TOP2A effects on transcription in gliomas: implications for personalized therapy

 描述（由申请人提供）：胶质母细胞瘤（GBM）是所有原发性脑肿瘤中最常见和最恶性的一种，浸润大脑并具有致死性。胶质母细胞瘤可以被重新诊断，也可以由低级别胶质瘤的进展引起。所有GBM都接受相同的非治愈性治疗，包括手术，化疗和放疗。鉴于GBM的分子特征高度可变，这些肿瘤已根据基因表达模式分为亚型。这些表达模式及其潜在机制可能提供了一种独特的肿瘤易感性，这种可能性尚未探索。在神经胶质瘤进展过程中，许多控制前神经GBM亚型基因表达的转录因子似乎受到拓扑异构酶IIA（TOP2A）的调节，拓扑异构酶IIA是一种自早期阶段以来由这些肿瘤表达的酶。TOP2A参与细胞周期进程、DNA修复和转录。TOP2A通过产生瞬时双链DNA断裂（DSB）使DNA去螺旋并增强染色质可及性。在干细胞中，TOP2A调节组蛋白3赖氨酸4甲基化（H3K4me2）位点的转录，但其在GBM中的作用尚不清楚。依托泊苷是一种诱导细胞毒性DSB的TOP2A靶向药物，对GBM患者的一个子集有效。使用小鼠模型，我表明，依托泊苷直接进入这些脑肿瘤的交付导致更高的疗效比全身交付所达到的浓度。然而，依托泊苷敏感性在GBM之间的差异仍然无法解释。我建议研究TOP2A对GBM的转录调控作用，以及它与依托泊苷敏感性的关系。为此，我将使用细胞系研究组蛋白甲基化标记H3K4me2是否影响TOP2A与肿瘤基因组的结合，以及这如何影响基因表达。使用人类GBM样本，我将比较TOP2A升高的样本与缺乏这种酶的样本。我还将探索不同肿瘤等级的TOP2A活性。使用神经导航引导，我将研究在GBM的可切除部分的TOP2A的差异，并在肿瘤边缘，以探讨是否可以用足叶乙甙靶向浸润脑的肿瘤细胞。TOP2A DNA结合和DSB对于这种酶的转录调节及其与依托泊苷的靶向是常见的，可能使受TOP2A转录调节的肿瘤对这种药物敏感。我将探讨这种关系的GBM细胞系与不同的依托泊苷敏感性。为了探索TOP2A在进展中的作用，我将研究TOP2A在转基因小鼠胶质瘤中不同时间的转录调控。为了探索依托泊苷在胶质瘤中引起的转录改变，我将研究依托泊苷治疗12小时后的肿瘤和复发。综上所述，我建议描述TOP2A在GBM中的转录调控及其在进展中的作用。我将确定个性化肿瘤内依托泊苷治疗的分子标准，并了解依托泊苷对转录调控的破坏如何影响复发。TOP2A作为浸润性肿瘤的靶标也将 被探索。总体目标是设计一个个性化的肿瘤内依托泊苷试验GBM。

 DESCRIPTION (provided by applicant): Glioblastomas (GBMs), the most common and malignant of all primary brain tumors, infiltrate the brain and are lethal. GBM can be diagnosed de novo, or result from progression of lower-grade gliomas. All GBMs undergo the same non-curative treatment, including surgery, chemotherapy and radiation. Given that molecular characteristics are highly variable across GBMs, these tumors have been classified into subtypes based on gene expression patterns. These expression patterns and their underlying mechanisms might provide a unique tumoral vulnerability, a possibility not yet explored. Many transcription factors that control gene expression on proneural GBM subtype during glioma progression appear to be modulated by Topoisomerase IIA (TOP2A), an enzyme expressed by these tumors since early stages. TOP2A is involved in cell cycle progression, DNA repair and transcription. TOP2A de-coils DNA and enhances chromatin accessibility by creating transient double strand-DNA breaks (DSB). In stem cells, TOP2A regulates transcription at loci with histone 3 lysine 4 methylation (H3K4me2), but its role in GBM is unknown. Etoposide, a TOP2A-targeting drug that induces cytotoxic DSB is efficacious for a subset of GBM patients. Using mouse models, I showed that delivery of etoposide directly into these brain tumors led to higher efficacy than the concentration achievable by systemic delivery. However, differences in etoposide sensitivity across GBMs remain unexplained. I propose to study TOP2A's role on transcriptional regulation on GBM, and how it relates to etoposide sensitivity. To do this, using cell lines I will investigate whether histone methylation mark H3K4me2 influences TOP2A binding to the tumor genome, and how this affects gene expression. Using human GBM specimens, I will compare samples with elevated TOP2A to those lacking this enzyme. I will also explore TOP2A activity across different tumor grades. Using neuro-navigation guidance, I will investigate the differences in TOP2A at the resectable portion of GBM, and at the tumor margins to explore whether tumor cells infiltrating the brain can be targeted with etoposide. TOP2A DNA binding and DSB are common to transcriptional regulation by this enzyme and its targeting with etoposide, potentially rendering tumors that are transcriptionally regulated by TOP2A susceptible to this drug. I will explore this relationship on GBM cell lines with variable etoposide sensitivity. To explore the role of TOP2A on progression, I will study TOP2A transcriptional regulation at different times in transgenic mouse gliomas. To explore the transcriptional alterations caused by etoposide in gliomas, I will study tumors following 12 hours of etoposide treatment, and at recurrence. In summary, I propose to characterize transcriptional regulation by TOP2A in GBM and its role on progression. I will identify the molecular criteria for personalizing intratumoral etoposide therapy and understand how disruption of transcriptional regulation with etoposide influences recurrence. TOP2A as a target for infiltrating tumor also will be explored. The overarching goal is to design a personalized intratumoral etoposide trial for GBMs.