in HCV genotype 1 infection.

Hepatocellular carcinoma (HCC) is one of the most common malignant tumors worldwide. The major etiologies and risk factors for HCC development are well defined and some of the multiple steps involved in hepatocarcinogenesis have been elucidated in recent years. The therapeutic options fall into five main categories: (1) surgical interventions, incl. liver transplantation, (2) percutaneous interventions, incl. ethanol injection and radiofrequency thermal ablation, (3) transarterial interventions, (4) radiation therapy and (5) drugs as well as gene and immune therapies. Because of the poor survival of the majority of patients, HCC prevention as well as early diagnosis and the development of novel systemic therapies for advanced disease are of paramount importance. In this context, recent data indicate that the 'targeted therapy' with monoclonal antibodies (mabs) or small molecule tyrosine kinase inhibitors (nibs) and other drugs seem to be effective to some degree. New technologies, including gene expression profiling and proteomic analyses, should allow to further elucidate the molecular events underlying HCC development and to identify novel diagnostic markers as well as therapeutic and preventive targets.

Elevated serum amino-transferase levels may be associated with liver injury. Testing for aspartate aminotransferase (AST) or alanine aminotransferase (ALT) is part of many routine screening approaches. The aim of this manuscript was to scrutinize the evidence for using ALT testing as a primary screening parameter for liver diseases. We conclude that (i) elevated serum ALT levels indicate a high specificity and a reasonable sensitivity liver injury, (ii) 10 - 25 % of German adults have elevated ALT levels, (iii) ALT values are increased in the majority but not all patients with acute and chronic liver disease (iv) elevated ALT-values are associated with an increased risk of liver-specific mortality, (v) elevated ALT values are also a risk factor for non-hepatic diseases including diabetes mellitus type 2, metabolic syndrome, cardiovascular diseases and malignancies, (vi) many liver diseases identified by an ALT screening can be treated successfully including prevention of development of clinical endpoints, (vii) an ALT-screening is very likely to be cost-effective although studies are needed for Germany to support this conclusion.

Hepatitis B and C viruses (HBV and HCV) cause chronic hepatitis and hepatocellular carcinoma (HCC) by poorly understood mechanisms. We show that cytokines lymphotoxin (LT) alpha and beta and their receptor (LTbetaR) are upregulated in HBV- or HCV-induced hepatitis and HCC. Liver-specific LTalphabeta expression in mice induces liver inflammation and HCC, causally linking hepatic LT overexpression to hepatitis and HCC. Development of HCC, composed in part of A6(+) oval cells, depends on lymphocytes and IKappa B kinase beta expressed by hepatocytes but is independent of TNFR1. In vivo LTbetaR stimulation implicates hepatocytes as the major LT-responsive liver cells, and LTbetaR inhibition in LTalphabeta-transgenic mice with hepatitis suppresses HCC formation. Thus, sustained LT signaling represents a pathway involved in hepatitis-induced HCC.

Although there is strong evidence that hepatitis C virus (HCV) is one of the leading causes of hepatocellular carcinoma (HCC), there is still much to understand regarding the mechanism of HCV-induced transformation. While liver fibrosis resulting from long-lasting chronic inflammation and liver regeneration resulting from immune-mediated cell death are likely factors that contribute to the development of HCC, the direct role of HCV proteins remains to be determined. In vitro studies have shown that HCV expression may interfere with cellular functions that are important for cell differentiation and cell growth. However, most studies were performed in artificial models which can only give clues for potential mechanisms that need to be confirmed in more relevant models. Furthermore, the difficulty to identify HCV proteins and infected liver cells in patients, contributes to the complexity of our current understanding. For these reasons, there is currently very little experimental evidence for a direct oncogenic role of HCV. Further studies are warranted to clarify these issues.

The natural history of hepatitis C virus (HCV) infection is determined by a complex interplay between host genetic, immunological and viral factors. This review highlights genes involved in innate and adaptive immune responses associated with different outcomes of HCV infection. For example, an association of HCV clearance with certain HLA alleles has been demonstrated. The mechanisms responsible for these associations have been linked to specific T cell responses for some particular alleles (e.g., HLA-B27). Genetic associations involved in T cell regulation and function further underline the role of the adaptive immune response in the natural history of HCV infection. In addition, some genes involved in innate NK cell responses demonstrate the complex interplay between components of the immune system necessary for a successful host response to HCV infection.

We report here the synthesis and preliminary evaluation of novel 1-(4-methoxyphenethyl)-1H-benzimidazole-5-carboxylic acid derivatives 6(a-k) and their precursors 5(a-k) as potential chemotherapeutic agents. In each case, the structures of the compounds were determined by FTIR, (1)H NMR and mass spectroscopy. Among the synthesized molecules, methyl 1-(4-methoxyphenethyl)-2-(4-fluoro-3-nitrophenyl)-1H-benzimidazole-5-carboxylate (5a) induced maximum cell death in leukemic cells with an IC(50) value of 3 microM. Using FACS analysis we show that the compound 5a induces S/G2 cell cycle arrest, which was further supported by the observed down regulation of CDK2, Cyclin B1 and PCNA. The observed downregulation of proapoptotic proteins, upregulation of antiapoptotic proteins, cleavage of PARP and elevated levels of DNA strand breaks indicated the activation of apoptosis by 5a. These results suggest that 5a could be a potent anti-leukemic agent.