Bacterial Vaccine Antigen Discovery

细菌疫苗抗原的发现

• 批准号: 7034669
• 负责人: LAWRENCE C PAOLETTI
• 金额: $ 29.1万
• 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 2008-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7034669
• 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）：细菌疫苗发现的新方向可能来自于宿主-微生物相互作用的研究，特别是通过使用新描述的技术：动态体外附着和入侵系统（DIVAS）。 DIVAS 的开发目的是研究细菌附着和侵袭，细胞处于特定和受控的生长、代谢和营养水平条件下。使用 DIVAS 和 B 族链球菌 (GBS) III 型菌株进行的实验结果证实了早期的发现，即荚膜多糖对于呼吸道上皮细胞的侵袭并不重要。此外，GBS仅在快速生长而不是相对较慢的生长速度时才侵入这些细胞，并且它们仅在有利于侵入的生长条件下表达多种蛋白质。在本提案中，我们试图检验以下假设：参与真核细胞侵袭的 GBS 蛋白是保护性免疫的新的重要靶标。 GBS 是新生儿败血症和脑膜炎的主要原因，并且在非妊娠成年人和患有基础疾病的老年人中越来越普遍。用 GBS 多糖制备的蛋白结合疫苗已成功进行临床前和临床试验。具有毒力特性的 GBS 蛋白已被描述，一些具有疫苗潜力的蛋白已在临床前进行了测试。 在本提案中，我们将使用 DIVAS 来识别有利于细菌附着/侵入真核细胞的生理条件，并使用 GBS 作为模型病原体。我们将分离并鉴定仅在侵入条件下表达的 GBS 膜蛋白（Sp. Aim 1）。将构建缺乏这些蛋白质基因的 GBS 突变体并测试 DIVAS 中的侵袭性和小鼠中的毒力（Sp. Aim 1）。其中一些蛋白质将直接从 GBS 中纯化，或克隆和重组表达，并作为候选疫苗在小鼠中进行测试（Sp. Aim 2）。 这些利用这种独特的疫苗抗原发现方法的研究结果可以直接应用于其他细菌病原体。