Small molecule induced KRAS activation triggers ferroptosis in mutant KRAS expressing PDAC

Pancreatic ductal adenocarcinoma (PDAC) is the most prevalent histological subtype of pancreatic cancer with limited treatment options and poor prognosis. More than 90% of PDAC harbor mutations in the oncogene KRAS and are dependent on downstream pro-survival and/or proliferative factors such as AKT, MEK and MAPK. As such, targeting these downstream pathways has been employed as a therapeutic strategy, however, recent success in developing KRAS-specific inhibitors has added an important dimension in the management of PDAC. The clinical efficacy of the direct KRAS inhibitors has been less than optimal, which circumvents the need for identifying new therapeutic strategies. To that end, we recently described a novel approach using a small molecule that induced KRAS activation-dependent, ROS-mediated, execution of mutant KRAS driven cancer cells ( Iskandar, K. et al. Autophagy 2024 ). Here we investigated the effect of this small molecule (merodantoin; C1) on mutant KRAS expressing PDAC. Results show that C1 selectively targets PDAC cells (cytotoxicity, colony formation and spheroid formation) harboring G12 KRAS mutation compared to wild-type KRAS expressing cells. Mechanistically, C1 treatment led to significant increase in both intracellular and mitochondrial reactive oxygen species (ROS), increase in labile iron levels due to increased transferrin receptor 1 (TfR1) and decreased ferritin heavy chain (FTH1), and disruption of the glutathione-based antioxidant system by reducing cystine uptake and decreasing Glutathione peroxidase 4 (GPX4) expression. These changes together triggered lipid peroxidation, a key sign of ferroptosis. Experiments using ferroptosis inhibitors, iron chelators, and ROS scavengers restored cell viability, confirming that C1 triggered cell death in PDAC cells by ferroptosis. Importantly, genetic and pharmacological modulation of KRAS provide evidence that mutant KRAS is upstream of ferroptotic cascade. Moreover, KRAS mutations were shown to make PDAC cells more sensitive to ferroptosis by naturally lowering their antioxidant defences and iron storage capacity. This was evident in the reduced baseline expression of GPX4 and SLC7A11, along with decreased cystine uptake in mutant KRAS expressing cell lines compared to wild-type controls. Beyond in vitro validation, C1 demonstrated strong effectiveness in patient-derived PDAC organoids and significantly limited tumor growth in a orthotopic PDAC mouse model. Evidence presented here strongly argue in support of targeting ferroptosis vulnerability of KRAS-mutant PDAC as a novel treatment strategy.