ABSTRACT
We studied a needle-free jet injection delivery of an experimental mRNA vaccine encoding the receptor-binding domain of the SARS-CoV-2 S protein (mRNA-RBD). Immunization of BALB/c mice with mRNA-RBD by a needle-free jet injector induced high levels of antibodies with virus-neutralizing activity and a virus-specific T-cell response. The immune response was low in the group of mice that received intramuscular injection of mRNA-RBD. The effectiveness of this simple and safe method of mRNA delivering has been demonstrated. Thus, jet injection of mRNA vaccine can be a good alternative to lipid nanoparticles.
Subject(s)
Antibodies, Neutralizing , Antibodies, Viral , COVID-19 Vaccines , COVID-19 , Mice, Inbred BALB C , SARS-CoV-2 , Spike Glycoprotein, Coronavirus , Animals , SARS-CoV-2/immunology , SARS-CoV-2/genetics , Mice , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/genetics , Antibodies, Viral/immunology , COVID-19 Vaccines/immunology , COVID-19 Vaccines/administration & dosage , Antibodies, Neutralizing/immunology , COVID-19/prevention & control , COVID-19/immunology , COVID-19/virology , Injections, Jet , mRNA Vaccines , RNA, Messenger/genetics , RNA, Messenger/immunology , Injections, Intramuscular , Female , Humans , T-Lymphocytes/immunology , Vaccines, Synthetic/immunology , Vaccines, Synthetic/administration & dosageABSTRACT
The removal of double-stranded RNA (dsRNA) contaminants during in vitro mRNA synthesis is one of the technological problems to be solved. Apparently, these contaminants are the result of the T7 RNA polymerase side activity. In this study, we used a modified method of mRNA purification based on the selective binding of dsRNA to cellulose in ethanol-containing buffer. It was shown both in vivo and in vitro that the cellulose-purified mRNA preparation leads neither to activation of the lymphocyte inflammatory marker CD69 nor to increased release of IFNα in mice, and does not contain impurities detectable by antibodies to dsRNA.
Subject(s)
RNA, Double-Stranded , RNA, Messenger , Animals , RNA, Messenger/genetics , RNA, Messenger/metabolism , RNA, Double-Stranded/genetics , RNA, Double-Stranded/metabolism , Mice , Antigens, Differentiation, T-Lymphocyte/metabolism , Antigens, Differentiation, T-Lymphocyte/genetics , Antigens, CD/genetics , Antigens, CD/metabolism , Lectins, C-Type/metabolism , Lectins, C-Type/genetics , DNA-Directed RNA Polymerases/metabolism , DNA-Directed RNA Polymerases/genetics , Interferon-alpha/genetics , Interferon-alpha/metabolism , Interferon-alpha/biosynthesis , Viral Proteins/metabolism , Viral Proteins/geneticsABSTRACT
A promising approach to the development of new means for preventing infection caused by tick-borne encephalitis virus can be DNA vaccines encoding polyepitope T-cell immunogens. A DNA vaccine pVAX-AG4-ub encoding an artificial polyepitope immunogen that includes cytotoxic and T-helper epitopes from the NS1, NS3, NS5, and E proteins of the tick-borne encephalitis virus has been obtained. The developed construct ensured the synthesis of the corresponding mRNAs in transfected eukaryotic cells. Immunization of mice with pVAX-AG4-ub induced the formation of a virus-specific T-cell response providing 50% protection from lethal infection with the virus.
Subject(s)
Encephalitis Viruses, Tick-Borne , Vaccines, DNA , Viral Vaccines , Animals , Mice , Encephalitis Viruses, Tick-Borne/genetics , Vaccines, DNA/genetics , Viral Vaccines/genetics , T-Lymphocytes , ImmunizationABSTRACT
An artificial T-cell immunogen consisting of conserved fragments of different proteins of the SARS-CoV-2 virus and its immunogenic properties were studied in BALB/c mice. To create a T-cell immunogen, we used an approach based on the design of artificial antigens that combine many epitopes from the main proteins of the SARS-CoV-2 virus in the one molecule. The gene of the engineered immunogen protein was cloned as part of the pVAX1 plasmid in two versions: with an N-terminal ubiquitin and without it. The obtained plasmids were analyzed for their ability to provide the synthesis of the immunogen protein in vitro and in vivo. It has been shown that protein product of the created artificial genes is actively processed in HEK293T cells and induces cellular immunity in mice.
Subject(s)
COVID-19 , T-Lymphocytes , Humans , Mice , Animals , HEK293 Cells , SARS-CoV-2/genetics , EpitopesABSTRACT
The spread of the monkeypox virus infection among humans in many countries outside of Africa, which started in 2022, is now drawing the attention of the medical and scientific communities to the fact that immunization against this infection is sorely needed. According to current guidelines, immunization of people with the first-generation smallpox vaccine based on the vaccinia virus (VACV) LIVP strain, which is licensed in Russia, should be performed via transepidermal inoculation (skin scarification, s.s.). However, the long past experience of using this vaccination technique suggests that it does not ensure virus inoculation into patients' skin with enough reliability. The procedure of intradermal (i.d.) injection of a vaccine can be an alternative to s.s. inoculation. The effectiveness of i.d. vaccination can depend on the virus injection site on the body. Therefore, the aim of this study was to compare the development of the humoral and cellular immune responses in BALB/c mice immunized with the LIVP VACV strain, which was administered either by s.s. inoculation or i.d. injection into the same tail region of the animal. A virus dose of 105 pfu was used in both cases. ELISA of serum samples revealed no significant difference in the dynamics and level of production of VACV-specific IgM and IgG after i.d. or s.s. vaccination. A ELISpot analysis of splenocytes from the vaccinated mice showed that i.d. administration of VACV LIVP to mice induces a significantly greater T-cell immune response compared to s.s. inoculation. In order to assess the protective potency, on day 45 post immunization, mice were intranasally infected with lethal doses of either the cowpox virus (CPXV) or the ectromelia virus (ECTV), which is evolutionarily distant from the VACV and CPXV. Both vaccination techniques ensured complete protection of mice against infection with the CPXV. However, when mice were infected with a highly virulent strain of ECTV, 50% survived in the i.d. immunized group, whereas only 17% survived in the s.s. immunized group. It appears, therefore, that i.d. injection of the VACV can elicit a more potent protective immunity against orthopoxviruses compared to the conventional s.s. technique.