Short-term (72 h drug exposure) was performed as previously described21 (link).
For long-term exposure, cells were seeded at low density in a six-well plate. Treatment started 24 h later, and refreshed twice weekly until 80% confluency. Cells were stained using crystal violet (Sigma-Aldrich) after fixation using 2% paraformaldehyde (Sigma-Aldrich, Zwijndrecht, The Netherlands).
KU-60019, Wortmannin, ETP-46464, VE-821, MK-8776 (SCH 900776), PF-477736, LY2603618/Rabusertib, and Palbociclib were purchased from Selleckchem (Munich, Germany), LY2606368/Prexasertib from Medchem (Sollentuna, Sweden). Adavosertib (MK-1775) from Biovision (Milpitas, USA). All were dissolved in a DMSO stock dilution (10 mM), except Palbociclib (10 mM in ddH2O). Assays contained <1% DMSO. All drug-survival assays depict the standard error of the mean (SEM) of three independent experiments in triplicate.
For long-term exposure, cells were seeded at low density in a six-well plate. Treatment started 24 h later, and refreshed twice weekly until 80% confluency. Cells were stained using crystal violet (Sigma-Aldrich) after fixation using 2% paraformaldehyde (Sigma-Aldrich, Zwijndrecht, The Netherlands).
KU-60019, Wortmannin, ETP-46464, VE-821, MK-8776 (SCH 900776), PF-477736, LY2603618/Rabusertib, and Palbociclib were purchased from Selleckchem (Munich, Germany), LY2606368/Prexasertib from Medchem (Sollentuna, Sweden). Adavosertib (MK-1775) from Biovision (Milpitas, USA). All were dissolved in a DMSO stock dilution (10 mM), except Palbociclib (10 mM in ddH2O). Assays contained <1% DMSO. All drug-survival assays depict the standard error of the mean (SEM) of three independent experiments in triplicate.