Targeting SPHK1 survival signaling synergizes with BH3 mimetics to induce cell death in AML. (A) MV4-11 cells were transduced with a doxycycline-inducible lentiviral vector to overexpress MCL1. Cells were then exposed to 1 µg/mL doxycycline and 24 hours postinduction treated with 30 μM of MP-A08 for 24 hours, and cell survival was determined by annexin V/PI staining. Results are mean ± SEM, n = 3. Immunoblot analysis of whole cell lysates with anti-MCL1 antibodies 24 hours postinduction confirmed doxycycline-inducible MCL1 expression. (B) MV4-11 cells were treated with increasing doses of MP-A08 in the presence (▪) or absence (●) of 10 nM ABT-737, and cell survival was quantified at 24 hours by annexin V/PI staining. Results are mean ± SEM, n = 3. (C) MV4-11 cells were treated with increasing doses of the S1PR2 antagonist JTE-013 or the S1PR1/3 antagonist VPC-23019 for 6 hours following which whole cell lysate were immunoblotted with the indicated antibodies. (D) MV4-11 cells were treated with increasing doses of JTE-013 (●) or VPC-23019 (▪), and cell survival was quantified at 48 hours by annexin V/PI staining. Results are mean ± SEM, n = 3. (E) MV4-11 cells were treated with increasing doses of JTE-013 in the presence (▪) or absence (●) of 10 nM ABT-737, and cell survival was quantified at 48 hours by annexin V/PI staining. Results are mean ± range. Significance was assessed by Student t test.