DNA vaccination in combination or not with lamivudine treatment breaks humoral immune tolerance and enhances cccDNA clearance in the duck model of chronic hepatitis B virus infection

Abstract

This study used a duck hepatitis B virus (DHBV) model to evaluate whether a novel DNA vaccination protocol alone or associated with antiviral (lamivudine) treatment was able to clear the intrahepatic covalently closed, circular viral DNA (cccDNA) pool responsible for persistence of infection. DHBV carriers received DNA vaccine (on weeks 6, 10, 13, 14, 28 and 35) targeting the large envelope and/or core proteins alone or combined with lamivudine treatment (on weeks 1-8) or lamivudine monotherapy. After 10 months of follow-up, a dramatic decrease in viraemia and liver DHBV cccDNA (below 0.08 cccDNA copies per cell) was observed in 9/30 ducks (30 %) receiving DNA mono-or combination therapy, compared with 0/12 (0 %) from lamivudine monotherapy or the control groups, suggesting a significant antiviral effect of DNA immunization. However, association with the drug did not significantly improve DHBV DNA vaccine efficacy (33 % cccDNA clearance for the combination vs 27 % for DNA monotherapy), probably due to the low antiviral potency of lamivudine in the duck model. Seroconversion to anti-preS was observed in 6/9 (67 %) ducks showing cccDNA clearance, compared with 1/28 (3.6 %) without clearance, suggesting a significant correlation (P,0.001) between humoral response restoration and cccDNA elimination. Importantly, an early (weeks 10-12) drop in viraemia was observed in seroconverted animals, and virus replication did not rebound following the cessation of immunotherapy, indicating a sustained effect. This study provides the first evidence that therapeutic DNA vaccination is able to enhance hepadnaviral cccDNA clearance, which is tightly associated with a break in humoral immune tolerance. These results also highlight the importance of antiviral drug potency and an effective DNA immunization protocol for the design of therapeutic vaccines against chronic hepatitis B.

In Vivo Proliferation of Hepadnavirus-Infected Hepatocytes Induces Loss of Covalently Closed Circular DNA in Mice

“…Understanding molecular mechanisms affecting stability and activity of the cccDNA in vivo may reveal new therapeutic targets for the development of novel antiviral strategies able to enhance cccDNA clearance. 11,20 Although previous studies have noted that cccDNA turnover is slow even following administration of potent polymerase inhibitors, [5][6][7]12 little is known about the kinetics of cccDNA decay in the context of liver regeneration. WM-HBV-infected PTHs generated in uPA chimeric mice do not undergo hepatocellular changes, such as lipid droplet accumulation, nor do they show any sign of damage after longterm residence in immunosuppressive uPA mice, as has been reported for human hepatocytes.…”

“…However, it has been proposed that cell division may favor dilution of the cccDNA, so that cccDNA-free cells can be generated while infected cells are forced to divide to compensate for the immunomediated loss of other infected cells. [9][10][11] Notably, studies in the duck model showed that antiviral therapy with polymerase inhibitors induced a greater cccDNA reduction in animals displaying higher hepatocyte proliferation rates. 12 A cccDNA decrease was also observed in hepatocytes chronically infected with woodchuck hepatitis virus when cell turnover was induced in vitro by addition of cellular growth factors and viral replication was suppressed by adefovir treatment.…”

Immunopathogenesis and Immunotherapy for Viral Hepatitis

“…In a model of duck hepatitis B virus infection, DNA vaccine encoding HBV large envelope and/or core protein was shown to induce reduction in not only viremia but also cccDNA in the liver in one third of ducks receiving DNA monotherapy or combination treatment along with lamivudine (Thermet et al, 2008). This finding is encouraging because clearance of cccDNA from the liver is the goal of treatment for HBV infection, but is difficult to achieve using IFN- or nucleoside analogs.…”

“…Ducks boosted with fowl poxvirus vectors expressing DHBc and Pre-S/S showed clearance of DHBV infection at a rate of 100%. Thermet et al, 2008; DNA vaccine encoding the DHBV large envelope and/or core protein was given 6 times with or without lamivudine in a DHBV model. Reduction of viremia and liver DHBV cccDNA was observed in 33% of ducks receiving DNA vaccine mono-or combination therapy.…”

Immune therapy for hepatitis B

“…HBsAg-based vaccines have shown HBV DNA negativity and seroconversion to anti-HBs in HBV TM (36,37). HBsAg-expressing DNA vaccines and combination of DNA vaccines with antiviral drugs have induced reduction or clearance of DHBV DNA and clearance of duck HBsAg (38,39). Immunizations of woodchucks with HBsAg-based vaccines or combination of vaccines and antiviral agents have also shown anti-HBs response and significant reductions of viral load (40).…”