Endolysins as tools to eradicate pneumococcal biofilms and development of protective immunity

内溶素作为根除肺炎球菌生物膜和发展保护性免疫的工具

• 批准号: 10587285
• 负责人: Norberto Gonzalez Juarbe
• 金额: $ 90.1万
• 依托单位: J. CRAIG VENTER INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-12-09 至 2023-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10587285
• 关键词: Address; Animal Model; Animals; Antibiotic Resistance; Antibiotics; Antibodies; Antibody Formation; Antibody Response; Antibody-mediated protection; Antigens; Antimicrobial Resistance; Apoptosis; Automobile Driving; Bacteremia; Bacteria; Bacterial Proteins; Bacteriophages; Benchmarking; Biochemical; Biochemistry; Biological; Biological Assay; Biophysics; Cell Death; Cell Death Induction; Cells; Cessation of life; Child; Coculture Techniques; Country; Developing Countries; Development; Disadvantaged; Disease; Effectiveness; Elderly; Enzymes; Epithelium; Extracellular Matrix; Fatality rate; Generations; Goals; Gram-Positive Bacteria; Health; Health Care Costs; Human; Immunity; Immunize; Immunologic Techniques; Immunologics; In Vitro; Individual; Inflammation; Lead; Lung; Lytic Phase; Macrophage; Mass Spectrum Analysis; Microbial Biofilms; Modeling; Molecular; Mus; N-Acetylmuramoyl-L-alanine Amidase; Nasopharynx; Necrosis; Nose; Oral mucous membrane structure; Organism; Otitis Media; Peptidoglycan; Pharyngeal structure; Phosphotransferases; Pneumococcal Colonization; Pneumococcal Infections; Pneumonia; Polymers; Polysaccharides; Population; Predisposition; Property; Proteins; Proteomics; Proxy; Research; Resistance; Role; Safety; Serotyping; Sinusitis; Specificity; Streptococcus pneumoniae; Streptococcus pneumoniae plY protein; Technology; Testing; Therapeutic; Tissues; Toxic effect; Toxin; Transgenic Mice; Vaccines; World Health Organization; antimicrobial; aspirate; cellular development; community acquired pneumonia; effectiveness evaluation; effectiveness testing; endolysin; extracellular; immunogenic; in vivo; member; mucosal vaccine; next generation; novel; opportunistic pathogen; oral commensal; point of care; prevent; resistant strain; response; single-cell RNA sequencing; sugar; tool; transcriptomics

Project Summary Streptococcus pneumoniae (Spn, the pneumococcus) are Gram-positive bacteria and the leading cause of community-acquired pneumonia worldwide. The World Health Organization estimates >1.6 million deaths are the result of Spn infection each year, with children and the elderly being the most susceptible populations. Major problems of pneumococcal disease include the acquisition of antimicrobial resistance and the global spread of resistant clones. In addition, these problems are magnified by the major disadvantages of the current capsular polysaccharide-based vaccines, such as serotype specificity and the resulting incomplete coverage. An emerging way to address the growing antimicrobial resistance problem is the use of bacteriophage endolysins. These enzymes are capable of degrading the bacterial peptidoglycan, killing and dispersing biofilm bacteria and its matrix. In preliminary studies, we have developed a chimeric derivative of the well characterized Cpl-1 endolysin that displays >100-fold increase in antimicrobial activity, termed ClyX-1. We also show the ability of this new endolysin to lyse planktonic Spn, and importantly we also observed that ClyX-1 was able to kill biofilm Spn, as well as disperse the biofilm matrix. In addition, we have shown that upon Spn nasopharyngeal colonization, activation of programmed necrosis, i.e. necroptosis, leads to development of antigen-specific antibodies. Of note, treatment of colonized mice with ClyX-1 further promoted necroptosis activation, suggesting endolysin treatment may enhance development of protective immunity against Spn. Herein, we aim to address three overall hypotheses: a) that endolysins are efficient pneumococcal anti-biofilm agents, b) that endolysins can be an effective way to prevent Spn colonization in a serotype-independent manner, and c) that intranasal treatment with endolysins promotes protective immunity, to prevent re-colonization and severe disease. We will use a combination of in vitro and in vivo studies with static and dynamic biofilms, mice, biochemistry (characterize and benchmark pneumococcal endolysins), transgenic mice (to define the role of programmed cell death in protective immunity), molecular and immunological techniques and next generation technologies (proteomics, single cell transcriptomics) to establish better understanding of the effects of endolysin treatments against pneumococcal disease in vivo and test their effectiveness in development of long-term serotype-independent protective immunity.

项目摘要 肺炎链球菌（Streptococcus pneumoniae，Spn，肺炎球菌）是革兰氏阳性菌，是引起肺炎的主要原因。 全球社区获得性肺炎据世界卫生组织估计， 每年都有大量的人感染Spn，儿童和老年人是最易感染的人群。主要 肺炎球菌疾病的问题包括获得抗微生物药物耐药性和肺炎球菌的全球传播。 抗性克隆此外，这些问题被当前的胶囊包装的主要缺点放大。 多糖类疫苗，如血清型特异性和由此产生的不完全覆盖。一个 解决日益增长的抗微生物剂耐药性问题的新兴方法是使用噬菌体内溶素。 这些酶能够降解细菌肽聚糖，杀死并分散生物膜细菌， 它的矩阵。在初步的研究中，我们已经开发了一个嵌合衍生物的特点Cpl-1 显示抗微生物活性增加>100倍的内溶素，称为ClyX-1。我们还展示了 这种新的内溶素溶解嗜酸性Spn，重要的是，我们还观察到ClyX-1能够杀死生物膜 Spn，以及分散生物膜基质。此外，我们已经表明，在Spn鼻咽 定植，程序性坏死的激活，即坏死性凋亡，导致抗原特异性 抗体的值得注意的是，用ClyX-1处理定殖小鼠进一步促进了坏死性凋亡激活，表明 内溶素治疗可增强针对Spn的保护性免疫的发展。在此，我们旨在解决 三个总体假设：a）内溶素是有效的肺炎球菌抗生物膜剂，B）内溶素 可以是以非依赖于细菌类型的方式预防Spn定植的有效方式，和c）鼻内 用内溶素治疗促进保护性免疫，以防止再定殖和严重疾病。我们将 使用静态和动态生物膜、小鼠、生物化学（表征）和生物降解的体外和体内研究的组合， 和基准肺炎球菌内溶素），转基因小鼠（以确定程序性细胞死亡在 保护性免疫），分子和免疫学技术以及下一代技术（蛋白质组学， 单细胞转录组学），以更好地了解内溶素治疗对 肺炎球菌疾病在体内和测试其有效性的发展，长期的非依赖型 保护性免疫