Removal of Double-Stranded RNA Contaminants During Template-Directed Synthesis of mRNA.

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.

Delivery of Experimental mRNA Vaccine Encoding the RBD of SARS-CoV-2 by Jet Injection.

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.

Problems of creating antibody phage libraries and their solutions.

Phage display has become an efficient, reliable and popular molecular technique for generating libraries encompassing millions or even billions of clones of divergent peptides or proteins. The method is based on the correspondence between phage genotype and phenotype, which ensures the presentation of recombinant proteins of known amino acid composition on the surface of phage particles. The use of affinity selection allows one to choose variants with affinity for different targets from phage libraries.

Immunogenic and Protective Properties of Recombinant Hemagglutinin of Influenza A (H5N8) Virus.

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.

DNA Vaccine Encoding the Artificial T-Cell Polyepitope Immunogen of Tick-Borne Encephalitis Virus.

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.

Artificial COVID-19 T-Cell Immunogen.

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 Search for Single-Domain Antibodies Interacting with the Receptor-Binding Domain of SARS-CoV-2 Surface Protein.

We performed a search for nanoantibodies that specifically interact with the receptor-binding domain (RBD) of the SARS-CoV-2 surface protein. The specificity of single-domain antibodies from the blood sera of a llama immunized with RBD of SARS-CoV-2 surface protein S (variant B.1.

mRNA Vaccine Platform: mRNA Production and Delivery.

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.

SARS-CoV-2 RBD Conjugated to Polyglucin, Spermidine, and dsRNA Elicits a Strong Immune Response in Mice.

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.

Immunogenicity of the DNA/Protein Combined Vaccine against COVID-19.

During the COVID-19 pandemic, the development of prophylactic vaccines, including those based on new platforms, became highly relevant. One such platform is the creation of vaccines combining DNA and protein components in one construct. For the creation of DNA vaccine, we chose the full-length spike protein (S) of the SARS-CoV-2 virus and used the recombinant receptor-binding domain (RBD) of the S protein produced in CHO-K1 cells as a protein component.

Optimization of In Vivo Electroporation Conditions and Delivery of DNA Vaccine Encoding SARS-CoV-2 RBD Using the Determined Protocol.

Vaccination against SARS-CoV-2 and other viral infections requires safe, effective, and inexpensive vaccines that can be rapidly developed. DNA vaccines are candidates that meet these criteria, but one of their drawbacks is their relatively weak immunogenicity. Electroporation (EP) is an effective way to enhance the immunogenicity of DNA vaccines, but because of the different configurations of the devices that are used for EP, it is necessary to carefully select the conditions of the procedure, including characteristics such as voltage, current strength, number of pulses, etc.

Can Modern Molecular Modeling Methods Help Find the Area of Potential Vulnerability of Flaviviruses?

Flaviviruses are single-stranded RNA viruses that have emerged in recent decades and infect up to 400 million people annually, causing a variety of potentially severe pathophysiological processes including hepatitis, encephalitis, hemorrhagic fever, tissues and capillaries damage. The family is represented by four genera comprising 89 known virus species. There are no effective therapies available against many pathogenic flaviviruses.

Methods to Produce Monoclonal Antibodies for the Prevention and Treatment of Viral Infections.

A viral threat can arise suddenly and quickly turn into a major epidemic or pandemic. In such a case, it is necessary to develop effective means of therapy and prevention in a short time. Vaccine development takes decades, and the use of antiviral compounds is often ineffective and unsafe.

Are Hamsters a Suitable Model for Evaluating the Immunogenicity of RBD-Based Anti-COVID-19 Subunit Vaccines?

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.

Model systems of human immunodef iciency virus (HIV-1) for in vitro eff icacy assessment of candidate vaccines and drugs against HIV-1.

HIV infection still remains a major challenge for healthcare systems of the world. There are several aspects on counteracting the HIV/AIDS epidemic. The f irst aspect covers preventive measures including educational campaigns on HIV/AIDS and promotion of a healthy lifestyle, protected sex, and pre-exposure prophylaxis of vulnerable groups.

Self-Assembled Particles Combining SARS-CoV-2 RBD Protein and RBD DNA Vaccine Induce Synergistic Enhancement of the Humoral Response in Mice.

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.

Comparative Immunogenicity of the Recombinant Receptor-Binding Domain of Protein S SARS-CoV-2 Obtained in Prokaryotic and Mammalian Expression Systems.

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.

Immunogenic Properties of the DNA Construct Encoding the Receptor-Binding Domain of the SARS-CoV-2 Spike Protein.

The development of preventive vaccines became the first order task in the COVID-19 pandemic caused by SARS-CoV-2. This paper reports the construction of the pVAX-RBD plasmid containing the Receptor-Binding Domain (RBD) of the S protein and a unique signal sequence 176 which promotes target protein secretion into the extracellular space thereby increasing the efficiency of humoral immune response activation. A polyglucine-spermidine conjugate (PGS) was used to deliver pVAX-RBD into the cells.

[Immunogenic Properties of the DNA Construct Encoding the Receptor-Binding Domain of the SARS-CoV-2 Spike Protein].

The development of preventive vaccines became the first order task in the COVID-19 pandemic caused by SARS-CoV-2. This paper reports the construction of the pVAX-RBD plasmid containing the Receptor-Binding Domain (RBD) of the S protein and a unique signal sequence 176 which promotes target protein secretion into the extracellular space thereby increasing the efficiency of humoral immune response activation. A polyglucine-spermidine conjugate (PGS) was used to deliver pVAX-RBD into the cells.

Construction and Immunogenicity of Modified mRNA-Vaccine Variants Encoding Influenza Virus Antigens.

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.

mRNA technology as one of the promising platforms for the SARS-CoV-2 vaccine development.

After the genome sequence of SARS-CoV-2 (Severe acute respiratory syndrome-related coronavirus 2) was published and the number of infected people began to increase rapidly, many global companies began to develop a vaccine. Almost all known approaches to vaccine design were applied for this purpose, including inactivated viruses, mRNA and DNA-vaccines, vaccines based on various viral vectors, synthetically generated peptides and recombinant proteins produced in cells of insects and mammals. This review considers one of the promising vaccine platforms based on messenger RNA.

Genetic Diversity and Drug Resistance Mutations in Reverse Transcriptase and Protease Genes of HIV-1 Isolates from Southwestern Siberia.

The analysis of a gene fragment encoding protease and part of reverse transcriptase was carried out for 55 sera collected in 2016 and 2018 from HIV-1-infected patients diagnosed in 2014-2018 living in the south of Western Siberia, Russia: Altai Territory ( = 11), Republic of Altai ( = 15), Kemerovo region ( = 18), and Novosibirsk region ( = 11). CRF63_02A was the dominant genetic form (>70%) in the Altai Territory and Kemerovo and Novosibirsk regions, with subsubtype A6 comprising <30% of samples. In the Altai Republic, subsubtype A6 was predominant (53%), with 33% of viruses belonging to CRF63_02A.

Delivery of mRNA Vaccine against SARS-CoV-2 Using a Polyglucin:Spermidine Conjugate.

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.

Cationic Polymers for the Delivery of the Ebola DNA Vaccine Encoding Artificial T-Cell Immunogen.

Background: According to current data, an effective Ebola virus vaccine should induce both humoral and T-cell immunity. In this work, we focused our efforts on methods for delivering artificial T-cell immunogen in the form of a DNA vaccine, using generation 4 polyamidoamine dendrimers (PAMAM G4) and a polyglucin:spermidine conjugate (PG).

Methods: Optimal conditions were selected for obtaining complexes of previously developed DNA vaccines with cationic polymers.