Measuring Signaling Pathway Dynamics During Tissue Growth in Hydrogels

项目概述：本文的研究受到肿瘤不再被感知的概念的启发 通过列举恶性肿瘤的基因突变肿瘤微环境有助于 影响癌症进展的关键信号。我们的长期目标是调节当地环境 这些研究旨在确定旁分泌信号， 异常生长虽然周围肿瘤微环境的组成已经与 疾病进展，用于鉴定源自间质的关键旁分泌因子的策略是有限的。 我们的方法是基于分泌组学的交叉点（即，来源于RNAseq）与大规模 使用转录活性细胞荧光（TRACER）测量以定量转录因子（TF），和 微RNA（miRNA）活性，以确定驱动观察到的表型的关键因素和途径。 巨噬细胞是在原发性肿瘤中以最高水平存在的免疫细胞。的部件 肿瘤及其环境决定了肿瘤相关巨噬细胞（TAM）的表型。的倾向 促进肿瘤生长和转移，或起杀瘤或抑瘤作用的TAM是致癌细胞 这个过程被称为免疫编辑。TAM作为肿瘤的函数的可变行为 属性可能有助于患者之间的差异结果，并且正在研究TAM 其预后价值。具体目标1将研究癌细胞和TAM之间的旁分泌通讯 这可能会影响TAMs促进或抑制侵袭性表型的能力。初步研究将调查 巨噬细胞/单核细胞群和癌细胞的双向教育。随后，乳房 上皮细胞或癌细胞将与不同表型的巨噬细胞（浸润前 单核细胞，组织驻留的巨噬细胞和TAM），并研究细胞间的进化通讯。 导致肿瘤促进或肿瘤抑制微环境的细胞。具体目标2将扩大 TRACER技术到单细胞水平（即，生物发光显微镜），以研究 最具侵袭性的癌细胞。肿瘤人群中的异质性越来越被认为是一种 是观察到的癌症表型的重要贡献者。这些研究捕获了活性TF/miRNA 与侵袭相关，并确定具有不同活性的因子，这些因子是肿瘤细胞分化表型的基础。 侵袭性细胞相对于群体内的非侵袭性细胞。捕捉动态的能力 许多TF/miRNA在单细胞分辨率下的活性可用于追踪驱动细胞命运的线索 这是通过其他方法无法实现的，并为癌症生物学提供了新的视角 和癌症免疫学最后，我们研究了旁分泌信号传导和TF/miRNA活性， 化疗和对肿瘤微环境的影响，因为代偿信号可能作为一种 治疗靶点，以提高目前癌症治疗的疗效。

Project Summary: The studies herein are inspired by the concept that tumors can no longer be perceived through enumeration of the genetic mutations within the malignancy. The tumor microenvironment contributes key signals that influence cancer progression. Our long-term goal is aimed at modulating the local environment to inhibit disease progression, and these studies are aimed at identifying paracrine signaling that may drive abnormal growth. While the composition of the surrounding tumor microenvironment has been connected to disease progression, strategies for identifying the key paracrine factors emanating from the stroma are limited. Our approach is based on the intersection of secretomics (i.e., derived from RNAseq) with large-scale measurements using a TRanscriptional Activity CEll aRray (TRACER) to quantify transcription factor (TF) and micro RNA (miRNA) activity in order to identify key factors and pathways driving the observed phenotype. Macrophages are the immune cells present at the greatest levels in a primary tumor. The components of the tumor and its environment determine the phenotype of tumor associated macrophages (TAMs). The propensity of the TAMs to promote tumor growth and metastasis, or act tumoricidal or tumorostatic, is an oncongenic cell process that has been termed immune editing. The variable behavior of TAMs as a function of the tumor properties may contribute to the differential outcomes among patients, and TAMs are being investigated for their prognostic value. Specific Aim 1 will investigate paracrine communication between cancer cells and TAMs that can impact the ability of TAMs to promote or inhibit invasive phenotypes. Initial studies will investigate the bi-directional education of macrophage/monocyte populations and the cancer cells. Subsequently, mammary epithelial cells or cancer cells will be co-cultured with macrophages of varying phenotypes (pre-infiltrating monocytes, tissue-resident macrophages, and TAMs) and investigate the evolving communication between the cells that leads to tumor-promoting or tumor-inhibiting microenvironments. Specific Aim 2 will extend the TRACER technology to the single cell level (i.e., bioluminescence microscopy) to investigate signaling within the most invasive cancer cells. Heterogeneity within the tumor population is increasingly appreciated as an important contributor to the observed cancer phenotype. These studies capture the active TFs/miRNA associated with invasion, and identify the factors with distinct activity that underlie the differential phenotype of the invasive cell relative to the non-invasive cells within the population. The ability to capture the dynamic activity of numerous TF/miRNAs at single cell resolution can be employed to track the cues driving cell fate decisions, which is not achievable through other methods and provides novel perspectives into cancer biology and cancer immunology. Finally, we investigate the paracrine signaling and TF/miRNA activity in the context of chemotherapy and impact on the tumor microenvironment, as compensatory signaling may serve as a therapeutic target to enhance the efficacy of current cancer treatments.