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Advantages of DNA vaccines including well-tolerance, safety and ability to induce antibody, cytotoxic T- lymphocytes, and T-helper immune responses make it more and more attractive in treatment of chronic virus infections. In order to successfully induce immune response, DNA vaccine needs to enter target cell for transcription and subsequent protein production. An ideal delivery carrier needs to be safe and effective in targeting specific cell, provide sufficient protection of DNA plasmid during transportation, as wel as low- responses to any existing self-immunity. Deficient viral particles, such as adenoviruses or retroviruses, offer an attractive option with great benefits in targeting and protection but majr drawbacks on strong self-immunity as well as the complexity in producing recombinant viral particles. Non-replicating virus-like particle derived from human hepatitis E virus (HEV-VLP) is empty icosahedral cage composed of 60 copies of recombinant HEV capsid proteins. This VLP is capable of encapsulating DNA vaccine in vitro, delivering DNA vaccine to epithelium cell at gastrointestinal tract, and inducing antigen-specific humoral and cellular immune responses. The simple procedure in DNA encapsulation makes HEV-VLP of particular interest as gene carrier; however, induction of self-immunity is still the hurdle in using HEV-VLP for therapeuticalvaccination. <P> Our studies on HEV-VLP crystal structure and antigenic structure reveal a structural modularity, with which HEV recombinant capsid proteins interplay between VLP assembly and antigen presentation. By carrying insertion of 15 amino acids at an antibody-binding site, the chimeric VLP reduces the reactivity to anti-HEV antibodies meanwhile retains the icosahedral assembly. This data suggests us a strategy to lower the reactivity of VLP to antibody- induced neutralization by structural alteration at antibody-binding sites, a mechanism that viruses have evolved to mediate their escape from host immune surveillance. <P> The proposed experiments in this application include 1) using cryo-electron microscopy and image reconstruction to identify HEV-VLP surface flexible loops that critical to antibody interactions; 2 inserting short peptide into the identified surface loops to create chimeric VLP with attenuate reactivity to HEV-VLP; 3) both wild type and chimeric VLPs will be evaluated in delivery of DNA vaccine encoding the surface antigen of hepatitis B virus, as potential gene carrier for treatment of chronic progressive hepatitis B. Because HEV is an enteric transmitted virus, HEV-VLP is able to transcytose the mucosal barrier at gastrointestinal tract and target specific mucosal region for antigen production. When a short peptide bearing mucosal adhesion ligand is inserted into surface antigenic loops, the chimeric VLP is hypothesized to have strong specificity in targeting mucosal region, leading to potent induction of mucosal immunity. In collaboration with Dr Kit Lam and Dr Christopher Walker, two experts respectively in drug discovery and HBV vaccine development, we expect to obtain sufficient data leading us to repeated use of HEV-VLPs as carrier for clinical gene therapy.
Public Health Relevance: <br/> Effective delivery of gene into target cells is the major challenge that remains to be overcome before clinical use of gene therapy as a safe and efficacious treatment of human genetic disorders as well as the chronic infections. Non-replicating hepatitis E virus-like particles (HEV-VLPs) encode structural modularity in the protein tertiary structure, which enables the icosahedral assembly of modified capsid protein, however, with reduced reactivity to antibody-induced neutralization. Proper insertion of mucosal-adhesion peptide at selected antigenic loops allows us to develop a safe and effective gene carrier for repeated use of HEV-VLPs in gene therapy.