IDH1突变在急性T淋巴细胞性白血病中的作用及其机制

IDH1 is a cytoplasmic enzyme that catalyzes the NADP-dependent conversion of isocitrate to α-ketoglutarate (αKG). Mutations in IDH1 at arginine 132 (R132) cause an enzymatic gain of function that results in the NADPH-dependent conversion of αKG to D-2-hydroxyglutarate (2HG). Somatic mutations in isocitrate dehydrogenase 1 (IDH1) are frequently observed in a number of malignancies. In cells and tissues of patients with IDH1 mutant tumors, 2HG builds up to high levels and is thought to contribute to tumorigenesis as an oncometabolite. T-cell IDH1 is a cytoplasmic enzyme that catalyzes the NADP-dependent conversion of isocitrate to α-ketoglutarate (αKG). Mutations in IDH1 at arginine 132 (R132) cause an enzymatic gain of function that results in the NADPH-dependent conversion of αKG to D-2-hydroxyglutarate (2HG). Somatic mutations in isocitrate dehydrogenase 1 (IDH1) are frequently observed in a number of malignancies. In cells and tissues of patients with IDH1 mutant tumors, 2HG builds up to high levels and is thought to contribute to tumorigenesis as an oncometabolite. T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy of developing T-cells. The genetic landscape of the disease has been characterized, and a large number of driver mutations have been identified. The most common genetic feature of T-ALL is the presence of activating mutations in Notch1. The concurrent mutation of DNA methyltransferase 3A (Dnmt3a) together with Idh1 mutation was also found in T-ALL patients. Previous work has shown that Idh1 mutations or Notch1 mutations alone are not sufficient to give rise to T-ALL. However, the molecular mechanisms are poorly understood. Therefore, this project will generate Idh1/Notch1- and Idh1/Dnmt3a-mutated mouse models to study how Idh1 mutation cooperates with other genetic mutations to drive T-ALL development and also carry out preclinical assessment of novel inhibitors of mutant IDH1 in T-ALL.

IDH1是一种细胞质酶，因R132突变造成酶功能获得型突变，进而促使NADPH依赖的αKG反应生成D-2-羟戊二酸(2HG) ；在许多癌症中常能观察到该突变。在携带IDH1-R132突变体的肿瘤细胞和组织中，2HG呈现高水平；并且，2HG作为一种癌性代谢物，大量积累可能促进肿瘤发生发展。急性T淋巴细胞白血病（T-ALL）是源于T细胞的侵袭性恶性肿瘤；许多驱动突变能促使其发生发展，以Notch1激活突变最为常见，也见于Dnmt3a与Idh1并发突变。我们前期研究发现，单独的Idh1或Notch1突变均不足于导致T-ALL发生发展，但其机制尚不清楚。我们拟构建携带Idh1突变与Notch1或Dnmt3a突变的并发突变小鼠模型，阐明IDH1-R132与Notch1或Dnmt3a突变协同作用导致T-ALL发生发展的机制，同时评估IDH1-R132的新型抑制剂对T-ALL的疗效。

异柠檬酸脱氢酶（Isocitrate Dehydrogenase1，IDH1）在第132位精氨酸突变造成新功能获得型酶从而促使α-酮戊二酸反应生成被称为“癌代谢物” (oncometabolite) 的D-2-羟戊二酸(2HG)。在许多恶性肿瘤中，常常观察到IDH1的突变尤其在造血系统中。但IDH1突变对血液系统肿瘤的影响和作用机制有待进一步明确。我们前期的研究工作已经发现IDH1-R132突变小鼠发生发展的急性T淋巴细胞白血病（T cell-acute lymphoblastic leukemia, T-ALL）携带功能活化型Notch1突变。所以本课题研究IDH1-R132和Notch1双突变对T-ALL发生发展的作用和分子机制。我们选择携带Notch1突变的T-ALL细胞系Molt-4，利用CRISPR/Cas9系统的基因组编辑功能对IDH1-R132定点突变。通过细胞克隆，DNA测序和酶切电泳分析鉴定获得了携带IDH1(R132H+/-)杂合体的多个单细胞克隆。与文献报道和预期相吻合，IDH1(R132H+/-)突变细胞相较于野生型细胞2-HG水平显著升高。在功能上，突变细胞增殖和迁移能力显著下降并伴随凋亡增加。通过添加外源性2-HG（Octyl-(R)-2HG）实验，证明这些表型变化是由突变体细胞分泌的2-HG引起的。在动物体内水平上，我们通过免疫缺陷小鼠动物肿瘤生长曲线模型揭示IDH1(R132H+/-)突变抑制急性T淋巴细胞白血病肿瘤的生长。所以我们的研究结果揭示IDH1(R132H+/-)突变通过2-HG抑制了T-ALL细胞增殖及成瘤能力，这一发现对急性T淋巴细胞白血病病人的治疗和预后都具有指导意义。