Realgar induces apoptosis by inhibiting glycolysis via regulating STAT3 in myelodysplastic syndrome.

ETHNOPHARMACOLOGICAL RELEVANCE

Myelodysplastic syndrome (MDS) is a hematologic malignancy that presents a unique opportunity for traditional Chinese medicine (TCM) to demonstrate its distinctive value in treatment. Realgar, a component of TCM, has shown notable potential in alleviating clinical symptoms and improving the prognosis of MDS patients. However, the precise mechanisms underlying the treatment of MDS with realgar, particularly its effects on apoptosis-related pathways, remain poorly understood.

AIM OF THE STUDY

This study aimed to investigate the pro-apoptotic effects of realgar on MDS cells and to elucidate the underlying molecular mechanisms.

MATERIALS AND METHODS

We explored the targets and pathways of realgar's action on MDS using public databases, network pharmacology, and RNA sequencing. Various techniques were employed, including cell transfection, Cell Counting Kit-8 (CCK8) assay, Cellular Thermal Shift Assay (CETSA), Western blot (WB), quantitative real-time polymerase chain reaction (qRT-PCR), apoptosis and glycolysis assays, extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) measurements, dual-luciferase reporter assays, and immunofluorescence, to investigate the regulatory mechanisms involving STAT3, glycolysis, and apoptosis. Hematoxylin and eosin (HE) staining was utilized to assess realgar's toxicity. Apoptosis and hemogram changes were analyzed to evaluate the therapeutic effect of realgar on MDS transgenic mice.

RESULTS

Analysis of public data indicated that apoptosis-related genes are downregulated in MDS patients. Through network pharmacology, CETSA, qRT-PCR, WB, apoptosis assays, and STAT3 overexpression cell transfection, we discovered that realgar inhibits STAT3 expression. Further investigation using RNA sequencing suggested that glycolysis may be involved in this regulatory process. ECAR, OCR, glycolysis assays, WB, apoptosis assays, and glycolysis inhibitor experiments demonstrated that glycolytic function was inhibited. Additionally, GLUT1 expression was significantly decreased, and GLUT1 was found to directly bind to STAT3. In MDS mice, realgar treatment enhanced levels of white blood cells, red blood cells, hemoglobin, and platelets, and increased apoptosis levels.

CONCLUSION

Our findings reveal that realgar exerts a significant pro-apoptotic effect on MDS cells in both in vivo and in vitro models. Further analysis demonstrated that realgar regulates the STAT3 pathway, leading to GLUT1-mediated glycolysis alterations that ultimately induce apoptotic pathways, as represented by BCL2. These discoveries hold significant implications for the basic research and clinical diagnosis and treatment of MDS.