MTHFD2调控肿瘤代谢在结直肠癌发生发展中的作用与机制

Methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) serves as a key enzyme of one-carbon metabolism, in which oxidation of methylene tetrahydrofolate to 10-formyl-tetrahydrofolate is coupled to reduction of NADP+ to NADPH. With high-throughput reverse phase protein microarray technology analysis, our recent study show that high MTHFD2 expression was most significantly associated with overall survival in colorectal patients with poor prognosis. Howerver, the function and regulated mechanism of MTHFD2 is still unknown. Based on the important role of MTHFD2 in cancer metabolism, the preliminary experiment results show that MTHFD2 inhibition decrased the colorectal cell proliferation in vitro and tumorigenicity ability in vivo. Also, depletion of MTHFD2 resulted in decreased cellular NADPH/NADP+ and GSH/GSSG ratios and increased cell sensitivity to oxidative stress. Based on these findings, we aim to investigate 1) the function and clinical significance of MTHFD2 on colonrectal tumorigenesis, 2) how MTHFD2 regulates NADPH homeostasis to promote tumour cell survival during energy stress, 3) and to explore the signalling pathway and underlying molecular mechanism involed in MTHFD2 regulation. Our study may reveal a novel function of MTHFD2, provide a new predictive marker and therapeutic target for colorectal cancer diagnosis and treatment.

亚甲基四氢叶酸脱氢酶（MTHFD2）是细胞一碳单位代谢过程中以NADP+为氢受体催化亚甲基四氢叶酸生成甲酰基四氢叶酸，并生成NADPH的关键酶。我们近期通过高通量反相蛋白芯片技术分析发现，在结直肠癌肿瘤组织中MTHFD2表达与总生存联系最显著，MTHFD2高表达的结直肠患者预后明显较差，但具体功能和机制不明。我们的预实验结果表明，下调MTHFD2的表达抑制了结直肠癌细胞体外增殖和体内成瘤的能力，且MTHFD2可维持结直肠癌细胞内NADPH介导的氧化还原稳态，减少活性氧引发的细胞凋亡。基于此，本项目拟探讨：1）MTHFD2对结直肠癌发生发展的影响及其临床意义; 2）明确MTHFD2在能量应激的条件下调控NADPH水平和氧化还原稳态促进细胞存活的作用及其机制；3）探索调控MTHFD2异常表达的相关信号通路和分子机制，期望为结直肠癌的诊疗提供新的预测指标和治疗靶点。

肿瘤细胞中的ROS是一把“双刃剑”，适度升高的ROS可以促进细胞的生长增殖，而ROS过度增加能引起蛋白质和脂质分子氧化损伤。ROS水平主要决定于ROS产生和抗氧化系统的动态平衡，而NADPH是细胞内抗氧化防御系统的重要组成成分，作为还原性谷胱甘肽（GSH）的辅酶在细胞应对氧化应激防御活性氧损伤方面起着重要的作用。因此，阐明肿瘤细胞及肿瘤微环境中NADPH代谢的调控机理，开发靶向代谢通路的治疗策略，对包括结直肠癌在内的恶性肿瘤具有重要的理论意义和临床应用价值。. 本项目通过系统分析生成NADPH分子相关限速酶的表达，并通过实验证实叶酸代谢通路中的MTHFD2是维持结直肠癌NADPH稳态的关键分子。MTHFD2在细胞中以NADP+为氢受体，催化亚甲基四氢叶酸（CH2-THF）生成甲酰基四氢叶酸（CHO-THF）和NADPH，在细胞核酸合成和应激反应中发挥着关键作用。该研究进一步通过临床样本分析发现MTHFD2在结直肠癌组织中表达上调，且高表达MTHFD2的结直肠患者预后明显较差。在低氧或悬浮培养的条件下，MTHFD2可以维持细胞内NADPH和GSH的水平，抵御活性氧诱发的细胞死亡，促进肿瘤在裸鼠体内的增殖和转移。同时证实新型的叶酸类似物LY345899，作为MTHFD2的抑制剂在体内外实验中都表现出较好的抗肿瘤活性，可抑制结直肠癌患者的肿瘤组织（PDX）在裸鼠体内的生长和转移。机制研究发现，癌基因Kras可以通过下游的Akt 及ERK信号通路，激活转录因子c-Myc，上调MTHFD2的mRNA和蛋白的表达水平。. 该工作首次揭示了叶酸代谢通路中的关键酶MTHFD2在维持结直肠癌细胞内的NADPH稳态及防御活性氧损伤，促进肿瘤的恶性进展过程中的关键作用及调控机制；同时通过体内外实验证明MTHFD2可作为结直肠癌新的预后预测标志物和潜在的治疗靶点，其抑制剂具有潜在的药物开发价值。