Targeting SPHK1 induces AML cell death. AML cell lines (A), primary AML blasts (B), or normal bone marrow–derived CD34⁺ cells (C) were plated in the indicated concentrations of MP-A08, and cell survival was determined by annexin V staining. (D) FACS-purified primary human CD34⁺/CD38⁻/CD123⁺ LSPCs were treated with a dose response of MP-A08 for 24 hours, and cell survival was determined by annexin V staining. Results are mean ± range. MV4-11 cells were transduced with doxycycline-inducible short hairpin RNA (shRNA) constructs to knockdown SPHK1 and a nontargeting control shRNA. Cells were incubated with 1 μg/mL doxycycline (Dox) and 48 hours postinduction quantitative RT-PCR (mean ± standard deviation, n = 3) (E), SPHK1 activity assays (mean ± range) (F), immunoblot analysis of whole cell lysates were performed and quantified by laser densitometry (ratio of SPHK1/ACTB) (G), and analysis of cell survival was determined by annexin V staining (mean ± standard error of the mean [SEM], n = 3) (H). Significance was assessed by Student t test.