Microfluidic Techniques for the Molecular and Functional Analysis of Gene Express

用于基因表达分子和功能分析的微流控技术

• 批准号: 7662667
• 负责人: STEPHEN R QUAKE
• 金额: $ 44.58万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7662667
• 关键词: Antibodies; Area; Automation; Behavior; Bioinformatics; Biological; Biological Assay; Blood Vessels; CD47 gene; Cell Count; Cell Culture Techniques; Cell model; Cells; Chromatin; Chromatin Structure; Chromatin Structure Alteration; Complement; Culture Media; DNA; Data; Data Analyses; Devices; Diagnosis; Disease; Environment; Epigenetic Process; Fibroblasts; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genomics; Growth; Growth Factor; Heterogeneity; Histones; Human; Human Resources; In Vitro; Individual; Investigation; Laboratories; Ligands; Malignant Neoplasms; Measurement; Mesenchymal; Methylation; Microfluidic Microchips; Microfluidics; Miniaturization; Modification; Molecular; Mus; Normal tissue morphology; Outcome; Pathway Analysis; Pathway interactions; Patients; Play; Polycomb; Population; Post-Translational Protein Processing; Proliferating; Property; Proteins; Protocols documentation; Radiopharmaceuticals; Recruitment Activity; Regulation; Regulatory Pathway; Resistance; Role; Schedule; Screening procedure; Sequence Analysis; Signal Transduction Pathway; Solid; Solid Neoplasm; Stem cells; Stratification; Stromal Cells; Support System; Supporting Cell; System; Techniques; Technology; Testing; Therapeutic; Therapeutic Agents; Time; Video Microscopy; Xenograft procedure; base; cancer cell; cancer stem cell; cancer therapy; cell behavior; cell growth; cell motility; chromatin immunoprecipitation; design; high throughput screening; in vitro Assay; in vitro testing; in vivo; insight; leukemia; malignant breast neoplasm; matrigel; neoplastic cell; new therapeutic target; research study; response; self-renewal; stem cell population; therapeutic target; time use; tool; transcription factor; tumor; tumor growth; tumorigenic

Cancer stem cells in solid tumors have been recently isolated and appear to be primarily responsible for the growth and spread of the disease. The presence of a stem cell population in a tumor has implications for the diagnosis and treatment of cancer, as it is these cancer stem cells that must be targeted to achieve a cure. Preliminary evidence demonstrates that there is a host of genes differentially expressed by the cancer stem cells and their non-tumorigenic progeny. Many of these genes are thought to play a role in essential cancer functions including proliferation, survival, self renewal and resistance to standard therapeutics. Uncovering the true functional stratification of the superficially uniform population of stem cells in leukemia, breast cancer and other solid cancers is a challenge which requires new kinds of measurements at the single-cell level. Based on recent discoveries by the Clarke and Weissman groups, we believe that gene expression in normal stem cells, as well as cancer stem cells is significantly regulated at the epigenetic level. Epigenetic regulation of gene expression is in part controlled by the posttranslational modification of histone proteins, as well as methylation of DMA, both of which result in the alteration of chromatin structure. These modifications are examined using Chromatin immunoprecipitation (ChIP) assays. Epigenetic analysis is a necessary complement to gene expression analysis in order to understand the control of normal and cancer stem cell self-renewal and discover new therapeutic targets. The microfluidic tools developed by the Quake lab for single cell gene expression (aim1), chip (aim 2) and high throughput in vitro cell culture (aim 3) will allow the Clarke and Weissman laboratories to perform gene expression and epigenetic analyses on rare, purified cells from model mice and primary human cancer or cancer xenografts. These assays will all use microfluidic platforms which have already been developed and validated in the Quake laboratory or are commercially available. New systems will also be designed as needed in the course of this project. Microfluidic epigenetic and genetic assays will allow the study of highly purified, homogeneous, rare cell populations that were previously inaccessible with the standard techniques. Microfluidic cell culture platforms will allow finding appropriate conditions to culture cancer stem cells in vitro and test new therapeutic targets

实体瘤中的癌症干细胞最近已经被分离出来，并且似乎主要负责肿瘤的生长。 疾病的生长和传播。肿瘤中干细胞群的存在对肿瘤的发生有影响。 癌症的诊断和治疗，因为它是这些癌症干细胞，必须有针对性地实现治愈。 初步的证据表明，有一个主机的基因差异表达的癌干 细胞及其非致瘤后代。这些基因中的许多被认为在原发性癌症中发挥作用 功能包括增殖、存活、自我更新和对标准治疗剂的抗性。揭开 在白血病、乳腺癌、乳腺癌、 和其他实体癌症是一项挑战，需要在单细胞水平上进行新型测量。 根据Clarke和Weissman小组最近的发现，我们认为， 正常干细胞以及癌症干细胞在表观遗传水平上受到显著调节。后生 基因表达的调节部分由组蛋白的翻译后修饰控制，如 以及DNA甲基化，这两者都导致染色质结构的改变。这些修改 使用染色质免疫沉淀（ChIP）测定来检查。表观遗传分析是一个必要的 补充基因表达分析，以了解正常和癌症干细胞的控制 自我更新和发现新的治疗靶点。地震实验室开发的微流体工具， 单细胞基因表达（aim 1）、芯片（aim 2）和高通量体外细胞培养（aim 3）将允许 克拉克和韦斯曼实验室进行基因表达和表观遗传学分析的稀有，纯化 来自模型小鼠和原发性人癌症或癌症异种移植物的细胞。这些分析都将使用微流体 已经在Quake实验室开发和验证的平台或商业平台 available.在本项目实施过程中，还将根据需要设计新的系统。微流控表观遗传 基因分析将允许研究高度纯化的，同质的，稀有的细胞群， 这是以前用标准技术无法达到的。微流体细胞培养平台将允许发现 体外培养癌症干细胞和测试新治疗靶点的合适条件