Sepin-1 Inhibitory Effect on Viability

Cells were seeded into 96‐well plates (3000 cells/well), and viability was determined using Cell Counting Kit‐8 (CCK8; Dojindo). At each time point, cells were incubated with CCK8 for 2 hours, and the absorbance was measured using Microplate Manager v6.0 (Bio‐Rad). For assessing the Sepin‐1 inhibitory effect, 24 hours after seeding, the medium was changed to medium containing Sepin‐1 (for each concentration) or dimethyl sulfoxide (0.5%; vehicle control). The IC₅₀, which is defined as the drug concentration at which cell viability is reduced by half, was determined by GraphPad Prism 7 (GraphPad Software) as described previously.²⁰ (link) Three sets of independent experiments were performed for each time point.