Tissue Factor as a Key Determinant of IDH1 Mutant versus IDH1 Wild-type Glioma Thrombosis and Malignancy

项目总结/摘要 浸润性神经胶质瘤是成人中最常见的原发性脑肿瘤类型， 美国每年。大约20-30%的浸润性胶质瘤含有异柠檬酸突变 脱氢酶1（IDH 1 mut）。IDH 1 mut导致整体DNA超甲基化，这有助于胶质瘤的发生。 然而，IDH 1 mut胶质瘤的侵袭性明显低于缺乏这种突变的胶质瘤。目前还不清楚 DNA超甲基化是如何导致这种独特的表型的。神经胶质瘤经常在肿瘤内产生血凝块（血栓）。 肿瘤和整个血流。长期以来，这些血栓被认为是不良预后的预测因子。 结果。我们最近报道IDH 1 mut胶质瘤产生的血栓少得多;我们的数据强烈表明， 甲基化诱导的F3（编码组织因子（TF）的基因）的抑制是原因。TF是 强效促凝血剂，当由癌症产生和释放时，会导致静脉血栓栓塞（VTE）。这 衰弱现象发生在~25%的胶质瘤患者中，但当IDH 1 mut存在时，则不会发生。除了 触发血栓形成，TF结合并激活蛋白酶激活受体2（PAR 2），一种跨膜受体 由癌细胞表达，通过多种细胞内途径发出信号以促进肿瘤恶性。我们 数据显示：（i）在所有直接参与血液凝固的基因中，F3 mRNA水平最强， （ii）IDH 1 mut胶质瘤中F3甲基化显著高于IDH 1 wt胶质瘤; (iii)TF蛋白水平在IDH 1 mut胶质瘤中始终低于IDH 1 wt胶质瘤;（iv）循环TF在IDH 1 mut胶质瘤中低于IDH 1 wt胶质瘤。 IDH 1 mut神经胶质瘤患者比IDH 1 wt神经胶质瘤患者;（v）高循环TF与VTE风险增加相关;（vi） 具有内源性IDH 1 mut的患者源性胶质瘤细胞比IDH 1 wt产生更小和更少的静脉血栓 （vii）在IDH 1 wt胶质瘤中抑制TF大大降低了它们的体外和体外增殖能力， 体内恶性肿瘤;（viii）胶质瘤表达低TF的患者的中位生存期为 胶质瘤高表达TF的患者，独立于IDH 1 mut。因此，我们假设甲基化- 诱导的TF抑制是IDH 1 mut低血栓形成和低恶性的关键决定因素， 表型在目的1中，我们将最终确定F3超甲基化是IDH 1 mut表达的机制。 抑制TF表达。在目标2中，我们将在一系列患者来源的IDH 1 wt中调节TF的表达。 和IDH 1 mut胶质瘤细胞，观察对细胞培养中肿瘤诱导的血栓形成和恶性肿瘤的影响， 在移植的小鼠中。在目标3中，我们将使用分子和药理学方法来研究治疗方法。 在胶质瘤中阻断TF-PAR 2信号传导的潜力。此外，我们将前瞻性地评估 确定循环TF水平，沿着其他临床、血液和组织生物标志物， 胶质瘤患者VTE风险的第一个预测模型。总的来说，这项研究将大大促进我们的 IDH 1 mut肿瘤生物学的理解，它将告知关于新的治疗和诊断策略， 改善神经胶质瘤患者的护理。

PROJECT SUMMARY/ABSTRACT Infiltrative glioma is the most common type of primary brain tumor in adults, causing over 17,000 deaths in the United States every year. Approximately 20-30% of infiltrative gliomas contain mutations in isocitrate dehydrogenase 1 (IDH1mut). IDH1mut causes global DNA hypermethylation, which contributes to gliomagenesis. Yet, IDH1mut gliomas are significantly less aggressive than gliomas lacking this mutation. It has remained unclear how DNA hypermethylation leads to this unique phenotype. Gliomas often produce blood clots (thrombi) within the tumor and throughout the bloodstream. These thrombi have long been known to be predictors of poor outcome. We recently reported that IDH1mut gliomas produce far fewer thrombi; our data strongly indicate that methylation-induced suppression of F3, the gene encoding Tissue Factor (TF), is the reason why. TF is a powerful procoagulant that, when produced and released by cancers, causes venous thromboemboli (VTE). This debilitating phenomenon occurs in ~25% of glioma patients, but never when IDH1mut is present. In addition to triggering thrombosis, TF binds and activates protease-activated receptor 2 (PAR2), a transmembrane receptor expressed by cancer cells that signals through multiple intracellular pathways to promote tumor malignancy. Our data show that: (i) among all genes that directly participate in blood clotting, F3 mRNA levels have the strongest inverse relationship with IDH1mut; (ii) F3 methylation is significantly higher in IDH1mut gliomas than IDH1wt gliomas; (iii) TF protein levels are consistently lower in IDH1mut gliomas than IDH1wt gliomas; (iv) circulating TF is lower in patients with IDH1mut gliomas than IDH1wt gliomas; (v) high circulating TF correlates with increased VTE risk; (vi) patient-derived glioma cells with endogenous IDH1mut produce smaller and fewer venous thrombi than IDH1wt gliomas in xenograft mouse models; (vii) suppression of TF in IDH1wt gliomas greatly reduces their in vitro and in vivo malignancy; (viii) patients whose gliomas express low TF have more than double the median survival of patients whose gliomas express high TF, independent of IDH1mut. Thus, we hypothesize that methylation- induced suppression of TF is a critical determinant of the less thrombogenic, and less malignant, IDH1mut phenotype. In Aim 1, we will conclusively establish that F3 hypermethylation is the mechanism by which IDH1mut suppresses TF expression. In Aim 2, we will modulate the expression of TF in a series of patient-derived IDH1wt and IDH1mut glioma cells, observing the effects on tumor-induced thrombosis and malignancy in cell cultures and in engrafted mice. In Aim 3, we will use molecular and pharmacologic approaches to investigate the therapeutic potential of blocking TF-PAR2 signaling in gliomas. Further, we will prospectively evaluate the utility of determining circulating TF levels, along with other clinical, blood-based, and tissue-based biomarkers, to create the first predictive model of VTE risk in glioma patients. In total, this research will greatly advance our understanding of IDH1mut tumor biology, and it will inform regarding novel treatment and diagnostic strategies for improving glioma patient care.