Studies of the SARS-CoV-2 Spike Protein

SARS-CoV-2 刺突蛋白的研究

• 批准号: 10926406
• 负责人: JAY A BERZOFSKY
• 金额: $ 214.72万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10926406
• 关键词: 2019-nCoV; ACE2; Adjuvant; Affect; Air Movements; Animal Model; Animals; Antibodies; Antibody Specificity; Antibody titer measurement; Antigens; Appearance; B-Lymphocyte Epitopes; Binding; Bioinformatics; Body Weight decreased; Bronchoalveolar Lavage Fluid; CD8-Positive T-Lymphocytes; COVID-19; COVID-19 prevention; COVID-19 susceptibility; COVID-19 vaccine; Cancer cell line; Cell Line; Cells; Cholesterol; Collaborations; Coupled; DNA Vaccines; Data; Dendritic Cells; Diet; Disease; Dose; Electroporation; Engineering; Epithelial Cells; FDA Emergency Use Authorization; Female; Formulation; Gender; Granulocyte-Macrophage Colony-Stimulating Factor; HIV vaccine; Hamsters; Human; Human Cell Line; Immune; Immune response; Immunity; Immunize; Immunoglobulin A; Immunoglobulin G; In Vitro; Industrialization; Infection; Infection prevention; Interferon Type I; Interleukin-12; Interleukin-15; Licensing; Ligands; Lung; Macaca; Macaca mulatta; Malignant neoplasm of lung; Medical center; Messenger RNA; Methods; Modeling; Monitor; Morbidity - disease rate; Mucosal Immunity; Mucous Membrane; Mus; Myeloid Cells; Nasal cavity; Natural Immunity; Non-Small-Cell Lung Carcinoma; Nose; Obesity; Omega-3 Fatty Acids; Oropharyngeal; Persons; Production; Proteins; Public Health; Publishing; RNA vaccine; Reagent; Recombinants; Respiratory Mucosa; Risk Reduction; Role; Route; SARS-CoV-2 B.1.1.529; SARS-CoV-2 B.1.617.2; SARS-CoV-2 infection; SARS-CoV-2 spike protein; SARS-CoV-2 variant; SIV Vaccines; Serum; Site; South African; T cell response; T-Lymphocyte; TLR3 gene; TMPRSS2 gene; Testing; Texas; Time; Touch sensation; Transgenic Mice; Universities; Vaccinated; Vaccination; Vaccines; Variant; Vascular Endothelial Cell; Viral; Viral Vaccines; Virion; Virus; Work; airway epithelium; aluminum sulfate; chemokine; comparison control; coronavirus disease; dimer; disorder control; disorder prevention; experience; gender difference; health goals; immunogenicity; immunopathology; improved; in vivo; indicated prevention; lung cancer cell; male; mucosal vaccine; nanoparticle; neutralizing antibody; nonhuman primate; preclinical study; prevent; receptor; receptor binding; respiratory challenge; response; transmission process; vaccine candidate; vaccine trial; variants of concern; viral transmission

In vivo vaccine studies in macaques and hamsters: Rhesus macaques have been primed IM with S1 spike protein in different adjuvants and boosted systemically with spike in alum or mucosally intranasally with spike in nanoparticles with IL-15 and TLR ligand adjuvants. We have found and published that nanoparticles containing S1 spike protein delivered intranasally can boost macaques primed IM with S1 in alum and result in better protection against respiratory challenge with SARS-CoV-2 than can the IM vaccine alone even though the S1-binding and neutralizing antibody levels are lower. Other mechanisms must play a role and we have found correlations with mucosal IgA, dimeric IgA, and type I interferon production in the lung, and certain types of myeloid cells. We then tested a mucosal nanoparticle boost with the B1.351 (South African) variant S1 protein to protect against this SARS-CoV-2 variant in macaques. The beta variant was the most difficult to neutralize before the appearance of the omicron variant, which had not been identified at the time we started this second study. The beta variant mucosal nanoparticle vaccine, given 1 full year after the animals had last been boosted systemically or mucosally with the original Wuhan strain, induced a 3-log increase in both IgG binding antibody in both the serum and bronchoalveolar lavage fluid (BAL, representing response in the lung). Further, it boosted the titer to the original Wuhan strain as much as to the beta variant. Neutralizing antibody titers were also similar against both virus variants. IgA and dimeric IgA to both strains were also increased. This suggests a role for original antigenic sin in determining the fine specificity of antibodies at the time of first primary vaccination. When challenged, the animals were well protected against intranasal challenge with the beta variant SARS-CoV-2. Thus, a variant intranasal vaccine can induce strong protective immunity in the lungs and nasal cavity and eliminate virus from these sites. These studies suggest that a human intranasal nanoparticle COVID-19 vaccine given to people who had been previously immunized systemically with one of the approved vaccines, could improve protection against infection and reduce the risk of forward transmission to others by reducing intranasal virus, which is especially a problem with the delta and omicron variants This second NHP study was also published. In addition, three more studies have been carried out in a hamster model, as hamsters get COVID disease more like humans. The intranasal vaccine was able to markedly reduce weight loss in the immunized animals compared to controls, implying the prevention of disease, and reduce virus particles in the lungs. It could also more effectively reduce virus in the oropharynx of hamsters than the S1 protein in alum, an important accomplishment that would be expected to reduce forward transmission. To test this hypothesis, we have now done a forward transmission study in hamsters. In this study, we also examined priming with one of the licensed mRNA vaccines to compare. Groups of hamsters were vaccinated first IM with the Moderna mRNA spike protein vaccine. Then half of them were boosted with the same mRNA vaccine IM, but the other half were boosted with the nasal nanoparticle S1 vaccine incorporating the CpG/PolyI:C & IL15 adjuvants. All the animals were challenged with SARS-CoV-2, including a group that was not immunized at all. Then, they were cohoused with naive hamsters separated only by a screen that allowed air flow between them but did not allow touching. The naive hamsters were then monitored for infection. The intranasal nanoparticle vaccine boost was substantially superior to a second IM dose of the mRNA vaccine in preventing forward transmission to the co-housed naive hamsters! This finding is critical as it demonstrates that an intranasal boost with an effective vaccine such as the one we have developed is needed to prevent COVID-19 infections from spreading, the key public health goal. Gender effects: In the hamsters, we observed that female hamsters were better protected against SARS-CoV-2 infection and disease than males, but reagents are not available to study the hamster immune response in detail. To determine the mechanism, we carried out similar studies in mice and confirmed the gender difference in mice as well. Studies are in progress to identify the mechanistic differences in immune response between the male and female mice, which could be important in optimizing efficacy of human vaccines. In vivo studies in mice: In wild type B6 mice, we have immunized with recombinant spike protein S1, S1+S2, or RBD in several different adjuvants to determine the best formulation. The best combination so far is S1 antigen with IL-15 + ligands for TLR3 and 9, for both antibody and T cell responses, but the runner up was S1 with GM-CSF and IL-12. Studies are in progress to determine which components contribute the most to protection and whether they induce qualitatively different types of immune responses. The DNA vaccine with spike protein coupled to a chemokine has been constructed and initial results show that it can induce a strong CD8 T cell response. Human cell lines: We have received the immortalized human lung epithelial cell lines, which express ACE2, from John Minna at UTSW, as well as some of his non-small-cell lung cancer cell lines that also express ACE2. We have obtained an antibody to ACE2 to verify expression. Initial results show that omega-3 fatty acids and cholesterol differentially affect ACE2 expression on lung cancer cells as well as TMPRSS2 expression and may help explain how diet and obesity as well as lung cancer can affect susceptibility to SARS-CoV-2.

猕猴和仓鼠中的体内疫苗研究：猕猴在不同的佐剂中用S1尖峰蛋白进行了IM，并在校友或粘液鞘内与IL-15和TLR辅助剂的纳米颗粒中的尖峰在校内进行尖峰。我们发现并发表了含有鼻内递送的S1尖峰蛋白的纳米颗粒可以在校友中促进与S1发出的猕猴，并在SARS-COV-2中与SARS-COV-2相比，即使S1结合和中和抗体的水平较低，也可以单独使用SARS-COV-2的呼吸挑战。其他机制必须发挥作用，我们发现与肺中的粘膜IgA，二聚体IgA和I型干扰素产生以及某些类型的髓样细胞相关。然后，我们用B1.351（南非）变体S1蛋白测试了粘膜纳米颗粒的增强，以防止猕猴中的SARS-COV-2变体。在Omicron变体出现之前，Beta变体是最难中和的，在我们开始第二项研究时尚未确定。 β变异粘膜纳米粒子疫苗在动物上次被全身或粘膜上促进原始武汉菌株后整整1年，诱导了血清和支气管肺泡灌洗液中的IgG结合抗体的3-gog增加（代表肺中的BAL）。此外，它与beta变体一样多地提高了原始武汉菌株。中和抗体滴度与两个病毒变体也相似。 IgA和二聚体IgA对两种菌株也增加了。这表明原始抗原犯罪在首次初次疫苗接种时确定抗体的精细特异性中的作用。当受到挑战时，动物受到β变体SARS-COV-2的保护，免受鼻内挑战。因此，鼻内疫苗可以诱导肺部和鼻腔中强烈的保护性免疫，并从这些部位消除病毒。这些研究表明，人类鼻内纳米颗粒共证疫苗以前曾对先前已通过一种认可的疫苗进行全身免疫的人进行免疫，可以改善免受感染的保护，并通过减少Delta和Omicron verianiants的第二次NHP研究的问题来改善鼻内病毒的前进风险。此外，随着仓鼠更像人类的仓鼠疾病，仓鼠模型中又进行了三项研究。与对照组相比，鼻内疫苗能够显着减少免疫动物的体重减轻，这意味着预防疾病并减少肺部病毒颗粒。与明矾中的S1蛋白相比，它还可以更有效地降低仓鼠口咽的病毒，这是一个重要的成就，可以减少前进的传播。为了检验这一假设，我们现在在仓鼠进行了前进的传输研究。在这项研究中，我们还检查了一种有执照的mRNA疫苗进行比较的启动。首先，使用Moderna mRNA尖峰蛋白疫苗接种仓鼠。然后，其中一半用相同的mRNA疫苗IM增强，但另一半通过纳入纳米粒子S1疫苗促进了CPG/Polyi：C＆IL15佐剂。所有动物都受到SARS-COV-2的挑战，其中包括根本没有免疫的团体。然后，它们与幼稚的仓鼠共同居住，仅被屏幕隔开，屏幕使空气流动，但不允许触摸。然后监测天真的仓鼠感染。鼻内纳米颗粒疫苗的提升基本上优于第二次IM剂量的mRNA疫苗，以防止向前传播到共同开设的天真仓鼠！这一发现至关重要，因为它表明，需要采用有效的疫苗（例如我们开发的疫苗）来提高鼻内促进，以防止199年感染传播，这是关键的公共卫生目标。性别影响：在仓鼠中，我们观察到，女性仓鼠比男性更好地保护了SARS-COV-2感染和疾病，但不能详细研究仓鼠免疫反应。为了确定机制，我们在小鼠中进行了类似的研究，并确认了小鼠的性别差异。正在进行研究以确定雄性和雌性小鼠之间免疫反应的机械差异，这对于优化人类疫苗的功效可能很重要。体内研究小鼠：在野生型B6小鼠中，我们已用重组尖峰蛋白S1，S1+S2或RBD免疫，以确定最佳配方。到目前为止，最好的组合是抗体和T细胞反应的S1抗原，具有TLR3和9的IL-15 +配体，但具有GM-CSF和IL-12的亚军为S1。正在进行研究以确定哪些组成部分对保护最大的作用以及它们是否诱导了质性不同类型的免疫反应。已经构建了与趋化因子耦合的尖峰蛋白的DNA疫苗，并最初的结果表明，它可以诱导强大的CD8 T细胞反应。人类细胞系：我们从UTSW的John Minna以及他的一些非小细胞肺癌细胞系中收到了不朽的人类肺上皮细胞系，这些细胞表达ACE2，并从John Minna中表达了ACE2。我们已经获得了与ACE2抗体验证表达的抗体。最初的结果表明，omega-3脂肪酸和胆固醇在肺癌细胞以及TMPRSS2表达上差异影响ACE2的表达，并可能有助于解释饮食和肥胖以及肺癌如何影响SARS-COV-2的易感性。

项目成果

Protection from COVID-19 disease in hamsters vaccinated with subunit SARS-CoV-2 S1 mucosal vaccines adjuvanted with different adjuvants.

• DOI: 10.3389/fimmu.2023.1154496
• 发表时间: 2023
• 期刊: Frontiers in immunology
• 影响因子: 7.3

Potential SARS-CoV-2 Immune Correlates of Protection in Infection and Vaccine Immunization.

• DOI: 10.3390/pathogens10020138
• 发表时间: 2021-01-30
• 期刊: Pathogens (Basel, Switzerland)
• 作者: Sui Y;Bekele Y;Berzofsky JA
• 通讯作者: Berzofsky JA

SARS-CoV-2 Spike Protein Suppresses ACE2 and Type I Interferon Expression in Primary Cells From Macaque Lung Bronchoalveolar Lavage.

• DOI: 10.3389/fimmu.2021.658428
• 发表时间: 2021
• 期刊: Frontiers in immunology
• 影响因子: 7.3
• 作者: Sui Y;Li J;Venzon DJ;Berzofsky JA
• 通讯作者: Berzofsky JA

IL-7 in SARS-CoV-2 Infection and as a Potential Vaccine Adjuvant.

• DOI: 10.3389/fimmu.2021.737406
• 发表时间: 2021
• 期刊: Frontiers in immunology
• 影响因子: 7.3
• 作者: Bekele Y;Sui Y;Berzofsky JA
• 通讯作者: Berzofsky JA

SARS-CoV-2 mucosal vaccine protects against clinical disease with sex bias in efficacy.

SARS-CoV-2 粘膜疫苗可预防具有性别偏见的临床疾病。

• DOI: 10.1016/j.vaccine.2023.11.059
• 发表时间: 2024
• 期刊: Vaccine
• 影响因子: 5.5
• 作者: Sui,Yongjun;Andersen,Hanne;Li,Jianping;Hoang,Tanya;Minai,Mahnaz;Nagata,BiancaM;Bock,KevinW;Alves,DerronA;Lewis,MarkG;Berzofsky,JayA
• 通讯作者: Berzofsky,JayA