Figure 4. Pharmacological inhibition of nucleotide metabolism modulates ferroptosis.

(A) Protein expression in HT-1080 Control cells treated with mycophenolic acid (MPA, 5 μM), pyrazofurin (Pyf, 5 μM), or Nutlin-3 (10 μM). (B) Cell death in HT-1080^N Control, p53^KO and p21^KO cells pretreated for 48 h with mycophenolic acid (5 μM, left) or pyrazofurin (5 μM, right) ± buthionine sulfoximine (BSO, 100 μM), then treated with erastin2 (Era2, 1 μM, 24 h). (C) Protein expression in HT-1080 Control cells treated with DMSO, gemcitabine (Gem, 200 nM), hydroxyurea (HU, 5 mM), or nutlin-3 (10 μM). (D) Cell death in HT-1080^N Control, p53^KO or p21^KO cells pretreated with hydroxyurea (5 mM), gemcitabine (200 nM), or nutlin-3 (10 μM), then ± era2 for 24 h. (E) Cell death in HT-1080 Control cells pretreated with gemcitabine (Gem, 200 nM) ± buthionine sulfoximine (100 μM), then treated with Era2 (1 μM, 24 h). (F) Cell death in HT-1080 Control cells pretreated with ± gemcitabine (200 nM) for 48 h then treated ± ML162 (1 μM) ± ferrostatin-1 (1 μM). (G) Metabolic flux assay using LC-MS detection of glutathione in HT-1080 Control, p53^KO or p21^KO cells pretreated with DMSO or gemcitabine (200 nM) for 48 h. After pretreatment, medium containing Era2 (1 μM) and U-¹³C Serine was added to cells for 8 h. De novo synthesized glutathione is labeled as an m+2 isotopologue (yellow). Data information: In (B, D, E, F, G), data are mean ± SD of at least three independent experiments.