FLT3-ITD-TKD double mutation showed poor response to AC220 comparing to FLT3-ITD. The human FLT3-ITD-TKD double mutation was generated by inducing TKD mutation into the FLT3-ITD sequence using the site-directed mutagenesis approach. WISH showing normal expression of pu.1 (A,E,I), mpo (B,F,J), cebpα (C,G,K), l-plastin (D,H,L) in uninjected control (A-D), FLT3-ITD-TKD embryos treated with DMSO (E-H) and AC220 (I-L) from 6 to 36 hpf. (M-P) Comparison of pu.1 (M), mpo (N), cebpα (O), and l-plastin (P) expansion in FLT3-ITD-TKD embryos treated with DMSO or AC220 from 6 to 36 hpf. There was no statistically significant effect of AC220 treatment (P > .05). Scale bars represent 500 μm.