Production and characterization of norovirus virus-like particles vaccine candidates in a genetically modified...

The development of an effective vaccine against noroviruses remains a major public health priority. Norovirus GII.4 capsid protein VP1 as a promising vaccine candidate was produced by the methylotrophic yeast Ogataea minuta production system. It was intracellularly expressed and subsequently purified by immobilized metal affinity chromatography and anion exchange chromatography, yielding 13.4 mg of highly purified VP1 protein from 50 mL of culture supernatant. The formation of VP1-based virus-like particles (VLPs) that retained their structure even after freeze-thawing was confirmed by transmission electron microscopy. The VP1 protein purified in the form of VLPs displayed strong antigenicity and specific, dose-dependent binding to histo-blood group antigens, as determined by the enzyme-linked immunosorbent assay (ELISA). Immunogenicity studies in BALB/c mice demonstrated that intramuscular administration induced robust serum IgG responses across all tested doses, with no significant dose-dependent differences. Furthermore, mucosal administration intranasally or sublingually induced systemic IgG, systemic IgA, and mucosal IgA responses. These responses were significantly enhanced by the lipid A adjuvant. These findings showed that the O. minuta production system is capable of producing immunogenic Norovirus VLPs as a vaccine candidate.