Formate overflow drives toxic folate trapping in MTHFD1 inhibited cancer cells

Alanna C. Green, Petra Marttila, Nicole Kiweler, Christina Chalkiadaki, Elisée Wiita, Victoria Cookson, Antoine Lesur, Kim Eiden, François Bernardin, Karl S.A. Vallin, Sanjay Borhade, Maeve Long, Elahe Kamali Ghahe, Julio J. Jiménez-Alonso, Ann Sofie Jemth, Olga Loseva, Oliver Mortusewicz, Marianne Meyers, Elodie Viry, Annika I. JohanssonOndřej Hodek, Evert Homan, Nadilly Bonagas, Louise Ramos, Lars Sandberg, Morten Frödin, Etienne Moussay, Ana Slipicevic, Elisabeth Letellier, Jérôme Paggetti, Claus Storgaard Sørensen, Thomas Helleday*, Martin Henriksson*, Johannes Meiser*

*Corresponding author for this work

Research output: Contribution to journalArticleResearchpeer-review

9 Citations (Scopus)


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.

Original languageEnglish
Pages (from-to)642-659
Number of pages18
JournalNature Metabolism
Issue number4
Early online date3 Apr 2023
Publication statusPublished - 20 Apr 2023


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