Development of novel diabetes autoantibody assays based on luciferase reporters

基于荧光素酶报告基因的新型糖尿病自身抗体测定的开发

基本信息

  • 批准号:
    7962864
  • 负责人:
  • 金额:
    $ 7.65万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2010
  • 资助国家:
    美国
  • 起止时间:
    2010-06-01 至 2012-05-31
  • 项目状态:
    已结题

项目摘要

DESCRIPTION (provided by applicant): Type 1A diabetes (T1D) is characterized by an absolute insulin insufficiency resulting from autoimmune destruction of pancreatic beta cells, and is the most common autoimmune endocrine disease afflicting children in the US. It is responsible for significant societal and health care costs, and is increasing at a rate of approximately 2 - 5% per year. Diabetes autoantibodies are currently the most robust biomarkers of disease, and are central to diagnosis, and for stratification of those genetically at risk. Autoantibodies to insulin (IAA) are typically the first to develop, especially in young children, and their measurement may enable identification of individuals who are most likely to benefit from prophylactic insulin therapy. This makes a reliable assay for IAAs particularly important. However the presently used assay performs poorly in standardization tests, and is clearly in need of improvement. Our ongoing studies of autoantibodies to zinc transporter 8 (ZNT8A) have demonstrated that significant improvements to assay specificity and sensitivity can be achieved by making modifications to the probe used for their detection, Our hypothesis is that we can apply the experience we have gained from optimizing the ZNT8A assay to develop an improved procedure for measuring IAAs, based upon the use of novel luciferase containing probes. Our preliminary data indicate that IAAs can be detected using a fusion protein between human preproinsulin (hPPI) and a modified Gaussia princeps luciferase (GLuc) that has enhanced light emission stability. The single specific aim of the current proposal is to optimize this hPPI- GLuc based IAA assay. Initially this will involve a systematic investigation of key features of the probe, such as inclusion of multiple domains, potential for excision of the C peptide, and sequence of the insulin component (human v porcine). Subsequently we will optimize probe production and basic features of the assay procedure such as length and temperature of the incubation between diabetic sera and the probe, and the method of collection of the resulting immune complexes. We believe that the current proposal will lead to the development of an assay for IAAs that has at least equivalent specificity and sensitivity to the currently used mIAA, with the greater utility of not involving the use of radio-isotopes. The basic probe design can also be adapted to incorporate other protein antigens, and hence our proposal could have significant utility for the diagnosis of a variety of other autoimmune diseases, in addition to T1D. PUBLIC HEALTH RELEVANCE: Type 1A insulin dependent diabetes mellitus is one of the most frequent chronic diseases of children and young adults, and carries a high risk of devastating complications in later life. Autoantibodies are the most robust biomarkers of disease, and our proposal will develop improved procedures for measuring autoantibodies to insulin, which currently show poor reproducibility. Insulin is one of the major molecular targets in type 1A diabetes, especially in young children, and our proposal will facilitate diagnosis, and the identification of individuals who may benefit the most from preventative insulin therapy.
描述(由申请人提供):1A型糖尿病(T1D)的特征是由于自身免疫破坏胰腺β细胞而导致的绝对胰岛素不足,是美国儿童最常见的自身免疫性内分泌疾病。它造成了巨大的社会和卫生保健费用,并以每年约2%至5%的速度增长。糖尿病自身抗体是目前最可靠的疾病生物标志物,是诊断和遗传风险分层的核心。胰岛素自身抗体(IAA)通常是首先产生的,特别是在幼儿中,其测量可以识别最有可能从预防性胰岛素治疗中受益的个体。这使得对IAAs的可靠测定尤为重要。然而,目前使用的分析方法在标准化测试中表现不佳,显然需要改进。我们正在进行的锌转运蛋白8 (ZNT8A)自身抗体的研究表明,通过对用于检测的探针进行修改,可以显著提高检测的特异性和灵敏度。我们的假设是,我们可以应用我们从优化ZNT8A检测中获得的经验来开发一种改进的方法来测量IAAs,基于使用新的含荧光素酶的探针。我们的初步数据表明,可以使用人胰岛素前原(hPPI)和改进的高斯原理荧光素酶(GLuc)之间的融合蛋白检测IAAs,该蛋白增强了发光稳定性。当前建议的单一特定目标是优化这种基于hPPI- GLuc的IAA测定。最初,这将涉及对探针的关键特征的系统调查,例如包含多个结构域,C肽切除的可能性以及胰岛素成分的序列(人与猪)。随后,我们将优化探针的生产和检测过程的基本特征,如糖尿病血清与探针之间的孵育时间和温度,以及收集所得免疫复合物的方法。我们认为,目前的建议将导致开发一种IAAs检测方法,其特异性和灵敏度至少与目前使用的mIAA相当,并且不涉及使用放射性同位素的更大效用。该探针的基本设计也可用于结合其他蛋白抗原,因此我们的建议可能对除T1D外的各种其他自身免疫性疾病的诊断具有重要的实用价值。

项目成果

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HOWARD W DAVIDSON其他文献

HOWARD W DAVIDSON的其他文献

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{{ truncateString('HOWARD W DAVIDSON', 18)}}的其他基金

Multimodal analysis of the "honeymoon period" in autoimmune diabetes
自身免疫性糖尿病“蜜月期”的多模态分析
  • 批准号:
    10595074
  • 财政年份:
    2022
  • 资助金额:
    $ 7.65万
  • 项目类别:
Multimodal analysis of the "honeymoon period" in autoimmune diabetes
自身免疫性糖尿病“蜜月期”的多模态分析
  • 批准号:
    10443339
  • 财政年份:
    2022
  • 资助金额:
    $ 7.65万
  • 项目类别:
Analysis of diabetogenic human T cell receptors
致糖尿病的人类 T 细胞受体分析
  • 批准号:
    8311935
  • 财政年份:
    2011
  • 资助金额:
    $ 7.65万
  • 项目类别:
Development of novel diabetes autoantibody assays based on luciferase reporters
基于荧光素酶报告基因的新型糖尿病自身抗体测定的开发
  • 批准号:
    8075000
  • 财政年份:
    2010
  • 资助金额:
    $ 7.65万
  • 项目类别:
Active tolerosomes, a novel therapy for type 1 diabetes
活性耐受体,一种治疗 1 型糖尿病的新疗法
  • 批准号:
    7027696
  • 财政年份:
    2005
  • 资助金额:
    $ 7.65万
  • 项目类别:
Active tolerosomes, a novel therapy for type 1 diabetes
活性耐受体,一种治疗 1 型糖尿病的新疗法
  • 批准号:
    6925814
  • 财政年份:
    2005
  • 资助金额:
    $ 7.65万
  • 项目类别:
Cloning of molecular targets of CMI in type 1 diabetes
1型糖尿病CMI分子靶点的克隆
  • 批准号:
    8038522
  • 财政年份:
    1996
  • 资助金额:
    $ 7.65万
  • 项目类别:
Cloning of molecular targets of Cell Mediated Autoimmunity in Type 1 Diabetes
1 型糖尿病细胞介导的自身免疫分子靶标的克隆
  • 批准号:
    8636443
  • 财政年份:
    1996
  • 资助金额:
    $ 7.65万
  • 项目类别:
Cloning of molecular targets of Cell Mediated Autoimmunity in Type 1 Diabetes
1 型糖尿病细胞介导的自身免疫分子靶标的克隆
  • 批准号:
    8448325
  • 财政年份:
    1996
  • 资助金额:
    $ 7.65万
  • 项目类别:
Cloning of molecular targets of Cell Mediated Autoimmunity in Type 1 Diabetes
1 型糖尿病细胞介导的自身免疫分子靶标的克隆
  • 批准号:
    8039796
  • 财政年份:
    1996
  • 资助金额:
    $ 7.65万
  • 项目类别:

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X-ray Structural Studies of Antigen-Antibody Complex Toward Malaria Vaccine Development.
用于疟疾疫苗开发的抗原抗体复合物的 X 射线结构研究。
  • 批准号:
    01044086
  • 财政年份:
    1989
  • 资助金额:
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  • 项目类别:
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