Imaging the native 3D architecture of pancreatic and breast tumor patient tissue at single-cell resolution

以单细胞分辨率对胰腺和乳腺肿瘤患者组织的天然 3D 结构进行成像

• 批准号: 10461140
• 负责人: Tao Ju
• 金额: $ 36.03万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-03 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10461140
• 关键词: 3-Dimensional; Address; Adult; Architecture; Atlases; Breast; Breast Cancer Patient; CD8B1 gene; CXCL14 gene; Cancerous; Cell Nucleus; Cells; Cessation of life; Collagen; Complex; Cystic Fibrosis Transmembrane Conductance Regulator; DNA; Data; Data Set; Detection; Diagnosis; Extracellular Matrix; FBXW7 gene; Fluorescence Microscopy; Genomics; Growth; Human; Image; Imaging Techniques; Immune; Immunofluorescence Immunologic; In Situ Hybridization; Inter-tumoral heterogeneity; Label; Lymphatic; Malignant Neoplasms; Malignant neoplasm of pancreas; Maps; Methods; Microscopy; Molecular; Neoplasm Metastasis; Optics; PECAM1 gene; Pancreas; Pancreatic Ductal Adenocarcinoma; Patients; Population; Proteomics; Quality of life; RNA; Reporter; Research; Resolution; Route; Sampling; Solid; Solid Neoplasm; Spatial Distribution; Specimen; Speed; Stains; Stromal Cells; Techniques; Technology; Thinness; Three-Dimensional Imaging; Tissue Sample; Tissues; Transcript; Tumor Tissue; United States; Universities; Vimentin; Visualization; Washington; Woman; Work; analysis pipeline; breast malignancies; cell type; cellular imaging; computational pipelines; computational suite; computerized tools; dimensional analysis; experimental study; human tissue; imaging approach; improved; innovation; insight; lymphatic vasculature; men; molecular imaging; mortality; neoplastic cell; pancreatic ductal adenocarcinoma cell; pancreatic neoplasm; protein biomarkers; protein expression; reconstruction; single cell sequencing; spatial relationship; therapeutic target; therapy resistant; triple-negative invasive breast carcinoma; tumor; tumor heterogeneity; tumor microenvironment; two-dimensional

PROJECT SUMMARY Cancer remains the second-leading cause of adult death in the United States, yet the mechanisms by which cancerous growths are initiated and how crucial transitions such as metastasis and therapeutic resistance occur are not well understood. Significant progress has been made in the multiplexed analysis of solid tumors through the use of single-cell sequencing and spatio-molecular mapping techniques. However, despite the unique insights into tumor heterogeneity these methods have afforded, they are limited to two-dimensional (2D) thin analyses which provide little information on the native three-dimensional architecture of the tumor. Serial reconstruction can provide some three-dimensional context, but such approaches are both inefficient in terms of sample throughput and are inherently destructive to the tissue architecture. We propose a cross-disciplinary, approach to quantitatively characterize the native three-dimensional architecture of human solid tumor tissue from triple negative breast cancer (TNBC) and pancreatic ductal adenocarcinoma (PDAC) patients using a combination of the state-of-the art, yet mature technologies of tissue clearing, immunofluorescence and in situ hybridization labeling, and high-resolution lightsheet fluorescence microscopy. Analysis of these tissue volumes will enable the elucidation of the spatial interactome of different tumor, immune and stromal cells that give rise to tumor heterogeneity as well as their interactions with components of the tumor microenvironment. Our Specific Aims are (1) to image the three-dimensional spatial distribution of tumor, immune and stromal cells in relation to vasculature, lymphatics and the extracellular matrix in native solid tumor tissue from human pancreatic and breast malignancies using a combination of tissue clearing, immunofluorescence, in situ hybridization and 3D lightsheet microscopy, and (2) to develop a computational pipeline to build three-dimensional spatial maps of protein expression and RNA transcript localization in intact solid tumor tissue from human pancreatic and breast malignancies. The innovation of the proposed work lies in our cross-disciplinary strategy of combining the cutting- edge tissue clearing, multiplexed labelling and high-resolution lightsheet microscopy to better understand the native three-dimensional architecture of solid tumors. Specifically, by classifying and quantitating previously unknown three-dimensional features of solid tumor tissue we will be able to relate the spatial organization of the tumor to genomic and proteomic data taken from the same specimen. The significance of this proposal is that successful three-dimensional characterization of human tumor tissues will enable key insights to be derived on both intra- and inter-tumor heterogeneity thus facilitating the identification of cell-cell and cell-microenvironment interactions that could serve as potential therapeutic targets thus providing new routes to treatments to decrease patient mortality and improve quality of life.

项目摘要 癌症仍然是美国成人死亡的第二个领先原因，但其机制是 开始癌性生长，以及如何发生转移和治疗性抗性等关键过渡 不太了解。通过对实体瘤的多路复用分析取得了重大进展 单细胞测序和空间分子映射技术的使用。但是，尽管有独特的 对肿瘤异质性的见解，这些方法提供了，它们仅限于二维（2D）薄 分析很少提供有关肿瘤本地三维结构的信息。系列 重建可以提供一些三维环境，但是这种方法在 样品吞吐量，对组织结构具有固有的破坏性。我们建议一个跨学科， 定量表征人类实体瘤组织的天然三维结构的方法 使用A的三重阴性乳腺癌（TNBC）和胰腺导管腺癌（PDAC）患者 最先进的，但成熟的组织清除，免疫荧光和原位的技术结合 杂交标记和高分辨率灯表荧光显微镜。这些组织体积的分析 将阐明产生的不同肿瘤，免疫和基质细胞的空间相互作用组 肿瘤异质性以及它们与肿瘤微环境组成部分的相互作用。我们的具体 目的是（1）与肿瘤，免疫和基质细胞的三维空间分布相关 来自人类胰腺的天然实体瘤组织中的脉管系统，淋巴机和细胞外基质 乳房恶性肿瘤结合了组织清除，免疫荧光，原位杂交和3D 灯表显微镜和（2）开发计算管道以构建三维空间图 蛋白质表达和RNA转录本在人类胰腺和乳房中完整的实体瘤组织中的定位 恶性肿瘤。 拟议工作的创新在于我们结合削减的跨学科策略 - 边缘组织清除，多路复用标记和高分辨率灯表显微镜，以更好地了解 实体瘤的本地三维结构。具体而言，通过对先前进行分类和定量 实体瘤组织的未知三维特征我们将能够关联 从同一标本中获取的基因组和蛋白质组学数据。该提议的重要性是 成功的人类肿瘤组织的三维表征将使关键见解得出 肿瘤内和肿瘤间的异质性，从而促进了细胞环境和细胞 - 微环境的鉴定 可以用作潜在治疗靶点的相互作用，从而提供新的治疗途径以减少 患者死亡率和改善生活质量。