Investigating ABCB1, ABCG2, ABCC1, and ABCC10 Mediated Drug Resistance

The MTT assay was carried out to determine the cytotoxicity of BMS-599626 in parental and ABCB1-, ABCG2-, ABCC1- and ABCC10-overexpressing sublines. Cells were trypsinized, resuspended and 5 × 10³ cells per well were seeded in a 96-well plate. After 24 h, the cells were incubated with or without drugs for 72 h preceded by a 2 h treatment with either BMS-599626 or Ko143. MTT (3 mg/mL) was added to the cells and the cells were further incubated for another 4 h. Subsequently, the supernatant was removed and dimethyl sulfoxide (DMSO) was added to dissolve the formazan crystals. The absorbance was read in an Opsys microplate reader (Dynex Technologies, Chantilly, VA) at an absorbance of 570 nm. The concentration at which 50% of cell growth was inhibited (IC₅₀) was determined as illustrated previously [8 (link),18 (link)]. The ratio of the IC₅₀ of the resistant cells to the IC₅₀ of sensitive cells yielded the resistant folds (Rf). The concentrations of BMS-599626 used for the reversal studies were 100 and 300 nM. Ko143 was used as positive control for inhibition of ABCG2. Cytotoxicity assays were conducted in triplicates and each assay was run at least three times.