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    Home > Active Ingredient News > Infection > Cell and Nature double publications to see how Ta helps the development of new nanoparticle COVID-19 vaccine

    Cell and Nature double publications to see how Ta helps the development of new nanoparticle COVID-19 vaccine

    • Last Update: 2021-10-11
    • Source: Internet
    • Author: User
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    As of September 26, 2021, SARS-CoV-2 has caused more than 230 million confirmed COVID-19 cases worldwide, including as many as 4.
    751 million deaths
    .

    Although China has fully controlled the large-scale spread of the new crown epidemic, there are still some imported cases and sporadic local cases
    .

    The development of a powerful vaccine against the new coronavirus has become a top priority for the prevention and control of the current epidemic
    .

    At present, there are more than 200 candidate vaccines in the world under development, and 22 new crown vaccines in China have entered the clinical trial stage
    .

    In the early stage of the COVID-19 outbreak, the research and development of new crown vaccines mostly focused on inactivated vaccines, nucleic acid vaccines and adenovirus vaccines with relatively mature technical routes
    .

    Subunit vaccines need to explore effective and feasible immunogens, so the early stage is relatively slow.
    However, due to the relatively simple process of subunit vaccines and higher potency and safety, it has recently become the mainstream direction for further development of specific vaccines in the world.
    As many as 80 recombinant subunit vaccines are in preclinical and clinical research
    .

    Professor Zhang Hui’s research group from the Institute of Human Virology, Sun Yat-sen University is engaged in the research of new nanoparticle vaccines.
    Different from the inactivated vaccines, attenuated vaccines and adenovirus vaccines that have mature technical routes in the past, nanoparticle vaccines are multi-antigen recombinant protein vaccines.
    , It can display 24 identical or different new coronavirus antigens on the surface of a single nano-vaccine at the same time
    .

    On November 25, 2020, the Sun Yat-sen University team published a study titled Nanoparticle Vaccines Based on the Receptor Binding Domain (RBD) and Heptad Repeat (HR) of SARS-CoV-2 Elicit Robust Protective Immune Responses in the Cell sub-Journal Immunity.
    Thesis
    .

    Zhang Hui’s team carried out a wild-type SARS-CoV-2 respiratory tract infection on a humanized mouse model immunized with the nanoparticle vaccine, and found that the lungs of the unimmunized mice were severely damaged, the virus replicated rapidly, and some mice They died one after another three days after infection
    .

    The mice immunized with the nanoparticle vaccine are in good health, with no lung damage, and no residual virus can be detected in 11 organs and tissues, including the lungs
    .

    It can be seen that the neutralizing antibodies induced by the nanoparticle vaccine can significantly protect the body from the new coronavirus infection
    .

    In terms of safety, the nanoparticle vaccine developed by Sun Yat-sen University has relatively high safety
    .

    The experimental results showed that the nanoparticle vaccine did not cause side effects such as damage to other organs in mice.
    The team also tested whether the antibodies induced by the mice might cause antibody-dependent infection enhancement (ADE), and found that none of the antibodies induced by the nanoparticle vaccine ADE function
    .

    During the initial immunogen screening process, Professor Zhang Hui’s team fully considered the possible lack of immunogenicity caused by a single immunization with monomeric antigens, and the possible synergistic effect of helper T cells (Tfh) and germinal center B cells (GC B) Low
    .

    The team applied Ferritin's nanoparticle display technology to the development of new crown vaccines, using Ferritin's ability to self-assemble into 24-mers to densely display RBD antigens on the surface of nanoparticles
    .

    At the same time, in order to fully mobilize the synergistic immunity of T cells, the team also introduced HR antigens rich in T cell epitopes onto the surface of the nanoparticle vaccine, and displayed the dual antigens of RBD and HR on the surface of the nanoparticle through the SpyTag/SpyCatcher (ST/SC) system.
    (Figure 1C)
    .

    Ferritin is expressed by the bacterial system, RBD and HR are expressed by the eukaryotic CHO-S system, and purified using Ni Sepharose excel (Cytiva)
    .

    Subsequently, different ratios of RBD and HR were incubated with Ferritin, and quickly assembled in a common buffer environment to form nanoparticles containing RBD or RBD/HR
    .

    It is purified by Superose 6 Increase 10/300 GL molecular sieve (Cytiva) and concentrated by ultrafiltration to finally produce a nanoparticle vaccine (Figure 1B)
    .

    Nanoparticles containing only SC-Ferritin peaked at 12-15mL; RBD-Ferritin nanoparticles peaked at 11-14mL; RBD/HR-Ferritin nanoparticles peaked at 8-13mL (Figure 1E)
    .

    The purity and uniformity of the nanoparticles were verified by Western blotting, molecular sieve (SEC), and transmission electron microscopy (TEM)
    .

    Figure 1 The construction and purification of nanoparticle vaccines
    .
    Subsequently, the author tested the humoral immune response of nanoparticle vaccines .

    Compared with protein vaccines, these two nano-vaccines produced stronger B cell immune responses after immunizing BALB/c mice with specific antibody levels about 2 orders of magnitude higher
    .

    Since SARS-CoV-2 recognizes and enters target cells through RBD binding hACE2, the author then used Biacore to study whether the coupling of nanoparticles affects the binding of RBD and hACE2
    .

    The specific Biacore experiment was completed by teacher Zhang Xuanhong on the platform of Sun Yat-sen University School of Medicine using Biacore T100
    .

    Use a CM5 chip to couple hACE2 (2 μg/mL, in 10mM acetate buffer, pH 4.
    5)
    .

    Nanoparticles containing only Ferritin (negative control), RBD monomer (positive control), RBD-Ferritin nanoparticles and RBD-HR-Ferritin nanoparticles were diluted into different concentration gradients for injection (30 μL/min) for 120 seconds, and dissociated 200s
    .

    Biacore results showed that the binding of RBD-Ferritin and RBD-HR-Ferritin nanoparticles to hACE2 was in the same order of magnitude
    .

    After that, the author used Flow cytometry for serum suppression experiments, FRNT cell experiments for neutralization experiments, and further studied the humoral immune response of the nanoparticle vaccine
    .

    Figure 2 Biacore tests the binding of different nanoparticles and RBD monomers to hACE2.
    Next, the author used ELISpot (enzyme-linked immune absorbent spot) and ICCS (intracellular cytokine staining) assays to test the T cell immune response of the nanoparticle vaccine
    .

    Compared with protein vaccines, these two nano-vaccines produced stronger T cell immune response after immunizing BALB/c mice, and the immune response was mainly Th1, but there was no significant difference in Th2 immune response (IL-4 level was equivalent), indicating Nanoparticle vaccines will not produce vaccine-related enhancement of respiratory diseases
    .

    The analysis of dendritic cells and macrophages showed that these two nano-vaccines can promote a more effective antigen presentation process
    .

    Based on the ex vivo test of human PBMC, similar results were obtained, and the nanoparticle vaccine induced a stronger T cell and B cell immune response
    .

    The researchers also found that serum immunized with nanoparticle vaccines does not enhance the infection of target cells through FcγR receptors, that is, there is no antibody-mediated infection enhancement (ADE)
    .

    The serum of the positive control group, that is, the serum of animals infected with ZIKV, showed a significant increase in infection
    .

    Finally, the study also systematically analyzed the lesions and damages of more than 10 organs and tissues of the mouse model immunized with the nanoparticle vaccine, as well as the body temperature, weight, blood routine and urine routine of the rhesus monkey from 1 week before immunization to 12 weeks after immunization.
    Changes in other physiological and biochemical indicators (Figure 3)
    .

    It was found that the organs of the individual animals immunized with the nanoparticle vaccine were normal, and all indicators fluctuated within the normal range, which provided a valuable research basis for in-depth clinical practice
    .

    Figure 3 Nanoparticle vaccine preparation and animal immune model.
    In the whole article, ÄKTA is supplemented by Ni Sepharose excel, Superose 6 Increase 10/300 GL molecular sieve, which helps the preparation and quality control of nanoparticle vaccines, and Biacore helps the molecular level of nanoparticle vaccines.
    Combined with activity analysis, combined with a series of cell and animal experiments, we finally explored and developed a new dual-antigen nanoparticle vaccine against the new coronavirus
    .

    The neutralizing antibody titer, T/B cell synergistic immune response and cross-protection of the nanoparticle vaccine are significantly higher than that of the subunit monomer vaccine, which greatly expands the current vaccine research and development technology route
    .

    In-depth exploration of the mechanism by which the nanoparticle vaccine works and the development of preclinical safety assessment will provide a stronger guarantee for the vaccine to be launched on the market
    .

    Figure 4 A review of the overall idea of ​​the article.
    In August 2021, Professor Zhang Hui’s team made persistent efforts and re-published an article in the Nature sub-Journal Cellular & Molecular Immunology, screening and optimizing the key “connection elements” in nanoparticle vaccine delivery vehicles, using GvTagOpti/ The SdCatcher (Gv/Sd) system replaces the industry-used SpyTag/SpyCatcher (ST/SC) system (Figure 1A, C), and compared the differences between Gv/Sd and ST/SC with Biacore.
    The relevant experiment was conducted by Zhang Xuanhong in Sun Yat-sen University Medicine The hospital platform uses Biacore T100 to complete
    .

    Recombinant SdCatcher-Ferritin and SpyCatcher-Ferritin were fixed on the CM5 chip by amino coupling
    .

    The half-diluted GvTagOpti-RBD and SpyTag-RBD concentration gradients (0.
    1875-6 μM) were injected with HEPS buffer
    .

    The results show that the affinity of GvTagOpti-RBD to SdCatcher-Ferritin is 3 times higher than that of SpyTag-RBD to SpyCatcher-Ferritin (Figure 5)
    .

    The natural protein isopeptide bond "connection element" with completely independent intellectual property rights screened and optimized from various bacteria not only enhances antigen presentation, but is also suitable for industrialized production, and overcomes the problem of the international industry
    .

    Figure 5 Biacore tests the difference between the GvTagOpti/SdCatcher system and the SpyTag/SpyCatcher system.
    Benefits are here! The AKTA and Biacore instruments used in this research are Cytiva products.
    For this reason, Cytiva invites you to fill out the survey questionnaire (you can fill in by identifying the QR code) to obtain the "Biacore Chips and Consumables Selection Guide", and you can win a gift! We will draw 30 of them to give out Cytiva customized SWISSGEAR backpacks
    .

    The winning result will be announced in the comment section of this article on the 29th
    .

     Biacore, for a better life Reference: Nanoparticle Vaccines Based on the Receptor Binding Domain (RBD) and Heptad Repeat (HR) of SARS-CoV-2 Elicit Robust Protective Immune ResponsesImmunity.
    2020 Dec 15;53 (6):1315-1330.
    e9.
    Improvement of a SARS-CoV-2 vaccine by enhancing the conjugation efficiency of the immunogen to self-assembled nanoparticlesCell Mol Immunol.
    2021 Aug;18(8):2042-2044.
    More Biacore Please refer to the extended reading for articles that help the development of the new crown vaccine: (slide up and down to view) BioNTech & Pfizer (mRNA vaccine) BNT162b vaccines protect rhesus macaques from SARS-CoV-2 Nature.
    2021 Apr;592(7853):283-289.
    Globe Biotech Limited (MRNA vaccine) An mRNA-based vaccine candidate against SARS-CoV-2 elicits stable immuno-response with single doseVaccine.
    2021 Jun 23;39(28):3745-3755.
    State Key Laboratory of Biotherapy, West China Hospital, Sichuan University (Asia Unit vaccine) A vaccine targeting the RBD of the S protein of SARS-CoV-2 induces protective immunityNature.
    2020 Oct;586(7830):572-577.
    Beijing Institute of Life Sciences, Chinese Academy of Sciences (Subunit Vaccines) A ​​Universal Design of Betacoronavirus Vaccines against COVID-19, MERS, and SARSCell.
    2020 Aug 6;182(3):722-733.
    e11.
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