![Biomarker modulation by Aurora and FLT3 inhibitors in MOLM-13-RES cells. (A) MOLM-13-RES cells were arrested in mitosis by overnight incubation with 50 ng/mL nocodazole and treated with the indicated concentrations of CCT241736, CCT137690, MLN518, or AMG900 for 2.5 hours. Cell lysates were analyzed for STAT5 and HH3 phosphorylation by immunoblotting using specific antibodies. Total STAT5 and total HH3 were used as controls. The bar graphs (bottom panels) show compound cellular potency after normalization to the control total proteins. (B) BaF3FLT3-ITD and BaF3FLT3-ITD-F691L cells were treated for 2.5 hours with increasing concentrations of CCT241736 (20 nM to 10 μM) and with 50 nM AC220. Cell lysates were prepared, FLT3 was immunoprecipitated using a specific FLT3 antibody, and its phosphorylation was analyzed by immunoblotting with phospho-tyrosin–specific antibodies. The bar graph (bottom panel) shows CCT241736 and AC220 cellular potency after normalization to the control total proteins. (C) Fold change in viability IC50 with 10 ng/mL FL. MOLM-13 cells were treated with increasing concentrations of cytarabine, CCT241736, AC220, or MLN518 with or without 10 ng/mL FL for 72 hours. The IC50 values for viability were measured using MTS. Experiments were performed in parallel on 3 separate occasions. Error bars represent the mean change (± standard error of the mean [SEM]) in viability IC50 with 10 ng/mL FL compared with viability IC50 with no FL treatment. *P < .05 on paired Student t test analysis.](https://ash.silverchair-cdn.com/ash/content_public/journal/bloodadvances/4/7/10.1182_bloodadvances.2019000986/1/advancesadv2019000986f2.png?Expires=1720846947&Signature=aEqA7pXyZ4JtNXrND9L0NWPaI815C7lXoZXCVNFCgH0Gt9L8Zj8iVBmGnzBWLnSHP4ft-0yIB66HpW8~Pf8tmS8wIxOqLyb-jMCLV6fgP9jtzWZ2bI~wrYiCaex~cyWLTJDsbI-~7steeliOIox5HqGdULQbloVOAS~zz8jpJhzzzcReLX449H~nsA5l078Gh6w0R63dX-FMDAhdx9J8wmBWd0V7FlQ~csmxExQ4rCJ29m2jsKqONBscIvptwUQJ-jkSht7Nc6raUN95TuvR6LUP98k2F8MoSrCwZ~bWhiXbqP1yd903mRJ-Shb7F7gAjCqRm8TZ4Ly7m2tOPYzzmQ__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Biomarker modulation by Aurora and FLT3 inhibitors in MOLM-13-RES cells. (A) MOLM-13-RES cells were arrested in mitosis by overnight incubation with 50 ng/mL nocodazole and treated with the indicated concentrations of CCT241736, CCT137690, MLN518, or AMG900 for 2.5 hours. Cell lysates were analyzed for STAT5 and HH3 phosphorylation by immunoblotting using specific antibodies. Total STAT5 and total HH3 were used as controls. The bar graphs (bottom panels) show compound cellular potency after normalization to the control total proteins. (B) BaF3FLT3-ITD and BaF3FLT3-ITD-F691L cells were treated for 2.5 hours with increasing concentrations of CCT241736 (20 nM to 10 μM) and with 50 nM AC220. Cell lysates were prepared, FLT3 was immunoprecipitated using a specific FLT3 antibody, and its phosphorylation was analyzed by immunoblotting with phospho-tyrosin–specific antibodies. The bar graph (bottom panel) shows CCT241736 and AC220 cellular potency after normalization to the control total proteins. (C) Fold change in viability IC50 with 10 ng/mL FL. MOLM-13 cells were treated with increasing concentrations of cytarabine, CCT241736, AC220, or MLN518 with or without 10 ng/mL FL for 72 hours. The IC50 values for viability were measured using MTS. Experiments were performed in parallel on 3 separate occasions. Error bars represent the mean change (± standard error of the mean [SEM]) in viability IC50 with 10 ng/mL FL compared with viability IC50 with no FL treatment. *P < .05 on paired Student t test analysis.