Colony formation assays were performed as previously described11 (link). Enzalutamide, LY3009120, Dabrafenib, Trametinib, sch772984 were obtained from Medkoo Biosciences, all drugs were dissolved in DMSO and stored at − 20 °C. Used seeding densities were 20,000 (LNCaP) or 10,000 (22rv1, CWR-R1) cells/well in 6-well plates, and drugs were added as indicated the next day. For 12-well assays, the used seeding densities were 10,000 (LNCaP), or 5000 (CWR-R1, 22rv1, PC3). The growth medium, containing vehicle or drugs, was refreshed every 36–48 h. After 12–14 days of growth in presence of the drugs, when the control cells reached ~ 90% confluency, all cells were fixed in 2% formaldehyde and stained with 0.1% crystal violet.
For quantification of the growth assays, crystal violet was extracted by incubating the stained plates with 5% acetic acid for 1 h at room temperature. The solution, containing the crystal violet, was transferred to a 96-well plate and measured using the Envision 2104 Multilabel Reader (PerkinElmer). Growth assays were performed at least three times for each experiment. Therefore, when quantified data is shown, bars represent the average data from at least three independent experiments ± SEM. P-values are indicated with ***p < 0.001, **p < 0.01 and *p < 0.05 (two-tailed t-test).
For quantification of the growth assays, crystal violet was extracted by incubating the stained plates with 5% acetic acid for 1 h at room temperature. The solution, containing the crystal violet, was transferred to a 96-well plate and measured using the Envision 2104 Multilabel Reader (PerkinElmer). Growth assays were performed at least three times for each experiment. Therefore, when quantified data is shown, bars represent the average data from at least three independent experiments ± SEM. P-values are indicated with ***p < 0.001, **p < 0.01 and *p < 0.05 (two-tailed t-test).