Venetoclax and hypomethylating agents synergize to increase cell death and metabolic remodeling in acute B-lymphoblastic leukemia cells

Introduction: Overexpression of anti-apoptotic protein Bcl-2 and hypermethylation are hallmarks of acute lymphoblastic leukemia (ALL) and can be pharmacologically addressed by venetoclax (VEN) and hypomethylating agents (HMA) such as azacytidine (AZA) or decitabine (DEC). Combined VEN and HMA application was recently successfully implemented into the clinical treatment regimen of acute myeloid leukemia but has so far not been investigated in ALL.

Methods: We therefore analyzed the anti-leukemic potential of VEN + HMA in four ALL cell lines and identified potential modes of synergy to overcome mono-drug-induced resistance using proliferation, metabolism, methylation and apoptotic protein expression assays. Single cell RNAseq of a VEN-treated PDX model was used to gain deeper insights into metabolic reprograming.

Results: All substances influenced proliferation and induced Apoptosis in a subset of cell lines. Combined VEN and HMA application resulted in significantly reduced metabolic activity. In contrast, no synergistic effects were observed regarding the Bcl-2 protein and methyltransferase expression or global methylation. Single cell RNAseq revealed that VEN interferes with both main energy supply routes, Oxidative Phosphorylation as well as glycolysis, to impede the cells' metabolism and mitochondrial activity. The addition of HMA, especially DEC, increased anti-metabolic effects, leading to a strong reduction of respiration, ATP production and proton leakage. AZA-induced metabolic suppression and overall anti-leukemic activity alone and in combination with VEN was generally weaker compared to DEC.

Conclusion: Altogether, we herein demonstrate that combined VEN and HMA application acts synergistically and significantly reduces the leukemic burden in ALL cell lines via impairment of tumor cell metabolism and mitochondrial function.