Combined Effects of 5-Azacytidine and Oleuropein on miR-149-3p, miR-375, miR-574-5p Expression and Apoptosis in HL-60 and THP-1 Cell Lines

Abstract Background Acute myeloid leukemia (AML) is an aggressive hematological malignancy characterized by the rapid expansion of immature myeloid cells and poor clinical outcomes. Despite conventional treatments, including chemotherapy and hematopoietic stem cell transplantation, relapse and resistance remain significant challenges. Epigenetic alterations, particularly dysregulated DNA methylation and microRNA (miRNA) expression, play a crucial role in AML pathogenesis. Objective This study aimed to evaluate the synergistic effects of azacitidine, a DNA methyltransferase inhibitor, and oleuropein, a natural polyphenol with anticancer properties, on AML cell lines (THP-1 and HL-60). Methods AML cells were treated with azacitidine, oleuropein, and their combination. Cell proliferation was assessed using MTT assays, apoptosis was analyzed via flow cytometry (Annexin V-FITC/PI staining), and miRNA expression levels were quantified using real-time PCR. Results Both azacitidine and oleuropein reduced cell viability and induced apoptosis in a dose- and time-dependent manner. Notably, the combination treatment significantly enhanced apoptosis, with a 2.5-fold increase in Annexin V-positive HL-60 cells at 72 hours. Furthermore, the treatment modulated miRNA expression, upregulating miR-149-3p and miR-375 while downregulating miR-574-5p. Conclusion The synergistic effects of oleuropein and azacitidine suggest a potential therapeutic strategy for AML by targeting epigenetic mechanisms and miRNA pathways. Further in vivo studies and clinical trials are necessary to validate these findings and optimize treatment protocols.