A Novel Prophylaxis Strategy Using Liposomal Vaccine Adjuvant CAF09b Protects against Influenza Virus Disease.

The SARS-CoV-2 pandemic caused a massive health and societal crisis, although the fast development of effective vaccines reduced some of the impact. To prepare for future respiratory virus pandemics, a pan-viral prophylaxis could be used to control the initial virus outbreak in the period prior to vaccine approval. The liposomal vaccine adjuvant CAF®09b contains the TLR3 agonist polyinosinic:polycytidylic acid, which induces a type I interferon (IFN-I) response and an antiviral state in the affected tissues. When testing CAF09b liposomes as a potential pan-viral prophylaxis, we observed that intranasal administration of CAF09b liposomes to mice resulted in an influx of innate immune cells into the nose and lungs and upregulation of IFN-I-related gene expression. When CAF09b liposomes were administered prior to challenge with mouse-adapted influenza A/Puerto Rico/8/1934 virus, it protected from severe disease, although the virus was still detectable in the lungs. However, when CAF09b liposomes were administered after influenza challenge, the mice had a similar disease course to controls. In conclusion, CAF09b may be a suitable candidate as a pan-viral prophylactic treatment for epidemic viruses, but must be administered prior to virus exposure to be effective.

The antigenicity of SARS-CoV-2 Delta variants aggregated 10 high-frequency mutations in RBD has not changed sufficiently to replace the current vaccine strain.

Emerging SARS-CoV-2 variants are the most serious problem for COVID-19 prophylaxis and treatment. To determine whether the SARS-CoV-2 vaccine strain should be updated following variant emergence like seasonal flu vaccine, the changed degree on antigenicity of SARS-CoV-2 variants and H3N2 flu vaccine strains was compared. The neutralization activities of Alpha, Beta and Gamma variants' spike protein-immunized sera were analysed against the eight current epidemic variants and 20 possible variants combining the top 10 prevalent RBD mutations based on the Delta variant, which were constructed using pseudotyped viruses. Meanwhile, the neutralization activities of convalescent sera and current inactivated and recombinant protein vaccine-elicited sera were also examined against all possible Delta variants. Eight HA protein-expressing DNAs elicited-animal sera were also tested against eight pseudotyped viruses of H3N2 flu vaccine strains from 2011-2019. Our results indicate that the antigenicity changes of possible Delta variants were mostly within four folds, whereas the antigenicity changes among different H3N2 vaccine strains were approximately 10-100-fold. Structural analysis of the antigenic characterization of the SARS-CoV-2 and H3N2 mutations supports the neutralization results. This study indicates that the antigenicity changes of the current SARS-CoV-2 may not be sufficient to require replacement of the current vaccine strain.

Hemagglutinin Nanoparticulate Vaccine with Controlled Photochemical Immunomodulation for Pathogenic Influenza-Specific Immunity.

Recently, viral infectious diseases, including COVID-19 and Influenza, are the subjects of major concerns worldwide. One strategy for addressing these concerns focuses on nasal vaccines, which have great potential for achieving successful immunization via safe, easy, and affordable approaches. However, conventional nasal vaccines have major limitations resulting from fast removal when pass through nasal mucosa and mucociliary clearance hindering their effectiveness. Herein a nanoparticulate vaccine (NanoVac) exhibiting photochemical immunomodulation and constituting a new self-assembled immunization system of a photoactivatable polymeric adjuvant with influenza virus hemagglutinin for efficient nasal delivery and antigen-specific immunity against pathogenic influenza viruses is described. NanoVac increases the residence period of antigens and further enhances by spatiotemporal photochemical modulation in the nasal cavity. As a consequence, photochemical immunomodulation of NanoVacs successfully induces humoral and cellular immune responses followed by stimulation of mature dendritic cells, plasma cells, memory B cells, and CD4+ and CD8+ T cells, resulting in secretion of antigen-specific immunoglobulins, cytokines, and CD8+ T cells. Notably, challenge with influenza virus after nasal immunization with NanoVacs demonstrates robust prevention of viral infection. Thus, this newly designed vaccine system can serve as a promising strategy for developing vaccines that are active against current hazardous pathogen outbreaks and pandemics.

RNA-dependent assembly of chimeric antigen nanoparticles as an efficient H5N1 pre-pandemic vaccine platform.

Highly pathogenic avian influenza viruses (HPAIVs) pose a significant threat to human health, with high mortality rates, and require effective vaccines. We showed that, harnessed with novel RNA-mediated chaperone function, hemagglutinin (HA) of H5N1 HPAIV could be displayed as an immunologically relevant conformation on self-assembled chimeric nanoparticles (cNP). A tri-partite monomeric antigen was designed including: i) an RNA-interaction domain (RID) as a docking tag for RNA to enable chaperna function (chaperna: chaperone + RNA), ii) globular head domain (gd) of HA as a target antigen, and iii) ferritin as a scaffold for 24 mer-assembly. The immunization of mice with the nanoparticles (~46 nm) induced a 25-30 fold higher neutralizing capacity of the antibody and provided cross-protection from homologous and heterologous lethal challenges. This study suggests that cNP assembly is conducive to eliciting antibodies against the conserved region in HA, providing potent and broad protective efficacy.

Broad neutralization of H1 and H3 viruses by adjuvanted influenza HA stem vaccines in nonhuman primates.

Seasonal influenza vaccines confer protection against specific viral strains but have restricted breadth that limits their protective efficacy. The H1 and H3 subtypes of influenza A virus cause most of the seasonal epidemics observed in humans and are the major drivers of influenza A virus-associated mortality. The consequences of pandemic spread of COVID-19 underscore the public health importance of prospective vaccine development. Here, we show that headless hemagglutinin (HA) stabilized-stem immunogens presented on ferritin nanoparticles elicit broadly neutralizing antibody (bnAb) responses to diverse H1 and H3 viruses in nonhuman primates (NHPs) when delivered with a squalene-based oil-in-water emulsion adjuvant, AF03. The neutralization potency and breadth of antibodies isolated from NHPs were comparable to human bnAbs and extended to mismatched heterosubtypic influenza viruses. Although NHPs lack the immunoglobulin germline VH1-69 residues associated with the most prevalent human stem-directed bnAbs, other gene families compensated to generate bnAbs. Isolation and structural analyses of vaccine-induced bnAbs revealed extensive interaction with the fusion peptide on the HA stem, which is essential for viral entry. Antibodies elicited by these headless HA stabilized-stem vaccines neutralized diverse H1 and H3 influenza viruses and shared a mode of recognition analogous to human bnAbs, suggesting that these vaccines have the potential to confer broadly protective immunity against diverse viruses responsible for seasonal and pandemic influenza infections in humans.

Intranasal Delivery of Inactivated Influenza Virus and Poly(I:C) Adsorbed Corn-Based Nanoparticle Vaccine Elicited Robust Antigen-Specific Cell-Mediated Immune Responses in Maternal Antibody Positive Nursery Pigs.

We designed the killed swine influenza A virus (SwIAV) H1N2 antigen (KAg) with polyriboinosinic:polyribocytidylic acid [(Poly(I:C)] adsorbed corn-derived Nano-11 particle based nanovaccine called Nano-11-KAg+Poly(I:C), and evaluated its immune correlates in maternally derived antibody (MDA)-positive pigs against a heterologous H1N1 SwIAV infection. Immunologically, in tracheobronchial lymph nodes (TBLN) detected enhanced H1N2-specific cytotoxic T-lymphocytes (CTLs) in Nano-11-KAg+Poly(I:C) vaccinates, and in commercial vaccinates detected CTLs with mainly IL-17A+ and early effector phenotypes specific to both H1N2 and H1N1 SwAIV. In commercial vaccinates, activated H1N2- and H1N1-specific IFNγ+&TNFα+, IL-17A+ and central memory T-helper/Memory cells, and in Nano-11-KAg+Poly(I:C) vaccinates H1N2-specific central memory, IFNγ+ and IFNγ+&TNFα+, and H1N1-specific IL-17A+ T-helper/Memory cells were observed. Systemically, Nano-11-KAg+Poly(I:C) vaccine augmented H1N2-specific IFNγ+ CTLs and H1N1-specific IFNγ+ T-helper/Memory cells, and commercial vaccine boosted H1N2- specific early effector CTLs and H1N1-specific IFNγ+&TNFα+ CTLs, as well as H1N2- and H1N1-specific T-helper/Memory cells with central memory, IFNγ+&TNFα+, and IL-17A+ phenotypes. Remarkably, commercial vaccine induced an increase in H1N1-specific T-helper cells in TBLN and naive T-helper cells in both TBLN and peripheral blood mononuclear cells (PBMCs), while H1N1- and H1N2-specific only T-helper cells were augmented in Nano-11-KAg+Poly(I:C) vaccinates in both TBLN and PBMCs. Furthermore, the Nano-11-KAg+Poly(I:C) vaccine stimulated robust cross-reactive IgG and secretory IgA (SIgA) responses in lungs, while the commercial vaccine elicited high levels of serum and lung IgG and serum hemagglutination inhibition (HI) titers. In conclusion, despite vast genetic difference (77% in HA gene identity) between the vaccine H1N2 and H1N1 challenge viruses in Nano-11-KAg+Poly(I:C) vaccinates, compared to over 95% identity between H1N1 of commercial vaccine and challenge viruses, the virus load and macroscopic lesions in the lungs of both types of vaccinates were comparable, but the Nano-11-KAg+Poly(I:C) vaccine cleared the virus from the nasal passage better. These data suggested the important role played by Nano-11 and Poly(I:C) in the induction of polyfunctional, cross-protective cell-mediated immunity against SwIAV in MDA-positive pigs.

Synthetic Biology-derived triterpenes as efficacious immunomodulating adjuvants.

The triterpene oil squalene is an essential component of nanoemulsion vaccine adjuvants. It is most notably in the MF59 adjuvant, a component in some seasonal influenza vaccines, in stockpiled, emulsion-based adjuvanted pandemic influenza vaccines, and with demonstrated efficacy for vaccines to other pandemic viruses, such as SARS-CoV-2. Squalene has historically been harvested from shark liver oil, which is undesirable for a variety of reasons. In this study, we have demonstrated the use of a Synthetic Biology (yeast) production platform to generate squalene and novel triterpene oils, all of which are equally as efficacious as vaccine adjuvants based on physiochemical properties and immunomodulating activities in a mouse model. These Synthetic Biology adjuvants also elicited similar IgG1, IgG2a, and total IgG levels compared to marine and commercial controls when formulated with common quadrivalent influenza antigens. Injection site morphology and serum cytokine levels did not suggest any reactogenic effects of the yeast-derived squalene or novel triterpenes, suggesting their safety in adjuvant formulations. These results support the advantages of yeast produced triterpene oils to include completely controlled growth conditions, just-in-time and scalable production, and the capacity to produce novel triterpenes beyond squalene.