Molecular Dissection of an Arntl2 induced pro-metastatic secretome

Arntl2 诱导的促转移分泌蛋白组的分子解剖

• 批准号: 9260764
• 负责人: Monte Meier Winslow
• 金额: $ 36.7万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-12 至 2021-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9260764
• 关键词: Adhesions; Benign; Biological; Biological Assay; CRISPR/Cas technology; Cancer Etiology; Cancer Patient; Cell Culture System; Cell Culture Techniques; Cell Death; Cell Line; Cells; Cessation of life; Clinical; Complex; Country; Data; Development; Disease; Dissection; Disseminated Malignant Neoplasm; Distant; Environment; Epithelial Cells; Gene Expression Profiling; Genes; Genetic; Genetic Engineering; Genetically Engineered Mouse; Goals; Growth; Health; Human; In Vitro; Individual; Knowledge; Lung Adenocarcinoma; Malignant Neoplasms; Malignant neoplasm of lung; Mediating; Metastatic Adenocarcinoma; Methods; Modeling; Molecular; Mus; Neoplasm Metastasis; Organ; Outcome; Output; Pathway interactions; Patient-Focused Outcomes; Patients; Phenotype; Positioning Attribute; Primary Neoplasm; Process; Proteins; Recombinant Proteins; Role; Sampling; Signal Transduction; Site; Solid Neoplasm; System; Testing; Transcriptional Regulation; Transplantation; United States; Woman; autocrine; cDNA Expression; cancer cell; cancer type; clinical practice; experimental study; fitness; genome editing; in vivo; knock-down; lentiviral-mediated; loss of function; men; metastatic process; mouse model; neoplastic cell; new therapeutic target; novel; novel therapeutics; overexpression; prevent; public health relevance; response; screening; therapeutic target; transcription factor; treatment strategy; tumor; tumor progression; tumorigenesis

 DESCRIPTION (provided by applicant): Lung cancer is the leading cause of cancer deaths in both men and women in the United States, with over 155,000 patients dying each year in this country alone. Several factors contribute to the poor outcome of lung cancer patients, but, as in most solid tumors, the ability of cancer cells to leave the primary tumor and establish inoperable metastases is a major impediment to successful therapy. Metastasis thus represents a major clinical challenge that is driven by as yet poorly understood cell state alterations. By integratin gene expression analyses on murine models of metastatic lung adenocarcinoma and human lung adenocarcinomas we identified the transcription factor Arntl2 as a key driver of lung adenocarcinoma metastatic ability. Arntl2 appears to drive metastatic fitness by controlling the expression of a complex pro-metastatic secretome that has the ability to greatly increase clonal growth potential. In Aim1, we will functionally interrogate Arntl2 function in human and mouse lung adenocarcinoma cell lines. We will perform gain- and loss-of-function experiments and fully assess the cellular phenotypes driven by Arntl2. Transplantation assays and quantification of initial adhesion, proliferation, and cell death within the metastatic site in vivo will elucidate te cellular consequence of high Arntl2 expression. In Aim2, we will investigate which Arntl2-regulated secreted factors cooperate to drive clonal growth and metastatic ability. We will integrate screening of recombinant proteins in cell culture, gain- and loss-of-function experiments in cell culture and in vivo, and therapeutically target pathways downstream of key pro-metastatic secreted proteins to better understand the importance of autocrine metastatic niche factors. In Aim3, we will use novel methods for CRISPR/Cas9-mediated genome editing and lentiviral-mediated cDNA expression to test the requirement and sufficiency of Arntl2 and Arntl2- regulated genes to promote step of the metastatic cascade in autochthonous mouse models of human lung cancer. Given the immense clinical impact of metastatic cancer and the current gap in understanding the molecular underpinnings of this disease state, both clinical practice and patient outcome would be greatly impacted by any new therapies that might result from the fundamental knowledge gained from our proposed analyses. By combining quantitative methods and powerful in vivo methods, we hope to uncover general principles that govern tumor progression and metastatic spread and ultimately reveal novel therapeutic targets across the continuum of cancer progression.

 描述（由申请人提供）：肺癌是美国男性和女性癌症死亡的主要原因，仅在这个国家每年就有超过155，000名患者死亡。有几个因素导致肺癌患者的预后不佳，但是，与大多数实体瘤一样，癌细胞离开原发肿瘤并建立不可手术转移的能力是成功治疗的主要障碍。因此，转移代表了一个主要的临床挑战，这是由尚未充分了解的细胞状态改变驱动的。通过对转移性肺腺癌和人肺腺癌的鼠模型的整合素基因表达分析，我们鉴定了转录因子Arntl 2作为肺腺癌转移能力的关键驱动因子。Arntl 2似乎通过控制复杂的促转移分泌蛋白质组的表达来驱动转移适应性，所述促转移分泌蛋白质组具有极大地增加克隆生长潜力的能力。在Aim 1中，我们将在人类和小鼠肺腺癌细胞系中功能性地询问Arntl 2功能。我们将进行功能获得和丧失实验，并充分评估Arntl 2驱动的细胞表型。移植测定和体内转移位点内的初始粘附、增殖和细胞死亡的定量将阐明高Arntl 2表达的细胞后果。在Aim 2中，我们将研究哪些Arntl 2调节的分泌因子协同驱动克隆生长和转移能力。我们将整合细胞培养中重组蛋白的筛选，细胞培养和体内功能获得和丧失实验，以及关键促转移分泌蛋白下游的治疗靶向途径，以更好地了解自分泌转移小生境因子的重要性。在Aim 3中，我们将使用CRISPR/Cas9介导的基因组编辑和慢病毒介导的cDNA表达的新方法来测试Arntl 2和Arntl 2调节的基因在人类肺癌的本地小鼠模型中促进转移级联步骤的需要和充分性。鉴于转移性癌症的巨大临床影响以及目前在理解这种疾病状态的分子基础方面的差距，临床实践和患者结局将受到任何新疗法的极大影响，这些新疗法可能来自我们提出的分析中获得的基础知识。通过结合定量方法和强大的体内方法，我们希望揭示控制肿瘤进展和转移扩散的一般原则，并最终揭示癌症进展连续体中的新治疗靶点。