Use of Organotypic and Mammary Gland Models to Investigate the Outcomes of Clonal

使用器官型和乳腺模型研究克隆的结果

• 批准号: 7617421
• 负责人: Joan Siefert Brugge
• 金额: $ 38.44万
• 依托单位: WHITEHEAD INSTITUTE FOR BIOMEDICAL RES
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-02-01 至 2014-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7617421
• 关键词: Adherent Culture; Behavior; Biological; Biological Models; Biology; Breast; Carcinoma in Situ; Cell Proliferation; Cell model; Cells; Classification; Clone Cells; Development; Dissociation; Down-Regulation; Ductal; Epithelial Cells; Event; Experimental Models; Gene Activation; Gene Expression; Genes; Growth; Human; In Situ Lesion; In Vitro; Individual; Invaded; Laboratories; Lentivirus Vector; Mammary gland; Modeling; Mus; Mutation; Normal Cell; Oncogenes; Oncogenic; Organoids; Outcome; Pathogenesis; Pathway interactions; Play; Population; Process; Proliferating; Role; Structure; Structure of thyroid parafollicular cell; Study models; Tissues; breast tumorigenesis; cancer initiation; carcinogenesis; cell behavior; gene induction; genetic manipulation; in vitro Model; in vivo; insight; interest; malignant breast neoplasm; neoplastic cell; three dimensional structure; tumor; tumor initiation; vector

Our laboratory has utilized a three-dimensional culture model to investigate the phenotypic effects of genes implicated in breast tumorigenesis on the biology of mammary epithelial cells. This model system has revealed interesting cell-biological behaviors that are not detectable in standard monolayer cultures and has provided important mechanistic insights into processes and pathways that appear to play important roles in tumor initiation and progression in the mammary gland (e.g, filling of the lumen, escape from growth arrest, loss of polarity, etc). In the studies proposed in this application, we will further expand the application of these models for studies of tumor pathogenesis by examining the fate of single mammary epithelial cells that carry genetic alterations associated with breast cancer initiation or progression (e.g. expression of oncogenes or downregulation of tumor suppresors). Since tumors evolve from clonal alterations in isolated, individual cells, rather than globally in all cells within such a tissue, the proposed experimental models will more closely mimic the natural events associated with spontaneous tumor initiation and progression, and allow us to examine the influence of normal cells and the architectural organization of mammary structures on expression of phenotypic changes provoked by oncogenic insults. We will use the well-characterized MCF-10A immortalized human mammary epithelial cell model as well as other in vitro and in vivo mammary epithelial cell models that we are currently developing. The studies will involve systematic comparisons of the fate of cells subjected to distinct oncogenic insults in proliferating versus growth-arrested structures, in single cells versus the total cell population, and in immortalized versus primary cell three- dimensional structures. Findings from the in vitro models will be evaluated in vivo using inducible expression of cDNAs or shRNAs in single cells within the mammary gland. These studies promise to provide important information on (i) the fate of cell clones carrying oncogenic alterations, (ii) the role of epithelial cells within the microenvironment on the outcome of oncogene insults, (iii) the mechanisms of escape from suppressive influences of the microenvironment, and (iv) requirements for conferring a competitive advantage to tumor cells within tissue-like structures.

我们的实验室利用了三维培养模型来研究基因的表型效应 与乳腺肿瘤发生有关乳腺上皮细胞的生物学。该模型系统具有 揭示了在标准单层培养物中无法检测到的有趣的细胞生物学行为，并且具有 为过程和途径提供了重要的机械见解 乳腺的肿瘤启动和进展（例如，管腔填充，从生长停滞中逃脱， 失去极性等）。在本应用程序中提出的研究中，我们将进一步扩展 这些模型用于研究肿瘤发病机理，通过检查单乳腺上皮细胞的命运 携带与乳腺癌开始或进展相关的遗传改变（例如 肿瘤基因或肿瘤替代物的下调）。由于肿瘤是由于分离的克隆改变而演变的，所以 提出的实验模型将 更紧密地模仿与自发性肿瘤起始和进展相关的自然事件，以及 允许我们检查正常细胞和乳腺结构的结构组织的影响 关于表型变化的表达，由致癌性侮辱引起。我们将使用良好的特征 MCF-10A永生的人类乳腺上皮细胞模型以及其他体外和体内乳腺 我们目前正在开发的上皮细胞模型。研究将涉及系统比较 细胞的命运受到在增殖与生长和生长结构中的明显致癌侮辱 单细胞与总细胞群体，在永生中与原代细胞三维和原代细胞中 结构。从体外模型的发现将在体内使用cDNA或可诱导表达的体内评估 乳腺内单细胞中的shRNA。这些研究有望提供重要信息 （i）带有致癌改变的细胞克隆的命运，（ii）上皮细胞在 微环境关于癌基因侮辱的结果，（iii）逃脱的机制 微环境的影响以及（iv）赋予肿瘤竞争优势的要求 组织样结构内的细胞。