Abstract
Development of a prophylactic vaccine against hepatitis C virus (HCV) has been hampered by the extraordinary viral diversity and the poor host immune response. Scaffolding, by grafting an epitope onto a heterologous protein scaffold, offers a possible solution to epitope vaccine design. In this study, we designed and characterized epitope vaccine antigens for the antigenic sites of HCV envelope glycoproteins E1 (residues 314–324) and E2 (residues 412–423), for which neutralizing antibody-bound structures are available. We first combined six structural alignment algorithms in a “scaffolding meta-server” to search for diverse scaffolds that can structurally accommodate the HCV epitopes. For each antigenic site, ten scaffolds were selected for computational design, and the resulting epitope scaffolds were analyzed using structure-scoring functions and molecular dynamics simulation. We experimentally confirmed that three E1 and five E2 epitope scaffolds bound to their respective neutralizing antibodies, but with different kinetics. We then investigated a “multivalent scaffolding” approach by displaying 24 copies of an epitope scaffold on a self-assembling nanoparticle, which markedly increased the avidity of antibody binding. Our study thus demonstrates the utility of a multi-scale scaffolding strategy in epitope vaccine design and provides promising HCV immunogens for further assessment in vivo.
Highlights
384–410) is a major target for the antibody response[17,18,19,20,21,22]
Broadly effective vaccines have not been developed for viral pathogens such as human immunodeficiency virus type-1 (HIV-1), hepatitis C virus (HCV), and influenza[69]
Protein engineering has been applied to the development of novel diagnostic and therapeutic agents, for which cysteine knots[70,71], designed ankyrin repeat proteins (DARPins)[72,73], and other unique protein folds have been used as scaffolds to display a functional site
Summary
384–410) is a major target for the antibody response[17,18,19,20,21,22]. Recently, several E2-directed broadly neutralizing antibodies (bnAbs) have been identified[23,24,25,26,27]. Kong et al reported a 2.65 Å crystal structure of the E2 core domain (E2c) in complex with a human bnAb AR3C35, which showed a stark difference from the class II fusion protein model found in other Flaviviridae viruses[36] This finding was confirmed by another E2c structure determined by Khan et al.[13] It is foreseeable that this ever-accumulating structural information will enable an in-depth understanding of HCV biology as well as structure-based design of epitope vaccines. Scaffolding was applied to the antigenic site II of the fusion (F) protein of respiratory syncytial virus (RSV)[43], and neutralizing antibodies were induced by a scaffolded epitope in macaques[44] Based on these studies, a general strategy consisting of epitope identification, scaffolding, particulate presentation of the designed antigen, animal immunization, and quantitative analysis of immune response has been proposed for epitope vaccine design and evaluation[45]. We expect that the antigenic epitope scaffolds and nanoparticles will provide promising candidates for animal studies, while the new scaffolding methods will find many applications in epitope vaccine design
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