MTS assay

SK-MEL-5, SK-MEL-28 and SK-MEL-31 cells (1 × 10³ cells/well) were seeded into 96-well plates in 100 µL of 10% FBS/Eagle’s MEM. The cells were treated with 4HD or XAG. After incubation for 1, 2, 3 or 4 days, NHEM cells (1 × 10⁴ cells/well) were seeded into 96-well plates for determining cytotoxicity. After an overnight incubation, cells were treated with different concentrations (5, 10 or 20 µM) of 4HD or XAG or vehicle control and incubated for 1 or 2 days. Then 20 µL of the CellTiter 96 Aqueous One Solution (Promega, Madison, WI) were added to each well and cells were then incubated for an additional 1 h at 37°C. Absorbance was measured at an optical density of 492 and 690 nm using the Thermo Multiskan plate-reader (Thermo Fisher Scientific, Waltham, MA).