Bacterial Vaccine Antigen Discovery

细菌疫苗抗原的发现

• 批准号: 6873703
• 负责人: LAWRENCE C PAOLETTI
• 金额: $ 29.8万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6873703
• 关键词: SDS polyacrylamide gel electrophoresis; Streptococcus agalactiae; bacteria infection mechanism; bacterial antigens; bacterial genetics; bacterial polysaccharides; bacterial proteins; bacterial vaccines; drug discovery /isolation; enzyme linked immunosorbent assay; host organism interaction; immunogenetics; immunomodulators; laboratory mouse; membrane proteins; microorganism culture; microorganism immunology; molecular cloning; mutant; newborn animals; polymerase chain reaction; protein purification; recombinant proteins; respiratory epithelium; tissue /cell culture; virulence

DESCRIPTION (provided by applicant): New directions in bacterial vaccine discovery may arise from studies of host-microbe interactions, particularly through the use of a newly described technology: the dynamic in vitro attachment and invasion system (DIVAS). DIVAS was developed to study bacterial attachment and invasion with cells held at specific and controlled conditions of growth, metabolism, and nutrient levels. Results from experiments performed with DIVAS and group B Streptococcus (GBS) type III strains substantiated earlier findings that capsular polysaccharide is not critical for invasion of respiratory epithelial cells. Moreover, GBS invaded these cells only when held at a fast as opposed to a relatively slower rate of growth and they expressed several proteins solely under growth conditions conducive for invasion. In this proposal, we seek to test the hypothesis that GBS proteins involved with invasion of eukaryotic cells are new and important targets of protective immunity. GBS is a major cause of neonatal sepsis and meningitis, and is increasingly prevalent among non-pregnant adults and the elderly with underlying illnesses. Preclinical and clinical trials have been successfully performed with protein conjugate vaccines prepared with GBS polysaccharides. GBS proteins with virulence properties have been described and some with vaccine potential have been tested preclinically. In this proposal, we will use DIVAS to identify physiological conditions conducive for bacterial attachment/invasion of eukaryotic cells using GBS as a model pathogen. We will isolate and identify GBS membrane proteins expressed solely under invasive conditions (Sp. Aim 1). GBS mutants lacking genes for these proteins will be constructed and tested for invasiveness in DIVAS and virulence in mice (Sp. Aim 1). Several of these proteins will be purified directly from GBS, or cloned and recombinantly expressed, and tested as vaccine candidates in mice (Sp. Aim 2). Findings from these studies utilizing this unique approach to vaccine antigen discovery could be directly applied to other bacterial pathogens.

描述（由申请人提供）：细菌疫苗发现中的新方向可能是由宿主 - 微生物相互作用的研究引起的，尤其是通过使用新描述的技术：动态体外附着和入侵系统（DIVAS）。 DIVA的开发是为了研究细菌的依恋和侵袭，并在生长，代谢和营养水平的特定和受控条件下保持细胞。用Divas和B组链球菌（GBS）进行的实验结果证实了早期发现的结果，即囊囊多糖对于侵袭呼吸道上皮细胞并不是至关重要的。此外，GBS仅在速度而不是相对较慢的生长速率时才入侵这些细胞，并且它们仅在有助于侵袭的生长条件下表达了几种蛋白质。在此提案中，我们试图检验以下假设：与侵袭真核细胞有关的GBS蛋白是保护性免疫的新靶标。 GBS是新生儿败血症和脑膜炎的主要原因，在非怀孕的成年人和患有潜在疾病的老年人中越来越普遍。临床前试验和临床试验已通过用GBS多糖制备的蛋白质结合疫苗成功进行。已经描述了具有毒力特性的GBS蛋白，并且有些具有疫苗电位的蛋白质已临床上测试。 在此提案中，我们将使用DIVAS来确定有利于使用GBS作为模型病原体的真核细胞的细菌附着/侵袭的生理条件。我们将隔离并鉴定仅在侵入性条件下表达的GBS膜蛋白（Sp。AIM1）。缺乏这些蛋白质基因的GBS突变体将被构造并测试小鼠的Divas和毒力中的侵入性（Sp。AIM1）。这些蛋白质中的几种将直接从GB中纯化，或者克隆并重组表达，并在小鼠中作为疫苗候选物进行测试（Sp。AIM2）。 这些研究利用这种独特方法进行疫苗抗原发现的发现可以直接应用于其他细菌病原体。