Imatinib

WT‐, M351T‐, Y253F‐ or T315I‐BCR‐ABL‐transformed clones of mouse pro‐B Baf3 cells (Baf3p210 cells) were gifted from Brian J. Druker, Oregon Health and Science University Cancer Institute.18 WT‐BCR‐ABL positive human ALL cell line TCCY was established as reported previously.19 To establish imatinib‐resistant clone having T315I‐mutated BCR‐ABL, the WT‐BCR‐ABL‐harboring TCCY cells were treated with imatinib by gradually increasing the concentration (3‐20 μM). The dead cells were washed out every 3 to 4 days, and the resistant subclones were isolated by limiting dilution. Cells were tested for Mycoplasma contamination by using a MycoAlert Mycoplasma Detection Kit (LT07‐118; Lonza, Rockland, ME, USA). Cells were cultured under an atmosphere of 95% air and 5% CO₂ at 37°C in RPMI‐1640 medium (189‐02025; Invitrogen, Carlsbad, CA, USA) supplemented with 10% heat‐inactivated FBS (Sigma‐Aldrich, St Louis, MO, USA). Chemical structure and synthesis of AIC‐47 were reported previously.8 AIC‐47 and imatinib (I0936; Tokyo Chemical Industry, Tokyo, Japan) were dissolved in DMSO and added to the cell culture medium at a final concentration of DMSO (<0.3%), which showed no significant effect on the growth and differentiation of the cells (data not shown). Viable cell numbers were measured by carrying out the Trypan‐blue dye‐exclusion test.