N-dihydrogalactochitosan serves as an effective mucosal adjuvant for intranasal vaccine in combination with recombinant viral proteins against respiratory infection.

Mucosal vaccinations for respiratory pathogens provide effective protection as they stimulate localized cellular and humoral immunities at the site of infection. Currently, the major limitation of intranasal vaccination is using effective adjuvants capable of withstanding the harsh environment imposed by the mucosa. Herein, we describe the efficacy of using a unique biopolymer, N-dihydrogalactochitosan (GC), as a nasal mucosal vaccine adjuvant against respiratory infections. Specifically, we mixed GC with recombinant SARS-CoV-2 trimeric spike (S) and nucleocapsid (NC) proteins to intranasally vaccinate K18-hACE2 transgenic mice, in comparison with Addavax (AV), an MF-59 equivalent. In contrast to AV, intranasal application of GC induces a robust, systemic antigen-specific antibody response and increases the number of T cells in the cervical lymph nodes. Moreover, GC+S+NC-vaccinated animals were largely resistant to the lethal SARS-CoV-2 challenge and experienced drastically reduced morbidity and mortality, with animal weights and behavior returning to normal 22 days post-infection. In contrast, animals intranasally vaccinated with AV+S+NC experienced severe weight loss, mortality, and respiratory distress, with none surviving beyond 6 days post-infection. Our findings demonstrate that GC can serve as a potent mucosal vaccine adjuvant against SARS-CoV-2 and potentially other respiratory viruses. STATEMENT OF SIGNIFICANCE: We demonstrated that a unique biopolymer, N-dihydrogalactochitosan (GC), was an effective nasal mucosal vaccine adjuvant against respiratory infections. Specifically, we mixed GC with recombinant SARS-CoV-2 trimeric spike (S) and nucleocapsid (NC) proteins to intranasally vaccinate K18-hACE2 transgenic mice, in comparison with Addavax (AV). In contrast to AV, GC induces a robust, systemic antigen-specific antibody response and increases the number of T cells in the cervical lymph nodes. About 90 % of the GC+S+NC-vaccinated animals survived the lethal SARS-CoV-2 challenge and remained healthy 22 days post-infection, while the AV+S+NC-vaccinated animals experienced severe weight loss and respiratory distress, and all died within 6 days post-infection. Our findings demonstrate that GC is a potent mucosal vaccine adjuvant against SARS-CoV-2 and potentially other respiratory viruses.

Development of inCVAX, In situ Cancer Vaccine, and Its Immune Response in Mice with Hepatocellular Cancer.

Manipulation of immune system toward the rejection of established cancers has become the standard of care in some patients. Here we propose the development of an in situ autologous cancer vaccine, inCVAX, for the treatment of hepatocellular cancer (HCC). inCVAX is based on the induction of local immunogenic cancer cell death combined with local dendritic cell stimulation by intratumoral injection of the immune-activator N-dihydro-galacto-chitosan (GC). In a first set of experiments, cellular and molecular studies were performed to investigate the effect of inCVAX on immune activation in a murine model of HCC that we previously developed. Once large tumors were formed in mice, the tumor is surgically exposed and a laser fiber was inserted into the center of an individual tumor mass. Using a 10 mm diffuser tip, laser irradiation of 1.5 W was applied to heat the tumor at different durations (6-10 min) to assess tolerability of photothermal application at different temperatures. The laser application was followed by immediate injection of GC, and each mouse received one laser treatment and one GC injection. ELISA was used to assess the level of cytokines; immunohistochemical staining was conducted to analyze the effect of inCVAX on immune cell tumor-filtration and expression of tumor-specific antigens (TSAs) and tumor-associated antigens (TAAs). Results indicate that survival correlated to thermal exposure. At lower temperatures the photothermal effect was sufficient to induce tumor necrosis, but without obvious complication to the mice, although at these temperatures the treatment didn't alter the level of TSAs and TAAs, so further optimization is suggested. Nevertheless, in response to the inCVAX treatment, cytotoxic cytokine IFN-γ was significantly increased, but suppressive cytokine TGF-ß was dramatically reduced. Furthermore, inCVAX prompted tumor infiltration of CD3+, CD4+, and CD8+ T cells; but modulated macrophage subsets differently. In conclusion, while the protocol needs further optimization, it would appear that inCVAX for the treatment of HCC activates an immune response in tumor-bearing mice, which in turn may have potential for the treatment of HCC.