RESUMO
INTRODUCTION: The COVID-19 pandemic caused by SARS-CoV-2 has created serious health problems worldwide. The most effective way to prevent the occurrence of new epidemic outbreaks is vaccination. One of the modern and effective approaches to vaccine development is the use of virus-like particles (VLPs). The aim of the study is to develop a technology for production of VLP based on recombinant SARS-CoV-2 proteins (E, M, N and S) in insect cells. MATERIALS AND METHODS: Synthetic genes encoding coronavirus proteins E, M, N and S were used. VLP with various surface proteins of strains similar to the Wuhan virus, Delta, Alpha and Omicron were developed and cloned into the pFastBac plasmid. The proteins were synthesized in the baculovirus expression system and assembled into VLP in the portable Trichoplusia ni cell. The presence of insertion in the baculovirus genome was determined by PCR. ELISA and immunoblotting were used to study the antigenic activity of VLP. VLP purification was performed by ultracentrifugation using 20% sucrose. Morphology was assessed using electron microscopy and dynamic light scattering. RESULTS: VLPs consisting of recombinant SARS-CoV-2 proteins (S, M, E and N) were obtained and characterized. The specific binding of antigenic determinants in synthesized VLPs with antibodies to SARS-CoV-2 proteins has been demonstrated. The immunogenic properties of VLPs have been studied. CONCLUSION: The production and purification of recombinant VLPs consisting of full-length SARS-CoV-2 proteins with a universal set of surface antigens have been developed and optimized. Self-assembling particles that mimic the coronavirus virion induce a specific immune response against SARS-CoV-2.
Assuntos
Baculoviridae , COVID-19 , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Vacinas de Partículas Semelhantes a Vírus , Animais , SARS-CoV-2/genética , SARS-CoV-2/imunologia , SARS-CoV-2/metabolismo , Vacinas de Partículas Semelhantes a Vírus/imunologia , Vacinas de Partículas Semelhantes a Vírus/genética , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Humanos , COVID-19/virologia , COVID-19/imunologia , Baculoviridae/genética , Baculoviridae/metabolismo , Vacinas contra COVID-19/imunologia , Anticorpos Antivirais/imunologia , Proteínas M de Coronavírus/genética , Proteínas M de Coronavírus/imunologia , Proteínas do Nucleocapsídeo de Coronavírus/imunologia , Proteínas do Nucleocapsídeo de Coronavírus/genética , Proteínas do Nucleocapsídeo de Coronavírus/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas da Matriz Viral/genética , Proteínas da Matriz Viral/imunologia , FosfoproteínasRESUMO
INTRODUCTION: In Russia, almost half of the cases of acute intestinal infections of established etiology in 2022 are due to rotavirus infection (RVI). There is no specific treatment for rotavirus gastroenteritis. There is a need to develop modern, effective and safe vaccines to combat rotavirus infection that are not capable of multiplying (replicating) in the body of the vaccinated person. A promising approach is to create vaccines based on virus-like particles (VLPs). OBJECTIVE: Study of the safety and immunogenicity of a vaccine against rotavirus infection based on virus-like particles of human rotavirus A in newborn minipigs with multiple intramuscular administration. MATERIALS AND METHODS: Newborn minipigs were used as an animal model in this study. The safety of the tested vaccine was assessed based on thermometry data, clinical examination, body weight gain, clinical and biochemical blood parameters, as well as necropsy and histological examination. When studying the immunogenic properties of the Gam-VLP-rota vaccine in doses of 30 and 120 µg, the cellular, humoral and secretory immune response was studied. RESULTS: The results of assessing the general condition of animals during the immunization period, data from clinical, laboratory and pathomorphological studies indicate the safety of the vaccine against human rotavirus infection based on VLP (Gam-VLP-rota) when administered three times intramuscularly. Good local tolerance of the tested vaccine was demonstrated. The results of the assessment of humoral immunity indicate the formation of a stable immune response after three-time immunization with Gam-VLP-rota, stimulation of the production of antigen-specific IgG antibodies and their functional activity to neutralize human rotavirus A. It was shown that following the triple immunization with the minimum tested concentration of 30 µg/dose, animals developed a cell-mediated immune response. The results of the IgA titer in blood serum and intestinal lavages indicate the formation of both a systemic immunological response and the formation of specific secretory immunity to human rotavirus A. CONCLUSION: Thus, three-time intramuscular immunization of minipigs with the Gam-VLP-rota vaccine forms stable protective humoral and cellular immunity in experimental animals. Evaluated vaccine is safe and has good local tolerability.
