Targeting Lung Cancer Stem Cells with Trimebutine: Synergistic Antitumor Activity in Combination with Cisplatin

Lung cancer recurrence and therapeutic resistance are driven by cancer stem cells (CSCs), yet clinically available CSC-targeted therapies remain lacking. To identify actionable anti-CSC agents, we screened 1018 FDA-approved drugs using a 3D sphere culture system enriching lung CSC-like cells. Trimebutine, a gastrointestinal motility regulator targeting G-protein-coupled receptors (GPCRs), was identified as a potent agent with over 49.5-fold higher selectivity toward lung CSC-like cells (SI > 49.55) compared with standard therapies such as cisplatin (SI = 1.39) and gefitinib (SI = 1.00). In H460-SP spheres, trimebutine markedly impaired sphere-forming capacity, reduced cell viability, and downregulated key CSC surface markers (CD133, CD44) alongside stemness regulators (c-MYC, EpCAM, BMI1, OCT3/4, NANOG, SOX2, SMAD3). Mechanistically, trimebutine suppressed overexpressed calcium and potassium channels (CACNA1F, CACNA1B, CACNA1E, CACNA1H, KCNH2), attenuating downstream YAP/TAZ–TEAD signaling within the Hippo pathway to trigger apoptosis. In combination with cisplatin, trimebutine exhibited strong synergistic efficacy (Chou–Talalay combination index, CI = 0.015–0.228) in H460-SP spheres and patient-derived lung tumor organoids, suppressing cell viability, stemness marker suppression, and apoptotic cleavage of caspase-3 and PARP compared to monotherapies. Overall, trimebutine targets lung CSC populations by disrupting GPCR–ion channel–Hippo signaling crosstalk, providing a strong preclinical rationale for trimebutine–cisplatin combination strategies to overcome drug resistance and recurrence in lung cancer.