Wang, X.; Ding, L.; Sun, S.; Loo, S. K.; Chen, L.; Li, T.; Wang, M.-T.; Pennathur, A.; Huang, Y.; Gao, S.-J.

Cytosolic arginine sensor for mTORC1 Subunit 1 (CASTOR1) functions as a key regulator of mechanistic target of rapamycin complex 1 (mTORC1) signaling. Despite its frequent dysregulation in cancers via mechanisms such as KSHV microRNA-mediated inhibition or AKT-driven phosphorylation and degradation, the impact of CASTOR1 loss on tumor initiation and progression remains poorly understood. Here, we identify CASTOR1 as a critical tumor suppressor in non-small cell lung cancer (NSCLC) by demonstrating that its genetic ablation amplifies tumorigenesis in a KRAS-driven genetically engineered mouse model (GEMM;LSL-KRASG12D). CASTOR1 deficiency markedly enhances lung tumor incidence, accelerates tumor progression, and increases proliferative indices in KRASG12D-driven tumors (KRASG12D;C1KO) compared to CASTOR1 wild type (WT) tumors (KRASG12D;C1WT). Advanced-stage tumors exhibit elevated phosphorylated CASTOR1 (pCASTOR1) and reduced total CASTOR1 levels, suggesting active degradation during tumorigenesis. Mechanistically, CASTOR1 loss amplifies mTORC1 signaling, as evidenced by heightened phosphorylation of downstream effectors 4EBP1 and S6, while also augmenting AKT and ERK activation, uncovering a crosstalk between the PI3K/AKT/mTORC1 and KRAS/ERK pathways. Furthermore, CASTOR1 ablation induces genome instability, which may contribute to enhanced tumor incidence and progression. Importantly, CASTOR1 deficiency confers resistance to KRASG12D-specific inhibitors, while over half of KRASG12D;C1WT tumors also display resistance. Organoids derived from KRASG12D;C1KO and KRASG12D;C1WT tumors reveal a correlation between KRAS inhibitor resistance and hyperactivation of mTORC1, with mTORC1 and PI3K inhibitors sensitizing resistant tumors to KRASG12D-targeted therapies. These findings position CASTOR1 as a novel tumor suppressor that modulates mTORC1 and KRAS signaling to constrain NSCLC progression. Our study further highlights the therapeutic potential of combining mTORC1 or ERK inhibitors with KRAS-targeted therapies for NSCLC characterized by hyperactive KRAS signaling and impaired CASTOR1 activity.\n\nHighlightsO_LICASTOR1 functions as a tumor suppressor in NSCLC by limiting KRAS-driven tumor initiation and progression.\nC_LIO_LICASTOR1 is frequently lost or inactivated in wild-type tumors during tumor progression, contributing to advanced-stage malignancies.\nC_LIO_LICASTOR1 deficiency amplifies mTORC1 signaling and enhances PI3K/AKT and KRAS/ERK crosstalk, driving tumorigenesis and resistance to KRAS-specific inhibitors.\nC_LIO_LICombining mTORC1 or PI3K inhibitors with KRAS-targeted therapies effectively overcomes resistance in KRAS-driven NSCLC.\nC_LI