Novel Approaches to Modeling and Treating IDH1 Mutant Glioma

项目总结 异柠檬酸脱氢酶1(IDH1)基因热点突变在低级别胶质瘤中高度复发 和继发性胶质母细胞瘤(GBM)，并编码突变的IDH1-R132H酶。这种酶 显示新的活性，将2-氧代戊二酸(2OG)转化为肿瘤性代谢物(R)-2-羟基戊二酸 ((R)-2HG)。(R)-2HG在IDH1突变型胶质瘤中积聚到毫摩尔水平，并通过以下方式促进胶质瘤的形成 竞争性地调节依赖于20G的酶来调节神经胶质细胞的转化。尽管这些 这些发现深刻地重塑了我们对这些疾病分子发病机制的理解， 这些见解尚未转化为新的、有效的胶质瘤治疗策略的开发。 病人。突变的idh酶的直接抑制剂在早期临床前和临床上表现出较低的活性。 IDH突变型胶质瘤与它们对IDH突变型白血病显示的强大疗效相关的研究。这些 研究结果突显了临床治疗IDH1突变胶质瘤的替代治疗策略的必要性。我 假设可以通过利用IDH1的合成致死性来开发新的治疗策略- 癌基因R132H。我用同基因idh1突变体和野生型进行了化学合成致死性筛选。 并发现多种嘧啶合成抑制剂优先杀死IDH1突变细胞。 我的初步数据显示，(R)-2HG介导了对依赖于2OG的支链氨基酸的抑制 转氨酶BCAT1和BCAT2影响氮进入嘧啶生物合成途径。 因此，在目标1中，我将测试BCAT活性降低是否代表潜在的分子机制 突变型IDH1诱导的嘧啶合成抑制剂超敏反应。我会继续评估IDH1是否- R132H突变是对嘧啶合成抑制剂反应的预测生物标志物 这些药物对一组IDH1突变和野生型胶质瘤神经球系的细胞毒性。最后，我 将测试一种新型的嘧啶生物合成酶二氢罗酸脱氢酶的脑穿透抑制物 DHODH单独和联合放射治疗IDH1突变型胶质瘤的异种移植模型。另一个 阻碍开发IDH1突变型胶质瘤新的有效治疗策略的主要障碍是 这种疾病的基因工程小鼠(GEM)模型很少，可以用来评估这种疾病 战略。在目标2中，我建议建立第一个基于Crispr/Cas9的突变型胶质瘤GEM模型 IDH1。我已经成功地在小鼠身上产生了突变的IDH1驱动的III级间变性星形细胞瘤，我将 优化我的方法，在这个宝石模型中增加星形细胞瘤的透视率。我还将修改我的战略，以 建立IDH1突变型和野生型GBM的等基因GEM模型，可用于未来的体内研究 IDH1-R132H癌基因的合成致死性。如果成功，这些项目将为 用于IDH1突变型胶质瘤治疗的嘧啶合成抑制剂的临床试验和GEM模型的建立 这一疾病支持未来对新设计的治疗策略的测试。

PROJECT SUMMARY Hot-spot mutations in the isocitrate dehydrogenase 1 (IDH1) gene are highly recurrent in lower grade gliomas (LGGs) and secondary glioblastomas (GBMs) and encode the mutant IDH1-R132H enzyme. This enzyme displays neomorphic activity, converting 2-oxoglutarate (2OG) to the oncometabolite (R)-2-hydroxyglutarate ((R)-2HG). (R)-2HG accumulates to millimolar levels in IDH1 mutant gliomas and promotes gliomagenesis by competitively modulating 2OG-dependent enzymes that regulate glial cell transformation. Although these discoveries have profoundly reshaped our understanding of the molecular pathogenesis of these diseases, these insights have not yet translated to the development of new, effective therapeutic strategies for glioma patients. Direct inhibitors of the mutant IDH enzyme have shown poor activity in early preclinical and clinical studies of IDH mutant glioma relative to the robust efficacy they display against IDH mutant leukemias. These findings highlight the clinical need for alternative therapeutic strategies to treat IDH1 mutant gliomas. I hypothesize that novel treatment strategies can be developed by exploiting synthetic lethality with the IDH1- R132H oncogene. I performed a chemical synthetic lethality screen using isogenic IDH1 mutant and wild-type glioma cell lines and found that multiple pyrimidine synthesis inhibitors preferentially killed IDH1 mutant cells. My preliminary data shows that (R)-2HG-mediated inhibition of the 2OG-dependent branched chain amino acid transaminases BCAT1 and BCAT2 impairs nitrogen incorporation into the pyrimidine biosynthesis pathway. Therefore, in Aim 1 I will test whether reduced BCAT activity represents the molecular mechanism underlying mutant IDH1-induced pyrimidine synthesis inhibitor hypersensitivity. I will go on to assess whether the IDH1- R132H mutation is a predictive biomarker for response to pyrimidine synthesis inhibitors by testing the cytotoxicity of these agents against a panel of IDH1 mutant and wild-type glioma neurosphere lines. Finally, I will test a novel brain-penetrant inhibitor of the pyrimidine biosynthesis enzyme dihydroorotate dehydrogenase (DHODH) alone and in combination with radiotherapy in xenograft models of IDH1 mutant glioma. Another major impediment to the development of new, effective therapeutic strategies for IDH1 mutant glioma is a paucity of genetically-engineered mouse (GEM) models of this disease that can be used to evaluate such strategies. In Aim 2 I propose to establish the first Crispr/Cas9-based GEM models of glioma driven by mutant IDH1. I have successfully produced mutant Idh1-driven Grade III anaplastic astrocytomas in mice and I will optimize my approach to increase astrocytoma penetrance in this GEM model. I will also modify my strategy to generate isogenic GEM models of IDH1 mutant and wild-type GBM that can be used for future in vivo studies of synthetic lethality with the Idh1-R132H oncogene. If successful, these projects will establish rationale for clinical testing of pyrimidine synthesis inhibitors for IDH1 mutant glioma therapy and generate GEM models of this disease to support future testing of newly devised treatment strategies.