FGF2 protection of K562 cells is mediated by FGFR3. K562 cells were cultured in media without and with 10 ng/mL FGF2 and exposed to a titration of FGFR inhibitors. (A) PD173074: 25, 50, or 100 nM or (B) AZD1480: 0.5, 1, or 2 μM and in combination with 1 μM IM as indicated. Viability was measured by MTS assay after 48 hours. Cells not exposed to IM were normalized to untreated K562 cells. IM-treated cells were compared with their respective untreated control (eg, media alone, FGF2, 25 nM PD173074). (C) Blocking FGF2 Ab (Millipore bFM-1) at 100 μg/mL, 10 μg/mL, or 1 μg/mL was added to media with or without 10 ng/mL FGF2 and preincubated for 1 hour at 37°C. K562 cells were then added with or without 1 μM IM and viability measured by MTS. Viability is presented as the percent of untreated control for each condition. (D) K562 cells were electroporated with siRNAs targeting FGFR1, FGFR2, FGFR3, FGFR4, and a nonspecific (NS) control. After 48 hours, the cells the cells were pelleted and resuspended in media with or without FGF2 ± 1 μM IM and viability assessed by MTS after 48 hours. At 48 hours, a portion of the cells were lysed and analyzed by western blot for FGFR3 to evaluate protein expression (lower panel and supplemental Figure 2 for quantitative polymerase chain reaction). *Indicates P < .01 by Student t test. (E) K562 cells were electroporated with pools of siRNAs targeting the tyrosine kinome as described previously^12,13  and incubated with 10 ng/mL FGF2 and 1 μM IM. Viability was assessed after 72 hours by MTS; the dark gray horizontal line indicates 2 standard deviations below the mean for all siRNAs. ABL1 and FGFR3 siRNA are denoted as the only 2 siRNA pools that reduced viability below 2 standard deviations.