Demethylzeylasteral alleviates myeloid leukemia through PERK/eIF2α/ATF4/CHOP-mediated cell apoptosis

Sep 30, 2026

Uploaded by Jacob

Myeloid leukemia requires urgently developed therapeutic drugs, and demethylzeylasteral (DML), a terpenoid extract from Tripterygium, was investigated for its effects on leukemia cell proliferation and apoptosis. DML reduced viability and triggered G0/G1 arrest and apoptosis with ER-stress signatures, including increased intracellular calcium, ROS generation, ATP depletion, and unfolded protein accumulation. Mechanistic work showed PERK/eIF2α/ATF4/CHOP pathway activation and an irrecoverable UPR leading to apoptotic death, validated by inhibitors, shRNA, RT-qPCR, and Western blot, with in vivo survival prolongation in an NB4-derived xenograft model.

LanguageEnglish

CategoryHealthcare

FormatPDF

Pages18

Reading Time45 min

AccessFree to download

Background

Study aim

Methods

Cell assays and measurements

Transcriptome and pathway analysis

In vivo xenograft evaluation

Results

Screening and validation

PERK/eIF2α/ATF4/CHOP activation and apoptosis

Therapeutic impact in vivo

Conclusions

Key findings

Demethylzeylasteral alleviates myeloid leukemia through PERK/eIF2α/ATF4/CHOP-mediated cell apoptosis Xue Li1, Xianchao He1, Leyan Zhang2, Zhigang Li3, Yuanyuan Liu3 1Medical School of Guizhou University, Guiyang, China; 2International Medical College, Chongqing Medical University, Chongqing, China; 3Department of Radiological Medicine, School of Basic Medical Sciences, Chongqing Medical University, Chongqing, China Contributions: (I) Conception and design: X Li, Y Liu; (II) Administrative support: None; (III) Provision of study materials or patients: None; (IV) Collection and assembly of data: X Li, X He, L Zhang, Z Li; (V) Data analysis and interpretation: X Li, Y Liu; (VI) Manuscript writing: All authors; (VII) Final approval of manuscript: All authors. Correspondence to: Yuanyuan Liu, PhD. Department of Radiological Medicine, School of Basic Medical Sciences, Chongqing Medical University, No. 1 Yixueyuan Road, Yuzhong District, Chongqing 400016, China. Email: [liuyuanyuan@cqmu.edu.cn](liuyuanyuan@cqmu.edu.cn). Background: Myeloid leukemia, as a malignant tumor of hematologic system, urgently requires discovery of novel therapeutic drugs. Demethylzeylasteral (DML), a natural terpenoid primary extract from Tripterygium, has demonstrated antitumor properties across various cancer types. However, its functions in myeloid leukemia have not been clearly elucidated. This study aimed to explore the effects of DML on the proliferation and apoptosis of myeloid leukemia cells, and elucidate its mechanism. Methods: Cell viability was detected by Cell Count Kit-8 (CCK-8) method. Post-DML treatment, cell cycle, apoptosis, intensity of unfolded proteins, adenosine triphosphate (ATP), intracellular calcium, and reactive oxygen species (ROS) levels were assessed by flow cytometry. Additionally, transcriptome sequencing and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were conducted to uncover pathways linked to gene enrichment in the DML treatment group, which were validated using small molecular inhibitors, short hairpin RNA (shRNA) interference, flow cytometry, reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis. In vivo, the anti-leukemia activity of DML was evaluated in a NB4 cell-derived xenograft model using NOD-PrkdcscidIl2rgem1/Smoc (M-NSG) mice. Results: A library of 65 terpenoids was screened against myeloid leukemia cell lines (K562, NB4, THP-1), identifying DML through screening and validation using CCK-8 assays. Comprehensive in vitro analyses revealed that DML treatment significantly reduced cell viability while inducing G0/G1 phase arrest and apoptosis, accompanied by characteristic endoplasmic reticulum (ER) stress markers including elevated intracellular calcium, ROS generation, ATP depletion, and unfolded protein accumulation. Transcriptomic profiling combined with mechanistic validation through shRNA knockdown and pharmacological inhibitors demonstrated that DML activated the PERK/eIF2α/ATF4/CHOP signaling axis, initiating an irrecoverable unfolded protein response (UPR) that culminated in apoptotic cell death. The therapeutic relevance of these findings was substantiated in vivo, where DML administration significantly prolonged survival in an M-NSG mouse xenograft model engrafted with NB4 leukemia cells. Conclusions: These results systematically establish that DML induces apoptosis in myeloid leukemia cells directly through PERK/eIF2α/ATF4/CHOP signaling pathway in response to ER stress. Keywords: Demethylzeylasteral (DML); myeloid leukemia; endoplasmic reticulum stress (ER stress); PERK/ eIF2α/ATF4/CHOP signaling; unfolded protein response (UPR) Submitted Jul 26, 2025. Accepted for publication Nov 05, 2025. Published online Dec 18, 2025. doi: 10.21037/tcr-2025-1629 View this article at: [https://dx.doi.org/10.21037/tcr-2025-1629](https://dx.doi.org/10.21037/tcr-2025-1629) © AME Publishing Company. Transl Ca

FAQs

What is the main therapeutic mechanism by which DML targets myeloid leukemia cells?

DML activates the PERK/eIF2α/ATF4/CHOP signaling axis, initiating an irrecoverable unfolded protein response (UPR) in the context of ER stress, which culminates in apoptotic cell death.

How was DML’s effect on leukemia cells evaluated in vitro?

Cell viability was assessed using CCK-8, while apoptosis, cell cycle changes, and multiple ER-stress-related readouts (ATP, calcium, ROS, unfolded protein accumulation) were measured by flow cytometry. Mechanistic validation used transcriptome profiling along with shRNA and pharmacological inhibitors, supported by RT-qPCR and Western blot.

Does DML show anti-leukemia activity in vivo?

Yes. In an NB4 cell-derived xenograft model in M-NSG mice, DML administration significantly prolonged survival, supporting its therapeutic relevance beyond in vitro findings.

Sep 30, 2026

Uploaded by Jacob