Neuropeptide Y (NPY) as a hypoxia-driven metastatic factor

 描述（由申请人提供）：转移性恶性肿瘤的治疗受到对疾病传播基础的肿瘤-宿主相互作用理解不足的阻碍。特别是，患者整体健康状况影响癌症进展的机制仍不清楚。我们提出了一种交感神经递质，神经肽Y（NPY），作为一种新的转移因子，它可以全身释放，由于交感系统激活或分泌的肿瘤。其中，神经母细胞瘤（NB）和尤文肉瘤（ES）这两种小儿肿瘤的NPY含量特别丰富。引人注目的是，在这两种恶性肿瘤中，患者的高血清NPY与转移和骨受累相关。此外，我们已经发现，NPY刺激肿瘤细胞的运动性和侵袭性，而肽及其受体的表达在侵袭性和转移性细胞中显著增强。NPY在骨转移中特别高表达，而富含NPY的ES细胞在原位异种移植模型中引起频繁的骨转移。值得注意的是，在NB和ES中，NPY的这些转移作用由缺氧诱导的Y2 R/Y 5 R途径介导。同样的受体也存在于内皮细胞上并介导NPY的血管生成作用。基于此，我们假设NB和ES的传播是由NPY的常见的、与疾病无关的作用驱动的，例如1）肿瘤细胞增殖、运动性和侵袭性的增加; 2）血管生成; 3）通过阻断成骨细胞分化和刺激溶骨因子的释放来促进骨侵袭。相同的机制可以有助于其他肿瘤的传播，没有内源性NPY表达，当全身NPY水平升高，由于高交感神经活性。因此，我们推测，NPY轴可以成功地针对防止转移。在目前的研究中，我们将专注于破译这些普遍的过程，而不是肿瘤特异性特征，使用NB和ES作为富含NPY的肿瘤模型。为此，我们将1）确定转移形成的NPY依赖性阶段; 2）确定NPY作用的细胞和分子机制; 3）评估NPY受体作为潜在的治疗靶点。最初，将使用动物模型测试NPY敲低或过表达对转移形成的影响。对于ES，将通过动脉闭塞使原位异种移植物经受缺氧，以评估NPY作为缺氧诱导的肿瘤恶性程度增加的介质。在确定了转移的NPY依赖步骤后，我们将使用体外测定法来确定其作用机制和介导受体，以重现转移的阶段。我们还将确定NPY诱导的信号通路激活，基因表达和肿瘤细胞分泌的因子的变化。最后，我们将在体外和体内测试阻断所鉴定的受体和途径是否抑制NPY的转移作用。这项研究将是第一个测试NPY的转移特性，并评估其在肿瘤转移中的作用。 作为抗转移治疗靶点的价值。虽然该项目将重点关注高风险的肿瘤， 我们的数据将为将来研究系统性、交感神经源性NPY在癌症进展中的作用提供基础。

 DESCRIPTION (provided by applicant): The treatment of metastatic malignancies is hindered by a poor understanding of the tumor-host interactions underlying disease dissemination. Particularly, the mechanisms by which the overall patient's well-being impacts cancer progression remain unclear. We propose a sympathetic neurotransmitter, neuropeptide Y (NPY), as a novel metastatic factor, which can be released systemically due to sympathetic system activation or secreted from tumors. Among them, two pediatric tumors, neuroblastoma (NB) and Ewing sarcoma (ES), are particularly rich in NPY. Strikingly, in both malignancies, high serum NPY in patients associates with metastases and bone involvement. Moreover, we have found that NPY stimulates motility and invasiveness of tumor cells, while expression of the peptide and its receptors is markedly enhanced in invasive and metastatic cells. NPY is particularly highly expressed in bone metastases, while NPY-rich ES cells give rise to frequent bone metastases in orthotopic xenograft model. Notably, in both NB and ES, these metastatic actions of NPY are mediated by the hypoxia-inducible, Y2R/Y5R pathway. The same receptors are also present on endothelial cells and mediate angiogenic actions of NPY. Based on this, we hypothesize that the dissemination of NB and ES is driven by the common, disease-independent actions of NPY, such as 1) an increase in tumor cell proliferation, motility and invasiveness; 2) angiogenesis; 3) promoting bone invasion by blocking osteoblast differentiation and stimulating the release of osteolytic factors. The same mechanisms can contribute to the dissemination of other tumors, without endogenous NPY expression, when systemic NPY levels are elevated due to high sympathetic activity. Thus, we postulate that that NPY axis may be successfully targeted to prevent metastasis. In the current study, we will focus on deciphering these universal processes, rather than tumor-specific features, using NB and ES as models of NPY-rich tumors. To this end, we will 1) Identify NPY-dependent stages of metastases formation; 2) Determine cellular and molecular mechanisms of NPY actions, and 3) Assess NPY receptors as potential therapeutic targets. Initially, the effect of NPY knock-down or overexpression on metastases formation will be tested using animal models. For ES, orthotopic xenografts will be subjected to hypoxia by arterial occlusion to assess NPY as a mediator of hypoxia-induced increase in tumor malignancy. Having determined NPY-dependent steps of metastases, we will identify mechanisms of its actions and mediating receptors using in vitro assays recapitulating stages of metastases. We will also determine NPY-induced changes in signaling pathway activation, gene expression and factors secreted from tumor cells. Lastly, we will test if blocking the identified receptors and pathways inhibits metastatic actions of NPY in vitro and in vivo. This study will be the first to test metastatic properties of NPY and assess its value as a target for anti- metastatic therapies. While this project will focus on tumors with high NPY expression, our data will provide a foundation for future studies on the role of systemic, sympathetic neuron-derived NPY in cancer progression.