(P22) A DNA-based immunotherapy induces receptor-blocking antibodies that can neutralize HBV in humanized mice
Panagiota Maravelia (1), Lars Frelin (1), Neetu Jagya (1), Lieven Verhoye (2), Georg Verch (3), Yi Ni (3), Gustaf Ahlén (1), Stephan Urban (3), Philip Meuleman (2), Matti Sallberg (1)
1) Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden 2) Laboratory of Liver Infectious Diseases, University of Gent, Belgium 3) Molecular Virologie, University of Heidelberg, Heidelberg, Germany
Background: In chronic hepatitis B and D virus (HBV/HDV) infection the virus evades host immune responses by inducing a severe T cell dysfunction/tolerance, and by secreting HBV surface antigen (HBsAg) to inhibit HBsAg-specific neutralizing antibodies. As the infectious HBV virions, in contrast to circulating HBsAg, predominantly display the PreS1/2 proteins, the presence of PreS1 antibodies can block viral entry and spread of infection. To circumvent, rather than to overcome or break, immunological tolerance in chronic HBV, we used the large HDV capsid antigen (HDAg) as a heterologous carrier of T cell help. Thus, genetic immunization with HDAg, linked to PreS1 sequences, will prime healthy T cells that support B cell activation and production of neutralizing PreS1 specific antibodies also in a host with chronic HBV.
Methods: We designed various combinations of DNA vaccine candidates that induce receptor-blocking antibodies to the preS1 domain of HBV and HDV, and specific T cells. The DNA vaccines are composed of PreS1 sequences fused to HDAg genotype (gt) 1 and gt2 sequences, which were injected in C57BL/6 and BALB/c mice or rabbits by intramuscular immunization followed by in vivo electroporation. Induction of antibodies and specific T cells were analysed by ELISA and ELISpot. The elicited antibodies were further evaluated for their ability to neutralize HBV by an in vitro neutralization assay.
Results: We found that the HDAg-induced T cells are genotype specific in both C57BL/6 and BALB/c mice, suggesting that the vaccine needs to contain both HDV gt1 and gt2 sequences. Upon vaccination of mice and rabbits, broadly cross-reactive preS1-antibodies were effectively induced that recognized HBV of genotypes A to F. Importantly, the vaccine-primed high levels of PreS1 antibodies in mice and rabbits that effectively neutralized HBV in vitro. Antibodies purified from vaccinated and non-vaccinated rabbits were tested for their ability to prevent receptor-binding and infection in uPA-SCID mice with humanized livers. Only immunoglobulin from vaccinated rabbits showed protection against HBV infection.
Conclusion: The current therapeutic vaccine candidate, using a strategy of circumventing immunological tolerance by inducing healthy heterologous T cells, induced functional and neutralizing HBV- and HDV- specific immune responses. We show that our DNA-based vaccination strategy may represent an important constituent for future combination immunotherapies against chronic HBV and possibly also HDV infection.