Mechanisms associated with organotropic metastasis

摘要 大多数有实体瘤的癌症患者死于转移性疾病，器质性传播是一个研究不足的方面。 对于转移，迫切需要新的洞察力。流行的“种子和土壤”概念认为 为了支持潜伏的肿瘤的生存和生长，必须在遥远的器官(土壤)中有一个允许的环境。 细胞(种子)。建议在非转移器官中诱导允许土壤以改变转移路线；然而， 缺乏严格的实验证据和机理分析。纤维性组织改变，包括 炎症，提供转移的线索，与来自骨髓来源的细胞(BMDCs)的信号一起， 肿瘤分泌体和循环细胞外小泡。我们的初步数据显示器官嗜性转移 不仅依赖于次级器官中允许的基质重塑和BMDCs，而且还 取决于器官特异性血管连接对血管内皮屏障功能的破坏 蛋白质。我们的发现导致了一个中心假设，即血管异质性在功能上有助于 转移的器官趋向性‘。我们建议进行研究，以揭示不同的纤维化小生境 影响器官特有的血管床变化，导致器官趋化性转移。初步研究 血管生成素-2(Ang-2)被认为是肺转移的介质。我们的目标是解开 Ang-2依赖于肺而不是肾脏或肝脏，这也会在 纤维性环境。利用单细胞RNAseq和CyTOF，我们将确定细胞和分子靶点 Ang-2在转移前环境中的表达。初步研究表明血管紧张素-2可导致肺血管渗漏 不影响肾或肝血管的血管系统，从而引导转移到肺。释放的Exosome 被纤维化的器官还会增加血管的通透性和在肺部的转移定植，而不影响 肾或肝的血管构筑。纤维化器官的单细胞RNAseq以及基因工程小鼠 (GEMS)，将被用来解开组织特异性干扰Ang-2在乳腺癌中的限速作用 转移。使用实验室中产生的新宝石，我们将追踪特定血统的转移产生- 通过蛋白质组学分析，诱导外体并鉴定器官嗜性的决定因素。我们的预赛 研究表明，Ang-2通过抑制仅在肺中发现的claudin-5来破坏血管屏障 血管系统，与肾和肝血管不同，后者有多个多余的内皮细胞 克拉丁斯。整合内皮特异性缺失claudin-5和claudin-5报告小鼠的小鼠模型， 随着器官特异性内皮细胞在功能丧失和功能获得实验中的分子图谱，我们将 阐明特定的Claudins在器官亲和性转移中的作用。将进行分子研究以确定 血管紧张素-2抑制Claudin-5的机制及血管调节作用的研究 渗透性可以改变转移路线，而不考虑癌症特定的器官偏好。成功完成 拟议的研究将为转移机制和治疗意义提供新的见解。

ABSTRACT Most cancer patients with solid tumors die of metastatic disease and organotropic spread is an understudied aspect of metastasis, for which new insights are urgently required. The prevailing ‘seed and soil’ concept posits that permissive environment in a distant organ (soil) is necessary to support the survival and growth of lurking tumor cells (seeds). Induction of permissive soil in a non-metastatic organ is proposed to re-route metastasis; however, rigorous experimental evidence and mechanistic analyses are lacking. Fibrotic tissue alterations, including inflammation, provide cues for metastasis, together with the signals from bone marrow-derived cells (BMDCs), the tumor secretome, and circulating extracellular vesicles. Our preliminary data suggest organotropic metastasis is not solely dependent on permissive matrix remodeling and BMDCs in the secondary organs, but is also contingent on the disruption of vascular endothelial barrier function imposed by organ-specific vascular junction proteins. Our findings lead to a central hypothesis that ‘vascular heterogeneity functionally contributes to organotropism of metastasis’. We propose studies to unravel the mechanisms, by which distinct fibrotic niches effect organ-specific changes in the vascular beds, leading to organotropic metastasis. Preliminary studies identified angiopoetin-2 (Ang-2) as a putative mediator of lung metastasis. We aim to unravel the mechanisms of Ang-2 dependent tropism to the lung but not the kidney or liver, which also generate high Ang-2 levels in the fibrotic setting. Using single-cell RNAseq and CyTOF, we will determine the cellular and molecular targets of Ang-2 in the pre-metastatic milieu. Preliminary studies show Ang-2 induces vascular leakage in the lung vasculature without impacting kidney or liver vessels, thus directing metastasis to the lung. Exosomes released by the fibrotic organs also increase vascular permeability and metastatic colonization in the lung, without affecting kidney or liver vasculature. Single-cell RNAseq of fibrotic organs, as well as genetically engineered mice (GEMs), will be used to unravel the rate limiting effect of tissue-specific disruption of Ang-2 in breast cancer metastasis. Using novel GEMs generated in the lab, we will trace lineage-specific production of metastasis- inducing exosomes and identify the determinants of organotropism via proteomic analysis. Our preliminary studies show Ang-2 disrupts vascular barriers through repression of claudin-5 that is found exclusively in the lung vasculature, in contrast with the kidney and liver vessels, which present with multiple, redundant endothelial claudins. Integrating mouse models with endothelial-specific deletion of claudin-5 and claudin-5 reporter mice, and with molecular profiling of organ-specific endothelial cells in loss- and gain-of-function experiments, we will elucidate functions of specific claudins in organotropic metastasis. Molecular studies will be performed to identify putative mechanism of Ang-2 mediated suppression of claudin-5 and test whether manipulation of vascular permeability can re-route metastasis regardless of cancer-specific organ predilection. Successful completion of the proposed studies will provide new insights into mechanism of metastasis and therapeutic implications.