Fig. 3: FGFR2-mediated reprogramming of glucose metabolism requires NF-κB.

a Downregulated transcriptional regulators in ICC models upon FGFR (±ERBB) inhibition based on the TRRUST database. Treatments: ICC13-7: 75 nM futibatinib or DMSO (24 h); ICC10-6 and ICC21: DMSO, 100 nM infigratinib, 100 nM afatinib, or the combination (4 h); PDX-MG212: 1 mg per kg pemigatinib or vehicle (11 days); PDX-MG69: 25 mg per kg futibatinib or vehicle (14 days). b Relative mRNA levels of NF-κB targets in ICC13-7 cells treated with 75 nM futibatinib or DMSO for 4 h (n = 3 biological replicates). c Normalized NF-κB-dependent luciferase activity in ICC13-7 cells treated 24 h with vehicle or 100 nM infigratinib (n = 3 biological replicates). d, e Immunoblot of NF-κB pathway proteins in ICC13-7 cells treated with DMSO or 100 nM infigratinib for the indicated times (d), and in MG69 treated with vehicle or infigratinib for 10 days (e). f–h Phosphoproteomics on ICC13-7 cells treated with vehicle or FGFRi. f Schematic of study. Created with BioRender.com. g Pathway enrichment analysis (Hallmarks database) shows that futibatinib (75 nM, 4 h) downregulates Ser/Thr phosphorylation of NF-κB components in ICC13-7 cells. h Volcano plot of Ser/Thr phosphoproteome changes in ICC13-7 cells treated with 75 nM futibatinib for 4 h. Red dots: significantly downregulated phosphosites on NF-κB components. i Relative mRNA expression of the indicated glycolytic genes in ICC13-7 cells transfected with siRNA against RELA or control (n = 3 biological replicates) (i), and in ICC21 cells treated with 5 µM TPCA-1 or DMSO for 24 h (n = 3 biological replicates) (j). k, l Immunoblot of ICC13-7 cells treated with siRNA against RELA or control (k), or with 5 µM TPCA-1 or DMSO for 24 h (l). m–o Relative changes of glucose uptake (n = 4 biological replicates) (m), lactate secretion (n = 8 samples for siScramble; n = 4 samples for siRELA) (n), and ECAR (n = 6 samples for siScramble; n = 7 samples for siRELA) (o) in ICC13-7 cells treated with siRNA against RELA or control. p, q Relative changes in glucose uptake (n = 4 biological replicates) (p) and ECAR (n = 4 samples for DMSO; n = 3 samples for TPCA-1) (q) in ICC13-7 cells treated with 5 µM TPCA-1 or DMSO for 24 h. For mouse experiments, n = 3 per group. Phosphoproteomics was performed in duplicate. Bar graphs: data represent means ± SD. Student’s t-test (two-tailed) was performed. Western blots were repeated three times. Source data are provided as a Source Data file.