AML1-ETO GE-HTS screen identifies 2 top classes of hits. (A) Principal component analysis performed with compounds that were statistically significant by at least 2 discrete metrics for the AML1-ETO abrogation signature. The position of hits relative to controls with and without AML1-ETO expression is indicated. The points labeled “compound” represent the average of all chemicals on each individual screening plate. The hit compound datapoints represent the mean of 3 replicates. All other data points represent the mean of samples per individual screen plate: vehicle (16), Ctrl RNA (8), and AE RNAi (8). Validated hits included 2 primary drug classes: corticosteroids and DHFR antagonists. (B) Induction of AML1-ETO abrogation signature in Kasumi-1 cells treated with indicated dose of methotrexate for 72 hours. Each RNAi control or cell line control data point represents the mean of 16 samples, while the vehicle- and compound-treated points represent the mean of 3 samples. (C) Induction of abrogation signature in Kasumi-1 cells treated with methylprednisolone for 72 hours. Each RNAi control or cell line control data point represents the mean of 16 samples, while the vehicle- and compound-treated samples represent the mean of 3 samples. (D) Weighted summed score for the AML-ETO signature upon treatment of Kasumi-1 cells with 80 nM methotrexate (MTX) or 0.5 μM methylprednisolone (MPD) for 24 to 72 hours. There were 16 replicates for drug treatment and RNAi controls and 32 for DMSO controls.