Tsung Andrew J, Guda Maheedhara R, Asuthkar Swapna, Labak Collin M, Purvis Ian J, Lu Yining, Jain Neha, Bach Sarah E, Prasad Durbaka V R, Velpula Kiran K
Department of Cancer Biology and Pharmacology, University of Illinois College of Medicine at Peoria, Peoria, IL, USA.
Department of Neurosurgery, University of Illinois College of Medicine at Peoria, Peoria, IL, USA.
Oncotarget. 2017 Jul 4;8(27):44398-44409. doi: 10.18632/oncotarget.17897.
Glioblastoma (GBM) remains one of the most lethal and difficult-to-treat cancers of the central nervous system. The poor prognosis in GBM patients is due in part to its resistance to available treatments, which calls for identifying novel molecular therapeutic targets. In this study, we identified a mediator of Notch signaling, HEY1, whose methylation status contributes to the pathogenesis of GBM. Datamining studies, immunohistochemistry and immunoblot analysis showed that HEY1 is highly expressed in GBM patient specimens. Since methylation status of HEY1 may control its expression, we conducted bisulphite sequencing on patient samples and found that the HEY1 promoter region was hypermethylated in normal brain when compared to GBM specimens. Treatment on 4910 and 5310 xenograft cell lines with sodium butyrate (NaB) significantly decreased HEY1 expression with a concomitant increase in DNMT1 expression, confirming that promoter methylation may regulate HEY1 expression in GBM. NaB treatment also induced apoptosis of GBM cells as measured by flow cytometric analysis. Further, silencing of HEY1 reduced invasion, migration and proliferation in 4910 and 5310 cells. Furthermore, immunoblot and q-PCR analysis showed the existence of a potential positive regulatory loop between HEY1 and p53. Additionally, transcription factor interaction array with HEY1 recombinant protein predicted a correlation with p53 and provided various bonafide targets of HEY1. Collectively, these studies suggest HEY1 may be an important predictive marker for GBM and potential target for future GBM therapy.
胶质母细胞瘤（GBM）仍然是中枢神经系统中最致命且最难治疗的癌症之一。GBM患者预后不佳部分归因于其对现有治疗的抗性，这就需要确定新的分子治疗靶点。在本研究中，我们鉴定出一种Notch信号传导介质HEY1，其甲基化状态与GBM的发病机制有关。数据挖掘研究、免疫组织化学和免疫印迹分析表明，HEY1在GBM患者标本中高表达。由于HEY1的甲基化状态可能控制其表达，我们对患者样本进行了亚硫酸氢盐测序，发现与GBM标本相比，HEY1启动子区域在正常脑中发生了高甲基化。用丁酸钠（NaB）处理4910和5310异种移植细胞系可显著降低HEY1表达，同时DNMT1表达增加，证实启动子甲基化可能调节GBM中HEY1的表达。通过流式细胞术分析测量，NaB处理还诱导了GBM细胞凋亡。此外，沉默HEY1可降低4910和5310细胞的侵袭、迁移和增殖。此外，免疫印迹和q-PCR分析表明HEY1和p53之间存在潜在的正调控环。此外，与HEY1重组蛋白的转录因子相互作用阵列预测了与p53的相关性，并提供了HEY1的各种可靠靶点。总的来说，这些研究表明HEY1可能是GBM的重要预测标志物和未来GBM治疗的潜在靶点。
