SREBP-driven lipogenesis coupled with ferroptosis resistance as a therapeutic target in PTEN-loss extrahepatic cholangiocarcinoma.

Extrahepatic cholangiocarcinoma (eCCA) is an aggressive biliary tract cancer (BTC) with limited therapeutic options and actionable genetic alterations. Although PTEN loss and PI3K pathway activation are common in eCCA, their roles in disease pathogenesis and therapeutic vulnerability remain unclear. Therefore, we aimed to establish clinically relevant mouse models of PTEN loss-driven eCCA and identify novel therapeutic strategies. To investigate eCCA development and the underlying mechanisms, we generated CK19-CreERT-based genetically engineered mouse models with biliary epithelial cell-specific PTEN deletion combined with either TGFβR2 loss or oncogenic Kras activation. PTEN loss alone was insufficient for tumorigenesis, whereas combined deletion of PTEN and TGFβR2 induced periductal infiltrating-type eCCA originating from the peribiliary glands. In contrast, PTEN loss and oncogenic Kras activation predominantly generated intraductal-growing tumors with broader biliary involvement. These included intrahepatic cholangiocarcinoma (iCCA) and gallbladder cancer (GBC). RNA sequencing revealed that SCAP/SREBP-mediated lipogenesis was consistently activated in the PTEN loss-driven eCCA models. Genetic ablation of SCAP markedly suppressed eCCA development (p < 0.0001) but exacerbated iCCA, indicating the need for alternative therapeutic approaches. SREBP-driven lipogenesis increased the synthesis of polyunsaturated phospholipids and concomitantly induced GPX4, thereby conferring resistance to ferroptosis. GPX4 inhibition suppressed eCCA (p = 0.002) and GBC development without exacerbating iCCA. Consistently, PTEN-deficient human BTC cell lines, including CRISPR-Cas9-engineered models, exhibited increased vulnerability to GPX4 inhibition. Furthermore, clinical eCCA samples (n = 182) confirmed the coordinated activation of the PI3K, SREBP, and ferroptosis pathways. Distinct oncogenic alterations shape the histological and anatomical diversity of BTC. PTEN loss-driven eCCA critically depends on SCAP/SREBP-mediated lipid reprogramming and GPX4-dependent ferroptosis resistance, indicating the therapeutic potential of ferroptosis induction. This study provides a scientific rationale for the stratification of extrahepatic cholangiocarcinoma (eCCA) based on oncogenic context. Our findings demonstrate that PTEN loss is associated with shared lipid metabolic features, whereas additional genetic alterations determine histological growth patterns and anatomical distribution. We identified ferroptosis resistance as a key downstream feature of PTEN-deficient eCCA. Although further validation is required, these findings provide a theoretical basis for further exploration of ferroptosis-based therapeutic approaches for genetically defined subsets of biliary tract cancer.