Enhanced magnitude and breadth of neutralizing humoral response to a DNA vaccine targeting the DHBV envelope protein delivered by in vivo electroporation

Université Lyon 1, France.
Virology (Impact Factor: 3.32). 03/2012; 425(1):61-9. DOI: 10.1016/j.virol.2012.01.001
Source: PubMed


We explored in the duck hepatitis B virus (DHBV) model the impact of electroporation (EP)-mediated DNA vaccine delivery on the neutralizing humoral response to viral preS/S large envelope protein. EP enhanced the kinetics and magnitude of anti-preS response compared to the standard needle DNA injection (SI). Importantly, EP dramatically enhanced the neutralizing potency of the humoral response, since antibodies induced by low DNA dose (10 μg) were able to highly neutralize DHBV and to recognize ten antigenic regions, including four neutralization epitopes. Whereas, SI-induced antibodies by the same low DNA dose were not neutralizing and the epitope pattern was extremely narrow, since it was limited to only one epitope. Thus, EP-based delivery was able to improve the dose efficiency of DNA vaccine and to maintain a highly neutralizing, multi-specific B-cell response, suggesting that it may be an effective approach for chronic hepatitis B therapy at clinically feasible DNA dose.

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    • "Since HBV has an extremely narrow host range infecting only humans and chimpanzee, the closely related DHBV represents a reference and a very useful model to analyze gene function, viral replication and to evaluate novel antiviral strategies [27], [28], [29], [30], [31], [32], [33]. In search for new anti-HBV approaches, we have investigated in this model the ability of Peptide Nucleic Acids (PNAs) targeting the HBV signal ε to inhibit viral reverse transcription. "
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    ABSTRACT: Cationic cell-penetrating peptides (CPPs) and their lipid domain-conjugates (CatLip) are agents for the delivery of (uncharged) biologically active molecules into the cell. Using infection and transfection assays we surprisingly discovered that CatLip peptides were able to inhibit replication of Duck Hepatitis B Virus (DHBV), a reference model for human HBV. Amongst twelve CatLip peptides we identified Deca-(Arg)(8) having a particularly potent antiviral activity, leading to a drastic inhibition of viral particle secretion without detectable toxicity. Inhibition of virion secretion was correlated with a dose-dependent increase in intracellular viral DNA. Deca-(Arg)(8) peptide did neither interfere with DHBV entry, nor with formation of mature nucleocapsids nor with their travelling to the nucleus. Instead, Deca-(Arg)(8) caused envelope protein accumulation in large clusters as revealed by confocal laser scanning microscopy indicating severe structural changes of preS/S. Sucrose gradient analysis of supernatants from Deca-(Arg)(8)-treated cells showed unaffected naked viral nucleocapsids release, which was concomitant with a complete arrest of virion and surface protein-containing subviral particle secretion. This is the first report showing that a CPP is able to drastically block hepadnaviral release from infected cells by altering late stages of viral morphogenesis via interference with enveloped particle formation, without affecting naked nucleocapsid egress, thus giving a view inside the mode of inhibition. Deca-(Arg)(8) may be a useful tool for elucidating the hepadnaviral secretory pathway, which is not yet fully understood. Moreover we provide the first evidence that a modified CPP displays a novel antiviral mechanism targeting another step of viral life cycle compared to what has been so far described for other enveloped viruses.
    PLoS ONE 11/2012; 7(11):e48721. DOI:10.1371/journal.pone.0048721 · 3.23 Impact Factor
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    • "Screening of induced antibodies in ducks for pre-S B-cell epitopes recognition The antibody response of individual duck sera to DHBV pre-S linear epitopes were analyzed using the PEPSCAN method (Khawaja et al., 2012). Briefly, a pepset of 77 9-mers peptides with an overlap of 7 amino acids, spanning the entire DHBV pre-S protein (residues 1–166) was synthesized by Mimotopes peptide company (UK) in the format Biotin-SGSG-PEPTIDE-amide. Each duck serum taken at week 40 was tested for peptide binding as described previously in details (Khawaja et al., 2012). "
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    ABSTRACT: This preclinical study investigated the therapeutic efficacy of electroporation (EP)-based delivery of plasmid DNA (pDNA) encoding viral proteins (envelope, core) and IFN-γ in the duck model of chronic hepatitis B virus (DHBV) infection. Importantly, only DNA EP-therapy resulted in a significant decrease in mean viremia titers and in intrahepatic covalently closed circular DNA (cccDNA) levels in chronic DHBV-carrier animals, compared with standard needle pDNA injection (SI). In addition, DNA EP-therapy stimulated in all virus-carriers a humoral response to DHBV preS protein, recognizing a broader range of major antigenic regions, including neutralizing epitopes, compared with SI. DNA EP-therapy led also to significant higher intrahepatic IFN-γ RNA levels in DHBV-carriers compared to other groups, in the absence of adverse effects. We provide the first evidence on DNA EP-therapy benefit in terms of hepadnaviral infection clearance and break of immune tolerance in virus-carriers, supporting its clinical application for chronic hepatitis B.
    Virology 08/2012; 433(1):192-202. DOI:10.1016/j.virol.2012.07.014 · 3.32 Impact Factor
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    ABSTRACT: DNA vaccines have evolved greatly over the last 20 years since their invention, but have yet to become a competitive alternative to conventional protein or carbohydrate based human vaccines. Whilst safety concerns were an initial barrier, the Achilles heel of DNA vaccines remains their poor immunogenicity when compared to protein vaccines. A wide variety of strategies have been developed to optimize DNA vaccine immunogenicity, including codon optimization, genetic adjuvants, electroporation and sophisticated prime-boost regimens, with each of these methods having its advantages and limitations. Whilst each of these methods has contributed to incremental improvements in DNA vaccine efficacy, more is still needed if human DNA vaccines are to succeed commercially. This review foresees a final breakthrough in human DNA vaccines will come from application of the latest cutting-edge technologies, including "epigenetics" and "omics" approaches, alongside traditional techniques to improve immunogenicity such as adjuvants and electroporation, thereby overcoming the current limitations of DNA vaccines in humans.
    Journal of Biotechnology 09/2012; 162(2-3). DOI:10.1016/j.jbiotec.2012.08.012 · 2.87 Impact Factor
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