BACKGROUND Diffuse midline glioma (DMG) is a pediatric malignancy with poor prognosis. Most children die less than one year after diagnosis. Recently, mutations in histone H3 have been identified and are believed to be oncogenic drivers. Targeting this epigenetic abnormality using histone deacetylase (HDAC) inhibitors such as panobinostat (PS) is therefore a novel therapeutic option currently evaluated in clinical trials. METHODS BH3 profiling revealed engagement in an irreversible apoptotic process of glioma cells exposed to PS confirmed by annexin-V/propidium iodide staining. Using proteomic analysis of 3 DMG cell lines, we identified 2 proteins deregulated after PS treatment. We investigated biological effects of their downregulation by silencing RNA but also combinatory effects with PS treatment in vitro and in vivo using a chick embryo DMG model. Electron microscopy was used to validate protein localization. RESULTS Scaffolding proteins EBP50 and IRSp53 were upregulated by PS treatment. Reduction of these proteins in DMG cell lines leads to blockade of proliferation and migration, invasion, and an increase of apoptosis. EBP50 was found to be expressed in cytoplasm and nucleus in DMG cells, confirming known oncogenic locations of the protein. Treatment of glioma cells with PS together with genetic or chemical inhibition of EBP50 leads to more effective reduction of cell growth in vitro and in vivo. CONCLUSION Our data reveal a specific relation between HDAC inhibitors and scaffolding protein deregulation which might have a potential for therapeutic intervention for cancer treatment.

中文翻译：

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HDAC抑制诱导对儿童神经胶质瘤的肿瘤进展至关重要的支架蛋白的表达：着重于EBP50和IRSp53。

背景弥漫性中线神经胶质瘤（DMG）是一种儿童恶性肿瘤，预后较差。大多数儿童在诊断后不到一年就死亡。最近，已经鉴定出组蛋白H3中的突变，并且认为其是致癌的驱动因素。因此，使用组蛋白脱乙酰基酶（HDAC）抑制剂（例如panobinostat（PS））靶向这种表观遗传异常是当前在临床试验中评估的一种新型治疗选择。方法BH3谱图揭示了膜联蛋白-V /碘化丙锭染色证实了暴露于PS的神经胶质瘤细胞的不可逆凋亡过程。使用3个DMG细胞系的蛋白质组学分析，我们确定了PS处理后2种蛋白被解除调控。我们研究了通过沉默RNA来下调其下调的生物学效应，以及使用鸡胚DMG模型在体内和体外与PS处理的组合效应。电子显微镜用于验证蛋白质定位。结果PS处理可上调支架蛋白EBP50和IRSp53。DMG细胞系中这些蛋白质的减少导致增殖，迁移，侵袭和凋亡增加的阻断。发现EBP50在DMG细胞的细胞质和细胞核中表达，证实了该蛋白的已知致癌部位。PS对神经胶质瘤细胞的治疗以及对EBP50的遗传或化学抑制作用可导致更有效地减少体外和体内细胞的生长。