Treatment with NSC348884 induces apoptosis in mutant but not wild type NPM1 expressing AML cells. (A) HL-60 and OCI-AML3 cells were treated with the indicated concentrations of NSC348884 for 8 hours. At the end of treatment, total cell lysates were prepared and immunoblot analyses were performed for total PARP and cleaved PARP. The expression levels of β-actin in the lysates served as the loading control. Alternatively, evidence of apoptosis was demonstrated by DNA laddering assay. (B) HL-60 and OCI-AML3 cells were treated with 3μM NSC348884 for the indicated times. Then, cells were stained with annexin V and propidium iodide and the percentages of apoptotic cells were measured by flow cytometry. Alternatively, the percentages of nonviable cells were determined by trypan blue dye uptake in a hemocytometer. (C) HL-60 and OCI-AML3 cells were treated with the indicated concentrations of NSC348884 for 8 hours. Then, cells were cytospun onto glass slides and morphologic features of apoptosis were observed by light microscopy at 40× magnification. Representative images of untreated and NSC348884 treated HL-60 and OCI-AML3 cells are displayed.