Highly stable gold nanoparticle-antigen conjugates with self-adjuvanting property for induction of robust antigen-specific immune responses

Abstract

Poor long-term stability and formation of irreversible aggregates when subjected to a freeze-drying process greatly limits the clinical application of gold nanoparticles (GNPs) as a vaccine carrier. In this study, we synthesized a GNP-antigen conjugate with high colloidal stability by using a thiolated polyethylene glycol (PEG) linker to conjugate a model antigen (ovalbumin; OVA) onto the GNP surface (i.e. GNP-OVA) and demonstrated this conjugate had self-adjuvanting properties to augment antigen-specific immune responses. The synthesized GNP had an average hydrodynamic size of 13.8 ± 2.1 nm (n = 3); after conjugation of OVA, the diameter increased to 28.6 ± 7.3 nm (n = 3). The obtained GNP-OVA can maintain a stable dispersion state in aqueous solutions for at least 12 months and withstand stresses during lyophilization without creating irreversible aggregates. Compared with OVA alone or a mixture of PEG-functionalized GNP (GNP-PEG) and OVA (i.e. GNP-PEG/OVA), the chemical conjugation of OVA onto GNP-PEG substantially increased antigen uptake and upregulated major histocompatibility complex class II expression in macrophages. This indicated that the GNP can function as not only an adjuvant to promote the phagocytic activity of macrophages but also a carrier to deliver the conjugated antigen into the immune cells for the enhancement of its antigen presentation capability. Importantly, OVA-specific immunoglobulin G levels in the mice immunized with GNP-OVA were 4.1 and 2.9 times higher than those in the mice injected with OVA and GNP-PEG/OVA, respectively. These results demonstrated that the GNP-antigen conjugate exhibited remarkable stability either in liquid or freeze-dried form, which makes it attractive for further pharmaceutical applications. Moreover, covalently linking antigens onto the GNP surface was enabled to enhance the immunogenicity of antigens and boost immune responses, showing the potential of the GNP conjugation strategy for vaccine development.

Keywords: Adjuvant; Colloidal stability; Freeze-drying; Immune activation; Nanovaccine.