Bioinformatics-Guided Discovery of miRNAs Involved in Apoptosis Modulated by Parthenolide Combined with Vincristine in The NALM6 Cell Line

Abstract

Objective: Acute lymphoblastic leukemia (ALL) is a highly heterogeneous leukemia. Despite the current improvement inconventional chemotherapy and high survival rates, the outcomes remain challenging. Sesquiterpen extracted from theTanacetum parthenium, parthenolide, is a potential anticancer agent that can modulate the expression of miRNAs and induceapoptosis. The objective of this study was to investigate the effect of parthenolide in combination with vincristine and alone onthe apoptosis rate and expression of miR-125b-5p, miR-181b-5p, and miR-17-5p in the NALM6 cell line.Materials and Methods: In this experimental study, cell viability and metabolic activity were determined through MTTassay and PI staining. Flow cytometry was applied to evaluate the rate of apoptosis. The expression of miRNAs wasassessed using real-time polymerase chain reaction. Bioinformatic analyses, including Cytoscape, RNAhybrid, andsignaling pathway analysis were employed to investigate the association of miR-17-5p, miR-181b-5p and miR-125b-5p with apoptosis. Further, molecular docking served to validate the modulation of these miRNAs by parthenolide andvincristine treatment.Results: The MTT assay indicated that 7.7 μM of parthenolide decreased the metabolic activity to 50% after 48 hours. PIstaining analysis indicated that at concentrations below the half maximal inhibitory concentration, parthenolide caused50% cell death. Flow cytometric analysis indicated that parthenolide (1.925 μM) in combination with vincristine (1.2 nM)induced apoptosis in 83.2% of the cells. Real-time quantitative reverse transcription polymerase chain reaction (qRTPCR)analysis showed significant changes in the expression levels of miR-17-5p, miR-125b-5p, and miR-181b-5p.Moreover, the combination therapy downregulated the expression of miRNAs significantly. This was consistent with ourbioinformatic analysis demonstrating that the studied miRNAs are regulators of apoptosis. Finally, molecular dockingvalidated the modulation of the miRNAs by parthenolide and vincristine.Conclusion: Parthenolide in combination with vincristine triggers apoptosis at a high rate in the NALM6 cell line.Moreover, this combination therapy can decrease the expression of miR-17-5p, miR-181b-5p, and miR-125b-5p.

Keywords

• apoptosis
• b-acute lymphoblastic leukemia
• bioinformatics
• mirnas
• parthenolide