MEK inhibition reduces proliferation in Nras-mutant AML without inducing differentiation. (A) Serial white blood cell counts in secondary transplant recipients of myeloid neoplasms #6730 and #8064 treated with PD901 (red line, n = 8) or a control vehicle (black line, n = 10), with error bars representing the 95% confidence interval (*statistical significance by unpaired t test). Next, mice transplanted with AML #6695 were treated with 4 daily doses of vehicle (n = 4) or PD901 (n = 5) starting 10 days posttransplant. (B-C) PD901 treatment effectively reduced white blood cell counts (P = .045) and spleen size (P = .0057). (D) Normalized viable cell count after 4 days of growth in liquid culture with increasing doses of PD901 in 5 independent N-Ras^G12D myeloid neoplasms (n ≥ 3 mice per leukemia). (E) Western blot analysis of granulocyte macrophage colony-stimulating factor (GM-CSF) stimulated pERK and phosphorylated Akt (pAkt) in BM cells isolated from transplant recipients of Nras^G12D myeloid neoplasms and WT BM control. BM from moribund mice was assayed for level of pERK, total ERK, pAkt, total Akt, and β-actin at B (basal conditions), following S (starvation), and subsequent GM-CSF stimulation in the absence or presence of increasing concentrations of PD901 (0.01, 0.1, and 1 μM). (F) Mice engrafted with AML #6695 were treated with PD901 (n = 5) or vehicle (n = 4) for 4 days (as in panels B and C) and total BM was stained for c-Kit, Mac-1 (CD11b), and Gr-1 surface expression. WT untreated BM is shown as a control. Representative plots are shown, and numbers represent the mean percentage for each gate across replicates.