A Cellular and Molecular Analysis of the Intravasation Step in Tumor Metastasis

肿瘤转移中浸润步骤的细胞和分子分析

• 批准号: 7915828
• 负责人: JAMES P QUIGLEY
• 金额: $ 34.47万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7915828
• 关键词: Animals; Biological Assay; Blood; Blood Vessels; Carcinoma; Cell Communication; Cell Line; Cell physiology; Cells; Cessation of life; Chick Embryo; Clinical; Complement; Databases; Distal; Down-Regulation; Embryo; Evaluation; Event; Exhibits; Extravasation; Fibroblast Growth Factor 2; Fibroblasts; Gelatinase B; Goals; Human; In Vitro; Individual; Inflammatory; Intervention; Knowledge; Laboratories; Link; Malignant Epithelial Cell; Malignant Neoplasms; Matrix Metalloproteinases; Measurement; Mesenchymal; Modeling; Modification; Molecular; Molecular Analysis; Monitor; Mus; Neoplasm Metastasis; Organ; Outcome; Pancreatic carcinoma; Pathologic Processes; Physiological; Population; Primary Neoplasm; Procedures; Process; Property; Prostate; Proteins; Proteomics; Reporting; Research; Role; Site; Stromal Invasion; Survival Rate; Testing; Tissues; Tumor Cell Invasion; Variant; angiogenesis; cancer type; chorioallantoic membrane; comparative; congenic; fibrosarcoma; in vivo; insight; intravital microscopy; mouse model; neoplastic cell; novel; public health relevance; repository; tumor

DESCRIPTION (provided by applicant): A number of the specific steps in tumor dissemination have been extensively modeled, and molecularly dissected. However, one early step in the metastatic cascade, namely intravasation, the entry of escaping tumor cells into the vasculature, has been relatively understudied. One of the reasons for this is that up to now human tumor cell phenotypic variants that exhibit substantial differences in their intravasation ability have not been available for comparative analysis. Our laboratory reported the selection and isolation of two variants of a human fibrosarcoma cell line (HT-1080) that dramatically differ (50-100 fold) in their intravasation rate, yielding a similar >50 fold differential in their metastatic capabilities. These two tumor dissemination variants, HT- hi/diss and HT-lo/diss, were selected and monitored in the chick embryo model, where primary human tumors developing on the embryo's chorioallantoic membrane (CAM), recapitulate the multi-step tumor dissemination process and form micro metastatic foci in a number of secondary organs. We have also tested the two fibrosarcoma variants in different mouse metastasis assays and confirmed the substantial differential in their tumor dissemination capabilities. These two congenic, intravasation variants are thus suitable for a comparative analysis of early metastatic events. Therefore, in Specific Aim 1 we propose to molecularly dissect three physiological/pathological processes which determine the outcome of tumor cell intravasation events, i.e. induction of angiogenesis, stromal invasion and vasculotropism. We will analyze the functional role of select molecules contributing to the events involved in tumor cell intravasation including; inflammatory cell MMP-9; FGF-2; uPA; and tumor cell MMP-14. Unique assays for tumor cell interaction with blood vessels will be used to identify contributory molecules involved in the vasculotropic event. Mouse models for tumor cell dissemination and angiogenesis will be used to complement the chick embryo models for examining the identified molecules in the two fibrosarcoma variants. There exists however, a distinct lack of other tumor cell intravasation variants to compare and contrast and this is especially relevant for carcinomas, the major cancer in the human population. Therefore for Specific Aim 2 we propose to identify specific proteins which contribute mechanistically to the intravasation step in carcinoma dissemination by employing carcinoma variants selected in vivo for differential rates of intravasation. We will generate pairs of high and low intravasating variants from human prostate, colon and pancreatic carcinoma cell lines. Functional proteomic approaches will be applied both in vitro and in vivo to the selected carcinoma variants to verify and/or complement the cellular mechanisms and contributory molecules identified in fibrosarcoma-derived variants. The influence of mesenchymal fibroblasts and their products on the disseminating properties of the carcinoma variants will also be quantified. Novel mechanistic information about carcinoma cell intravasation will provide molecular links to a specific step in tumor dissemination, namely intravasation. PUBLIC HEALTH RELEVANCE: Deaths from cancer occur mainly because tumor cells spread from the primary tumor site to other vital organs and tissues. The tumor spread is usually through the vasculature. In order for tumor cells to escape from the primary tumor and enter the vasculature they alter some of their cellular processes by producing different levels of functioning molecules. The goal of the proposed research is to identify the relevant molecules and cellular processes that contribute to tumor cell spread so that clinical intervention can target those critical molecules and processes.

描述（由申请人提供）：肿瘤播散中的许多具体步骤已被广泛建模，并进行了分子解剖。然而，转移级联反应中的一个早期步骤，即内渗，即逃逸的肿瘤细胞进入脉管系统，一直相对研究不足。其原因之一是，迄今为止，在其内渗能力方面表现出实质性差异的人肿瘤细胞表型变体尚未用于比较分析。我们的实验室报道了选择和分离人纤维肉瘤细胞系（HT-1080）的两种变体，其在它们的内渗速率方面显著不同（50-100倍），在它们的转移能力方面产生类似的>50倍差异。在鸡胚模型中选择并监测这两种肿瘤播散变体HT-hi/diss和HT-lo/diss，其中在胚胎的绒毛尿囊膜（CAM）上发育的原发性人肿瘤重演了多步骤肿瘤播散过程并在许多次级器官中形成微转移灶。我们还在不同的小鼠转移试验中测试了两种纤维肉瘤变体，并证实了它们的肿瘤传播能力的实质性差异。因此，这两种同源的内渗变体适合于早期转移事件的比较分析。因此，在具体目标1中，我们提出从分子上剖析三个生理/病理过程，这些过程决定了肿瘤细胞内渗事件的结果，即诱导血管生成、基质侵袭和血管向性。我们将分析参与肿瘤细胞内渗的选定分子的功能作用，包括炎性细胞MMP-9、FGF-2、uPA和肿瘤细胞MMP-14。将使用肿瘤细胞与血管相互作用的独特测定来鉴定参与向血管性事件的贡献分子。肿瘤细胞播散和血管生成的小鼠模型将用于补充鸡胚模型，以检查两种纤维肉瘤变体中鉴定的分子。然而，存在明显缺乏其他肿瘤细胞内渗变体以进行比较和对比，这与癌症（人类群体中的主要癌症）尤其相关。因此，对于特定目标2，我们建议通过采用针对不同的内渗速率在体内选择的癌变体来鉴定在癌扩散中机械地有助于内渗步骤的特定蛋白质。我们将从人前列腺癌、结肠癌和胰腺癌细胞系中产生成对的高和低内渗变体。功能蛋白质组学方法将在体外和体内应用于选定的癌变体，以验证和/或补充纤维肉瘤衍生变体中鉴定的细胞机制和贡献分子。还将量化间充质成纤维细胞及其产物对癌变体的播散性质的影响。关于癌细胞内渗的新机制信息将提供与肿瘤扩散中的特定步骤即内渗的分子联系。公共卫生相关性：癌症导致的死亡主要是因为肿瘤细胞从原发肿瘤部位扩散到其他重要器官和组织。肿瘤通常通过脉管系统扩散。为了使肿瘤细胞逃离原发肿瘤并进入脉管系统，它们通过产生不同水平的功能分子来改变它们的一些细胞过程。这项研究的目的是确定有助于肿瘤细胞扩散的相关分子和细胞过程，以便临床干预可以针对这些关键分子和过程。