circ-GOT1.4调控GOT1表达致IDH1野生型和突变型胶质母细胞瘤糖代谢改变及预后差异的机制研究

Glioblastoma multiforme (GBM) has high malignancy and poor prognosis. GBM with mutant isocitrate dehydrogenase 1 (IDH1) has been found to have a better prognosis than with wild-type, but the mechanism is unclear. Bioinformatic analysis indicated that the expression of glutamate oxaloacetate transaminase 1 (GOT1), the key enzyme of glycometabolism in IDH1 mutant GBM was higher than that in IDH1 wild-type GBM, and the survival time of IDH1 mutant GBM with high expression of GOT1 was longer and cell proliferation was slower. It suggested that the high expression of GOT1 in IDH1 mutant GBM inhibited the glycometabolism and proliferation of GBM cells. Further analysis showed that circ-GOT1.4 could regulate GOT1 through competitive endogenous RNA (ceRNA) mechanism. Preliminary experiments confirmed that circ-GOT1.4 was over-expressed in IDH1 mutant GBM than in IDH1 wild-type GBM. Therefore, we propose the hypothesis that circ-GOT1.4, which is highly expressed in IDH1 mutant GBM, can up-regulate GOT1 by ceRNA, inhibit the glycometabolism and proliferation of GBM cells, and result in a better prognosis of mutant than that of wild type. In this study, we will take IDH1 mutant GBM cells and wild-type GBM cells as objects to study the mechanism of GOT1 affecting GBM glycometabolism and its regulation by circ-GOT1.4 in vivo and in vitro. We will reveal the reasons for the difference of prognosis between IDH1 mutant and wild-type GBM cells, and provide new ideas and targets for clinical adjuvant therapy.

胶质母细胞瘤（GBM）恶性程度高、预后差，其中异柠檬酸脱氢酶1（IDH1）突变型GBM预后好于野生型，但机制不明。预实验证实IDH1突变型中糖代谢关键酶谷氨酸草酰乙酸转氨酶1（GOT1）表达高于野生型，高表达GOT1的IDH1突变型GBM生存时间长、细胞增殖慢，提示IDH1突变型GBM中GOT1高表达抑制肿瘤细胞糖代谢和增殖，后还发现circ-GOT1.4可通过竞争性内源RNA（ceRNA）机制调控GOT1，且其表达在IDH1突变型较高。由此我们提出IDH1突变型中高表达的circ-GOT1.4通过ceRNA上调GOT1，抑制肿瘤糖代谢和增殖，造成突变型预后优于野生型的假说。本课题拟以IDH1突变型和野生型GBM细胞为对象，通过体内外实验对GOT1影响GBM糖代谢与其受circ-GOT1.4调控的机制进行研究，揭示IDH1突变型和野生型GBM预后差异原因，为临床辅助治疗提供新思路和靶点。

胶质母细胞瘤（GBM）恶性程度高，易复发，预后差。目前，最大限度手术全切除合并术后同步放化疗是标准治疗手段，但实际疗效仍不理想，死亡率高。GBM的增殖、复发与异常糖代谢有关，异柠檬酸脱氢酶1（IDH1）是糖代谢中的关键酶，且已知IDH1突变型GBM预后好于IDH1野生型，但具体机制不明，故揭示两者预后差异原因，有助于为临床辅助治疗提供新思路。前期实验已证实谷氨酸草酰乙酸转氨酶1（GOT1）在IDH1突变型GBM中的表达高于IDH1野生型，而circ-GOT1.4在IDH1突变型GBM中表达高于IDH1野生型，且其可通过竞争性内源RNA（ceRNA）机制调控GOT1。在此基础上，本项目按计划首先执行了对GOT1 在IDH1 野生型和突变型GBM 中的表达、与肿瘤细胞增殖的关系以及和患者预后间关系的研究，其中包括了研究GOT1 在IDH1 野生型和突变型GBM 中的表达，检测两种不同病理分型GBM 中，GOT1 表达改变对关键代谢物质浓度的影响、肿瘤细胞增殖能力以及可能的主要代谢途径的改变。其次，本项目验证了 circ-GOT1.4 通过ceRNA 机制结合miR-4448，间接调控GOT1 表达，继而影响GBM 细胞糖代谢和增殖的具体机制。随后，在分别调控了IDH1野生型和突变型GBM细胞中circ-GOT1.4 和GOT1的表达后，将这些细胞分组种植于裸鼠颅内，检测了各组成瘤组织的增殖、代谢以及裸鼠生存时间的差异。一系列的研究结果揭示了GOT1的表达变化是导致IDH1野生型和突变型GBM预后差异的可能原因，明确了circ-GOT1.4调控GOT1表达的具体机制，验证了IDH1突变型中高表达的circ-GOT1.4通过ceRNA上调GOT1，抑制肿瘤糖代谢和增殖，造成突变型预后优于野生型的假说。这些成果为将来临床上判断和改善GBM患者预后提供新的理论依据和实验基础，为药物辅助治疗GBM提供了新的靶点。