Targeted therapies are frequently combined with standard cytotoxic drugs to enhance clinical response. Targeting the B-cell lymphoma 2 (BCL-2) family of proteins is an attractive option to combat chemoresistance in leukemia. Preclinical and clinical studies indicate modest single-agent activity with selective BCL-2 inhibitors (for example, venetoclax). We show that venetoclax synergizes with cytarabine and idarubicin to increase antileukemic efficacy in a TP53-dependent manner. Although TP53 deficiency impaired sensitivity to combined venetoclax and chemotherapy, higher-dose idarubicin was able to suppress MCL1 and induce cell death independently of TP53. Consistent with an MCL1-specific effect, cell death from high-dose idarubicin was dependent on pro-apoptotic Bak. Combining higher-dose idarubicin with venetoclax was able to partially overcome resistance in Bak-deficient cells. Using inducible vectors and venetoclax to differentially target anti-apoptotic BCL-2 family members, BCL-2 and MCL1 emerged as critical and complementary proteins regulating cell survival in acute myeloid leukemia. Dual targeting of BCL-2 and MCL1, but not either alone, prolonged survival of leukemia-bearing mice. In conclusion, our findings support the further investigation of venetoclax in combination with standard chemotherapy, including intensified doses of idarubicin. Venetoclax should also be investigated in combination with direct inhibitors of MCL1 as a chemotherapy-free approach in the future.

Growth and survival of hematopoietic cells is regulated by growth factors and cytokines, such as interleukin 3 (IL-3). When cytokine is removed, cells dependent on IL-3 kill themselves by a mechanism that is inhibited by overexpression of Bcl-2 and is likely to be mediated by proapoptotic Bcl-2 family members. Bad and Bim are 2 such BH3-only Bcl-2 family members that have been implicated as key initiators in apoptosis following growth factor withdrawal, particularly in IL-3-dependent cells. To test the role of Bad, Bim, and other proapoptotic Bcl-2 family members in IL-3 withdrawalinduced apoptosis, we generated IL-3-dependent cell lines from mice lacking the genes for Bad, Bim, Puma, both Bad and Bim, and both Bax and Bak. Surprisingly, Bad was not required for cell death following IL-3 withdrawal, suggesting changes to phosphorylation of Bad play only a minor role in apoptosis in this system. Deletion of Bim also had no effect, but cells lacking Puma survived and formed colonies when IL-3 was restored. Inhibition of the PI3 kinase pathway promoted apoptosis in the presence or absence of IL-3 and did not require Bad, Bim, or Puma, suggesting IL IntroductionRegulation of apoptosis by growth factors and cytokines is critical to maintain health and avoid neoplastic disease. During development and in adulthood, excess and unwanted cells are removed when cells that do not receive sufficient growth factor signals activate their death program by default. 1 On the other hand, the ability of a mutant cell to survive in the absence of growth factors is an important step leading to its malignant transformation. Loss of cytokine signaling results in cell death by a mechanism characterized by activation of caspases that is regulated by the Bcl-2 family of proteins. 2,3 Some members of the Bcl-2 family, such as Bcl-2 and Bcl-xL, can prevent apoptosis induced by withdrawal of growth factor when overexpressed. 3,4 Bax and Bak, on the other hand, are proapoptotic Bcl-2 family members, and deletion of both renders cells highly resistant to apoptosis following growth factor withdrawal, 5,6 emphasizing the pivotal role these proteins play in this pathway.Bad, Bim, and Puma belong to a third, proapoptotic subgroup of the Bcl-2 family, which share homology only in a BH3 domain, the so-called BH3-only proteins. These proteins promote apoptosis when overexpressed 7,8 probably by antagonizing the prosurvival antiapoptotic Bcl-2 family members. 9 Regulation of cell survival by cytokines has often been studied in interleukin 3 (IL-3)-dependent hematopoietic cell lines such as FDC-P1, FL.5, and Ba/F3 cells. [10][11][12] Several studies have implicated the BH3-only protein Bad as an important mediator of apoptosis following IL-3 withdrawal. When phosphorylated by kinases such as AKT, Bad binds to the tau form of 14-3-3 protein. 13,14 In the absence of IL-3, unphosphorylated Bad is able to bind to, and inhibit, 11,15 suggesting this is the mechanism by which cytokines regulate survival of hematopoietic cells. Consistent with this model, ex...