Aptamer-Based Detection of Cardiac Biomarker Glycosylation States Using APT-SNAP

使用 APT-SNAP 基于适体的心脏生物标志物糖基化状态检测

• 批准号: 8648358
• 负责人: Mary Szatkowski Ozers
• 金额: $ 31.74万
• 依托单位: PROTEOVISTA, LLC
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2017-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8648358
• 关键词: Address; Adopted; Affinity; Antibodies; Atherosclerosis; Biological Assay; Biological Markers; Brain natriuretic peptide; C-reactive protein; Cardiac; Cardiovascular Diseases; Cardiovascular system; Chronic Kidney Failure; Clinical; DNA; Detection; Devices; Diagnosis; Diagnostic; Diagnostic tests; Disease; Dyes; Fluorescence; Fluorescent Antibody Technique; Glass; Goals; Heart Diseases; Heart failure; Human; Longevity; Marketing; Medicine; Methodology; Modeling; Monitor; N-terminal; Nucleic Acids; Outcome; Patient Care; Patients; Pattern; Peptide antibodies; Phase; Physicians; Post-Translational Protein Processing; Price; Protein Array; Protein Binding; Protein Microchips; Proteins; Proteomics; RNA; Research; Sampling; Serum; Signal Transduction; Slide; Small Business Innovation Research Grant; Specificity; Spottings; Staging; Structure; Technology; Testing; Troponin C; Validation; aerobic respiration control protein; aptamer; base; cost effective; density; design; glycosylation; improved; insight; novel; point of care; point-of-care diagnostics; prohormone; protein function; prototype; public health relevance; sensor; three dimensional structure

DESCRIPTION (provided by applicant): Aptamer-Based Detection of Cardiac Biomarker Glycosylation States Using APT-SNAP Arrays PIs: Christopher L. Warren and Mary S. Ozers The presence of cardiac-related biomarkers in patient biospecimens can provide important insight into diagnosis of cardiovascular disease, its progression, and optimal therapies for treatment. Most of the cardiac biomarkers in clinical use detect heart failure, and the cardiovascular field is in great need of biomarkers that span the cardiovascular disease spectrum from developing atherosclerosis to late-stage disease. The clinically-accepted biomarker for heart failure, BNP, is modified by glycosylation, and the glycosylated form of BNP predominates in patients with heart failure and chronic renal failure. However, the leading diagnostic assay for BNP does not detect the glycosylated forms. Furthermore, robust technologies to accurately detect multiple biomarkers, especially glycosylated or other modified forms of biomarkers, are lacking. To address this significant diagnostic need for heart disease patients, we are developing the APT-SNAP (Aptamer Specificity and Affinity for Proteins) microarray, which displays millions of DNA and RNA aptamers, as a prototype high throughput device to design, discover, and optimize high affinity aptamers for heart disease biomarkers. Nucleic acid aptamers are composed of either DNA or RNA and can adopt a compact three-dimensional structure that recognizes a target with exceptional specificity and high affinity, making them useful as a sensor of cardiac biomarkers. In this proposal, aptamers will be identified on the APT-SNAP array for six major cardiac biomarkers as proof of principle. A key invention of the proposal is a novel methodology to synthesize high density RNA aptamers arrays. Identified aptamers for the cardiac biomarkers will be tested for their limit of detection n human serum. The APT-SNAP array will identify aptamers that recognize distinct glycosylation structures at specific residues of a biomarker, using NT-proBNP as a model biomarker. To accomplish this, we will synthesize a set of NT-proBNP proteins, each with a distinct glycosylation pattern, and iteratively design aptamers that specifically recognize the glycosylated BNP forms. To expand the universality of the assay beyond the limitations of antibodies currently used in many clinical assays, a fluorescent protein dye will be used to correlate array fluorescence intensity to biomarker concentration. The Specific Aims of this glycomics/proteomics-based proposal are: 1. Design novel high density DNA and RNA aptamer microarrays to identify aptamers for cardiac biomarkers. 2. Develop a panel of aptamers to detect specific glycosylated forms of cardiac biomarkers. The ultimate goals are to develop APT-SNAP as a biomarker discovery platform (Phase I), clinically correlate additional biomarkers (Phase II), and develop an affordable annual point-of-care diagnostic test (Phase IIB) for hundreds of cardiac biomarkers for physician use. These technologies will impact the ability to diagnose heart disease accurately, allowing for better treatment options, improved patient care, and longer lifespans.

描述（由申请人提供）：使用APT-SNAP阵列PIS的基于APTAMER的心脏生物标志物糖基化状态的检测：Christopher L. Warren和Mary S. Ozers在患者生物呼吸器中存在与心脏相关的生物标志物的存在，可以为心血管疾病的诊断提供重要的见解，以提供重要的洞察力。临床使用中的大多数心脏生物标志物检测心力衰竭，心血管局部非常需要生物标志物，这些生物标志物跨越了心血管疾病谱系，从发生动脉粥样硬化到晚期疾病。心力衰竭的临床认可的生物标志物通过糖基化修饰，BNP的糖基化形式在心力衰竭和慢性肾衰竭的患者中占主导地位。但是，BNP的主要诊断测定不能检测到糖基化形式。此外，缺乏可准确检测多种生物标志物，尤其是糖基化或其他修饰形式的生物标志物的强大技术。为了满足对心脏病患者的这种重大诊断需求，我们正在开发适当的SNAP（适体特异性和对蛋白质的亲和力）微阵列，该微阵列显示了数百万个DNA和RNA适体，作为一种原型高吞吐量，以设计，发现，发现，发现和优化心脏病生物标志物的高亲和力适应性。核酸适体由DNA或RNA组成，并且可以采用紧凑的三维结构，该结构识别具有特异性特异性和高亲和力的靶标，使其可作为心脏生物标志物的传感器。在此提案中，将在APT-SNAP阵列上为六个主要心脏生物标志物的APTAMER确定，以证明原理。该提案的关键发明是合成高密度RNA适体阵列的新方法。确定的心脏生物标志物的适体将测试其检测极限n人血清。 APT-SNAP阵列将使用NT-ProBNP作为模型生物标志物，识别在生物标志物的特定残基上识别不同糖基化结构的适体。为此，我们将合成一组NT-proBNP蛋白，每个蛋白具有不同的糖基化模式，并迭代设计适合剂，这些适应性特征专门识别糖基化的BNP形式。为了扩展分析的普遍性，超出了许多临床测定中当前使用的抗体的局限性，将使用荧光蛋白染料将阵列荧光强度与生物标志物浓度相关联。基于糖基因组学的建议的具体目的是：1。设计新型的高密度DNA和RNA适体微阵列，以识别心脏生物标志物的适体。 2。开发一个适体面板，以检测特定的心脏生物标志物的糖基化形式。最终的目标是开发适当的SNAP作为生物标志物发现平台（I期），在临床上相关联其他生物标志物（II阶段），并为数百个心脏生物标志物用于医师使用的价格合理的年度护理诊断测试（IIB期）。这些技术将影响准确诊断心脏病的能力，允许更好的治疗选择，改善患者护理和更长的寿命。