Endothelial-to-osteoblast transition in prostate cancer bone metastasis

项目摘要/摘要 骨转移是前列腺癌发病率和死亡率的主要因素。PCa骨 转移瘤是独特的成骨细胞，以新骨形成为特征，新骨形成促进肿瘤生长， 骨头因此，PCa细胞与其错误诱导的骨之间的双向相互作用在PCa中起着关键作用。 骨的进展。据推测，转移性前列腺癌通过刺激骨形成诱导新骨形成。 骨髓中成骨细胞的增殖。然而，我们最近证明， 内皮细胞可以通过一个很少被认识的过程产生成骨细胞，这个过程被称为内皮细胞- 成骨细胞（EC-至-OSB）转化，在该过程中产生具有独特性质的EC-OSB杂交细胞。我们 研究表明，肿瘤诱导的EC向OSB的转化是导致成骨细胞 前列腺癌骨转移。这一新的见解，令人惊讶的骨形成作用的EC-OSB杂交细胞在PCa 骨转移为靶向肿瘤微环境中的这些细胞提供了新的理论基础。 我们假设肿瘤诱导的EC向OSB转化产生EC-OSB杂交细胞，其提供了 旁分泌因子以支持骨中的转移性PCa生长。我们的目标是靶向EC-OSB杂交细胞， 改善骨转移的治疗。我们将：目标1。确定内皮细胞 细胞到成骨细胞（EC到OSB）的转变。EC向OSB的转化需要抑制血管生成 和内皮细胞中成骨细胞生成途径的激活。我们的初步结果表明，BMP 4 激活Notch-Hey1通路以抑制血管生成，激活p38 MAPK-β-catenin-OSX通路以刺激 成骨我们将研究BMP 4如何整合这两种途径来重新编程内皮细胞。目的 2.鉴定促进PCa进展的EC-OSB杂交细胞分泌因子（EC-OSB因子）。EC-OSB 通过iTRAQ（蛋白质）和RNAseq（基因）分析来检查杂交细胞分泌组。Tenascin C是 沿着与CTGF和多功能蛋白聚糖一起识别，它们一起可以构成"EC到OSB签名"。我们将 研究这些EC-OSB因子在体外和体内对PCa细胞活性的影响。"欧共体-OSB签字" 也将使用临床样品作为成骨细胞骨转移的生物标志物进行测试。目标3。发展 靶向EC-OSB杂交细胞以改善PCa骨转移的治疗结果的策略。欧盟委员会- OSB杂交细胞可能导致先前描述的从头治疗抗性。我们将研究我们是否 通过联合卡博替尼或卡巴他赛（靶向PCa）可以改善PCa骨转移的治疗结果 肿瘤）与Rad-223（靶向EC-OSB杂交细胞）。目标4。确定是否发生EC到OSB转换 在正常骨和肿瘤诱导骨中EC-OSB杂交细胞的分数。我们生成了一个新的Double 报告小鼠（col1-GFP/Tie2-cre/Rosa-tdTomato），其中EC为RFP+，OSB为GFP+，EC-OSB为GFP+。 RFP +/GFP+，以检查EC-OSB杂交细胞是否参与正常骨发育及其分数 在肿瘤诱发的骨骼中。

Project Abstract/Summary Bone metastases are the major contributing factors to prostate cancer (PCa) morbidity and mortality. PCa bone metastases are uniquely osteoblastic and characterized by new bone formation, which promotes tumor growth in bone. Thus, bi-directional interaction between the PCa cells and their mis-induced bone plays a critical role in PCa progression in bone. It has been assumed that metastatic PCa induces new bone formation by stimulating the proliferation of osteoblasts in the bone marrow. However, we recently demonstrated that tumor-associated endothelial cells could give rise to osteoblasts through a little-appreciated process known as endothelial cell-to- osteoblast (EC-to-OSB) conversion, generating in the process EC-OSB hybrid cells with unique properties. Our studies showed that tumor-induced EC-to-OSB conversion is one mechanism that leads to osteoblastic bone metastasis of PCa. This new insight into the surprising bone-forming role of EC-OSB hybrid cells in PCa bone metastasis provides a novel rationale to target these cells in the tumor microenvironment. We hypothesize that tumor-induced EC-to-OSB conversion generates EC-OSB hybrid cells, which provide paracrine factors to support metastatic PCa growth in bone. Our objective is to target EC-OSB hybrid cells for improving therapy for bone metastasis. We will: Aim 1. Determine the mechanisms underlying endothelial cell-to-osteoblast (EC-to-OSB) transition. EC-to-OSB conversion requires both the inhibition of angiogenesis and activation of osteoblastogenesis pathways in endothelial cells. Our preliminary results indicate that BMP4 activates the Notch-Hey1 pathway to inhibit angiogenesis and the p38MAPK-β-catenin-OSX pathway to stimulate osteogenesis. We will examine how BMP4 integrates these two pathways to reprogram the endothelial cells. Aim 2. Identify EC-OSB hybrid cell secreted factors (EC-OSB factors) that promote PCa progression. EC-OSB hybrid cell secretome was examined by both iTRAQ (protein) and RNAseq (gene) analyses. Tenascin C was identified along with CTGF and versican, which together may constitute an “EC-to-OSB signature”. We will investigate the effects of these EC-OSB factors on PCa cell activity in vitro and in vivo. The “EC-OSB signature” will also be tested as biomarkers for osteoblastic bone metastasis using clinical samples. Aim 3. Develop strategies that target EC-OSB hybrid cells to improve therapy outcomes for PCa bone metastasis. The EC- OSB hybrid cells may contribute to de novo therapy resistance described previously. We will examine whether we can improve therapy outcomes for PCa bone metastasis by combining cabozantinib or cabazitaxel (targets PCa tumor) with Rad-223 (targets EC-OSB hybrid cells). Aim 4. Determine whether EC-to-OSB transition occurs in normal bone and the fraction of EC-OSB hybrid cells in tumor-induced bone. We generated a new Double Reporter Mice (col11-GFP/Tie2-cre/Rosa-tdTomato), in which EC are RFP+, OSB are GFP+, and EC-OSB are RFP+/GFP+, to examine whether EC-OSB hybrid cells are involved in normal bone development and their fraction in tumor-induced bone.