Development and laboratory production of virus-like immune-stimulating complexes based on saponins and evaluation of their adjuvant potential using mice immunisation with influenza antigens
Abstract:The COVID-19 pandemic has exacerbated the public’s need for effective vaccines. Consequently, significant financial support has been provided to developers of a number of innovative vaccines, including the vaccines with saponin-based adjuvants. In 2021, the World Health Organisation recommended Mosquirix, the first malaria vaccine, which contains a saponin adjuvant. An anti-covid vaccine by Novavax is in the approval phase. A promising approach to vaccine development is presented by the use of virus-like immun… Show more
Introduction. Despite mass vaccination against the SARS-CoV-2 coronavirus, people in many countries worldwide continue to experience multiple pandemic outbreaks of the disease. Adaptive mutations in the viral genome may alter the pathogenic potential of the virus, affect the ability of the virus to escape immune system response and complicate the development of antiviral vaccines. Data on the neutralizing activity of sera obtained against previously circulating genetic variants of the virus in relation to the current SARS-CoV-2 strains may serve as a scientific basis for vaccine production. The aim of this work was to study cross-reactivity of SARS-CoV-2 coronavirus strains belonging to different genetic variants, which were isolated in the territory of the Russian Federation during 2020-2022 in the neutralization reaction using mouse hyperimmune sera. Materials and methods. Ten SARS-CoV-2 coronavirus strains belonging to different genetic variants (three non-VOC strains, alpha, beta, gamma, delta, delta+AY, omicron 1 and omicron 2) used in this work. BALBc mice immunized with inactivated concentrated antigen mixed with a 1:1 adjuvant, which was a virus-like immunostimulatory complex based on Quillaja soaponaria saponins (Quillaja saponaria). The antibody titer was determined in the neutralization reaction. Results. A decrease in the neutralizing ability of antibodies specific to viruses no VOC was shown to the beta variant and to a lesser extent to alpha and gamma variants. Neutralizing activity of sera specific to the delta variant against alpha and beta variants is reduced 4 -fold.Neutralizing activity of sera obtained to omicron variants 1 and 2 was reduced 18-fold with respect to the Wuhan variant, 12-fold for the gamma variant, and more than 30-fold for the delta variants; for the other variants it was even lower. Conclusions. The results obtained indicate the presence of cross-reactivity between strains of coronavirus belonging to genetic lines Wuhan, alpha, beta, gamma, it is weaker for delta variants. Mutations in the genome of the omicron variants resulted in a significant decrease in antigenic cross-reaction with earlier genetic variants of the coronavirus. The results obtained explain the low efficacy of vaccines based on the Wuhan strain, synthetic immunogens, and recombinant proteins based on it against omicron variants, which have caused an increase in the incidence of disease since early 2022, as well as cases of re-infection of people when infected with new genetic variants of coronavirus.
Introduction. Despite mass vaccination against the SARS-CoV-2 coronavirus, people in many countries worldwide continue to experience multiple pandemic outbreaks of the disease. Adaptive mutations in the viral genome may alter the pathogenic potential of the virus, affect the ability of the virus to escape immune system response and complicate the development of antiviral vaccines. Data on the neutralizing activity of sera obtained against previously circulating genetic variants of the virus in relation to the current SARS-CoV-2 strains may serve as a scientific basis for vaccine production. The aim of this work was to study cross-reactivity of SARS-CoV-2 coronavirus strains belonging to different genetic variants, which were isolated in the territory of the Russian Federation during 2020-2022 in the neutralization reaction using mouse hyperimmune sera. Materials and methods. Ten SARS-CoV-2 coronavirus strains belonging to different genetic variants (three non-VOC strains, alpha, beta, gamma, delta, delta+AY, omicron 1 and omicron 2) used in this work. BALBc mice immunized with inactivated concentrated antigen mixed with a 1:1 adjuvant, which was a virus-like immunostimulatory complex based on Quillaja soaponaria saponins (Quillaja saponaria). The antibody titer was determined in the neutralization reaction. Results. A decrease in the neutralizing ability of antibodies specific to viruses no VOC was shown to the beta variant and to a lesser extent to alpha and gamma variants. Neutralizing activity of sera specific to the delta variant against alpha and beta variants is reduced 4 -fold.Neutralizing activity of sera obtained to omicron variants 1 and 2 was reduced 18-fold with respect to the Wuhan variant, 12-fold for the gamma variant, and more than 30-fold for the delta variants; for the other variants it was even lower. Conclusions. The results obtained indicate the presence of cross-reactivity between strains of coronavirus belonging to genetic lines Wuhan, alpha, beta, gamma, it is weaker for delta variants. Mutations in the genome of the omicron variants resulted in a significant decrease in antigenic cross-reaction with earlier genetic variants of the coronavirus. The results obtained explain the low efficacy of vaccines based on the Wuhan strain, synthetic immunogens, and recombinant proteins based on it against omicron variants, which have caused an increase in the incidence of disease since early 2022, as well as cases of re-infection of people when infected with new genetic variants of coronavirus.
Scientific relevance. The use of recombinant antigens in vaccine production is limited because vaccines based on such antigens tend to have low immunogenicity. However, a COVID-19 vaccine that combines recombinant SARS-CoV-2 spike glycoprotein as its antigen and virus-like immune-stimulating complexes (ISCOMs) as its adjuvant (Nuvaxovid) induces a protective virus-neutralising response. The State Research Center of Virology and Biotechnology “Vector” (hereinafter, Vector) has developed the ISCOM adjuvant Matrix-V, which plays a key role in inducing virus-neutralising antibodies. Studying Matrix-V will provide for the wide use of recombinant antigens combined with this adjuvant in the development and production of novel Russian vaccines.Aim. This study aimed to evaluate the humoral immune responses of experimental animals to intramuscular injections of a complex combining the recombinant Wuhan-type SARS-CoV-2 spike RBD antigen and the virus-like ISCOM adjuvant containing Quillaja saponaria saponins.Materials and methods. The Matrix-V ISCOM adjuvant was produced using Vector’s proprietary technology, which involves cross-flow filtration through Sartorius VivaFlow cassettes. To determine the saponin and residual detergent concentrations in Matrix-V, the authors conducted high-performance liquid chromatography. Having produced the recombinant SARS-CoV-2 RBD antigen, the authors used electron microscopy to analyse the ultrastructure of the ISCOM–antigen complex. In the study of the ISCOM–antigen complex, 25 female Balb/c mice (5 groups) and 15 male and female outbred guinea pigs (3 groups) received two intramuscular injections with a 14-day interval. Serum tests relied on virus neutralisation (VN) and enzyme-linked immunosorbent assay (ELISA) methods and used antigens of 8 SARS-CoV-2 variants (State Collection of Viruses and Rickettsia, Vector). The authors used Statistica 10 to analyse the results.Results. Two injections of the SARS-CoV-2 RBD antigen (mice: 7 μg, guinea pigs: 1 μg) alone did not induce statistically significant virus-neutralising antibody responses, as shown by the VN results. Two injections of the SARS-CoV-2 RBD antigen (mice: 7 μg, guinea pigs: 1 μg) adjuvanted with Matrix-V (25 μg) resulted in geometric mean antibody titres of 1:83–1:178 (mice) and 1:174–1:587 (guinea pigs) in the VN tests with the Wuhan variant. One injection of the antigen (1 μg or 7 μg) with Matrix-V (25 μg) induced antibodies only in individual cases, as demonstrated by the VN and/or ELISA results. The most intensive immune response was observed in ELISA tests with the Delta variant after two injections of the Ecto-S-Wuhan (1 μg) and Matrix-V (25 μg) complex. Immune responses did not differ between the group that received two injections of the Ecto-S-Wuhan antigen (1 μg) without the ISCOM adjuvant and the negative control group (titres below 1:100; p=0.95). Two injections of the SARS-CoV-2 RBD antigen (7 μg) without the ISCOM adjuvant induced antibodies in mice (titres between 1:248 and 1:1477).Conclusions. Two intramuscular injections of the complex containing the recombinant SARS-CoV-2 RBD antigen and the Matrix-V ISCOM adjuvant induce virus-neutralising antibodies. The approach proposed by the authors has the potential for use in the development of immunobiological medicinal products to prevent and treat a wide range of infectious diseases.
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