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

 描述(申请人提供)：转移性恶性肿瘤的治疗因缺乏对肿瘤-宿主相互作用的了解而受到阻碍。特别是，整体患者的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，它可以通过交感神经系统的激活而系统释放或从肿瘤中分泌。其中，神经母细胞瘤(NB)和尤文肉瘤(ES)这两种儿科肿瘤中NPY含量尤其丰富。值得注意的是，在这两种恶性肿瘤中，患者的高血清NPY与转移和骨侵犯有关。此外，我们还发现NPY刺激肿瘤细胞的运动和侵袭，而在侵袭和转移细胞中，NPY及其受体的表达明显增强。NPY在骨转移瘤中的表达尤其高，而富含NPY的ES细胞在异种异种移植模型中可导致频繁的骨转移。值得注意的是，在NB和ES中，NPY的这些转移行为都是由低氧诱导的Y2R/Y5R途径介导的。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。在此基础上，我们假设NB和ES的扩散是由NPY常见的、与疾病无关的作用驱动的，例如：1)增加肿瘤细胞的增殖、运动和侵袭力；2)血管生成；3)通过阻止成骨细胞分化和刺激溶骨因子的释放来促进骨侵袭。当全身性NPY水平由于高交感神经活动而升高时，同样的机制可以促进其他肿瘤的扩散，而不是内源性NPY表达。因此，我们推测NPY轴可能被成功地靶向以防止转移。在目前的研究中，我们将专注于破译这些普遍的过程，而不是肿瘤的特定特征，使用NB和ES作为富含NPY的肿瘤的模型。为此，我们将1)确定NPY依赖的转移形成阶段；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。首先，将使用动物模型测试NPY下调或过表达对转移形成的影响。对于ES，原位异种移植物将通过动脉阻断接受低氧，以评估NPY作为低氧诱导的肿瘤恶性度增加的介质。在确定了NPY依赖的转移步骤之后，我们将通过体外实验总结转移的阶段，确定其作用机制和介导性受体。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因子的变化。最后，我们将在体外和体内测试阻断已识别的受体和途径是否能抑制NPY的转移作用。这项研究将是第一次测试NPY的转移特性并评估其 作为抗转移治疗靶点的价值。虽然这个项目将专注于高密度的肿瘤 NPY的表达，我们的数据将为未来研究系统交感神经元衍生的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.