An integrated human organ-on-chip ultrasensitive miRNA detection platform for novel biomarker discovery

用于新型生物标志物发现的集成人体器官芯片超灵敏 miRNA 检测平台

• 批准号: 9769864
• 负责人: Pinar Zorlutuna
• 金额: $ 38.57万
• 依托单位: UNIVERSITY OF NOTRE DAME
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9769864
• 关键词: Address; Animal Model; Benchmarking; Biological Assay; Biological Markers; Biosensing Techniques; Biosensor; Blood Circulation; Blood specimen; Cardiovascular Diseases; Cell Culture Techniques; Cells; Clinical; Coronary Arteriosclerosis; Culture Media; Detection; Development; Devices; Diagnostic; Disease; Disease model; Distress; Event; Fingerprint; Glucose; Goals; Health; Heart; Hematological Disease; Histologic; Human; Human Engineering; Hypoxia; Individual; Ischemia; Life; Ligation; Link; Liquid substance; Malignant Neoplasms; Methods; MicroRNAs; Microfluidic Microchips; Microfluidics; Modeling; Monitor; Myocardial; Myocardial Infarction; Myocardial tissue; Myocardium; Oxidative Stress; Patients; Performance; Physiological; Preparation; Property; Protocols documentation; RNA; Reperfusion Injury; Reperfusion Therapy; Role; Sampling; System; Techniques; Technology; Testing; Time; Tissue Engineering; Tissue Model; Tissue Sample; Tissues; Validation; Ventricular; base; biomarker development; biomarker discovery; circulating microRNA; clinical application; clinical translation; clinically relevant; conditioning; deep sequencing; disease phenotype; drug discovery; exosome; experimental study; human model; human subject; human tissue; induced pluripotent stem cell; innovation; ischemic conditioning; microRNA biomarkers; novel; novel marker; novel strategies; organ on a chip; percutaneous coronary intervention; personalized medicine; potential biomarker; predicting response; prognostic; real time monitoring; sensor; specific biomarkers; tool

Abstract Circulating miRNAs have proposed as specific biomarkers of disease states, including some of the most prevailing ones such as cardiovascular diseases and cancer. However using miRNAs as biomarkers is very challenging despite recent advances in high-throughput miRNA profiling. Various detection technologies, protocols, ligation and extraction/purification methods have led to varying miRNA profiling results of cells and biofluids under different conditions. Most importantly, all require days long sample-to-answer assay times, thus ruling them out for detection and monitoring of urgent, life threatening conditions such as myocardial infarction (MI). A rapid real-time, PCR-free miRNA-profiling device would be exceedingly valuable for precision, personalized medicine in years to come. However, it is very difficult to start even developing such a platform because of the limitations in testing models. Animal models often fail to predict responses in humans; and studies of human subjects do not readily allow for precise control over the disease events or temporal correlation of the disease state and biomarker expression dynamics. To address this challenge, in this study, we will develop an organ-on-a-chip device with an integrated attomolar (aM)-level miRNA sensing capability, which we will use for optimizing real-time monitoring of fluctuations in multiple miRNAs for novel biomarker discovery. As an immediate application, we will start with a human myocardium-on-chip (MoC) as a clinically relevant model and imitate the course of a heart attack. We hypothesize that using the MoC with ultrasensitive miRNA detection, we will discover a unique signature that indicates the onset of reperfusion injury during MI treatment. Finally, we will test the sensor device and the miRNA signature using clinical blood samples. Our microfluidic organ-on-a-chip platform will consist of four basic components: 1) the tissue engineered human MoC from human induced pluripotent stem cells (hiPSCs), 2) the exosome lysing unit, 3) the concentration unit for the lysed RNAs and 4) the detection unit for the miRNAs. In Aim 1, we will couple these components into a fully integrated microfluidic platform. First we will validate the clinical relevance of the MoC model by comparing with human tissue and blood samples. Then we will characterize and optimize the performance of a novel miRNA detection biosensor using MoC and benchmark it against established miRNA analysis techniques. In Aim 2 we will focus on multiplexing the sensing approach for the real-time detection of a panel of miRNAs, and 1) use the MoC to discover a miRNA signature to be used as a novel biomarker that captures the RI onset, as well as 2) to optimize the multiplexed sensor for faster clinical translation. In Aim 3 we will determine the diagnostic and prognostic capabilities of the novel biosensor and miRNA biomarker signature we developed in Aims 1 and 2 using the MoC model, with clinical samples from MI patients. Our long-term goal is to utilize this integrated platform to study exosomes and their RNA content to advance current understanding of their role in human health and to determine their potential as biomarkers for disease states.

摘要