Heterologous polysaccharide synthesis in attenuated Salmonella

减毒沙门氏菌中异源多糖的合成

• 批准号: 8836815
• 负责人: Qingke Kong
• 金额: $ 18.55万
• 依托单位: ARIZONA STATE UNIVERSITY-TEMPE CAMPUS
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8836815
• 关键词: Adjuvant; Adult; Animals; Antibodies; Antigen Targeting; Antigens; Arabinose; Attenuated; Bacterial Infections; Bacterial Polysaccharides; Bacterial Proteins; Binding Sites; Campylobacter jejuni; Carbohydrates; Carrier Proteins; Cell Membrane Permeability; Cell surface; Cells; Characteristics; Chemicals; Child; Cloning; Complex; Conjugate Vaccines; DNA; DNA Vaccines; Distal; Down-Regulation; Elderly; Engineering; Equilibrium; Gene Expression; Generations; Genes; Genetic; Goals; Gram-Positive Bacteria; Growth; Heterophile Antigens; Immune response; Immunity; Immunization; Immunoglobulin A; Immunoglobulin G; In Vitro; Infection; Injection of therapeutic agent; Legal patent; Length; Life; Link; Lipid A; Lipopolysaccharides; Lymphoid Tissue; Memory; Memory B-Lymphocyte; Methods; Modification; Mus; Needles; O Antigens; Operon; Oryctolagus cuniculus; Outcome; Plasmid Cloning Vector; Polysaccharides; Population; Property; Proteins; Quality of life; Recombinants; Safety; Salmonella; Salmonella typhimurium; Series; Serum; Site; Structure; Surface; System; T-Lymphocyte; Technology; Time; Tissues; Toxic effect; United States National Institutes of Health; Vaccines; Virulence; Virulent; Work; Yeasts; base; cell mediated immune response; cost; design; design and construction; dolichyl-diphosphooligosaccharide - protein glycotransferase; extracellular; genetic manipulation; glycosylation; homologous recombination; immunogenicity; improved; in vivo; innovation; macromolecule; mutant; novel; novel strategies; pathogen; pathogenic bacteria; periplasm; prevent; public health relevance; research study; response; sugar; vaccine development

DESCRIPTION (provided by applicant): Polysaccharide (PS) constitutes one of major classes of bio-macromolecules covering the surface of the bacterial pathogens, representing a major virulent determinant and immunogen. Several PS vaccines (including conjugated PS) have been developed to prevent the pathogen infections; these PS vaccines suffer from low yields, non-specific chemical conjugation, high costs, and delivery by injection. Recombinant attenuated Salmonella (RAS) are capable of delivering heterologous antigens, including PS, from a variety of pathogens, generating a range of immune responses including serum antibodies, mucosal IgA, and a panoply of cell-mediated immune responses at local and distal sites. These RAS are designed and constructed to deliver protein or DNA antigens. In this work, we undertake the construction of attenuated Salmonella strains designed specifically for delivery of bacterial PS antigens, including O-antigen polysaccharides (O-PS) from Gram-negative pathogens and capsular polysaccharides (C-PS) from Gram-positive pathogens. In our strains, these complex carbohydrates will be covalently linked to the Salmonella cell surface. Using our novel regulated delayed gene expression technology, all native Salmonella surface carbohydrates will be present at the time of immunization to facilitate Salmonella-host interactions necessary for optimal immunogenicity, but will be down-regulated in vivo, after colonization of host lymphatic tissues. Conversely, synthesis of heterologous carbohydrates will not occur until after the vaccine strain has colonized host tissues. Down-regulation of Salmonella carbohydrates will permit optimal expression and presentation of heterologous carbohydrates without interference or steric blocking by Salmonella surface structures. Linking the target polysaccharide antigens to the Salmonella cell surface provides the strong adjuvant effects of Salmonella LPS. In addition, these "biologically conjugated polysaccharide vaccines" will allow us to take advantage of the known ability of traditional, chemically conjugated polysaccharide antigens to stimulate both T-cell dependent and T-cell independent immune responses to the target antigen.

描述（申请人提供）： 多糖（PS）是覆盖细菌病原体表面的主要生物大分子之一，代表主要的毒力决定簇和免疫原。已开发出多种 PS 疫苗（包括结合 PS）来预防病原体感染；这些 PS 疫苗存在产量低、化学结合非特异性、成本高以及注射给药等问题。重组减毒沙门氏菌 (RAS) 能够传递来自多种病原体的异源抗原，包括 PS，产生一系列免疫反应，包括血清抗体、粘膜 IgA 以及局部和远端部位的一系列细胞介导的免疫反应。这些 RAS 的设计和构建是为了传递蛋白质或 DNA 抗原。在这项工作中，我们构建了专门设计用于传递细菌 PS 抗原的减毒沙门氏菌菌株，包括来自革兰氏阴性病原体的 O 抗原多糖 (O-PS) 和来自革兰氏阳性病原体的荚膜多糖 (C-PS)。在我们的菌株中，这些复杂的碳水化合物将共价连接到沙门氏菌细胞表面。使用我们新颖的调节延迟基因表达技术，所有天然沙门氏菌表面碳水化合物将在免疫时存在，以促进沙门氏菌-宿主相互作用，从而实现最佳免疫原性，但在宿主淋巴组织定植后，将在体内下调。相反，异源碳水化合物的合成只有在疫苗株定植宿主组织后才会发生。沙门氏菌碳水化合物的下调将允许异源碳水化合物的最佳表达和呈递，而不会受到沙门氏菌表面结构的干扰或空间阻断。将目标多糖抗原连接到沙门氏菌细胞表面可提供沙门氏菌 LPS 的强大佐剂作用。此外，这些“生物缀合多糖疫苗”将使我们能够利用传统化学缀合多糖抗原的已知能力来刺激对靶抗原的T细胞依赖性和T细胞非依赖性免疫反应。