For cell migration studies, 2 x 10 5 GBM6840 cells were seeded in 3 cm dishes and treated with 0, 2.5 or 5 µM ATV formulated with A-pBuOzi-A for 65 h. GBM6840 cells were trypsinized, resuspended in 1 mL DMEM containing 0.5 mg/mL soybean trypsin inhibitor, and washed once by centrifugation (300 × g, 5 min at RT). The cell suspension (2.5 x 10 4 cells/0.5 mL DMEM) was added to the upper compartment of a transwell (Falcon cell culture inserts for 24 well-plates, 8 µm pore size; ThermoFisher Scientific). The bottom side of the membrane was pre-coated with fibronectin (10 mg/mL; Sigma-Aldrich) to assure attachment of the transmigrated cells; epidermal growth factor (100 ng/mL; Sigma-Aldrich) was used as a chemoattractant in the lower transwell compartment. Cells were allowed to migrate for 8 h in a humidified tissue culture incubator (37 °C, 5 % CO 2 ) in the presence of 0, 2.5 or 5 µM ATV. Remaining cells on the upper side of the transwell membranes were removed with cotton swabs, and cells on the lower side of the membranes were fixed with 4 % (w/v) para-formaldehyde for 20 min. Nuclei were stained with 4ʹ,6-diamidine-2ʹ-phenylindole (DAPI, 1 µg/mL) in PBS, and transwells were mounted on coverslips using Immunomount (ThermoFisher Scientific). Cell nuclei were visualized on a Leica SP5 confocal microscope (Leica Microsystems) equipped with a 10× objective, and four images covering ≈ 75 % of the total transwell membrane surface area were taken for each transwell. Cell numbers were quantified semi-automatically using ImagePro-Plus software, version 7.0 (Media Cybernetics, Rockville, U.S.A.).