Background: acute myeloid leukemia (AML) is a highly heterogeneous hematological malignancy whose incidence increases with age. Although the traditional induction chemotherapy is effective, the accompanying toxic and side effects are strong, and relapse is still unavoidable after treatment. Since 2017, several new targeted drugs have been approved for the treatment of leukemia patients. Venetoclax is the only oral BCL-2 specific inhibitor approved to date for the treatment of chronic lymphocytic leukemia. However, its efficacy in AML patients is very limited, suggesting the need to find and establish safe and effective combination regimens. BCL-2 not only regulates mitochondria-dependent apoptosis, but also participates in autophagy by regulating type Ⅲ PI3K complex. At present, no studies have reported whether targeting VPS34, a key molecule in the type Ⅲ PI3K complex, can promote the anti-AML effect of venetoclax.
Objective: To investigate the function of VPS34 in AML and whether targeting VPS34 can promote the anti-AML effect of venetoclax, a BCL-2 inhibitor.
Methods: The expression level of VPS34 in various tumor tissues, including AML, and its correlation with BCL-2 were analyzed by online database. The effects of venetoclax and VPS34-specific inhibitor SAR405 alone or in combination on the viability of AML cell lines, primary cells from AML patients and peripheral blood mononuclear cells in healthy people were investigated by cell proliferation assay. The changes of cell apoptosis, cycle and mitochondrial membrane potential were detected by flow cytometry. The synergistic molecular mechanism of venetoclax combined with SAR405 was analyzed by transcriptome sequencing, fluorescent quantitative PCR and western blot. To construct a xenograft mouse model of AML and evaluate the therapeutic effect of venetoclax combined with SAR405 on AML.
Results: Using UALCAN、GEPIA database, VPS34 was found to be expressed in various tumor tissues, and the expression level was highest in AML. Moreover, VPS34 was positively correlated with BCL-2, with a correlation coefficient of 0.43. The results of cell proliferation experiments showed that SAR405 inhibited the viability of AML cell lines in a concentrate-dependent manner, but had no significant effect on the mononucleus of peripheral blood of healthy people, suggesting the feasibility and safety of targeting VPS34. More importantly, SAR405 significantly promoted venetoclax's action and synergistic effect in AML cell lines and primary tumors of AML patients. After transcriptome sequencing and analysis of the results, it was found that venetoclax combined with SAR405 significantly enriched the cell cycle regulated by PI3K/AKT and MAPK/ERK pathways compared with the combination of Venetoclax and SAR405 alone. Flow cytometry demonstrated that venetoclax combined with SAR405 induced apoptosis, disrupted mitochondrial membrane potential, and blocked the cell cycle in G1 phase. At the molecular level, apoptosis-regulating proteins such as PARP, Caspase-3, MCL-1, BCL-XL and cycle regulating molecules such as CDK2, CDK4, CDK6, c-Myc, p21 also showed corresponding changes. Meanwhile, venetoclax combined with SAR405 significantly inhibited the activation of PI3K/AKT and MAPA/ERK signals. Finally, in an in vivo mouse model transplanted with MV4-11 cells, venetoclax combined with SAR405 significantly reduced tumor load in mice, delayed the progression of AML, and extended survival time in mice.
Conclusion: The results of this study suggest that VPS34 is not only a potential target for AML treatment, but also an effective synergistic target of BCL-2, which provides a new idea and treatment strategy for clinical treatment of AML.
No relevant conflicts of interest to declare.
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