Abou-Antoun Tamara J, Nazarian Javad, Ghanem Anthony, Vukmanovic Stanislav, Sandler Anthony D
Department of Pharmaceutical Sciences, the School of Pharmacy, Lebanese American University, Byblos, Lebanon.
The Sheikh Zayed Institute for Pediatric Surgical Innovation, Children's National Health System, Washington, D.C., United States of America.
PLoS One. 2018 Jan 3;13(1):e0189711. doi: 10.1371/journal.pone.0189711. eCollection 2018.
Despite significant advances in cancer treatment and management, more than 60% of patients with neuroblastoma present with very poor prognosis in the form of metastatic and aggressive disease. Solid tumors including neuroblastoma are thought to be heterogeneous with a sub-population of stem-like cells that are treatment-evasive with highly malignant characteristics. We previously identified a phenomenon of reversible adaptive plasticity (RAP) between anchorage dependent (AD) cells and anchorage independent (AI) tumorspheres in neuroblastoma cell cultures. To expand our molecular characterization of the AI tumorspheres, we sought to define the comprehensive proteomic profile of murine AD and AI neuroblastoma cells. The proteomic profiles of the two phenotypic cell populations were compared to each other to determine the differential protein expression and molecular pathways of interest. We report exclusive or significant up-regulation of tumorigenic pathways expressed by the AI tumorspheres compared to the AD cancer cells. These pathways govern metastatic potential, enhanced malignancy and epithelial to mesenchymal transition. Furthermore, radio-therapy induced significant up-regulation of specific tumorigenic and proliferative proteins, namely survivin, CDC2 and the enzyme Poly [ADP-ribose] polymerase 1. Bio-functional characteristics of the AI tumorspheres were resistant to sutent inhibition of receptor tyrosine kinases (RTKs) as well as to 2.5 Gy radio-therapy as assessed by cell survival, proliferation, apoptosis and migration. Interestingly, PDGF-BB stimulation of the PDGFRβ led to transactivation of EGFR and VEGFR in AI tumorspheres more potently than in AD cells. Sutent inhibition of PDGFRβ abrogated this transactivation in both cell types. In addition, 48 h sutent treatment significantly down-regulated the protein expression of PDGFRβ, MYCN, SOX2 and Survivin in the AI tumorspheres and inhibited tumorsphere self-renewal. Radio-sensitivity in AI tumorspheres was enhanced when sutent treatment was combined with survivin knock-down. We conclude that AI tumorspheres have a differential protein expression compared to AD cancer cells that contribute to their malignant phenotype and radio-resistance. Specific targeting of both cellular phenotypes is needed to improve outcomes in neuroblastoma patients.
尽管癌症治疗和管理取得了重大进展，但超过60%的神经母细胞瘤患者表现出转移性和侵袭性疾病的预后极差。包括神经母细胞瘤在内的实体瘤被认为是异质性的，存在一群具有高度恶性特征且对治疗有逃避性的干细胞样细胞。我们之前在神经母细胞瘤细胞培养中发现了锚定依赖性（AD）细胞和锚定独立性（AI）肿瘤球之间可逆的适应性可塑性（RAP）现象。为了扩展我们对AI肿瘤球的分子特征描述，我们试图确定小鼠AD和AI神经母细胞瘤细胞的全面蛋白质组学图谱。将这两种表型细胞群体的蛋白质组学图谱相互比较，以确定差异蛋白质表达和感兴趣的分子途径。我们报告，与AD癌细胞相比，AI肿瘤球表达的致瘤途径有独特或显著上调。这些途径控制转移潜能、增强恶性程度以及上皮-间质转化。此外，放射治疗导致特定致瘤和增殖蛋白（即生存素、细胞周期蛋白依赖性激酶2（CDC2）和聚[ADP-核糖]聚合酶1）显著上调。通过细胞存活、增殖、凋亡和迁移评估，AI肿瘤球的生物功能特性对受体酪氨酸激酶（RTK）的舒尼替尼抑制以及2.5 Gy放射治疗具有抗性。有趣的是，血小板衍生生长因子BB（PDGF-BB）对血小板衍生生长因子受体β（PDGFRβ）的刺激在AI肿瘤球中比在AD细胞中更有效地导致表皮生长因子受体（EGFR）和血管内皮生长因子受体（VEGFR）的反式激活。舒尼替尼对PDGFRβ的抑制在两种细胞类型中都消除了这种反式激活。此外，48小时的舒尼替尼治疗显著下调了AI肿瘤球中PDGFRβ、MYCN、SOX2和生存素的蛋白质表达，并抑制了肿瘤球的自我更新。当舒尼替尼治疗与生存素敲低联合使用时，AI肿瘤球的放射敏感性增强。我们得出结论，与AD癌细胞相比，AI肿瘤球具有差异蛋白质表达，这有助于其恶性表型和放射抗性。需要对两种细胞表型进行特异性靶向治疗，以改善神经母细胞瘤患者的治疗结果。
