Chemical Probes Targeting Gliomas with IDH Mutation

描述（申请人提供）：异柠檬酸脱氢酶（IDH）是三羧酸循环中的关键酶之一，利用NADP+催化异柠檬酸转化为α-酮戊二酸（α-KG）。大量人类胶质瘤的DNA测序显示，约75%的中低级别胶质瘤和继发性多形性胶质母细胞瘤（GBM，一种高致死性脑肿瘤）携带IDH突变，其中IDH1 R132H突变占主导地位（约90%）。遗传学研究发现，IDH突变总是杂合的，这表明野生型酶对正常细胞和肿瘤细胞都是必需的，IDH突变是胶质瘤发生的早期和关键事件。突变蛋白的生化表征表明，它们在将异柠檬酸转化为α-KG方面没有活性。相反，所有的突变蛋白，如IDH1(R132H)，都具有新的酶活性：它们利用NADPH催化α-KG还原为d -2-羟基戊二酸（2-HG）。这种新功能是导致携带IDH突变的原发肿瘤细胞中2-HG水平显著升高（约100倍）的原因。越来越多的证据有力地支持2-HG是一种肿瘤代谢物，而突变型IDH是干预的目标。虽然高水平的2-HG对人体健康（尤其是中枢神经系统）非常有害，但抑制突变型IDH是否会阻断这类肿瘤细胞的生长或诱导其凋亡仍有待解决。这是因为到目前为止还没有突变型IDH的抑制剂。此外，RNA干扰（RNAi）也不适合用于本研究，因为它也会敲低野生型IDH酶，其功能对正常细胞和肿瘤细胞都至关重要。第一个具体目标是利用药物化学、蛋白质x射线晶体学和定量构效关系（QSAR）来开发有效的、选择性的IDH1（R132H）抑制剂。第二个特异性目的是测试在Aim 1中合成的化合物的体外生物活性以及所选化合物的药代动力学和毒理学特性。第三个特定目标是确定我们的强效IDH1（R132H）抑制剂在脑内异种移植小鼠模型中的体内抗肿瘤活性，并使用分子和细胞生物学方法确定这些新型抑制剂的作用机制。

DESCRIPTION (provided by applicant): Isocitrate dehydrogenase (IDH) is one of the key enzymes in the tricarboxylic acid cycle, catalyzing the conversion of isocitric acid to α-ketoglutaric acid (α-KG) using NADP+. DNA sequencing of a large number of human gilomas has revealed that ~75% of low - medium grade gliomas and secondary glioblastoma multiforme (GBM, a highly lethal form of brain tumor) carry IDH mutations, with IDH1 R132H mutation being predominant (~90%). Genetic investigations have found that IDH mutations are always heterozygous, suggesting the wild-type enzyme is essential for both normal and tumor cells, and IDH mutation is an early and critical event in the glioma tumorigenesis. Biochemical characterization of the mutant proteins revealed that they are inactive in converting isocitrate to α-KG. Rather, all of the mutant proteins, e.g., IDH1(R132H), possess a new enzymatic activity: they catalyze the reduction of α-KG to D-2-hydroxyglutaric acid (2-HG) using NADPH. This neo-function is responsible for the greatly elevated (>100x) level of 2-HG in primary tumor cells bearing IDH mutations. Growing evidence strongly supports 2-HG is an onco-metabolite and mutant IDH a target for intervention. Although it is clear that a high level of 2-HG is very harmfu to human health (especially central nervous system), whether inhibition of mutant IDH blocks the growth or induces apoptosis of this type of tumor cells remains to be answered. This is because there have been no inhibitors of mutant IDH to date. In addition, RNA interference (RNAi) is not appropriate for this study, since it will also knock down the wild-type IDH enzyme, whose function is essential for both normal and tumor cells. The first Specific Aim is to use medicinal chemistry, protein X-ray crystallography and quantitative structure activity relationship (QSAR) to develop potent and selective inhibitors of IDH1(R132H). The second Specific Aim is to test in vitro biological activities of compounds synthesized in Aim 1 as well as pharmacokinetic and toxicological properties of selected compounds. The third Specific Aim is to determine the in vivo antitumor activity of our potent IDH1(R132H) inhibitors in intra-cerebral xenograft mouse models as well as to use molecular and cell biology methods to identify the mechanisms of action of these novel inhibitors.