MEIS1 and SMC4 expression levels affect response to MI-3454 treatment. (A) MEIS1 and SMC4 mRNA expression in OCI-AML3, treated with DMSO control or MI-3454 (25 nM, n = 3) for 7 days; values are shown as fold change to housekeeping gene control, TBP. (B) Western blot analysis showing 2 experimental replicates of SMC4 protein expression in OCI-AML3 cells treated with 25 nM MI-3454 for 7 days vs DMSO control. LAMIN-B was used as endogenous control. (C) IC₅₀ calculation of MI-3454 inhibitor on Meis1 overexpressing NM1 cells vs endogenous Meis1 expressing ex vivo MLL-AF9 cells. (D) Colony number in the colony forming assay of NM1 cells treated with DMSO and MI-3454. (E) IC₅₀ calculation of MI-3454 inhibitor on NM1 cells transduced with scrambled control or shRNA against Smc4. (F) Colony forming assay of NM1 cells transduced with scrambled control and shRNA-mediated Smc4 knockdown, treated with DMSO and MI-3454 (n = 3). ∗P < .05; ∗∗P < .001; ∗∗∗P < .0001. DMSO, dimethyl sulfoxide; n, number of independent experiments; TBP, TATA-binding protein.