Novel Metabolic Biomarker for Autism Spectrum Disorder

描述（由申请人提供）：自闭症谱系障碍（ASD）影响美国1/110的儿童，但医生还没有有效的实验室测试可以确认临床诊断。该提案旨在开发一种快速可靠的代谢阵列来识别ASD患者。 使用从Biolog（海沃德，CA）获得的表型微阵列板，我们已经能够表明，与40名对照相比，在淋巴母细胞中，在色氨酸作为唯一能量来源的情况下，54/56（96.4%）名ASD患者的还原型烟酰胺腺嘌呤二核苷酸（NADH）产量持续降低。 该项目的第一个目的是使用定制的表型微阵列板，使用另外50名患者和另外50名对照的队列来验证我们的初步发现。新的平板将允许我们每个平板测试12个个体，每个个体仅使用16万个细胞。除了初步研究中的96份样本外，拟定的100份样本在单侧t检验中的统计功效为95.6%，显著性水平为0.01。 第二个目的是使用我们的测定来评估新鲜血液样品中色氨酸存在下的NADH产生。作为第一步，我们计划使用我们已经测试过的6名患者和6名对照，以复制在新鲜的淋巴母细胞系中观察到的结果。 白色细胞。这第一阶段将使我们能够确定，我们是否能够观察到相同的低水平的NADH生产色氨酸的存在下，在白细胞中观察到的淋巴母细胞。接下来，我们将测试12名ASD患者和12名对照的新鲜血液样本。为了减少输出变异性，我们将尝试对所有样本同时开始测试，协调对24个个体的采样。最后，我们将以盲法测试来自新个体的24名患者和24名对照血液样本，以建立该测试在ASD患者中的预测能力。对于实验的最后一部分，我们将在每个样品到达实验室后立即进行测试。我们将在每个个体的定制板上使用一排8个威尔斯孔，用铝箔覆盖板的其余部分，以保留其他行用于将来的检测。 如果我们在淋巴母细胞中的初步发现在白细胞中得到证实，我们将能够开发一种可靠，易于执行，廉价的实验室测试，可以在大约4-5天内提供诊断。这样的测试可以代表ASD的第一个生化筛查测试。 公共卫生相关性：该项目的目标是开发自闭症谱系障碍（ASD）的筛查工具。在我们的初步研究中，我们检测到与对照组相比，ASD患者在色氨酸存在下产生的NADH水平显着降低。我们建议利用相同的代谢分析方法，使用新鲜血液样本中的白细胞来确定ASD是否可以快速可靠地与对照区分开来。

DESCRIPTION (provided by applicant): Autism Spectrum Disorders (ASDs) affect 1/110 children in the United States, but physicians do not yet have an effective laboratory test which can confirm the clinical diagnosis. This proposal aims to develop a quick and reliable metabolic array to identify individuals with ASD. Using Phenotype Microarray plates obtained from Biolog (Hayward, CA), we have been able to show that Nicotinamide Adenine Dinucleotide, reduced form (NADH) production in the presence of tryptophan as only energy source in lymphoblastoid cells was consistently reduced in 54/56 (96.4%) patients with ASD when compared to 40 controls. The first aim of the proposed project is to validate our preliminary findings using a cohort of 50 additional patients and 50 additional controls employing customized Phenotype Microarray plates. The new plates will allow us to test 12 individuals per plate, using just 160,000 cells per individual. The proposed 100 samples, in addition to the 96 samples from the preliminary study, will give statistical power of 95.6% in a one- sided t-test at significance leve of 0.01. The second aim is to use our assay to evaluate the NADH production in the presence of tryptophan in fresh blood samples. As a first step, we plan to use 6 patients and 6 controls that we have already tested to replicate the results observed in lymphoblastoid cell lines in fresh white cells. This first stage will allow us to determine if we are able to observe the same low levels of NADH production in the presence of tryptophan in leukocytes as was observed in lymphoblasts. Next, we will test fresh blood samples from 12 patients with ASD and 12 controls. To reduce output variability, we will try to start the test at the same time for all the samples, coordinating the sampling of the 24 individuals. Finally, we will test 24 patient and 24 control blood samples from new individuals in a blinded fashion to establish the predictive power of this test in patients with ASD. For this last part of the experiment, we will test each sample as soon as it arrives at our laboratory. We will use a single row of 8 wells on the customized plates for each individual, covering the rest of the plate with aluminum foil, to preserve the other rows for future tests. If our preliminary findings in lymphoblasts are confirmed in leukocytes, we would be able to develop a reliable, easy to perform, inexpensive laboratory test which could provide a diagnosis in about 4-5 days. Such a test could represent the first biochemical screening test for ASDs. PUBLIC HEALTH RELEVANCE: The goal of this project is to develop a screening tool for Autism Spectrum Disorders (ASDs). In our preliminary study, we detected significantly lower levels of NADH generated in the presence of tryptophan in patients with ASD when compared to controls. We propose to utilize the same metabolic profiling approach using leukocytes from fresh blood samples to determine if ASDs can be quickly and reliably distinguished from controls.