Figure 3. Drug combination effects in CML cells.

(A) KU812 cells (upper panel) and K562 cells (lower panel) were incubated in control medium (Co), with various concentrations of ponatinib or bosutinib, or with a combination of both drugs at a fixed ratio (KU812 at 1:16; K562 at 1:100) at 37°C for 48 hours. Thereafter, ³H-thymidine was added for 16 hours, and uptake of ³H-thymidine was measured in a β-counter. Results are expressed as percent of medium control and represent the mean ± S.D. of quadruplicates. In case of KU812 cells, the drug combination was found to be highly synergistic, whereas in K562 cells, mostly additive effects were obtained. (B) Primary patient-derived blast cells were incubated in control medium (Co) or with increasing concentrations of hydroxyurea (HU), ponatinib, or bosutinib at 37°C and 5% CO₂ for 48 hours as indicated. Then, ³H-thymidine uptake was measured. Results are expressed as percent of control and represent the mean ± S.D. of triplicates. (C) KU812 cells were incubated with ponatinib (0.1 nM: ■-■, 0.5 nM: ▲−▲, 1.0 nM: ▼-▼, or control medium: •-•) at 37°C for 4 hours. Then, cells were washed and incubated in control medium (Co) or bosutinib at various concentrations as indicated for another 48 hours. Thereafter, ³H-thymidine uptake was measured. Results are expressed as percent of control and represent the mean ± S.D. of triplicates. (D) KU812 cells were incubated in control medium (Co), ponatinib (0.2 nM), bosutinib (10 nM), or a combination of both drugs at 37°C for 48 hours. Thereafter, the percentage of AnnexinV/PI-positive cells was determined by flow cytometry. Results represent the mean ± S.D. of 3 independent experiments. Asterisk (*): p < 0.05 compared to control.