2D Drug screen

Drugs (10 nmol/L, 100 nmol/L, and 1 μmol/L final concentration) were prealiquoted into 384-well 2D culture plates (Greiner Bio-One) using an ECHO550 acoustic liquid handler prior to seeding 1,200–2,400 cells/well in 50 μL of cell line–specific media. Cell viability was assessed using CellTiter-Glo (Promega; day 5). Assay quality was assessed using calculation of Z-prime per plate, and only plates with Z-prime of ≥0.5 were taken for further analysis. Three independent biological replicates (each replicate formed of six plates) were performed to reduce the risk of false hit identification. To generate a percentage of control (POC) score, the raw luminescence value for each well was normalized by dividing this value by the median value of the negative controls on the plate. The mean of the POC scores of the three independent biological replicates was used to generate a mean response score for each drug, at each concentration tested.