(A) CFU serial replating assay of mouse MIG-CRENpm1c/Dnmt3a mutant cell line (SIIIL12) with dimethyl sulfoxide (DMSO) or VTP-50469. Data represent the mean of three independent experiments. d7, day 7. (B) CFU assay of mouse SIIIL12 cells transduced with MSCV-PURO control (left) or Meis1-PURO (right) virus and grown in the presence of 10 nM VTP-50469. Data represent the mean of three independent experiments. (C) CFU assay of SIIIL12 cells electroporated with control or Meis1- or Rpa3-targeting single guide RNAs (sgRNAs) for Cas9-mediated KO. Data represent the mean of three independent experiments. (D) Chromatin immuno- precipitation sequencing (ChIP-seq) density plots showing changes in chromatin occupancy of Menin, MLL, and H3K4me3 and changes in mRNA expression in response to VTP-50469 in OCI-AML3 cells at the MEIS1 and HOXA loci. (E and F) MEIS1 (E) and HOXB5 (F) gene expression in OCI-AML3 cells transduced with control, sgMLL1, sgMLL2, and sgMenin. Data represent the mean of three independent experiments. One-way ANOVA was performed. Error bars indicate mean ± SD. ns, not significant; *P < 0.05; **P < 0.01; ***P <0.001.