Immune modulation in chronic HBV and HCV infection

Abstract

Chronic viral hepatitis is a major health problem worldwide. The two major viral causes for chronic hepatitis are the hepatitis B (HBV) and C viruses (HCV). HBV is a DNA virus with a rather limited genetic variability whereas HCV is an RNA virus with a high genetic variability. Thus, the two hepatotropic viruses have adopted distinct strategies to persist in humans. For HCV the high genetic variability is most likely of great importance in evading the host immune response. One mechanism proposed for persistence of HBV infection is the over production of viral proteins, which may block humoral responses and impair cell-mediated immune responses. In both infections the type of immune response that is evoked in the host is thought to be one factor determining the outcome of the infection. As an example, a balance between T helper-1 (Th 1) and Th2-like cells and cytokine production skewed towards Th1 has been proposed to favour clearance of HBV and HCV infections. Today only little is known on how different antiviral therapies affect the immune responses nod the Th1/Th2 balance. The aim of this thesis was to further characterise the humoral and cellular immune responses to HBV/HCV and to investigate the immune modulatory effects of current and experimental therapies for chronic viral hepatitis.

There is a limited knowledge on the importance of vim] variability in the interaction with the host CD4+ T cell response, We therefore developed a simple model using one of the smallest proteins of HCV, the non-structural (NS) protein 4A. We found that the combination of the viral genotype and the host MHC determined the ability to mount an NS4A-specific immune response. Only a few amino acid changes within a CD4+ T cell site were sufficient to convert a murine responder haplotype to a nonresponder. Thus, viral mutations within CD4+ T cell sites should be an effective way to escape or modulate the host immune response. The CD4+ T cell response in patients with chronic HCV infection has been studied previously, though in most studies only a few samples have been obtained from the same patient. We herein performed a close monitoring of the CD4+ T cell response against HCV NS3 during antiviral therapy. We found that transient CD4+ T cell responses to HCV NS3 developed during therapy, regardless of the outcome of therapy. These transient responses coincided with changes in the viral load, suggesting that the viral load might be affected by HCV-specific CD4+ T cells. The transient nature of these responses suggest that these cells most likely represent a temporary excess or activation of intra-hepatic T cells caused by changes in the viral replication due to the antiviral therapy. Excess T-cells are then redistributed or simply spill over in to the periphery. The function of HBV- and HCV-specific Th1 and Th2-like CD4+ T cells in vivo has been studied previously. These studies have suggested that a function of HBeAg may be to cross to the placenta to induce intra marine T cell tolerance. In HBeAg-Tg mice the degree of T cell tolerance to HBeAg is MHC dependent and mainly Th2-like HBeAg-specific T cells seem to evade tolerance. To evaluate some functional properties of HBeAg-specific Th2 cells an IL-4 producing HBc/eAg specific M clone (CNT7) was established. When the CNT7 clone was adopetively transferred into unprimed HBeAg-Tg H-2b mice a rapid production of anti-HBe was observed without signs of liverdisease. Thus, the Th2-mediated anti-HBe production in the HBeAg-Tg mice should be of value to study the effects of compounds that modulate the Th1/Th2 balance in vivo. The HBeAg-Tg mouse model was used to study the effects of ribavirin. Ribavirin has been shown to exert a synergistic effect in combination therapy with [alpha]IFN of chronic HCV infection. However, the mechanism of action has not been completely understood. Ribavirin mono-therapy of chronic HBV and HCV infections normalises the transaminase levels in many patients, but has no clear effects on the viremia. We investigated the effect of ribavirin in vitro and in vivo. We found that ribavirin promotes the development of [gamma]IFN producing Th1-like CD4+ T cells in vivo thereby skewing the Th1/Th2 balance. Thus, these data suggest that one beneficial effect of ribavirin in the treatment of chronic viral hepatitis may be a shift towards Th1-like immune responses. Thus, ribavirin might not be regarded strictly as an antiviral compound.

In the mid 1990s new antiviral compounds were introduced for the therapy of chronic HBV infections. These compounds have (his far mainly been used in mono therapies. We evaluated the use of a nucleoside analogue combination therapy in a patient with chronic replicative HBV infection and studied their effect on the endogenous CD4+ T cell responses. The regimen was well tolerated and clearly improved the well-being of the patient. We were able to detect a transient HBc/eAg-specific T cell proliferative and cytokine ([gamma]IFN and IL- 12) response in the periphery at four to six weeks from therapy start. This response coincided with the start of a stable decrease and normalisation of ALT levels. Thus, these data, together with those obtained in chronic HCV infections, suggest that the transient proliferative peak detected in the periphery may represent a redistribution or leakage of intra-hepatic CD4+ T cells. If so, this underscores the antiviral activity of Th1-like CD4+ T cells. Th1-like CD4+ T cells are considered to be important for control and clearance of HBV and HCV infections. We were therefore interested to analyse how these most effectively can be induced. The effectiveness of the two DNA based immunogens to prime CD4+ T cells were compared to regular immunisation with recombinant NS3 (rNS3) protein. rNS3 in adjuvant primed the most rapid and vigorous CD4+ T cell responses consisting of a mixed Th1 and Th2 population. In contrast, the CD4+ T cell responses primed by the NS3-expressing DNA vectors developed slower and were of a lower magnitude. However, NS3-specific humoral and CD4+ T cells primed by DNA immunisation were mainly restricted to a Th1-like phenotype. Thus, DNA based vectors, although quantitatively less effective may prim the desired type of CD4+ T cell responses. The present studies further emphasise the importance of CD4+ T cells in the control of chronic HBV and HCV infections. Compounds and strategies that modulate or enhance the endogenous CD4+ T cell responses may therefore be of importance in future therapies of chronic HBV and HCV infections.