A Therapeutic Approach for Acute Leukemias: Glycolysis Inhibition Revisited

Background: Recent evidence suggests that myeloid and lymphoid leukemia cells are dependent on aerobic glycolysis for ATP production, known as the Warburg effect. Early glycolysis inhibitors were effective but only at high mM concentrations. The novel third generation glycolysis inhibitor, 3-bromo-2-oxopropionate-1-propyl ester (3-BrOP), has shown efficacy in models of neuroblastoma, glioblastoma, colon carcinoma, and lymphoma in the mcM range and has improved oral bioavailability and biodistribution. We proposed that potent inhibition of glycolysis would lead to ATP depletion and induce growth arrest and cell death in acute lymphoblastic leukemia (ALL) and acute myelogenous leukemia (AML) cells. Objective: To determine the feasibility and effect of 3-BrOP on human AML, precursor-B, and T cell leukemia cell lines and patient samples.

Design/Methods: A panel of fifteen human pre-B ALL, T-ALL, and AML cell lines were treated with 3-BrOP to determine IC50s via cell counts, cell cycle, and Alamarblue. In addition, western blots and ATP assays were performed to determine caspase activation and ATP depletion.

Results: All cell lines demonstrate growth arrest, decreased alamar blue metabolism and significant apoptosis when treated with 3-BrOP. The five human precursor-B ALL cell lines had IC50s between 10–30mcM, while the T ALL cell lines had IC50s between 15–30mcM. The human AML cell line panel had IC50s between 20–40mcM. In addition, patient samples representing ALL (IC50 15 mcM) and AML (IC50 30mcM) were sensitive to 3-BrOP mediated cell death. Of note, caspase-3 cleavage was detected by 2 hours of exposure, and was nearly complete by 6 hours in both pre-B and T-ALL suggesting a potent an early pro-apoptotic signal. Overall, cell cycle analysis at 48 hours of treatment, showed the majority of cells had died (83–97%, p<0.001). The remaining cells were apoptotic with >80% of Nalm6 (pre-B ALL) and 72% of U937 (AML) cells having sub-G0 DNA content (p<0.001). As expected, exposure to 3-BrOP reduced intracellular ATP concentrations in a dose and time dependent manner. In Nalm6 cells (pre-B ALL), by 48 hours of exposure, the ATP levels were 50% of control at a dose of 15 mcM. At a dose of 22.5 mcM, the ATP level was 13% of control, and by 30 mcM there was no detectable ATP. Importantly, at higher doses (>30mcM) ATP levels were reduced significantly within 24 hours. These effects on ATP levels correlate with dose and time dependent viability.

Conclusions: These data demonstrate that a broad range of acute leukemia cell lines and patient samples appear to be dependent on glycolysis for ATP production and survival. The novel 3rd generation glycolysis inhibitor, 3-BrOP, induces cell death across all subtypes of human acute leukemia cell lines. Glycolysis inhibition via 3-BrOP represents a novel and clinically feasible therapeutic approach for acute leukemias and warrants further pre-clinical evaluation.