Chemoimmunotherapy of Chronic Hepatitis B Virus Infection in the Woodchuck Model Overcomes Immunologic Tolerance and Restores T-Cell Responses to Pre-S and S Regions of the Viral Envelope Protein

Abstract

Treatment of chronic hepatitis B virus (HBV) infection could combine potent antiviral drugs and therapeuticvaccines to overcome immunological tolerance and induce the recovery phenotype to protect against disease progression. Conventional vaccination of woodchucks chronically infected with the woodchuck hepatitis virus (WHV) elicited differential T-cell response profiles depending on whether or not carriers were treated with the potent antiviral drug clevudine (CLV), which significantly reduces viral and antigen loads. The differential T-cell responses defined both CLV-dependent and CLV-independent epitopes of the pre-S and S regions of the WHV envelope protein. Only combined treatment involving CLV and conventional vaccine therapeutically restored the T-cell response profile of chronic WHV carrier woodchucks to that seen in prophylactic vaccination and in recovery from acute WHV infection. The results have implications for mechanisms of immunological tolerance operating in chronic HBV infection and suggest that such combined chemoimmunotherapy may be useful for treatment of humans with chronic HBV infection. Chronic hepatitis B virus (HBV) infection is often associatedwith immunological tolerance to the virus characterized by hyporesponsive T helper (Th) cells, reduced numbers of cytolytic T lymphocytes, diminished Th1-type cytokine responses, and undetectable virus-neutralizing antibodies to viral envelope proteins (3,8,9,16,20,24,27,42, 46). When HBVspecific cellular immune responses sometimes become detectable in HBV carriers, they are often suboptimal and contribute more to disease progression than to viral clearance and recovery (8,16,27, 46). Immunological tolerance in chronic HBV infection (2,4,6,7,37,38) may arise theoretically from central or peripheral tolerance mechanisms (or both). Central tolerance could involve negative selection of antigen-specific T cells by thymic deletion in the presence of antigen (6,7,37,38). Peripheral tolerance may follow after positive selection of antigen-specific T cells and result from clonal anergy, immunological exhaustion, or altered regulation between Th1 and Th2 cells (1, 43). The development and/or maintenance of central and peripheral tolerance in chronic HBV infection may be a consequence of the high viral and antigen loads often observed in chronic carriers. The development of immunotherapeutics able to circumvent T-cell tolerance, alone or in combination with antiviral therapeutics, represents a further critical step toward the successful treatment of chronic HBV infection.Vaccines for HBV will ultimately interdict transmission of the virus and eradicate HBV-related diseases. However, there are currently more than 350 million chronic carriers of HBV worldwide who are at risk of developing chronic hepatitis, cirrhosis, and hepatocellular carcinoma (HCC) (47). Treatment options for chronic HBV infection are limited presently. Pegylated alpha interferon brings about sustained antiviral responses in approximately one-third of patients (25) but is associated with ...

Treatment of Chronic Viral Hepatitis in Woodchucks by Prolonged Intrahepatic Expression of Interleukin-12

“…Similar data have emerged from other studies showing that woodchucks with chronic WHV infection were able to mount a strong antigen-specific immune response fol- lowing vaccination only after prolonged viremia reduction with potent antiviral drugs given for several months (19). Interestingly, we found that PBMCs from woodchucks with high viremia levels showed an impairment in the induction of IFN-␥ following stimulation with IL-12, while this alteration was not observed in animals with low viremia levels.…”

“…The presence of serum WHV surface antigen (WHsAg) levels was determined by enzyme-linked immunosorbent assay (ELISA) (2,19). The WHV e antigen (WHeAg) concentration was determined using a commercial ELISA for HBVe antigen that recognizes woodchuck antigen (DiaSorin S.p.A., Saluggia, Italy).…”

Semliki Forest Virus Expressing Interleukin-12 Induces Antiviral and Antitumoral Responses in Woodchucks with Chronic Viral Hepatitis and Hepatocellular Carcinoma

“…For determining T-cell responses against WHV, in vitro stimulators were used at concentrations previously determined as optimal for cultures of woodchuck peripheral blood mononuclear cells (PBMCs) (31,32). Viral antigen stimulators consisted of native 22-nm WHsAg (2 g/ml) and synthetic peptides of WHsAg and WHcAg.…”

“…Viral antigen stimulators consisted of native 22-nm WHsAg (2 g/ml) and synthetic peptides of WHsAg and WHcAg. WHs peptide S18 (10 g/ml; SigmaGenosys, The Woodlands, TX) corresponded to amino acids 341 to 360 of WHsAg starting at the N terminus of the large surface protein (32). WHc peptide C100-119 (10 g/ml; Sigma-Genosys) corresponded to amino acids 100 to 119 of WHcAg starting at the N terminus of WHcAg (31).…”

Immunopathogenesis and Immunotherapy for Viral Hepatitis

“…Most of the mice showed reduction of HBV DNA level and disappearance of HBeAg and HBsAg. Menne et al, 2007; A combination of conventional vaccine on the base of the WHV large surface protein contained HBsAg with pre-S and clevudine significantly restored the Tcell response to Pre-S and S region in chronic WHV infection. Miller at al, 2008; One hour post-infection with DHBV, DNA vaccine expressing DHBc and Pre-S/S and entecavir were given simultaneously and continued for 14 days.…”