Vorinostat potentiates the lethality of MK-0457 in primary CML cells, while largely sparing normal bone marrow MNCs. (A) MNCs were isolated from bone marrow samples obtained from 2 patients with CML in chronic phase (patients 1 and 2) who had progressed after IM treatment, and from an additional patient (patient 3) who had not yet been treated, as well as from a normal bone marrow specimen. MNCs were exposed to 300 nM MK with or without 1.5 μM Vor for 48 hours, after which the percentage of 7AAD⁺ cells was determined by flow cytometry. (B) MNCs from patient 2 were treated with 100 to 500 nM MK with or without 1.5 μM Vor for 48 hours, after which cells were lysed and subjected to Western blot analysis to monitor caspase-3 cleavage. (C,D) Primary CD34⁺ CML cells isolated from bone marrow MNCs were obtained from patients 1 and 3. Cells were then stained with PE-conjugated CD34 antibody and subjected to flow cytometry to determine the purity of CD34⁺ cells. A representative result is shown (C). Numbers reflect the percentage of cells in the 2 top quadrants. Isolated primary CD34⁺ CML cells as well as normal CD34⁺ cells were exposed to 300 nM MK with or without 1.5 μM Vor for 24 to 48 hours, after which cells were stained with both DiOC₆ and 7AAD and subjected to flow cytometry. “Low” DiOC₆ uptake/7AAD⁻ (bottom left quadrant) corresponds to early apoptosis (mitochondrial damage, reflected by loss of Δψ_m) and “low” DiOC₆/7AAD⁺ (top left quadrant) corresponds to late apoptosis. Values reflect the percentage of cells in the corresponding quadrants. Results represent the means plus or minus SD for experiments performed in triplicate (A). (B) Each lane was loaded with 100 μg protein. The results of a representative experiment are shown; an additional experiment yielded equivalent results. Veh indicates vehicle.