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Dendritic Cell-Based Biosensor System

基于树突状细胞的生物传感器系统

基本信息

批准号：
7355641
负责人：
AKIRA TAKASHIMA
金额：
$ 28.32万
依托单位：
UNIVERSITY OF TOLEDO HEALTH SCI CAMPUS
依托单位国家：
美国
项目类别：
财政年份：
2003
资助国家：
美国
起止时间：
2003-09-01 至 2008-02-28
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Dendritic cells (DC) are strategically located at the environmental interface, serving as immunological sentinel and tissue-resident antigen presenting cells. Upon exposure to infectious microorganisms or "danger signals", DC produce pro-inflammatory mediators, undergo maturation, and migrate to lymph nodes to activate naive T cells. Thus, DC activation is the initial and key event linking innate to adaptive immunity. We will employ the DC-based biosensor system to study DC activation processes systematically and to develop novel DC-stimulating agents. Specific aims are: 1) To optimize the DC-based biosensor. We have developed a DC biosensor prototype by transfecting XS106 DC line with 13 luciferase reporters with different cis-enhancers and identified various microbial, biological, pharmacological, and physical agents that activate distinct transcription regulatory pathways in DC. We will optimize this system by isolating DC clones permanently expressing EGFP reporter constructs. 2) System validation. We will test the utility of the DC biosensor by testing currently known DC-stimulating agents and by screening a chemical compound library (>105 complexity) from NCI. Once validated, this technology will be transferred to the industry to facilitate their drug discovery effort. 3) To synthesize DC-stimulating peptoids. Peptoids (N-substituted glycine oligomers) have advantages over peptides in protease resistance, higher stability, and higher diversity. Peptoid libraries (>107 complexity) synthesized in split/pool fashions will be screened in bead-immobilized forms. "Hit" compounds will be transformed into soluble dimeric forms and modified by molecular shuffling to obtain DC-stimulating "lead" peptoids. 4) To synthesize DC-stimulating illumination. Using a tunable illumination device, we will search the universe of light by testing the impact of >107 different illumination. We will also test the concept that DC activation is inducible by photochemical cross-linking of surface receptors. 5) To determine biological consequences of DC activation by different agents. Newly identified DC-stimulating agents will be examined for their in vitro impact on gene expression profiles, surface phenotype, cytokine and chemokine production, and T cell-stimulating capacity of DC and for in vivo impact on Langerhans cells. We will analyze experimenta data using computational biology tools to define algorithms that govern DC activation processes. Our study wil provide new insights into DC biology and may lead to the development of novel adjuvants and therapeutics.

描述（由申请人提供）：树突状细胞（DC）战略性地位于环境界面，充当免疫哨兵和组织驻留抗原呈递细胞。当接触感染性微生物或“危险信号”时，DC 会产生促炎介质，成熟并迁移到淋巴结以激活幼稚 T 细胞。因此，DC 激活是连接先天性免疫和适应性免疫的初始关键事件。我们将利用基于 DC 的生物传感器系统系统地研究 DC 激活过程并开发新型 DC 刺激剂。具体目标是： 1）优化基于直流的生物传感器。我们通过用 13 个具有不同顺式增强子的荧光素酶报告基因转染 XS106 DC 系，开发了 DC 生物传感器原型，并鉴定了激活 DC 中不同转录调控途径的各种微生物、生物、药理学和物理制剂。我们将通过分离永久表达 EGFP 报告基因构建体的 DC 克隆来优化该系统。 2）系统验证。我们将通过测试当前已知的 DC 刺激剂并筛选来自 NCI 的化合物库（> 105 复杂性）来测试 DC 生物传感器的实用性。一旦经过验证，这项技术将被转移到业界，以促进他们的药物发现工作。 3) 合成DC刺激肽。 Peptoids（N-取代的甘氨酸寡聚体）比肽具有蛋白酶抗性、更高的稳定性和更高的多样性等优点。以分流/池方式合成的类肽文库（>107 复杂性）将以珠固定形式进行筛选。 “命中”化合物将转化为可溶性二聚体形式，并通过分子改组进行修饰，以获得刺激 DC 的“先导”肽。 4) 合成直流刺激照明。使用可调照明设备，我们将通过测试超过 107 种不同照明的影响来探索光的宇宙。我们还将测试 DC 激活可通过表面受体的光化学交联诱导的概念。 5) 确定不同试剂激活 DC 的生物学后果。将检查新鉴定的 DC 刺激剂对基因表达谱、表面表型、细胞因子和趋化因子产生以及 DC 的 T 细胞刺激能力的体外影响以及对朗格汉斯细胞的体内影响。我们将使用计算生物学工具分析实验数据，以定义控制 DC 激活过程的算法。我们的研究将为 DC 生物学提供新的见解，并可能导致新型佐剂和治疗方法的开发。