Fig 2: Low glucose and Mg2+ levels are required for anticancer activity by an allosteric IDH1 inhibitor (MiaPaCa-2 cells).a, ROS levels were detected by DCFDA assay in MiaPaCa-2 pancreatic cancer cells treated with AG-120 (1 µM) for 48 h under the indicated conditions and cultured in medium containing 2.5 mM glucose (n = 3 independent experiments). b,c, OCR in MiaPaCa-2 PDAC cells treated with vehicle or AG-120 cultured in medium containing either 0.80 mM Mg2+ and 2.5 mM glucose (b) or 0.08 mM Mg2+ and 2.5 mM glucose (c) for 30 h (representative experiments of three independent biological replicates with similar results are shown). d,e, Total pool size, including glucose-independent (m + 0) and glucose-dependent isotopologs (d) and isotopolog distribution (e) in cells cultured with unlabeled 2.5 mM glucose for 38 h followed by 2.5 mM [U-13C]glucose for an additional 10 h (n = 3 individual biological replicates). f, Relative clonogenic growth of cells treated with vehicle or AG-120 for 4 days under different levels of glucose and Mg2+ (n = 3 independent biological replicates). g, Relative clonogenic growth of indicated cells treated with vehicle or AG-120 for 4 days under 2.5 mM glucose and 0.08 mM Mg2+ (n = 3 independent biological replicates). Data provided as mean ± s.d. Pairwise comparisons were conducted using two-tailed, unpaired Student’s t-tests.Source data

Fig 3: IDH1-/- cells have impaired mitochondrial function.a, Oxygen consumption rates (OCR) in MiaPaCa-2 cells cultured under the indicated glucose conditions for 30 hours (a representative of three independent biological replicates with similar results is shown). b, ATP levels in MiaPaCa-2 cells cultured under the indicated conditions for 24 hours (n = 3 independent biological replicates). c, Basal OCR in Hs766T pancreatic cancer cells cultured in 1 mM glucose for 24 hours (a representative of two independent biological replicates with similar results is shown). Western blot of IDH1 expression (representative immunoblots of two independent biological replicates with similar results are shown). d, Mitochondrial mass and cell viability in MiaPaCa-2 cells cultured with 2.5 mM glucose for 30 hours (n = 4 independent biological replicates). e, Relative abundance of the indicated TCA-related metabolites in MiaPaCa-2 cells cultured with unlabeled 2.5 mM glucose for 38 hours followed by unlabeled αKG (4 mM) or sodium citrate (4 mM) for 10 hours (n = 3 individual biological replicates). f, g, Total pool size (f) and isotopologue distribution (g) of indicated metabolites in cells cultured with unlabeled 25 mM glucose for 38 hours followed by 25 mM [U-13C]glucose for an additional 10 hours (n = 3 individual biological replicates). Data are provided as mean ± s.d. (a-c,e-g) or mean ± s.e.m. (d). Pairwise comparisons were conducted using two-tailed, unpaired Student’s t-tests. Source data

Fig 4: The impact of magnesium on pancreatic cancer cell metabolism and allosteric inhibitor binding to IDH1.a, Wild-type IDH1 from the Protein Database. Mg2+ is believed to interact with D279 (Asp279) in the allosteric pocket (accession number: PDB 1T0L). Under high Mg2+ conditions, the cation outcompetes the allosteric inhibitor, to render the drug ineffective. Under low Mg+2 conditions, the drug inhibits wtIDH1. b, The effect of AG-120 on IDH2 activity (cell-based assay) in MiaPaCa-2 PC cells (a representative of three independent biological replicates with similar results is shown. c, Validation of the fluorescent probe binding to wtIDH1 (n = 2 independent experiments). d, e, ATP levels (d, n = 3 independent biological replicates) and oxygen consumption rates (e, a representative of three independent biological replicates with similar results is shown) in MiaPaCa-2 cells under the indicated conditions. f, Cell viability of PDAC cells, cultured in MgSO4 at the indicated concentrations and 25 mM glucose for five days (n = 3 independent biological replicates). g, The impact of the indicated conditions and treatments on genes involved in magnesium homeostasis in MiaPaCa-2 cells (n = 3 individual biological replicates). Data are provided as mean ± s.d. (b-f). Pairwise comparisons were conducted using two-tailed, unpaired Student’s t-tests. Source data

Fig 5: AG-120 activity against KPC cells.a, Sanger sequencing of amplicons correlating with codon 132 of the wtIDH1 gene in KPC murine pancreatic cancer cells (K8484). The reference murine wild-type sequence is shown. b, IDH1 activity of KPC cells treated with AG-120 for 24 hours in media containing 0.08 mM Mg2+ (n = 3 independent biological replicates). c, Relative clonogenic growth of KPC cells under the indicated conditions for 5 days (n = 3 independent biological replicates). d, Relative clonogenic growth of KPC cells with the indicated treatments, cultured with 1 mM glucose and 0.08 mM Mg2 for five days (n = 3 independent biological replicates). e, Growth of KPC subcutaneous xenografts in C57BL/6 J mice under the indicated treatment arms (vehicle, n = 5 tumors; AG-120, n = 4 tumors). f, g, CT/18F-FDG-PET images (f) and SUV values (g) of C57BL/6 J mice bearing orthotopic KPC tumors treated with vehicle or AG-120 (n = 5 mice per group). h, Survival analysis in tamoxifen-inducible KP-/-C (KrasG12D/+; Trp53lox/lox; Pdx1-Cre) mice treated with vehicle (n = 6 mice) or AG-120 (n = 7 mice). Data are provided as mean ± s.d. (b-d) or mean ± s.e.m. (e,g). Pairwise comparisons were conducted using two-tailed, unpaired Student’s t-tests. Longitudinal mixed models were fit for tumor size growth, and time by treatment interactions were assessed. Survival data are represented by Kaplan-Meier curves, and tests for treatment differences were conducted with Fleming-Harrington (0,1) weighted log rank test statistics. Source data