癌细胞通过上调一碳代谢（1C）来满足其对核苷酸供应的增加需求，包括酶甲烷四氢叶酸脱氢酶环化酶1和2（MTHFD1和MTHFD2）。TH9619是MTHFD1和MTHFD2中脱氢酶和环化酶活性的有效抑制剂，并且能选择性地杀死癌细胞。在这里，我们揭示了TH9619在细胞中靶向核MTHFD2但不抑制线粒体MTHFD2。因此，在TH9619的存在下，来自线粒体的甲酸溢出仍在继续。TH9619抑制线粒体甲酸释放下游发生的MTHFD1的活性，导致10-甲酰四氢叶酸的积累，我们称之为“叶酸陷阱”。这导致胸腺嘧啶耗尽并导致表达MTHFD2的癌细胞死亡。这种先前未表征的叶酸陷阱机制受生理性次黄嘌呤水平的加剧而恶化，次黄嘌呤阻断了新生嘌呤合成途径，并且还防止10-甲酰四氢叶酸用于嘌呤合成。这里描述的TH9619的叶酸陷阱机制与其他MTHFD1/2抑制剂和抗叶酸类药物不同。因此，我们的发现揭示了一种攻击癌症的方法，并揭示了1C代谢的调节机制。©2023年作者（们）。

Cancer cells fuel their increased need for nucleotide supply by upregulating one-carbon (1C) metabolism, including the enzymes methylenetetrahydrofolate dehydrogenase-cyclohydrolase 1 and 2 (MTHFD1 and MTHFD2). TH9619 is a potent inhibitor of dehydrogenase and cyclohydrolase activities in both MTHFD1 and MTHFD2, and selectively kills cancer cells. Here, we reveal that, in cells, TH9619 targets nuclear MTHFD2 but does not inhibit mitochondrial MTHFD2. Hence, overflow of formate from mitochondria continues in the presence of TH9619. TH9619 inhibits the activity of MTHFD1 occurring downstream of mitochondrial formate release, leading to the accumulation of 10-formyl-tetrahydrofolate, which we term a 'folate trap'. This results in thymidylate depletion and death of MTHFD2-expressing cancer cells. This previously uncharacterized folate trapping mechanism is exacerbated by physiological hypoxanthine levels that block the de novo purine synthesis pathway, and additionally prevent 10-formyl-tetrahydrofolate consumption for purine synthesis. The folate trapping mechanism described here for TH9619 differs from other MTHFD1/2 inhibitors and antifolates. Thus, our findings uncover an approach to attack cancer and reveal a regulatory mechanism in 1C metabolism.© 2023. The Author(s).