Screening Small-Molecule Inhibitors in AML

Bone marrow aspirates from three AML mice were combined, isolated, and cultured in RPMI containing 20% fetal bovine serum (FBS), streptomycin/penicillin, 50 µM 2-mercaptoethanol (RPMI-20), and supplemented with murine SCF (10 ng/mL) and IL-3 (10 ng/mL) (Peprotech or BioLegend). The cells were serially passaged for ~1 month. Inhibitor library screening to evaluate drug sensitivity was performed as previously described [5 (link)]. Briefly, cultured AML mouse-derived cells were counted and seeded into four 384-well plates at a concentration of 2000 cells/well. The cells were then subjected to titrations of 188 unique small-molecule inhibitors (SMIs) in culture for 72 h. MTS reagent (CellTiter96 AQueous One; Promega) was added and the optical density was read at 490 nm to assess viability.