B-cell innate and adaptive protective immunity to SARS-CoV-2

B 细胞对 SARS-CoV-2 的先天性和适应性保护性免疫

• 批准号: 10778521
• 负责人: John D Mountz
• 金额: --
• 依托单位: BIRMINGHAM VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10778521
• 关键词: 2019-nCoV; Adjuvant; Affinity; Alabama; Antibodies; Antibody Affinity; Antibody Response; Antigens; Antiviral Response; Autoimmune; Autoimmune Diseases; B cell repertoire; B-Cell Development; B-Lymphocyte Subsets; B-Lymphocytes; B-cell receptor repertoire sequencing; Bar Codes; Biological Assay; Biotechnology; Body Fluids; CD86 gene; COVID-19; COVID-19 outbreak; COVID-19 surveillance; Cells; Cloning; Complement 3d Receptors; Complex; Core Facility; Coronavirus; Critical Care; Data Collection; Dendritic Cells; Development; Disease; Endosomes; Event; Exhibits; Feasibility Studies; Flow Cytometry; Future; Genes; Genomics; Helper-Inducer T-Lymphocyte; Herd Immunity; Human; Hydroxychloroquine; ISG15 gene; ITGAX gene; Immune; Immune response; Immunity; Immunoglobulin D; Immunoglobulin G; Immunology; Individual; Infection; Innate Immune Response; Interferon Type I; Interferon Type II; Interferon alpha; Interferon-beta; Interferons; Label; Laboratories; Light; Mature B-Lymphocyte; Mediating; Microbiology; Mus; Nuclear Protein; Nucleoproteins; Nucleotides; Pathogenesis; Pathway interactions; Patients; Peripheral; Phenotype; Plasma; Plasma Cells; Probability; Production; Proteins; RNA; Recurrence; Research Personnel; Research Project Grants; Ribonucleoproteins; Role; SARS coronavirus; SARS-CoV-2 antibody; SARS-CoV-2 antiviral; SARS-CoV-2 immunity; SARS-CoV-2 infection; SARS-CoV-2 spike protein; Scientist; Serology; Signal Transduction; Social Distance; Specimen Handling; Symptoms; Systemic Lupus Erythematosus; T-Lymphocyte; TLR7 gene; Technology; Testing; Vaccination; Vaccine Design; Viral; Viral Genes; Virus; adaptive immune response; antiviral drug development; contagion; cytokine; expression vector; immunomodulatory therapies; interleukin-21; mid-career faculty; monocyte; neoantigens; pandemic disease; plasma cell development; prevent; programs; recruit; response; single cell analysis; single-cell RNA sequencing; transcriptomics; virology

The COVID-19 caused by the SARS-CoV-2 coronavirus infection is currently a global pandemic. The most important question is why some individuals produce rapid protective antibodies (Abs) and exhibit null or only mild symptoms, whereas others do not? A second urgent question is if a standard laboratory test can be developed to stratify subjects who would be most likely to develop rapid long-lived B cells that could produce protective Abs after infection or after vaccination? Based on the understanding of efficient Ab responses to other coronavirus, an uncomplicated infection would promote a response to the spike (S-protein) as well as to the viral nucleoprotein (vNP). This proposal will focus on the role of interferon-β (IFN-β) that enhances the B cell Ab response. Our previous studies in autoimmune systemic lupus erythematosus (SLE) patients have shown that high levels of B cell endogenous IFN-β enables development of a B cell repertoire skewed toward recognition of an endogenous ribonuclear protein (RNPs) which we propose then form the immune repertoire that is poised to respond to vNPs. In the case of SARS-CoV-2, the vNP is a positive strand RNA complex with a nuclear protein very similar to the natural repertoire that develops in humans. For the 2003 SARS-CoV, vNP was detected in multiple body fluids during early infection and IgG anti-NP has been shown to be long-lived and elevated in recovered patients. We have shown that type I IFN promotes long-lived plasma B cell development including T-cell mediated development of high affinity antibody to a neo-antigen. A similar mechanism is proposed for development of protective Ab response to the S-protein of SARS-CoV-2. At a mechanistic level, we propose that the high affinity response to S-protein also requires enhanced single stranded RNA-induced toll-like receptor 7 (TLR7) signaling and follicular T-cell cytokines especially type II IFN to propel B cell into long-lived plasma cells (PCs). Specific Aim 1 will determine if B cells from COVID-19 convalescent subjects with high endogenous IFN-β produce a strong anti-vNP and S-protein response. This will be investigated using fluorescent labeling of antigen strategy to identify vNP and S-protein-specific B cells and to phenotype these B cells by flow cytometry analysis. Specific Aim 2 will utilize fluorescent and nucleotide barcoded vNP and S-protein to enable analysis by 5' 10X Genomics sequencing to determine the program of type I and type II IFN response pathways that are associated with development of B cells producing protective Abs. These studies are made feasible by our past studies of B cell Ab development and responses from recovered subjects that exhibit high endogenous IFN-β and produce antibodies to vNP or S-proteins. Cloning, sequencing, production and testing of the Ab produced by the vNP and S-protein specific B cells will be carried out using 5' 10X Genomics and cloning of heavy and light chain regions into an expression vector by TWIST Biotechnologies. An outstanding team of investigators from the Birmingham VAMC has been assembled including Dr. John Mountz, with expertise in B cell development and Dr. John Kappes, with expertise in virology. We have recruited scientists with expertise in single-cell analysis and immunology to assist with the project. The studies are highly significant since they will lead to a better understanding and stratifications of subjects who did and did not generate an efficient Ab response of the SARS-CoV-2 virus after infection and to determine the underlying immune landscape required for an efficient response. The studies will be important in the design of vaccines to understand the combined role of the vNP and S-protein of SARS-CoV-2, as well as adjuvant that can promote an efficient interferons and TLR7-mediated B cell response. The studies will also have immediate applications to potential immunomodulatory therapy related to development of the anti-viral response such as a potential mechanism of hydroxychloroquine which can modulate TLR7 signaling and other aspects of viral processing upon entry into cells. Other immediate applications would be to guide development of both innate and adaptive immune responses to boost and enable a long-term B cell immunity.

由SARS-COV-2冠状病毒感染引起的Covid-19目前是全球大流行。最多 重要的问题是为什么有些人会产生快速保护的抗体（ABS）和暴露的null或仅 轻度症状，而其他症状却没有？第二个紧急问题是标准实验室测试是否可以 开发为分层的受试者，这些受试者最有可能发展可能产生的快速长寿命B细胞 感染后或疫苗后的保护性ABS？基于对有效的AB回应的理解 其他冠状病毒，一种简单的感染将促进对峰值（S蛋白）的反应以及对 病毒核蛋白（VNP）。该建议将重点关注干扰素-β（IFN-β）的作用，从而增强B 细胞AB响应。我们先前在自身免疫性全身性红斑狼疮（SLE）患者的研究中 表明高水平的B细胞内源性IFN-β使B细胞库的发展偏向于 识别内源性核糖核蛋白（RNP），然后我们提出，然后形成免疫库 这是毒回VNP的中毒。在SARS-COV-2的情况下，VNP是一个正链RNA复合物， 一种与人类发展的自然曲目非常相似的核蛋白。对于2003 SARS-COV，VNP 在早期感染期间在多种体液中检测到，IgG抗NP已显示为长期寿命 并在恢复的患者中升高。我们已经表明，I型IFN促进了长期寿命的血浆B细胞 开发，包括T细胞介导的对新抗原的高亲和力抗体的发育。类似 提出了用于发展受保护AB对SARS-COV-2 S蛋白的反应的机制。 机械水平，我们建议对S蛋白的高亲和力响应也需要增强 滞留的RNA诱导的Toll样受体7（TLR7）信号传导和卵泡T细胞因子，特别是II型IFN 将B细胞推入长寿命的浆细胞（PC）。特定的目标1将确定来自COVID-19的B细胞是否 具有高内源性IFN-β的疗养受试者产生了强烈的抗VNP和S蛋白质反应。这 将使用抗原策略的荧光标记来研究以鉴定VNP和S蛋白特异性B细胞 并通过流式细胞仪分析表型表型。特定的目标2将利用荧光和核苷酸 条形码的VNP和S蛋白通过5'10X基因组学测序进行分析，以确定程序的程序 I型和II型IFN响应途径与生成受保护的B细胞的发展有关 腹肌。我们过去对B细胞AB发展的研究以及来自 恢复了暴露高内源性IFN-β并产生对VNP或S-蛋白质的抗体的受试者。克隆， 将携带VNP和S蛋白特异性B细胞产生的AB的测序，生产和测试 使用5'10X基因组学以及将重和轻链区域克隆到表达矢量中。 生物技术。伯明翰VAMC的一支杰出的调查员团队已经组装 包括John Mountz博士，具有B细胞开发专业知识和John Kappes博士，并具有专业知识 病毒学。我们招募了具有单细胞分析和免疫学专业知识的科学家，以协助 项目。这些研究非常重要，因为它们将导致更好地理解和分层 感染后SARS-COV-2病毒和没有产生有效的AB反应的受试者 确定有效响应所需的潜在免疫景观。研究将很重要 在设计疫苗的设计中，以了解SARS-COV-2的VNP和S蛋白的联合作用，以及 调整可以促进有效的干扰素和TLR7介导的B细胞反应。研究也将 在与病毒的发展有关的潜在免疫调节疗法中立即应用 响应，例如可以调节TLR7信号和其他的羟氯喹的潜在机制 进入细胞时病毒加工的各个方面。其他直接申请将是指导开发 先天性和适应性免疫复杂都可以增强和启用长期B细胞免疫回报。