Assay of cell viability with two drugs

The EC₅₀ values obtained from single-drug cell viability assays were used to design subsequent drug combination experiments. Two-drug combination experiments were performed by using a 5 × 5 matrix in 96-well plates to interrogate 25 dosing pairs. Cells in complete growth medium were plated at 5,000 cells per well in a 96-well microplate, which was incubated for 24 hours at 37 °C. Cells were treated with kinase inhibitor for 1 h, followed by QBI-139. After 48 hours, cell viability was evaluated as described above.

Dose–response data for each drug alone and for two-drug combinations were determined experimentally. CI values were calculated using CalcuSyn 2.0 from Biosoft (Cambridge, UK). CI < 1, CI = 1, and CI > 1 indicate synergism, an additive effect, and antagonism, respectively. Each CI value was the mean of data from 3 biological replicates.