An effective COVID-19 vaccine against broad SARS-CoV-2 variants is still an unmet need. In the study, the vesicular stomatitis virus (VSV)-based vector was used to express the SARS-CoV-2 Spike ...protein to identify better vaccine designs. The replication-competent of the recombinant VSV-spike virus with C-terminal 19 amino acid truncation (SΔ19 Rep) was generated. A single dose of SΔ19 Rep intranasal vaccination is sufficient to induce protective immunity against SARS-CoV-2 infection in hamsters. All the clones isolated from the SΔ19 Rep virus contained R682G mutation located at the Furin cleavage site. An additional S813Y mutation close to the TMPRSS2 cleavage site was identified in some clones. The enzymatic processing of S protein was blocked by these mutations. The vaccination of the R682G-S813Y virus produced a high antibody response against S protein and a robust S protein-specific CD8
T cell response. The vaccinated animals were protected from the lethal SARS-CoV-2 (delta variant) challenge. The S antigen with resistance to enzymatic processes by Furin and TMPRSS2 will provide better immunogenicity for vaccine design.
A recombinant vaccine strain SL3261/pLT105 of attenuated
aroA
Salmonella enterica serovar Typhimurium SL3261 strain expressing a secreted dengue virus type 2 non-structural NS1 and
Yersinia
pestis F1 ...(Caf1) fusion protein, rNS1:Caf1, was generated. Immunological evaluation was performed by prime-boost vaccine regimen. Oral immunization of mice with 1
×
10
9
cfu of SL3261/pLT105 only induced low levels of NS1-specific antibody response and protective immunity following dengue virus challenge. The parenteral NS1 protein priming-oral
Salmonella boosting protocol enhanced both NS1-specific serum IgG response and protective efficacy as compared to mice immunized with each type vaccine alone. Addition of an antifungal antibiotic amphotericin B (AmB) to
Salmonella vaccine further enhanced the synergic effects of prime-boost vaccine regimen on the elicited NS1-specific serum IgG response and the protective efficacy. Together, the results demonstrated that the rNS1:Caf1 producing
Salmonella SL3261/pLT105 strain fails to provide effective protection as an oral vaccine alone despite co-administration of AmB as an adjuvant capable of enhancing the immune responses, and moreover, the protein priming-oral
Salmonella vaccine boosting approach in combination with AmB as an immunization regimen may have the potential to be further explored as an alternative approach for dengue vaccine development.
A recombinant vaccine strain SL3261/pLT105 of attenuatedaroASalmonella entericaserovar Typhimurium SL3261 strain expressing a secreted dengue virus type 2 non-structural NS1 andYersiniapestisF1 ...(Caf1) fusion protein, rNS1:Caf1, was generated. Immunological evaluation was performed by prime-boost vaccine regimen. Oral immunization of mice with 1x109cfu of SL3261/pLT105 only induced low levels of NS1-specific antibody response and protective immunity following dengue virus challenge. The parenteral NS1 protein priming-oralSalmonellaboosting protocol enhanced both NS1-specific serum IgG response and protective efficacy as compared to mice immunized with each type vaccine alone. Addition of an antifungal antibiotic amphotericin B (AmB) toSalmonellavaccine further enhanced the synergic effects of prime-boost vaccine regimen on the elicited NS1-specific serum IgG response and the protective efficacy. Together, the results demonstrated that the rNS1:Caf1 producingSalmonellaSL3261/pLT105 strain fails to provide effective protection as an oral vaccine alone despite co-administration of AmB as an adjuvant capable of enhancing the immune responses, and moreover, the protein priming-oralSalmonellavaccine boosting approach in combination with AmB as an immunization regimen may have the potential to be further explored as an alternative approach for dengue vaccine development.
Salmonella enterica serovar Typhimurium ATCC 13311 is virulent at a dose as low as 10(2) colony-forming units when administered intraperitoneally to BALB/c mice. In order to develop highly attenuated ...mutant strain through the combination of 2 phenotypically attenuated markers, we constructed a number of amino acid requiring auxotrophic strains of S. enterica serovar Typhimurium by means of UV-induced mutations. One of them, strain NDMC-B1, was highly attenuated for mice, with an LD50-value of 6 and 3 log units lower for mice than the wild-type strain and S. enterica serovar Typhimurium aroA strain, respectively. This strain still contained the Salmonella O- and H-antigens but had a requirement for cysteine and was unable to utilize citrate as its sole carbon source. NDMC-B1 colonized the gut-associated lymphoid tissue more efficiently than the wild-type strain, but its capacities to colonize spleen and liver were significantly reduced. Mice intraperitoneally or orally vaccinated with NDMC-B1 were highly protected against either an intraperitoneal challenge with 10(6) colony-forming units or an oral challenge with 10(9) colony-forming units of the wild-type strain. Taken together, the results illustrate that through the combination of 2 independently phenotypical attenuating markers, the requirement for cysteine and the inability to use citrate, we have successfully constructed a highly attenuated, stable, and immunogenic S. enterica serovar Typhimurium vaccine strain which can induce protective immunity in a mouse model against lethal challenge of wild-type strain.
Protein subunit vaccines have been used as prophylactic vaccines for a long time. The well‐established properties of these vaccines make them the first choice for the coronavirus disease 2019 ...(COVID‐19) outbreak. However, it is not easy to develop a protein vaccine that induces cytotoxic T lymphocyte responses and requires a longer time for manufacturing, which limits the usage of this vaccine type. Here, we report the combination of a recombinant spike (S)‐trimer protein with a DNA vaccine‐encoded S protein as a novel COVID‐19 vaccine. The recombinant S protein was formulated with different adjuvants and mixed with the DNA plasmid before injection. We found that the recombinant S protein formulated with the adjuvant aluminum hydroxide and mixed with the DNA plasmid could enhance antigen‐specific antibody titers, neutralizing antibody titers. We further evaluated the IgG2a/IgG1 isotype and cytokine profiles of the specific boosted T‐cell response, which indicated that the combined vaccine induced a T‐helper 1 cell‐biased immune response. Immunized hamsters were challenged with severe acute respiratory syndrome coronavirus 2, and the body weight of the hamsters that received the recombinant S protein with aluminum hydroxide and/or the DNA plasmid was not reduced. Alternatively, those that received control or only the DNA plasmid immunization were reduced. Interestingly, after the third day of the viral load in the lungs, the viral challenge could not be detected in hamsters immunized with the recombinant S protein in aluminum hydroxide mixed with DNA (tissue culture infectious dose < 10). The viral load in the lungs was 109, 106, and 107 for the phosphate‐buffered saline, protein in aluminum hydroxide, and DNA‐only immunizations, respectively. These results indicated that antiviral mechanisms neutralizing antibodies play important roles. Furthermore, we found that the combination of protein and DNA vaccination could induce relatively strong CD8+ T‐cell responses. In summary, the protein subunit vaccine combined with a DNA vaccine could induce strong CD8+ T‐cell responses to increase antiviral immunity for disease control.
During the sustained COVID-19 pandemic, global mass vaccination to achieve herd immunity can prevent further viral spread and mutation. A protein subunit vaccine that is safe, effective, stable, has ...few storage restrictions, and involves a liable manufacturing process would be advantageous to distribute around the world. Here, we designed and produced a recombinant spike (S)-Trimer that is maintained in a prefusion state and exhibits a high ACE2 binding affinity. Rodents received different doses of S-Trimer (0.5, 5, or 20 μg) antigen formulated with aluminum hydroxide (Alum) or an emulsion-type adjuvant (SWE), or no adjuvant. After two vaccinations, the antibody response, T-cell responses, and number of follicular helper T-cells (Tfh) or germinal center (GC) B cells were assessed in mice; the protective efficacy was evaluated on a Syrian hamster infection model. The mouse studies demonstrated that adjuvating the S-Trimer with SWE induced a potent humoral immune response and Th1-biased cellular immune responses (in low dose) that were superior to those induced by Alum. In the Syrian hamster studies, when S-Trimer was adjuvanted with SWE, higher levels of neutralizing antibodies were induced against live SARS-CoV-2 from the original lineage and against the emergence of variants (Beta or Delta) with a slightly decreased potency. In addition, the SWE adjuvant demonstrated a dose-sparing effect; thus, a lower dose of S-Trimer as an antigen (0.5 μg) can induce comparable antisera and provide complete protection from viral infection. These data support the utility of SWE as an adjuvant to enhance the immunogenicity of the S-Trimer vaccine, which is feasible for further clinical testing.
A series of recombinant human type 5 adenoviruses that express the full-length or membrane-truncated spike protein (S) of SARS-CoV-2 (AdCoV2-S or AdCoV2-SdTM, respectively) was tested the efficacy ...against SARS-CoV-2 via intranasal (i.n.) or subcutaneous (s.c.) immunization in a rodent model. Mucosal delivery of adenovirus (Ad) vaccines could induce anti-SARS-CoV-2 IgG and IgA in the serum and in the mucosal, respectively as indicated by vaginal wash (vw) and bronchoalveolar lavage fluid (BALF). Serum anti-SARS-CoV-2 IgG but not IgA in the vw and BALF was induced by AdCoV2-S s.c.. Administration of AdCoV2-S i.n. was able to induce higher anti-SARS-CoV-2 binding and neutralizing antibody levels than s.c. injection. AdCoV2-SdTM i.n. induced a lower antibody responses than AdCoV2-S i.n.. Induced anti-S antibody responses by AdCoV2-S via i.n. or s.c. were not influenced by the pre-existing serum anti-Ad antibody. Novelty, S-specific IgG1 which represented Th2-mediated humoral response was dominantly induced in Ad i.n.-immunized serum in contrast to more IgG2a which represented Th1-mediated cellular response found in Ad s.c.-immunized serum. The activation of S-specific IFN-ɣ and IL-4 in splenic Th1 and Th2 cells, respectively, was observed in the AdCoV2-S i.n. and s.c. groups, indicating the Th1 and Th2 immunity were activated. AdCoV2-S and AdCoV2-SdTM significantly prevented body weight loss and reduced pulmonary viral loads in hamsters. A reduction in inflammation in the lungs was observed in AdCoV-S via i.n. or s.c.-immunized hamsters following a SARS-CoV-2 challenge. It correlated to Th1 cytokine but no inflammatory cytokines secretions found in AdCoV-S i.n. -immunized BALF. These results indicate that intranasal delivery of AdCoV2-S vaccines is safe and potent at preventing SARS-CoV-2 infections.
The emergence of SARS-CoV-2 variants has raised concerns about the sustainability of vaccine-induced immunity. Little is known about the long-term humoral responses and spike-specific T cell memory ...to Omicron variants, with specific attention to BA.4/5, BQ.1.1, and XBB.1.
We assessed immune responses in 50 uninfected individuals who received varying three-dose vaccination combinations (2X AstraZeneca + 1X Moderna, 1X AstraZeneca + 2X Moderna, and 3X Moderna) against wild-type (WT) and Omicron variants at eight months post-vaccination. The serum antibody titers were analyzed by enzyme-linked immunosorbent assays (ELISA), and neutralizing activities were examined by pseudovirus and infectious SARS-CoV-2 neutralization assays. T cell reactivities and their memory phenotypes were determined by flow cytometry.
We found that RBD-specific antibody titers, neutralizing activities, and CD4+ T cell reactivities were reduced against Omicron variants compared to WT. In contrast, CD8+ T cell responses, central memory, effector memory, and CD45RA+ effector memory T cells remained unaffected upon stimulation with the Omicron peptide pool. Notably, CD4+ effector memory T cells even exhibited a higher proportion of reactivity against Omicron variants. Furthermore, participants who received three doses of the Moderna showed a more robust response regarding neutralization and CD8+ T cell reactions than other three-dose vaccination groups.
Reduction of humoral and CD4+ T cell responses against Omicron variants in vaccinees suggested that vaccine effectiveness after eight months may not have sufficient protection against the new emerging variants, which provides valuable information for future vaccination strategies such as receiving BA.4/5 or XBB.1-based bivalent vaccines.
Dengue virus (DENV) and Zika virus (ZIKV) are mosquito-borne flaviviruses that cause severe illness after infection. Currently, there are no specific or effective treatments against DENV and ZIKV. ...Previous studies have shown that tyrosine kinase activities and signal transduction are involved in flavivirus replication, suggesting a potential therapeutic strategy for DENV and ZIKV. In this study, we found that compound
can significantly reduce viral protein expression and viral titers in HEK-293, MCF-7, HepG2, and Huh-7 cells and exhibits superior therapeutic efficacy against flaviviral infection compared to other tyrosine kinase inhibitors. In addition, compound
can decrease endogenous HER2 activation and inhibit the phosphorylation of the HER2 downstream signaling molecules Src and ERK1/2, the levels of which have been associated with viral protein expression in MCF-7 cells. Moreover, silencing HER2 diminished DENV-2 and ZIKV expression in MCF-7 cells, which suggests that HER2 activity is involved in flavivirus replication. Furthermore, in DENV-2-infected AG129 mice, treatment with compound
increased the survival rates and reduced the viremia levels. Overall, compound
demonstrates therapeutic efficacy both in vitro and in vivo and could be developed as a promising antiviral drug against emerging flaviviruses or for concurrent DENV and ZIKV outbreaks.
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