Nanoparticle orientationally displayed antigen epitopes improve neutralizing antibody level in a model of porcine circovirus type 2

Ding, Peiyang; Zhang, Teng; Li, Yafei; Teng, Man; Sun, Yaning; Liu, Xiao; Chai, Shujun; Zhou, Enmin; Jin, Qianyue; Zhang, Gaiping

ell suited for biomedical applications. Importantly, AuNPs can be readily conjugated with peptides through the gold-thiol (Au-S) bond by simply mixing AuNPs with peptides that contain cysteine, thus offering a highly flexible and customizable approach to provide a scaffold that can be used to fix proteins or peptides to form a certain spatial orientation. 22, 23 According to bioinformatics analysis, mainly based on antigen crystal structure and the corresponding pathogenic characteristics, we selected the capsid (Cap) protein of PCV2 as a model antigen. PCV2, classified in the family Circoviridae and one of the smallest DNA viruses, is widespread and has caused an estimated € 5.76 billion economic loss for the EU. 24, 25 The Cap protein can form virus-like particles (VLPs) dividing the structure of Cap protein into two faces, neutralizing epitopes on the outer surface, while the unique cysteine residue is on the inner face ( Figure 1A and B). 26 The decoy epitope, an immunodominant region consisting of residues 169-180 (Cap (169-180)), exposed on the surface of the Cap protein monomer but buried in virus particles or VLPs, allows PCV2 to evade humoral immunity by inducing low-level NA and high-level non-protection antibodies. 13, 27 In the current study, we used AuNPs as an orientationalconnect carrier to display Cap protein (Cap-conjugated AuNPs [Cap-AuNPs]), with neutralizing epitopes exposed on the outside surface of the nanoparticles ( Figure 1C ). The immune responses to Cap-AuNPs were evaluated in mice. To examine the mechanisms of the antibody production, the efficiency of cellular uptake of Cap-AuNPs into RAW264.7 macrophage cells and DC2.4 dendritic cells and subsequent cytokine secretion were measured. Our study lays the foundation for the development of a human and animal subunit vaccine based on antigen structural analysis that utilizes nanoparticles as a fixed-point connection carrier for antigens with reasonable conformation.