Discovery of tepotinib as a novel LY75-targeting small-molecule inhibitor with anti-tumor activity in ovarian cancer via virtual screening

To identify small-molecule inhibitors targeting LY75 and evaluate their anti-tumor activity and underlying mechanisms in ovarian cancer. LY75 expression and prognostic relevance were analyzed using TCGA and HPA datasets. Structure-based virtual screening of the TargetMol compound library was performed using the LY75 crystal structure (PDB: 8K8H), followed by docking, MM/GBSA, PLIF, and ADMET analyses. Candidate compounds were evaluated by CCK-8, apoptosis, migration, protein stability, SPR binding, and xenograft assays. To address the known MET-inhibitory background of Tepotinib, c-MET expression, MET–LY75 co-expression, and pharmacological controls with capmatinib were assessed. LY75 was significantly upregulated in ovarian cancer tissues and high LY75 expression was associated with shorter progression-free survival. Tepotinib was identified as the leading candidate and preferentially inhibited LY75-high SKOV3 and OVCAR-8 cells, with IC₅₀ values of 16.31 µM and 18.91 µM, respectively, while showing weaker activity in LY75-low HO8910 cells. SPR confirmed direct binding between Tepotinib and LY75 (kinetic K _ D = 2.52 µM; steady-state K _ D = 3.74 µM). Tepotinib induced apoptosis, inhibited migration, and downregulated LY75 protein by accelerating its degradation through a ubiquitin-proteasome-independent mechanism. In vivo, oral Tepotinib markedly suppressed SKOV3 xenograft growth, achieving a tumor inhibition rate of 79.6%, and reduced LY75 expression in tumor tissues. MET-related analyses and capmatinib controls suggested that MET inhibition alone was insufficient to account for the full anti-tumor activity of Tepotinib. This study identifies Tepotinib as a repurposed small molecule with LY75-binding and LY75-downregulating activity in ovarian cancer. These findings support LY75 as a potential therapeutic target and provide a rationale for further investigation of Tepotinib-based targeted therapy in ovarian cancer.