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1.
Vaccines (Basel) ; 12(5)2024 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-38793789

RESUMO

The development of a safe and effective vaccine against avian influenza A virus (AIV) H5N8 is relevant due to the widespread distribution of this virus in the bird population and the existing potential risk of human infection, which can lead to significant public health concerns. Here, we developed an experimental pVAX-H5 DNA vaccine encoding a modified trimer of AIV H5N8 hemagglutinin. Immunization of BALB/c mice with pVAX-H5 using jet injection elicited high titer antibody response (the average titer in ELISA was 1 × 105), and generated a high level of neutralizing antibodies against H5N8 and T-cell response, as determined by ELISpot analysis. Both liquid and lyophilized forms of pVAX-H5 DNA vaccine provided 100% protection of immunized mice against lethal challenge with influenza A virus A/turkey/Stavropol/320-01/2020 (H5N8). The results obtained indicate that pVAX-H5 has good opportunities as a vaccine candidate against the influenza A virus (H5N8).

2.
Vaccines (Basel) ; 12(2)2024 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-38400127

RESUMO

In this study, we characterized recombinant hemagglutinin (HA) of influenza A (H5N8) virus produced in Chinese hamster ovary cells (CHO-K1s). Immunochemical analysis showed that the recombinant hemagglutinin was recognized by the serum of ferrets infected with influenza A (H5N8) virus, indicating that its antigenic properties were retained. Two groups of Balb/c mice were immunized with intramuscular injection of recombinant hemagglutinin or propiolactone inactivated A/Astrakhan/3212/2020 (H5N8) influenza virus. The results demonstrated that both immunogens induced a specific antibody response as determined by ELISA. Virus neutralization assay revealed that sera of immunized animals were able to neutralize A/turkey/Stavropol/320-01/2020 (H5N8) influenza virus-the average neutralizing titer was 2560. Immunization with both recombinant HA/H5 hemagglutinin and inactivated virus gave 100% protection against lethal H5N8 virus challenge. This study shows that recombinant HA (H5N8) protein may be a useful antigen candidate for developing subunit vaccines against influenza A (H5N8) virus with suitable immunogenicity and protective efficacy.

3.
Vaccines (Basel) ; 11(4)2023 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-37112720

RESUMO

Despite the rapid development and approval of several COVID vaccines based on the full-length spike protein, there is a need for safe, potent, and high-volume vaccines. Considering the predominance of the production of neutralizing antibodies targeting the receptor-binding domain (RBD) of S-protein after natural infection or vaccination, it makes sense to choose RBD as a vaccine immunogen. However, due to its small size, RBD exhibits relatively poor immunogenicity. Searching for novel adjuvants for RBD-based vaccine formulations is considered a good strategy for enhancing its immunogenicity. Herein, we assess the immunogenicity of severe acute respiratory syndrome coronavirus 2 RBD conjugated to a polyglucin:spermidine complex (PGS) and dsRNA (RBD-PGS + dsRNA) in a mouse model. BALB/c mice were immunized intramuscularly twice, with a 2-week interval, with 50 µg of RBD, RBD with Al(OH)3, or conjugated RBD. A comparative analysis of serum RBD-specific IgG and neutralizing antibody titers showed that PGS, PGS + dsRNA, and Al(OH)3 enhanced the specific humoral response in animals. There was no significant difference between the groups immunized with RBD-PGS + dsRNA and RBD with Al(OH)3. Additionally, the study of the T-cell response in animals showed that, unlike adjuvants, the RBD-PGS + dsRNA conjugate stimulates the production of specific CD4+ and CD8+ T cells in animals.

4.
Viruses ; 14(7)2022 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-35891430

RESUMO

The conventional live smallpox vaccine based on the vaccinia virus (VACV) cannot be widely used today because it is highly reactogenic. Therefore, there is a demand for designing VACV variants possessing enhanced immunogenicity, making it possible to reduce the vaccine dose and, therefore, significantly eliminate the pathogenic effect of the VACV on the body. In this study, we analyzed the development of the humoral and T cell-mediated immune responses elicited by immunizing mice with low-dose VACV variants carrying the mutant A34R gene (which increases production of extracellular virions) or the deleted A35R gene (whose protein product inhibits antigen presentation by the major histocompatibility complex class II). The VACV LIVP strain, which is used as a smallpox vaccine in Russia, and its recombinant variants LIVP-A34R*, LIVP-dA35R, and LIVP-A34R*-dA35R, were compared upon intradermal immunization of BALB/c mice at a dose of 104 pfu/animal. The strongest T cell-mediated immunity was detected in mice infected with the LIVP-A34R*-dA35R virus. The parental LIVP strain induced a significantly lower antibody level compared to the strains carrying the modified A34R and A35R genes. Simultaneous modification of the A34R gene and deletion of the A35R gene in VACV LIVP synergistically enhanced the immunogenic properties of the LIVP-A34R*-dA35R virus.


Assuntos
Vacina Antivariólica , Varíola , Vacínia , Animais , Camundongos , Camundongos Endogâmicos BALB C , Varíola/prevenção & controle , Vacina Antivariólica/genética , Vacinas Atenuadas/genética , Vaccinia virus
5.
Viruses ; 14(5)2022 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-35632800

RESUMO

Currently, SARS-CoV-2 spike receptor-binding-domain (RBD)-based vaccines are considered one of the most effective weapons against COVID-19. During the first step of assessing vaccine immunogenicity, a mouse model is often used. In this paper, we tested the use of five experimental animals (mice, hamsters, rabbits, ferrets, and chickens) for RBD immunogenicity assessments. The humoral immune response was evaluated by ELISA and virus-neutralization assays. The data obtained show hamsters to be the least suitable candidates for RBD immunogenicity testing and, hence, assessing the protective efficacy of RBD-based vaccines.


Assuntos
Vacinas contra COVID-19 , COVID-19 , Imunogenicidade da Vacina , Glicoproteína da Espícula de Coronavírus , Animais , COVID-19/prevenção & controle , Vacinas contra COVID-19/imunologia , Galinhas , Cricetinae , Modelos Animais de Doenças , Furões , Camundongos , Camundongos Endogâmicos BALB C , Coelhos , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinas de Subunidades Antigênicas/imunologia
6.
Int J Mol Sci ; 23(4)2022 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-35216301

RESUMO

Despite the fact that a range of vaccines against COVID-19 have already been created and are used for mass vaccination, the development of effective, safe, technological, and affordable vaccines continues. We have designed a vaccine that combines the recombinant protein and DNA vaccine approaches in a self-assembled particle. The receptor-binding domain (RBD) of the spike protein of SARS-CoV-2 was conjugated to polyglucin:spermidine and mixed with DNA vaccine (pVAXrbd), which led to the formation of particles of combined coronavirus vaccine (CCV-RBD) that contain the DNA vaccine inside and RBD protein on the surface. CCV-RBD particles were characterized with gel filtration, electron microscopy, and biolayer interferometry. To investigate the immunogenicity of the combined vaccine and its components, mice were immunized with the DNA vaccine pVAXrbd or RBD protein as well as CCV-RBD particles. The highest antigen-specific IgG and neutralizing activity were induced by CCV-RBD, and the level of antibodies induced by DNA or RBD alone was significantly lower. The cellular immune response was detected only in the case of DNA or CCV-RBD vaccination. These results demonstrate that a combination of DNA vaccine and RBD protein in one construct synergistically increases the humoral response to RBD protein in mice.


Assuntos
Vacinas contra COVID-19/química , Vacinas contra COVID-19/farmacologia , Imunidade Humoral/efeitos dos fármacos , Glicoproteína da Espícula de Coronavírus/química , Animais , Sítios de Ligação , Vacinas contra COVID-19/imunologia , Chlorocebus aethiops , Dextranos/química , Feminino , Células HEK293 , Humanos , Camundongos Endogâmicos BALB C , Domínios Proteicos , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Espermidina/química , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinas de DNA/genética , Vacinas de DNA/imunologia , Vacinas de DNA/farmacologia , Células Vero
7.
Vaccines (Basel) ; 10(1)2022 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-35062757

RESUMO

The receptor-binding domain (RBD) of the protein S SARS-CoV-2 is considered to be one of the appealing targets for developing a vaccine against COVID-19. The choice of an expression system is essential when developing subunit vaccines, as it ensures the effective synthesis of the correctly folded target protein, and maintains its antigenic and immunogenic properties. Here, we describe the production of a recombinant RBD protein using prokaryotic (pRBD) and mammalian (mRBD) expression systems, and compare the immunogenicity of prokaryotic and mammalian-expressed RBD using a BALB/c mice model. An analysis of the sera from mice immunized with both variants of the protein revealed that the mRBD expressed in CHO cells provides a significantly stronger humoral immune response compared with the RBD expressed in E.coli cells. A specific antibody titer of sera from mice immunized with mRBD was ten-fold higher than the sera from the mice that received pRBD in ELISA, and about 100-fold higher in a neutralization test. The data obtained suggests that mRBD is capable of inducing neutralizing antibodies against SARS-CoV-2.

8.
Viruses ; 13(8)2021 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-34452494

RESUMO

Mass vaccination has played a critical role in the global eradication of smallpox. Various vaccinia virus (VACV) strains, whose origin has not been clearly documented in most cases, have been used as live vaccines in different countries. These VACV strains differed in pathogenicity towards various laboratory animals and in reactogenicity exhibited upon vaccination of humans. In this work, we studied the development of humoral and cellular immune responses in BALB/c mice inoculated intranasally (i.n.) or intradermally (i.d.) with the VACV LIVP strain at a dose of 105 PFU/mouse, which was used in Russia as the first generation smallpox vaccine. Active synthesis of VACV-specific IgM in the mice occurred on day 7 after inoculation, reached a maximum on day 14, and decreased by day 29. Synthesis of virus-specific IgG was detected only from day 14, and the level increased significantly by day 29 after infection of the mice. Immunization (i.n.) resulted in significantly higher production of VACV-specific antibodies compared to that upon i.d. inoculation of LIVP. There were no significant differences in the levels of the T cell response in mice after i.n. or i.d. VACV administration at any time point. The maximum level of VACV-specific T-cells was detected on day 14. By day 29 of the experiment, the level of VACV-specific T-lymphocytes in the spleen of mice significantly decreased for both immunization procedures. On day 30 after immunization with LIVP, mice were infected with the cowpox virus at a dose of 46 LD50. The i.n. immunized mice were resistant to this infection, while 33% of i.d. immunized mice died. Our findings indicate that the level of the humoral immune response to vaccination may play a decisive role in protection of animals from orthopoxvirus reinfection.


Assuntos
Imunidade Adaptativa , Vírus da Varíola Bovina/fisiologia , Varíola Bovina/prevenção & controle , Reinfecção/prevenção & controle , Vaccinia virus/imunologia , Vacínia/imunologia , Vacinas Virais/administração & dosagem , Animais , Anticorpos Antivirais/imunologia , Varíola Bovina/imunologia , Varíola Bovina/virologia , Vírus da Varíola Bovina/genética , Vírus da Varíola Bovina/imunologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Reinfecção/imunologia , Reinfecção/virologia , Linfócitos T/imunologia , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/imunologia , Vacínia/virologia , Vaccinia virus/genética , Vaccinia virus/fisiologia , Vacinas Virais/imunologia
9.
Vaccines (Basel) ; 9(5)2021 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-34063689

RESUMO

Nucleic acid-based influenza vaccines are a promising platform that have recently and rapidly developed. We previously demonstrated the immunogenicity of DNA vaccines encoding artificial immunogens AgH1, AgH3, and AgM2, which contained conserved fragments of the hemagglutinin stem of two subtypes of influenza A-H1N1 and H3N2-and conserved protein M2. Thus, the aim of this study was to design and characterize modified mRNA obtained using the above plasmid DNA vaccines as a template. To select the most promising protocol for creating highly immunogenic mRNA vaccines, we performed a comparative analysis of mRNA modifications aimed at increasing its translational activity and decreasing toxicity. We used mRNA encoding a green fluorescent protein (GFP) as a model. Eight mRNA-GFP variants with different modifications (M0-M7) were obtained using the classic cap(1), its chemical analog ARCA (anti-reverse cap analog), pseudouridine (Ψ), N6-methyladenosine (m6A), and 5-methylcytosine (m5C) in different ratios. Modifications M2, M6, and M7, which provided the most intensive fluorescence of transfected HEK293FT cells were used for template synthesis when mRNA encoded influenza immunogens AgH1, AgH3, and AgM2. Virus specific antibodies were registered in groups of animals immunized with a mix of mRNAs encoding AgH1, AgH3, and AgM2, which contained either ARCA (with inclusions of 100% Ψ and 20% m6A (M6)) or a classic cap(1) (with 100% substitution of U with Ψ (M7)). M6 modification was the least toxic when compared with other mRNA variants. M6 and M7 RNA modifications can therefore be considered as promising protocols for designing mRNA vaccines.

10.
Vaccines (Basel) ; 9(2)2021 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-33494530

RESUMO

One of the key stages in the development of mRNA vaccines is their delivery. Along with liposome, other materials are being developed for mRNA delivery that can ensure both the safety and effectiveness of the vaccine, and also facilitate its storage and transportation. In this study, we investigated the polyglucin:spermidine conjugate as a carrier of an mRNA-RBD vaccine encoding the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. The conditions for the self-assembling of mRNA-PGS complexes were optimized, including the selection of the mRNA:PGS charge ratios. Using dynamic and electrophoretic light scattering it was shown that the most monodisperse suspension of nanoparticles was formed at the mRNA:PGS charge ratio equal to 1:5. The average hydrodynamic particles diameter was determined, and it was confirmed by electron microscopy. The evaluation of the zeta potential of the investigated complexes showed that the particles surface charge was close to the zero point. This may indicate that the positively charged PGS conjugate has completely packed the negatively charged mRNA molecules. It has been shown that the packaging of mRNA-RBD into the PGS envelope leads to increased production of specific antibodies with virus-neutralizing activity in immunized BALB/c mice. Our results showed that the proposed polycationic polyglucin:spermidine conjugate can be considered a promising and safe means to the delivery of mRNA vaccines, in particular mRNA vaccines against SARS-CoV-2.

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