Chz868

Experiments were performed in serum-deprived medium (0.1% FBS) and normal medium conditions, as indicated in the figure legends. Bemcentinib (gift from BerGenBio), ruxolitinib, and CHZ868 (both MedChem Express) were dissolved in dimethylsulfoxide and used in different concentrations as indicated in the figure legends. To assess cell viability, 4 × 10⁴ cells/well were seeded in 96 well plates and treated with the indicated drug and GAS6 concentrations, respectively. After 48 hours, cell numbers were assessed by water soluble tetrazolium salts assay (Roche Applied Science, Mannheim, Germany). To assess proliferation and apoptosis, 2 × 10⁵ cells/well were seeded in 24 well plates and treated with the indicated concentration of bemcentinib. Proliferation levels were measured by flow cytometry using bromodeoxyuridine/5-bromo-2′-deoxyuridine (BrdU) incorporation assays (Becton and Dickinson bromodeoxyuridine/5-bromo-2′-deoxyuridine ([BD]). Apoptosis levels were measured by flow cytometry using 7-aminoactinomycin D (7-AAD) staining (to exclude dead cells) and Annexin V-FITC. Events were captured using BD fluorescence- activated cell sorting Calibur.