Triptolide promotes ferroptosis by suppressing Nrf2 to overcome leukemia cell resistance to doxorubicin

Doxorubicin (DOX) is an extensively used chemo- therapeutic drug to treat leukemia. However, there remains a pivotal clinical problem of resistance to DOX in patients with leukemia. Erythroid 2-related factor 2 (Nrf2) is a master regulator of antioxidation response which serves a critical role in maintaining cellular oxidative homeostasis. However, whether Nrf2 is involved in DOX resistance is not totally clear. It is well-documented that triptolide, a widely used drug to treat autoimmune disorders, possesses anti-cancer activi- ties, yet whether triptolide affects leukemia cell sensitivity to DOX remains to be elucidated. The present study aimed to determine the role of triptolide-mediated downregulation of Nrf2 in regulating leukemia cell ferroptosis and resistance to DOX. For this purpose, low-dose DOX was used to establish DOX-resistant K562 cells and HL-60 cells. Nrf2 mRNA and protein expression were examined by quantitative PCR and western blotting assays. The effects of triptolide on leukemia cell viability, reactive oxygen species (ROS) levels, or lipid oxidation were determined by CCK8 assay, DCFH-DA assay, or BODIPY 581/591 C11 assay, respectively. The results show that Nrf2 expression was significantly upregulated in DOX-resistant leukemia cells and clinical leukemia samples. Silencing of Nrf2 significantly sensitized leukemia cells to DOX. Furthermore, it was demonstrated that triptolide inhib- ited Nrf2 expression and induced leukemia cell ferroptosis, as evidenced by increased ROS levels and lipid oxidation as well as decreased glutathione peroxidase 4 expression. Ectopic expression of Nrf2 significantly rescued triptolide-induced leukemia cell ferroptosis. Notably, the present study showed that triptolide re-sensitized DOX-resistant leukemia cells to DOX. In conclusion, the present study indicated that Nrf2 served a critical role in leukemia cell resistance to DOX and triptolide-induced ferroptosis and suggested a potential strategy of combination therapy using triptolide and DOX in leukemia treatment.