Vaccination is the most efficient way to prevent infectious diseases. mRNA-based vaccines is a new approach to vaccine development, which have several very useful advantages over other types of vaccines. Since mRNA encodes only the target antigen there is no potential risk of infection as in the case with attenuated or inactivated pathogens. The mode of action of mRNA-vaccines implies that their genetic information is expressed only in the cytosol, leaving very little possibility of mRNA integration into the host’s genome. mRNA-vaccines can induce specific cellular and humoral immune responses, but do not induce the antivector immune response. The mRNA-vaccine platform allows for easy target gene replacement without the need to change the production technology, which is important to address the time lag between the epidemic onset and vaccine release. The present review discusses the history of mRNA vaccines, mRNA vaccine production technology, ways to increase mRNA stability, modifications of the cap, poly(A)-tail, coding and noncoding parts of mRNA, target mRNA vaccine purification from byproducts, and delivery methods.

Background/Objectives: The development of a vaccine against highly pathogenic avian influenza viruses subtype A/H5 is an urgent task due to concerns about its pandemic potential. Methods: In this study, we have developed an experimental mRNA vaccine, mRNA-H5, encoding a modified hemagglutinin trimer of influenza virus A/turkey/Stavropol/320-01/2020 (H5N8). BALB/c mice were immunized with the mRNA-H5 vaccine using lipid nanoparticles (LNPs) and needle-free jet injection (JI). Subsequently, the immune response to vaccine was assessed using ELISA, microneutralization assay, and ICS methods, and a challenge study was conducted. Results: mRNA-H5 was shown to effectively stimulate specific humoral and T-cell immune responses. Moreover, mRNA-H5 delivered by LNPs and JI provided 100% protection of immunized mice against lethal challenge with homologous and heterologous strains of avian influenza virus (A/Astrakhan/3212/2020 (H5N8) and A/chicken/Magadan/14-7V/2022 (H5N1), respectively). Conclusions: The present results indicate that JI can be considered as an alternative to LNPs for mRNA delivery, and according to the literature, JI is safer than delivery using LNP. mRNA-H5 has potential as a vaccine against infection with highly pathogenic avian influenza A/H5 viruses with pandemic potential.

The development of a vaccine against highly pathogenic avian influenza A/H5 viruses is an urgent task due to concerns about its pandemic potential. Here, we have developed an experimental mRNA vaccine mRNA-H5, encoding a modified hemagglutinin trimer of influenza virus A/turkey/Stavropol/320-01/2020 (H5N8). Immunization of BALB/c mice with the obtained mRNA-H5 vaccine using lipid nanoparticles (LNPs) and needle-free jet injection (JI) has been shown to effectively stimulate specific humoral and T-cell immune responses. Moreover, mRNA-H5 delivered by LNPs and JI provided 100% protectivity of immunized mice against lethal challenge with homologous and heterologous strains of avian influenza virus (A/Astrakhan/3212/2020 (H5N8) and A/chicken/Magadan/14-7V/2022 (H5N1), respectively). The present results indicate that needle-free jet injection can be considered as an alternative to LNPs for mRNA delivery. mRNA-H5 is a promising candidate as a vaccine against highly pathogenic avian influenza A/H5 viruses with pandemic potential.

Background: Seasonal influenza remains a significant public health problem, and the constant antigenic drift of viruses requires regular vaccine updates. mRNA vaccines offer a promising platform for the development of new, effective influenza vaccines. Administration of the naked mRNA vaccine using a needle-free jet injection system further enhances its safety, reduces cost, and eliminates the need for lipid nanoparticles, which are traditionally used for mRNA delivery. Lyophilization of naked mRNA allows for long-term storage at +4 °C. Methods: We designed and produced an mRNA vaccine against seasonal influenza, designated mRNA-Vector-Flu, encoding the hemagglutinin (HA) of the A/Wisconsin/67/2022(H1N1)pdm09, A/Darwin/9/2021(H3N2), and B/Austria/1359417/2021 strains. The vaccine was lyophilized and stored for 1 month in a refrigerator (+4 °C). A comparative immunogenicity study was conducted between synthesized immediately before use prepared and lyophilized naked mRNA-Vector-Flu. The preparations were administered to BALB/c mice using a jet needleless injection twice, 3 weeks apart. Immunogenicity was assessed on day 35 of the study. Results: A comparative immunogenicity study of naked mRNA-Vector-Flu demonstrated that both the synthesized immediately before use prepared formulation and the lyophilized form, stored at +4 °C for a month, induced similar levels of virus-specific antibodies and generated a pronounced T-cell immune response. Conclusions: Delivery of the naked mRNA vaccine using a needle-free jet injection ensures a high-level immune response, which improves its safety, reduces its cost, and eliminates the need for lipid nanoparticles traditionally used for mRNA delivery. At the same time, lyophilization of the naked mRNA vaccine preserves its biological activity and ensures its storage for at least a month at +4 °C temperatures. Our results demonstrate that our proposed approach can be considered a promising direction for the development and improvement of the mRNA vaccine platform.

Avian influenza A virus subtype H5 is widespread in the bird population, and there is a risk of its transmission to humans, therefore, it is necessary to develop an effective vaccine to prevent virus spread.   The aim of the work was to develop prototypes of DNA vaccines encoding modified variants of the hemagglutinin trimer of highly pathogenic avian influenza A virus (H5N8) and its fragment (stem) and to study their immunogenic properties.   Material and methods. DNA constructs were designed, produced and purified, their expression was analyzed in eukaryotic cells using reverse transcription PCR and Western blot analysis. Laboratory animals (Syrian hamsters) were immunized with experimental DNA vaccines using the jet injection method, the antibody titer in their sera was determined using ELISA. The sera were also tested in the virus neutralization reaction in MDCK cell culture.   Results and discussion. Expression of target genes in experimental DNA vaccines was confirmed at the transcription and translation levels. Immunogenicity analysis showed that immunization of Syrian hamsters with pVAX-H5 and pVAX-H5 delT4 constructs encoding hemagglutinin ectodomain variants with and without the fibritin trimerizing domain of bacteriophage T4 using jet injection resulted in the formation of specific antibodies (average titer 1:3×104 and 1:5×103, respectively) with virus-neutralizing activity (average 50 % neutralizing titer 1:40). At the same time, specific antibodies were not detected in the group of animals immunized with the DNA vaccine encoding the hemagglutinin stalk.   Conclusions. In the course of the work, DNA vaccines encoding variants of the hemagglutinin trimer of the influenza virus subtype A (H5N8) and its fragment (stem) were designed and obtained. A comparison of the immunogenicity of experimental DNA vaccines was conducted and the most promising variant of the design – pVAX-H5, encoding a modified trimer of the hemagglutinin of the influenza virus with a trimerizing domain of the fibritin of the bacteriophage T4 – was established.

MRNA-Vaccine Platform: Features of Obtaining and Delivery of mRNA Платформа мРНК-вакцин: особенности получения и доставки мРНК