Preleukemic Npm1^cA/+ cells demonstrate enhanced sensitivity to RNA pol I inhibitors. (A) Relative proliferation of preleukemic Npm1^cA/+ vs Npm1^+/+ lin^– cells after 4 days of treatment with ActD at the indicated doses (MTS assay). (B) Representative CFU assays from preleukemic Npm1^cA/+ vs Npm1^+/+ lin^– cells after 24 hours pretreatment with 2.5 nM ActD. (C) Total number of colonies in CFU assays of preleukemic Npm1^cA/+ vs Npm1^+/+ lin^– cells after 24 h pretreatment with 2.5 nM of ActD. (D) Nucleolar morphology of Npm1^+/+ (top panel) or Npm1^cA/+ (bottom panel) lin^– cells after treatment with 1 nM ActD for 18 hours, highlighted using fluorescent staining for wild-type NPM1 (NPM1_WT) and nucleolin (NCL) (scale, 10 μm). (E-G) Relative proliferation of preleukemic Npm1^cA/+ vs Npm1^+/+ lin^– cells after 4 days of treatment with BMH-21 (E), CX-5461 (F), or cycloheximide (G) at the indicated doses (MTS assay). (A,C,E-G) Unpaired t test was used to calculate P values between groups. ∗P ≤ .05; ∗∗P ≤ .01; ∗∗∗P ≤ .001. (A,E-G) Absorbance values were normalized to DMSO-treated cells. Data represent 4 to 7 biological replicates per group (mean ± SD). (C) Values represent 4 biological replicates per group. (D) Representative example from 1 of 2 to 4 biological replicates. DAPI, 4′,6-diamidino-2-phenylindole.