Debulking different Corona (SARS-CoV-2 delta, omicron, OC43) and Influenza (H1N1, H3N2) virus strains by plant viral trap proteins in chewing gums to decrease infection and transmission.

Because oral transmission of SARS-CoV-2 is 3-5 orders of magnitude higher than nasal transmission, we investigated debulking of oral viruses using viral trap proteins (CTB-ACE2, FRIL) expressed in plant cells, delivered through the chewing gum. In omicron nasopharyngeal (NP) samples, the microbubble count (based on N-antigen) was significantly reduced by 20 µg of FRIL (p < 0.0001) and 0.925 µg of CTB-ACE2 (p = 0.0001). Among 20 delta or omicron NP samples, 17 had virus load reduced below the detection level of spike protein in the RAPID assay, after incubation with the CTB-ACE2 gum powder. A dose-dependent 50% plaque reduction with 50-100 ng FRIL or 600-800 µg FRIL gum against Influenza strains H1N1, H3N2, and Coronavirus HCoV-OC43 was observed with both purified FRIL, lablab bean powder or gum. In electron micrographs, large/densely packed clumps of overlapping influenza particles and FRIL protein were observed. Chewing simulator studies revealed that CTB-ACE2 release was time/dose-dependent and release was linear up to 20 min chewing. Phase I/II placebo-controlled, double-blinded clinical trial (IND 154897) is in progress to evaluate viral load in saliva before or after chewing CTB-ACE2/placebo gum. Collectively, this study advances the concept of chewing gum to deliver proteins to debulk oral viruses and decrease infection/transmission.

Vaccination with SARS-CoV-2 spike protein lacking glycan shields elicits enhanced protective responses in animal models.

A major challenge to end the pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is to develop a broadly protective vaccine that elicits long-term immunity. As the key immunogen, the viral surface spike (S) protein is frequently mutated, and conserved epitopes are shielded by glycans. Here, we revealed that S protein glycosylation has site-differential effects on viral infectivity. We found that S protein generated by lung epithelial cells has glycoforms associated with increased infectivity. Compared to the fully glycosylated S protein, immunization of S protein with N-glycans trimmed to the mono-GlcNAc-decorated state (SMG) elicited stronger immune responses and better protection for human angiotensin-converting enzyme 2 (hACE2) transgenic mice against variants of concern (VOCs). In addition, a broadly neutralizing monoclonal antibody was identified from SMG-immunized mice that could neutralize wild-type SARS-CoV-2 and VOCs with subpicomolar potency. Together, these results demonstrate that removal of glycan shields to better expose the conserved sequences has the potential to be an effective and simple approach for developing a broadly protective SARS-CoV-2 vaccine.

A Carbohydrate-Binding Protein from the Edible Lablab Beans Effectively Blocks the Infections of Influenza Viruses and SARS-CoV-2.

The influenza virus hemagglutinin (HA) and coronavirus spike (S) protein mediate virus entry. HA and S proteins are heavily glycosylated, making them potential targets for carbohydrate binding agents such as lectins. Here, we show that the lectin FRIL, isolated from hyacinth beans (Lablab purpureus), has anti-influenza and anti-SARS-CoV-2 activity. FRIL can neutralize 11 representative human and avian influenza strains at low nanomolar concentrations, and intranasal administration of FRIL is protective against lethal H1N1 infection in mice. FRIL binds preferentially to complex-type N-glycans and neutralizes viruses that possess complex-type N-glycans on their envelopes. As a homotetramer, FRIL is capable of aggregating influenza particles through multivalent binding and trapping influenza virions in cytoplasmic late endosomes, preventing their nuclear entry. Remarkably, FRIL also effectively neutralizes SARS-CoV-2, preventing viral protein production and cytopathic effect in host cells. These findings suggest a potential application of FRIL for the prevention and/or treatment of influenza and COVID-19.

Better influenza vaccines: an industry perspective.

Vaccination is the most effective measure at preventing influenza virus infections. However, current seasonal influenza vaccines are only protective against closely matched circulating strains. Even with extensive monitoring and annual reformulation our efforts remain one step behind the rapidly evolving virus, often resulting in mismatches and low vaccine effectiveness. Fortunately, many next-generation influenza vaccines are currently in development, utilizing an array of innovative techniques to shorten production time and increase the breadth of protection. This review summarizes the production methods of current vaccines, recent advances that have been made in influenza vaccine research, and highlights potential challenges that are yet to be overcome. Special emphasis is put on the potential role of glycoengineering in influenza vaccine development, and the advantages of removing the glycan shield on influenza surface antigens to increase vaccine immunogenicity. The potential for future development of these novel influenza vaccine candidates is discussed from an industry perspective.

Structure-function analysis of neutralizing antibodies to H7N9 influenza from naturally infected humans.

Little is known about the specificities and neutralization breadth of the H7-reactive antibody repertoire induced by natural H7N9 infection in humans. We have isolated and characterized 73 H7-reactive monoclonal antibodies from peripheral B cells from four donors infected in 2013 and 2014. Of these, 45 antibodies were H7-specific, and 17 of these neutralized the virus, albeit with few somatic mutations in their variable domain sequences. An additional set of 28 antibodies, isolated from younger donors born after 1968, cross-reacted between H7 and H3 haemagglutinins in binding assays, and had accumulated significantly more somatic mutations, but were predominantly non-neutralizing in vitro. Crystal structures of three neutralizing and protective antibodies in complex with the H7 haemagglutinin revealed that they recognize overlapping residues surrounding the receptor-binding site of haemagglutinin. One of the antibodies, L4A-14, bound into the sialic acid binding site and made contacts with haemagglutinin residues that were conserved in the great majority of 2016-2017 H7N9 isolates. However, only 3 of the 17 neutralizing antibodies retained activity for the Yangtze River Delta lineage viruses isolated in 2016-2017 that have undergone antigenic change, which emphasizes the need for updated H7N9 vaccines.