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Measuring signaling pathway dynamics during tissue growth in hydrogels

测量水凝胶中组织生长过程中的信号通路动态

基本信息

批准号：
8879162
负责人：
JACQUELINE SARA JERUSS
金额：
$ 33.28万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-09-10 至 2017-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8879162
关键词：
3-Dimensional Acinus organ component Adhesions Adhesives Binding Sites Biocompatible Materials Biological Assay Biology Breast Cancer Cell Breast Epithelial Cells Cell Communication Cell Culture Techniques Cell Line Cells Choristoma Clinical Trials Dependence Development Diagnostic Disease Progression Drug effect disorder Environment Epidermal Growth Factor Receptor Epithelial Estrogen receptor negative Estrogen receptor positive Extracellular Matrix Factor Analysis Fibroblasts Genetic Genomics Goals Growth Growth Factor Health Hydrogels Image In Vitro Intervention Lead Life Ligands Luciferases MCF10A cells Malignant Neoplasms Mammary gland Measures Mechanics Messenger RNA Methods Milk Modeling Noninfiltrating Intraductal Carcinoma Normal tissue morphology Oncogenes Pathway interactions Patients Pertuzumab Pharmaceutical Preparations Pharmacologic Substance Pharmacotherapy Phenotype Pluripotent Stem Cells Post-Translational Protein Processing Production Property Proteins Proteomics Reporter Reporter Genes Reporting Research Resistance Resistance development Role Signal Pathway Signal Transduction Structure System Techniques Technology Therapeutic Time Tissues Trastuzumab Tumor Biology anticancer research base bioluminescence imaging cancer initiation cell growth cellular development clinically relevant cytokine design in vitro Model in vivo lapatinib malignant breast neoplasm metaplastic cell transformation new technology novel overexpression patient population prevent response scaffold therapy resistant transcription factor tumor progression vector

项目摘要

DESCRIPTION (provided by applicant): Cancer initiation and metastatic progression are widely modeled in vitro, and biomaterials applied to these in vitro models can recapitulate the phenotypes that are observed in vivo. Our long-term goal is developing 3D systems for directed cell growth, and these studies are aimed at identifying pathways to target to prevent abnormal growth (e.g., cancer) or to promote the development of functional tissue replacements. We propose a cell array that reports on the large-scale dynamic activity of transcription factors (TFs for cells cultured in a hydrogel in order to investigate the relationship between oncogenes, material design (e.g., adhesion and degradation), and the active TFs, and the ultimate phenotype of the cells. Normal mammary epithelial cells (MECs) form round acini capable of milk production in permissive environments, whereas the combination of aberrant oncogene activity (e.g., ErbB2) and the presence of specific extracellular matrix environment can produce pre- invasive or invasive phenotypes. ErbB2 is overexpressed in 15-20% of patients with invasive breast cancer, and in 50% estrogen receptor (ER) negative and 12% of ER positive cases of ductal carcinoma in situ. The proposed studies focus on TFs that drive the transition from a normal to a pre-invasive phenotype (Aim 1), and subsequently from a pre-invasive phenotype to an invasive phenotype (Aim 2). TFs are key regulators of cell phenotype, as evidenced by their ability to generate pluripotent stem cells from fibroblasts, and they are the downstream targets of signaling pathways, which are the target of many pharmaceuticals. The quantification of TF activity is based on the parallel delivery of TF reporter constructs within an array, which is combined with bioluminescence imaging for large scale, dynamic quantification. Analysis of TF activity is distinct from the current genomics and proteomics approaches that quantify mRNA or proteins abundance respectively. These reporter constructs have typically been applied to few pathways at early time points, and our technology allows tracking of TF activity throughout development of normal and abnormal structures for cells cultured within hydrogels. We employ designer hydrogels to regulate adhesion and degradation in order to investigate the established dependence of phenotypic transitions on the matrix. In Aim 3, we investigate multiple drug therapies that target EGFR signaling, such as trastuzumab and lapatinib. While these compounds are being investigated clinically, their mechanism of action and the target patient population are unknown. Furthermore, many patients develop resistance to these compounds, and the dynamic analysis may identify compensatory pathways. The cellular response to therapeutics is dependent, in part, upon the microenvironmental context, and these studies are expected to identify conditions under which a therapeutic response is obtained. Taken together, we hypothesize that the dynamic TF activity will i) identify pathway signatures that correlate with normal and abnormal tissue growth, ii) provide fundamental design principles that relate the hydrogel design to active signaling pathways, and iii) identify mechanisms of drug action that may inform clinical trials.

描述(申请人提供)：癌症的起始和转移进展在体外被广泛模拟，应用于这些体外模型的生物材料可以概括体内观察到的表型。我们的长期目标是开发定向细胞生长的3D系统，这些研究旨在确定靶向的途径，以防止异常生长(例如癌症)或促进功能组织替代品的开发。为了研究癌基因、材料设计(如黏附和降解)、活性转录因子(TF)和细胞的最终表型之间的关系，我们提出了一种细胞阵列，它报告了在水凝胶中培养的细胞的大规模动态转录因子(TF)的活性。正常乳腺上皮细胞(MEC)在允许的环境中形成能够产奶的圆形腺泡，而癌基因异常活性(如ErbB2)和特定细胞外基质环境的组合可产生侵袭前或侵袭表型。ERBB2在15-20%的浸润性乳腺癌、50%的雌激素受体(ER)阴性和12%ER阳性的导管原位癌中过表达。建议的研究集中于驱动从正常到侵袭前表型(AIM 1)，以及随后从侵袭前表型到侵袭表型(AIM 2)的转归的转铁蛋白。转录因子是细胞表型的关键调节因子，从它们从成纤维细胞产生多能干细胞的能力中得到了证明，它们是信号通路的下游靶标，而信号通路是许多药物的靶标。TF活动的量化是基于在阵列，与生物发光成像相结合，进行大规模、动态定量。对转铁蛋白活性的分析不同于目前分别量化mRNA或蛋白质丰度的基因组学和蛋白质组学方法。这些报告结构通常在早期应用于少数途径，我们的技术允许在水凝胶中培养的细胞的正常和异常结构的整个发育过程中跟踪TF活性。我们使用设计者水凝胶来调节黏附和降解，以研究表型转变对基质的既定依赖性。在目标3中，我们研究了多种针对EGFR信号转导的药物疗法，如曲妥珠单抗和拉帕替尼。虽然这些化合物正在进行临床研究，但它们的作用机制和目标患者群体尚不清楚。此外，许多患者对这些化合物产生抗药性，动态分析可能确定代偿途径。细胞对治疗学的反应在一定程度上取决于微环境，这些研究有望确定获得治疗反应的条件。综上所述，我们假设动态转铁蛋白活性将：1)识别与正常和异常组织生长相关的通路信号；2)提供将水凝胶设计与主动信号通路联系起来的基本设计原则；3)识别可能为临床试验提供信息的药物作用机制。