Acute myeloid leukemia (AML) is a heterogeneous clonal hematopoietic malignancy with poor survival and frequent relapse. Recently, a posttranslational modification of proteins with small ubiquitin-like modifiers (SUMO) has been notably implicated in a wide spectrum of diseases, especially cancers. Ubc9, as the sole E2-conjugating enzyme in SUMOylation cascade, particularly has been associated with adverse clinical outcomes. 2-D08, a small molecular agent, functions by blocking the transfer of SUMO from the Ubc9 thioester to SUMO substrates without any effects on other individual steps in this process. However, both the effects and mechanisms of 2-D08 on AML cells are still unknown. In this study, we found that 2-D08 significantly suppressed cell viability and colony formation ability. Additionally, it induced mitochondrial-mediated apoptosis with dramatic accumulation of the reactive oxygen species (ROS), which could be almost completely rescued by the ROS scavenger N-acetylcysteine (NAC). Furthermore, we confirmed that the fatal accumulation of ROS was due to its aberrant generation instead of defective scavenging. In summary, our results suggest that 2-D08, as a specific SUMOylation inhibitor, induces ROS accumulation-mediated intrinsic apoptosis of AML cells possibly through deSUMOylation of NOX2. Therefore, 2-D08 might be a promising therapeutic agent for the treatment of AML in the future.

Acute myeloid leukemia (AML) is a heterogeneous hematopoietic disorder with a poor prognosis. The clinical significance of Leukemia stem cells (LSCs) plays an important role in the generation of AML and is the main cause of the recurrence after remission. Osteopontin (OPN), an extracellular matrix protein, has been implicated in hematopoietic malignancies. However, the specific role and the underlying mechanism of AML cell autocrined OPN in leukemia maintenance remain unknown. Here, we showed that knockdown of Opn expression significantly prolonged the survival of mice with MLL-AF9 cell-induced AML and markedly reduced the tumor burden. The LSCs from the Opn-knockdown groups exhibited decreased numbers and impaired function as determined by immunophenotype, colony-forming and limiting dilution assays. Further analysis revealed that Opn prevents LSCs from undergoing apoptosis and cell cycle arrest. Repression of OPN in human AML cell lines in vitro mimics the phenotypes observed in the mouse model. Overall, our data indicated that OPN is a potent therapeutic target for eradicating LSCs in AML.

Although great efforts have been made to improve available therapies, the mortality rate of acute myeloid leukemia (AML) remains high due to poor treatment response and frequent relapse after chemotherapy. Leukemia stem cells (LSCs) are thought to account for this poor prognosis and relapse. Phosphoinositide-dependent kinase 1 (PDK1) is a critical regulator of the PI3K/Akt pathway and has been shown to be frequently activated in leukemia. However, the role of PDK1 in the regulation of LSCs in AML is still not clear. Using a PDK1 conditional deletion MLL-AF9 murine AML model, we revealed that the deletion of PDK1 prolonged the survival of AML mice by inducing LSC apoptosis. This was accompanied by the increased expression of the pro-apoptotic genes Bax and p53 and the reduced expression of Stat5, which has been shown to be constitutively activated in leukemia. Thus, our findings suggest that PDK1 plays an essential role in maintaining LSCs. Further delineating the function of PDK1 in LSCs may provide a new strategy for the improved treatment of AML relapse.