Difluoromethylornithine modulates mutant K-RAS signaling pathways in pancreatic cancer cells

Abstract Oncogenic mutations in the K-RAS small GTPase occur in more than 90% of human PDACs and cause dysregulation of multiple metabolic pathways, including polyamine metabolism. The goal of this study was to determine whether blocking polyamine synthesis with the irreversible inhibitor α-difluoromethylornithine (DFMO) provides a valid approach to targeting metabolic dependency in mutant K-RAS pancreatic tumors, compared with direct suppression of the oncogenic K-RAS protein. The anti-tumor activity of DFMO was evaluated in vitro and in vivo in the MIA PaCa-2 pancreatic cell line ( K-RAS G12C ) and its isogenic clonally selected line M-27, which does not express a functional GTP-bound RAS protein. MIA PaCa-2 and M-27 cells had comparable intracellular polyamine levels, and DFMO treatment effectively suppressed total polyamine levels in both cell lines ( p < 0.01). DFMO reduced the protein levels of phosphorylated extracellular signal-regulated kinases 1 and 2 (ERK1/2) and RAS-like small GTPase A (RalA) but induced the phosphoinositide 3-phosphate kinase (PI3K) signaling pathway. Additionally, the intracellular levels of K-RAS and hypusinated eukaryotic initiation factor 5 A (eIF5A Hyp ) proteins were suppressed in DFMO-treated MIA PaCa-2 cells. DFMO therapy regimen (500 mg/kg/day by oral gavage) significantly improved survival rates of SCID mice with orthotopically implanted MIA PaCa-2 cell line (MIA PaCa-2 control vs. MIA PaCa-2 DFMO, p = 0.0184), with median survival time comparable to animals implanted with M-27 cell line (MIA PaCa-2 control vs. M-27 control, p = 0.0204). DFMO treatment did not significantly improve survival of mice bearing M-27 tumors (M-27 control vs. M-27 DFMO, p = 0.6055). These data indicate that DFMO treatment suppresses pancreatic tumorigenesis driven by the mutant K-RAS oncogene similarly to the targeted elimination of the active GTP-bound RAS protein.