TNT: Targeting and Triggering Adjuvancy of DNA Vaccines

TNT：DNA 疫苗的靶向和触发佐剂

• 批准号: 6585254
• 负责人: Michele Aileen Kutzler
• 金额: $ 4.16万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-06-01 至 2006-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6585254
• 关键词: CD antigens; CD4 molecule; CD44 molecule; CD8 molecule; MHC class I antigen; MHC class II antigen; T lymphocyte; active immunization; antigen antibody reaction; antigen presenting cell; cellular immunity; chemokine; enzyme linked immunosorbent assay; flow cytometry; human immunodeficiency virus 1; immunity; interferon gamma; interleukin 8; laboratory mouse; postdoctoral investigator; tumor necrosis factor alpha; vector vaccine; virus antigen

DESCRIPTION (provided by applicant): To control HIV-1, it is thought that the induction of cell-mediated immunity may be critical for interrupting the establishment of infection. This proposal aims to identify a specific immunization protocol in which vaccination with combinations of DNA constructs encoding the chemokines RANTES, IL-8, or CTACK, and/or a novel B7DeltaC-domain costimulatory protein results in an enhanced antigen specific cell mediated immune response to the model HIV-1 Gag antigens. These novel adjuvants were selected because together they should be able to "target and trigger" (TNT) the immune response in vivo. The goal of this application will be to determine the best method for generating a TNT effect, and secondly, to determine the specific cell phenotypes of the responsible mediators. Several HIV-1 Gag DNA immunization strategies including adjuvant combinations with simultaneous intramuscular injections, sequential prime/boost regime, or alternative immunization timing schedules will be examined. Cellular and humoral immune responses will be measured quantitatively through the use of antibody titer ELISA, IFN-g ELISPOT, and Flow Cytometric staining (intracellular stains for IFN-g and TNF-a, and surface staining for the T cell markers CD45RO, CD27, CCR7, CCR5, and MHC Class I and II Tetramers). Experiments will be carried out in a novel CHAD (transgenic for human MHC Class I and II) murine model system developed in our laboratory. In vivo expression and functional kinetics of antigen and adjuvant in muscle cells will be determined by an immunohistochemical assay, using the unique GeneGrip reporter plasmid encoding GFP. Taken together, this proposal aims to identify and characterize a unique TNT adjuvant combination for HIV-1 DNA vaccines resulting in enhanced antigen specific cell-mediated immune responses and improved HIV-1 DNA vaccine candidates.

描述（由申请人提供）：为了控制HIV-1，人们认为细胞介导的免疫力的诱导对于中断感染的建立至关重要。该提案旨在确定一种特定的免疫方案，在该方案中，与编码趋化因子咆哮，IL-8或CTACK的DNA构建体的疫苗接种和/或一种新型的B7DELTAC-核蛋白共刺激蛋白导致增强的抗原抗原细胞介导的对模型HIV-1 GAG-1 GAG抗原的免疫反应。选择了这些新型佐剂，因为它们应该能够“靶向和触发”（TNT）体内免疫反应。该应用的目的是确定产生TNT效应的最佳方法，其次，确定负责任介体的特定细胞表型。将检查几种HIV-1 GAG DNA免疫策略，包括同时进行肌肉内注射，顺序/增强状态或替代免疫计时时间表的辅助组合。细胞和体液免疫反应将通过使用抗体滴度ELISA，IFN-G ELISPOT和流式细胞仪染色（IFN-G和TNF-A的细胞内染色），并通过使用抗体滴度ELISA和TNF-A的细胞染色以及T细胞标记物的表面染色来定量测量。实验将在我们实验室开发的新型CHAD（人类MHC I和II类的转基因）中进行。肌肉细胞中抗原和辅助的体内表达和功能动力学将使用唯一的Genegrip Reporter质粒编码GFP来确定免疫组织化学测定。综上所述，该建议旨在识别和表征HIV-1 DNA疫苗的独特TNT辅助组合，从而增强抗原特异性细胞介导的免疫反应并改善HIV-1 DNA疫苗候选物。