High-throughput, Multiplexed Detection of miRNA Biomarkers in Single Cancer Cells

单癌细胞中 miRNA 生物标志物的高通量多重检测

• 批准号: 8726351
• 负责人: Patrick S Doyle
• 金额: $ 22.11万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8726351
• 关键词: Address; Advanced Development; Area; Basic Science; Biological Assay; Biological Markers; Biopsy; Buffers; Cancer Biology; Cell Line; Cells; Cellular biology; Chromosomes; Clinical; Collaborations; Complex; Cytolysis; DNA; DNA Sequence; Data Set; Detection; Development; Diagnosis; Differential Diagnosis; Evaluation; Functional RNA; Gene Expression; Gene Expression Regulation; Genome; Goals; Heterogeneity; Human; Immune; Ligation; Malignant Neoplasms; Measures; Messenger RNA; Metric; MicroRNAs; Microfluidics; Modeling; Molecular Profiling; Motivation; Nature; Neurosurgeon; Patients; Performance; Phase; Play; Primary Neoplasm; Process; Protein Analysis; Public Health; Regulator Genes; Research; Reverse Transcriptase Polymerase Chain Reaction; Role; Scheme; Specimen; Stratification; System; Technology; Time; Validation; Variant; anticancer research; base; cancer cell; cancer diagnosis; cancer genetics; clinical Diagnosis; commercialization; cost; density; design; innovative technologies; microchip; nanolitre; neoplastic cell; novel; public health relevance; research study; single cell proteins; stem cell differentiation; success; technology development; tool; tumor

DESCRIPTION (provided by applicant): MicroRNAs (miRNAs) are a class of small non-coding RNAs recently discovered as negative gene regulators. A large faction of miRNAs are located within the fragile regions of chromosomes, which are areas of the genome more tightly associated with human cancers, indicating the possible role of miRNAs in tumor development. It has been shown that miRNA expression profiles correlate with various cancers and the miRNA signatures are potentially unique cancer biomarkers to accurately diagnose and classify human cancers. However, human cancers are highly heterogeneous due in part to the high degree of intratumoral heterogeneity at the single-cell level, requiring the evaluation of a large number of single tumor cells, each of which the miRNA expression profile can be measured. Here we propose to combine a single-cell barcode chip platform and a novel ligation-based miRNA detection scheme to develop an innovative technology that for the first time offers both high-throughput and multiplexing for rapid miRNA detection in single cells without amplification and at low cost. More specifically, we will: (1) Design and validate a DNA barcode microarray for multiplexed detection of miRNA biomarkers without amplification, and (2) Integrate this barcode array with a nanoliter microfluidics chip to perform multiplexed detection of 12 miRNA biomarkers from a large number of single cancer cells. Our technology is a versatile platform to detect miRNA biomarkers implicated in a variety of human cancers. It can be further modified to detect other non-encoding RNAs and messenger RNA biomarkers from single cells. Moreover, the proposed microchip is inexpensive, scalable and easy to manufacture. It may find immediate applications in basic research and clinical stratification of human cancers.

描述（由申请人提供）：microRNA（miRNA）是最近被发现为阴性基因调节剂的一类小型非编码RNA。 miRNA的一个大派别位于染色体的脆弱区域内，这些区域是与人类癌症更紧密相关的基因组的区域，表明miRNA在肿瘤发育中的作用可能作用。已经表明，miRNA的表达谱与各种癌症相关，而miRNA特征是潜在独特的癌症生物标志物，可以准确诊断和分类人类癌症。然而，人类癌的高度异质性是由于单细胞水平高度高度的肿瘤内异质性，需要评估大量的单肿瘤细胞，每个肿瘤细胞都可以测量miRNA表达谱。在这里，我们建议将单细胞条形码芯片平台和一种新型的基于连接的miRNA检测方案结合起来，以开发一种创新的技术，该技术首次提供高通量和多路复用，用于在没有扩增和低成本的单个单元中快速miRNA检测。更具体地说，我们将：（1）设计和验证DNA条形码微阵列，用于对miRNA生物标志物的多路复用检测，而不会放大，并且（2）将此条形码阵列与纳米级微流体芯片相结合，以执行来自大量单个癌细胞的12 miRNA生物标志物的多路复用检测。我们的技术是一个多功能平台，可检测与各种人类癌症有关的miRNA生物标志物。可以进一步修改以检测单个细胞的其他非编码RNA和Messenger RNA生物标志物。此外，拟议的微芯片是便宜，可扩展且易于制造的。它可能会在人类癌症的基础研究和临床分层中立即应用。