Monocytic Leukemia Cell Resistance to Ferroptosis Is Overcome By Iron Chelation

Introduction:In the United States, acute myeloid leukemias (AML) have a mean 5-year survival rate of 32%, while the monocytic subtype (AMoL) has a mean 5-year survival rate of 27%¹. Therapies targeting induction of ferroptosis hold promise as new strategies for AML, however, resistance can be a problem². To investigate ferroptosis resistance, we have studied iron deficiency and excess in AMoL cells. Fundamentally it is problematic to have either too little iron (dysfunctional energy metabolism and DNA repair) or too much iron (increased reactive oxygen species (ROS) that induce ferroptosis). Dysregulated iron homeostasis, in favour of accumulation, is a central feature of highly proliferative leukemias, enabling the required energy metabolism and biosynthesis for unrestrained proliferation. Iron deprivation through the use of iron chelators and the induction of ferroptosis are two potential avenues of clinical antileukemic treatment.

Objective:To examine the effects of chronic states of iron deficiency and excess in THP-1 cells, an in vitro model of AMoL. Moreover, this study sought to examine the effects of these chronic iron states on THP-1 cell resistance to erastin, a compound that induces ferroptosis by inhibiting the uptake of cystine, a pivotal source of extracellular cysteine used in the synthesis of glutathione (GSH), and thereby, the function of glutathione peroxidase 4 (GPX4). Erastin is known to be largely ineffective as a ferroptosis-inducer in THP-1 cells and other leukemic cell lines.

Methods:THP-1 cells were cultured in various concentrations of ferric citrate or deferoxamine (DFO) for 96 hours to simulate chronic states of iron excess and deficiency, respectively. Following 72 hours of treatment, cells were administered 40 μM erastin for the final 24 hours of treatment. Cell death, metabolic activity, and intracellular GSH were quantified.

Results:20 μM, 40 μM, and 100 μM DFO induced marked, dose-dependent cell death. 40 μM DFO induced a 60.7% reduction in metabolic activity, relative to control cells (± 34.4%, p = 0.0082), while 100 μM DFO induced a mean 91.2% reduction in metabolic activity, relative to control cells (± 4.7%, p < 0.0001). THP-1 cell resistance to erastin was partially attenuated by ferric citrate treatment, evidenced by a small but consistent iron-dose dependent increase in cell death when co-treated with 40 μM erastin. 20 μM DFO is shown to act synergistically with 40 μM erastin in THP-1 cells, inducing a significant increase in cell death, and in the surviving cells, a mean 53.2% reduction in metabolic output relative to control cells (± 11.4%, p = 0.0029) and a mean 60.5% reduction in metabolic output relative to cells treated solely with 20 μM DFO (± 11.4%, p < 0.0001).

Conclusions:The present study demonstrates the robustness of THP-1 cells in states of iron excess and their sensitivity to iron deficiency. It also provides novel evidence of ferric citrate attenuating THP-1 cell resistance to erastin and, most notably, a synergistic lethality between an iron chelator and ferroptosis-inducer (DFO and erastin). This result suggests a novel avenue of antileukemic combination therapy that has potential to combat the poor prognoses of AMoL and acute leukemias, more broadly.

References:

¹SEER*Explorer: An interactive website for SEER cancer statistics. (2023). National Cancer Institute. https://seer.cancer.gov/statistics-network/explorer/

^2.Akiyama, H., Carter, B.Z., Andreeff, M., and Ishizawa, J. Molecular Mechanisms of Ferroptosis and Updates of Ferroptosis Studies in Cancers and Leukemia. (2023). Cells. Apr; 12(8): 1128. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10136912/

No relevant conflicts of interest to declare.