Sgc0946

Real-time cell analysis (RTCA) was performed with xCELLigence (ACEA Bioscience, SD, USA) as described in [14 (link)]. 2 × 10⁵ cells were used per well. The “normalized cell index” compares the change of impedance due to cellular coverage on a 16-well E-plate (Roche, Swiss) to a medium-only control. For inhibitor treatment, 5 μM SGC0946 (Sigma-Aldrich, MO, USA) or EPZ5676 (Absource Diagnostics, Germany), solved in dimethyl sulfoxide (DMSO, Sigma-Aldrich, MO, USA), was applied to CGNP 20 h after plating, for a duration of 4 h or 24 h. To CGN, the inhibitor was applied 4 h after plating, for the duration of 44 h. During RTCA, SGC0946 was replenished every 48 h with half a media change.

MN cells from the spleens of tumor‐bearing mice were cultured in Anne Kelso modified Dulbecco's modified Eagle's medium (Low glucose DMEM (Invitrogen), 4 g/l glucose, 36 mg/l folic acid, 116 mg/l l‐arginine HCl, 216 mg/l l‐glutamine, 2 g/l NaHCO₃, and 10 mM HEPES), supplemented with 20% fetal bovine serum (FBS) (Invitrogen), 0.1 mM l‐Asparagine (Sigma‐Aldrich), and 1% penicillin/streptomycin (Invitrogen). Cells were maintained at 37°C and 10% CO₂. OCI‐AML3 and MOLM13 were purchased from the DSMZ. HEK293T, MV4‐11, and Kasumi‐1 cells were purchased from ATCC. OCI‐AML3, MOLM13, and MV4‐11 were cultured in RPMI‐1640 (Gibco) supplemented with 20% FBS, 1% pen/strep, and 100 µM Glutamax (Invitrogen) at 37°C and 5% CO₂. HEK293T cells were cultured in DMEM (Gibco) supplemented with 10% FBS and 1% pen/strep. All cell lines in this study were routinely tested for Mycoplasma contamination. Cells were treated as indicated with AG636, PUGNAc (Sigma‐Aldrich), Cytarabine (Hospira), JQ1 (Bradner Lab), SGC0946 (Sigma‐Aldrich), Roscovitine (Assay Matrix), or DMSO (Sigma‐Aldrich). For the combination therapy, experiment drugs were refreshed every second day and cells were passaged every 2–4. To quantify the viable cells, cells were stained with 0.2 µg/ml DAPI for 15 min and analyzed by flow cytometry with CountBright™ Absolute Counting Beads (ThermoFisher).