Purpose: Hyaluronic acid (HA) nanogel has hyaluronic acid in the main chain and a hydrophobic group in the side chain, thus a hydrophilic, biocompatible material with low toxicity. A nano-ordered gel is spontaneously formed due to the hydrophobic interaction, in which it can load small molecules, peptides, and proteins. (1,2) HA nanogel can load various polypeptides and deliver them to lymph nodes by subcutaneous administration, which regresses tumors effectively as a cancer vaccine. (3) We investigated the effect of the HA nanogel / spike protein complex on infectious diseases COVID-19 as a model system. Methods: HA nanogel vaccine preparation: Protein (SARS-CoV-2 Spike protein) was incubated with HA nanogel at 37 C. Encapsulation of the protein was confirmed by size exclusion chromatography (SEC). Vaccination: Spike protein vaccine (200uL) with or without HA nanogel was administrated subcutaneously to mice twice. Evaluation: Antibody titer was evaluated by mouse IgG measurement kit for SARS-CoV-2 Spike protein. Results: An increase in antibody titer was confirmed in mouse serum after the second vaccination of the HA nanogel / spike protein formulation, and the effect was sustained for an extended period of one year or longer. Although it was infrequent, mice with decreased antibody titers were observed. These mice with low antibody titers were vaccinated a third time and the antibody titers increased again, and then antibody production was maintained for six months or more, which is considered sufficient immunological memory. Virus infection inhibition experiment using Vero E6 cells proved that the antibody had neutralizing activity. Furthermore, we confirmed that the HA nanogel complex was delivered to the lymph nodes by fluorescence observation. Conclusion: We investigated the application of HA nanogels for vaccines against COVID-19. HA nanogel vaccines showed high antibody titers and could be maintained for an extended period. In addition, it was cleared that HA nanogel complexes elicited sufficient immunological memory. These results indicated that HA nanogel vaccine is a promising vaccine formulation. References: 1. Macromol. Biosci. 2012, 12, 475 – 483; 2. WO2010-053140; 3. WO2020-158771 Figure 1. Antibody titers one year after the second vaccination
