Acute myeloid leukaemia is a highly heterogeneous blood cancer driven by the clonal expansion of immature myeloid cells, and long-term survival remains poor. Many patients achieve complete remission after standard induction chemotherapy with cytarabine and an anthracycline, yet the majority eventually relapse because the disease acquires drug resistance. Mounting evidence points to the bone marrow microenvironment as a central accomplice in this process. The new study, led by researchers at Fujian Medical University Union Hospital, focused on bone marrow stromal cells and asked precisely how these cells confer a survival advantage on leukaemic blasts under therapeutic stress.

The starting clue came from the team’s own earlier work showing that AML cells grown alongside stromal cells ramp up mitochondrial oxidative phosphorylation, the energy-generating process that leukaemic cells rely on heavily to meet their bioenergetic and biosynthetic demands. Elevated mitochondrial respiration, however, comes at a cost: it produces more reactive oxygen species, particularly within the mitochondria. Excessive mitochondrial ROS can trigger dysfunction and apoptosis, but a moderate increase can paradoxically activate adaptive antioxidant responses that enhance cell survival, a phenomenon known as mitohormesis. In the new experiments, AML cells co-cultured with stromal cells showed significantly higher intracellular ROS levels, yet their viability was unchanged, implying that compensatory antioxidant defences had been induced. Those cells displayed greater total antioxidant activity and increased tolerance to hydrogen peroxide, and the same pattern appeared in primary AML cells taken from patients.

To test whether antioxidant capacity was truly the determinant of chemoresistance, the researchers manipulated it chemically. N-acetyl-L-cysteine, a precursor of glutathione, the cell’s principal antioxidant molecule, mimicked the stromal effect by enhancing antioxidant activity and drug resistance. Conversely, 2-phenylethyl isothiocyanate, which depletes glutathione, abolished antioxidant function and sensitized AML cells to both oxidative stress and chemotherapy. These results established that stromal cells enhance the antioxidant capacity of leukaemic cells, thereby conferring resistance to oxidative damage and cytotoxic drugs.

The next question was which upstream signal mediates this adaptation. Because mitochondrial ATP production was known to rise in AML cells co-cultured with stromal cells, the team turned to purinergic receptors, sensors of extracellular nucleotides that regulate tumour cell survival, metabolism and therapy response. Their attention settled on P2RX7, a ligand-gated ion channel on the plasma membrane activated by extracellular ATP. P2RX7 is highly expressed in leukaemic blasts and leukaemia-initiating cells, and its elevated expression correlates with poor clinical outcomes. Functionally, it has been implicated in promoting leukaemic cell survival, proliferation and homing to the bone marrow niche. In the new study, P2RX7 protein was markedly upregulated in AML cells co-cultured with stromal cells, and the same upregulation appeared when primary AML cells were grown with mesenchymal stromal cells extracted from AML patients themselves.

Functionally, the picture was equally clear. Treating AML cells with BzATP, a P2RX7 agonist, reduced their sensitivity to cytarabine and daunorubicin, while the selective antagonist AZ10606120 blocked that effect and partially reversed the protection conferred by stromal co-culture. Using CRISPR/Cas9-mediated knockout and lentiviral overexpression, the researchers confirmed the causal role: genetic ablation of P2RX7 impaired both antioxidant capacity and chemoresistance, effects that could be rescued by glutathione supplementation, whereas overexpression enhanced both, an effect abolished by glutathione depletion. To directly link P2RX7-mediated antioxidant defence to protection against oxidative damage, the team measured 8-hydroxy-2′-deoxyguanosine, a marker of oxidative DNA damage. Cytarabine increased 8-OHdG levels, and this effect was further augmented in P2RX7-deficient cells. Co-culture with stromal cells failed to reduce 8-OHdG accumulation in P2RX7-null cells, indicating that stromal support is insufficient to compensate for P2RX7 loss. Overexpressing cells, in contrast, displayed reduced 8-OHdG levels after drug exposure.

The mechanistic core of the study concerns how P2RX7 imposes mitochondrial stress that ultimately licences a robust antioxidant programme. As a calcium-permeable channel, P2RX7 activation promotes cytosolic calcium influx, sodium influx and potassium efflux. Using the calcium chelator BAPTA-AM, the researchers showed that reducing cytosolic calcium decreased both mitochondrial calcium uptake and mitochondrial ROS levels. Pharmacological inhibitors of mitochondrial calcium uptake, SN-6 and Ru360, suppressed mitochondrial calcium loading and ROS, whereas CGP37157, which activates the mitochondrial calcium uniporter, increased both. BzATP induced rapid calcium influx in control AML cells but failed to elicit any rise in P2RX7-null cells, confirming the calcium signal is P2RX7 dependent. Overexpression of P2RX7 enhanced mitochondrial calcium accumulation and ROS, effects abolished by the uptake inhibitors. Critically, the mitochondria-targeted ROS scavenger mitoTEMPO sensitized P2RX7-overexpressing cells to chemotherapy, while the mitochondria-targeted pro-oxidant mitoPQ restored antioxidant activity and chemoresistance in P2RX7-deficient cells. This demonstrates that stromal cell-induced P2RX7 upregulation drives mitochondrial calcium overload and oxidative stress, which in turn activates adaptive antioxidant defences.

Downstream of mitochondrial stress, the study identifies the transcription factor Nrf2 as the essential effector. Under basal conditions, Nrf2 is bound by Kelch-like ECH-related protein 1, or Keap1, which promotes its continuous ubiquitin-proteasomal degradation and thereby maintains redox homeostasis. Upon oxidative stress, Nrf2 escapes Keap1-mediated degradation, accumulates and translocates to the nucleus, where it binds antioxidant response elements in the promoters of cytoprotective genes including CAT, NQO1 and HMOX1. In co-cultured AML cells, Nrf2 protein levels rose significantly while its messenger RNA remained unchanged, indicating post-transcriptional regulation. Genetic ablation of Nrf2 abolished the ability of P2RX7 overexpression to enhance antioxidant capacity and chemoresistance, establishing Nrf2 as the necessary downstream effector. ARE-driven luciferase reporter activity was impaired by P2RX7 knockout and enhanced by overexpression, with the latter effect suppressed by mitoTEMPO, confirming that mitochondrial oxidative stress is required for P2RX7-mediated Nrf2 activation.

How does P2RX7-driven mitochondrial stress stabilize Nrf2? The study identifies PGAM5, a protein anchored to the outer mitochondrial membrane, as the critical retrograde signalling hub. PGAM5 loss did not affect basal Nrf2 levels in monocultured cells but markedly attenuated the Nrf2 increase observed under stromal co-culture, and P2RX7 overexpression failed to induce Nrf2 accumulation in PGAM5-deficient cells. Neither mitoTEMPO nor mitoPQ altered Nrf2 levels in PGAM5-knockout cells, suggesting PGAM5 is essential for transducing mitochondrial ROS signals to Nrf2. PGAM5 deletion also abolished the ability of P2RX7 to enhance antioxidant capacity and chemoresistance. Co-immunoprecipitation experiments revealed that stromal co-culture promoted the interaction between PGAM5 and Keap1, whereas P2RX7 knockout impaired this interaction, and mitoPQ restored PGAM5-Keap1 binding in the absence of P2RX7. In PGAM5-deficient cells, P2RX7 overexpression was unable to disrupt the Keap1-Nrf2 complex. The relay model that emerges is one in which P2RX7-induced mitochondrial stress causes PGAM5 to engage Keap1, coinciding with reduced Keap1-Nrf2 binding, decreased Nrf2 ubiquitination, slower Nrf2 degradation, and activation of the Nrf2-ARE antioxidant programme.

Finally, the therapeutic implications were tested in vivo. In systemic AML mouse models established with HL-60 or U937 cells in immunodeficient NCG mice, and repeated in an MLL-AF9-driven murine AML model in C57BL/6 mice with normal immune systems, mice received vehicle, cytarabine, AZ10606120, or the combination for five consecutive days. Monotherapy with either agent had minimal impact on leukaemic burden, but combination therapy significantly reduced spleen and liver infiltration, decreased AML cell burden in the bone marrow, relieved hepatosplenomegaly, and significantly prolonged survival. Ex vivo analysis of residual bone marrow AML cells showed that AZ10606120 monotherapy reduced total antioxidant capacity and increased 8-OHdG accumulation, effects further exacerbated by combination treatment. The enhanced efficacy of cytarabine in vivo is therefore at least partially attributable to the ability of P2RX7 blockade to compromise antioxidant defences, rendering leukaemic cells more vulnerable to oxidative stress and chemotherapeutic damage. The researchers caution that clinical translation of P2RX7 inhibitors requires careful consideration of potential off-target effects given the receptor’s pleiotropic roles in immunity and normal tissue homeostasis, and note that highly selective antagonists and advanced delivery strategies such as liposomal or bone marrow-targeting systems could mitigate systemic exposure. They also emphasize that stromal cells interact with AML cells through multiple bidirectional mechanisms, including direct cell-cell contact, paracrine signalling and extracellular vesicles, raising the possibility of a feed-forward loop that reinforces the protective leukaemic niche. Future studies with larger, genotype-stratified primary AML cohorts and patient-derived xenograft models will be needed to determine whether this axis is preferentially active in specific molecular subtypes and whether it functionally intersects with other resistance pathways.

Subject of Research: Acute myeloid leukaemia

Article Title: Bone marrow stromal cells promote chemoresistance in acute myeloid leukaemia through P2RX7-mediated antioxidant defence

Article References: You, R., Wang, X., Zheng, X., Fang, W., Jiang, C., Liu, J., Xu, X., Tan, M., Cai, D., Hou, D., Zheng, J., Lin, X., & Huang, H. (2026). Bone marrow stromal cells promote chemoresistance in acute myeloid leukaemia through P2RX7-mediated antioxidant defence. Experimental & Molecular Medicine, 58(9), 3025-3041. https://doi.org/10.1038/s12276-026-01839-4

Image Credits: AI Generated

DOI: 10.1038/s12276-026-01839-4

Keywords: Bone, marrow, stromal, cells, promote, chemoresistance, acute, myeloid, leukaemia, P2RX7-mediated, antioxidant, defence