Quantifying proteins in plasma do democratize personalized medicine for patients with type 1 diabetes

摘要 1型糖尿病在美国影响着超过125万人，每年 发病率正在以令人震惊的3-4%的速度增长。美国人的情感和经济负担 疾病是压倒性的，我们目前无法预测或预防新病例。因为我们 更好地了解胰腺和胰腺的病理生理过程 高血糖的下游影响(和治疗期间的周期性低血糖)，更多 需要强大的生物标记物分析来提高研究结果的重复性，并 将这些发现转化为临床护理。一种技术，可以提供强大的、可转移的 测定蛋白质的分析方法是液-质联用法。 通过直接检测感兴趣的分析物，使用质谱学检测的分析可以 比免疫分析有更好的特异性，当与浓缩策略配合使用时，它们 也可能非常敏感。正如我们之前所演示的那样，很简单 协调质谱分析的结果，这对 一般是免疫分析。该提案旨在生成和验证可转让的小说 利用质谱学的力量进行蛋白质分析。我们的目标是利用一种新方法 用于浓缩胞外小泡和新的从头蛋白以进行亲和浓缩， 被称为迷你粘合剂，以帮助提高方法的敏感性。只要有可能，化验将是 如果浓缩需要抗体，它们将通过以下途径广泛分布 爱荷华州杂交会银行。编码迷你粘合剂的质粒将存放在Addgene。 来自方法开发的层析数据(特别是肽选择，它将使用 窄窗口数据独立采集，而不是依赖算法或数据- 不同的获取方法)以及来自方法验证的层析数据 通过Panorama分发，以及详细的标准操作程序。按照中的要求 RFA DK-21-031，产生的部分分析将针对胰高血糖素，其他片段 胰高血糖素原、胰岛素原及其片段、糖化可溶性CD59、胰淀素和 嗜铬颗粒。我们的目标优先排序委员会将帮助确定最重要的 将蛋白质添加到这个列表中，并集中我们的开发努力。

ABSTRACT Type 1 diabetes affects more than 1.25 million people in the United States and the annual incidence is increasing at an alarming rate of 3-4%. The emotional and financial burden of the disease is overwhelming and we currently have no way to predict or prevent new cases. As we gain a better understanding of the pathophysiological processes in the pancreas and the downstream effects of hyperglycemia (and periodic hypoglycemia during treatment), more robust biomarker assays are needed to improve the reproducibility of research findings and to translate those findings to clinical care. One technology that can provide robust, transferable assays for the measurement of proteins is liquid chromatography-tandem mass spectrometry. By directly detecting the analyte of interest, assays that use mass spectrometry detection can have better specificity than immunoassays and when paired with enrichment strategies, they can also be very sensitive. As we have demonstrated previously, it is straightforward to harmonize the results of mass spectrometric assays, which is significantly more difficult for immunoassays in general. This proposal aims to generate and validate novel transferable protein assays that harness the power of mass spectrometry. We aim to leverage a new method for the enrichment of extracellular vesicles and new de novo proteins for affinity enrichment, called minibinders, to help with sensitivity of the methods. Whenever possible, assays will be multiplexed and if antibodies are required for enrichment, they will be widely distributed through the Iowa Hybridoma Bank. Plasmids encoding minibinders will be deposited at Addgene. Chromatographic data from method development (particularly peptide selection, which will use narrow-window data-independent acquisition rather than relying on algorithms or data- dependent acquisition methods) as well as chromatographic data from method validation will be distributed via Panorama, along with detailed standard operating procedures. As requested in RFA DK-21-031, a portion of the assays produced will target glucagon, other fragments of proglucagon, proinsulin and its fragments, glycated soluble CD59, amylin, and the chromogranins. Our Target Prioritization Committee will help identify the most important proteins to add to this list and focus our development efforts.