In vitro and in silico evaluation of the synergistic effect of Cytarabine and Zataria multiflora Boiss extract on pre-B acute lymphoblastic leukemia cells (NALM-6 cell line).

BACKGROUND: Although cytarabine remains a cornerstone chemotherapeutic agent for Acute lymphoblastic leukemia (ALL), its clinical efficacy is frequently compromised by systemic toxicity and resistance mechanisms, notably telomerase activation via hTERT upregulation. This study investigates the synergistic potential of Zataria multiflora extract (ZME) combined with cytarabine to induce apoptosis in pre-B ALL cells (NALM-6) and explores the underlying molecular mechanisms. METHODS: In vitro experiments were conducted using the NALM-6 cell line. Cell viability was assessed via Trypan blue exclusion and MTT assays. Apoptosis was quantified using Annexin V/PI flow cytometry. The mRNA expression levels of BAX, BCL2, and hTERT were evaluated by qRT-PCR. Additionally, in silico molecular docking was performed to analyze the interactions between the primary active components of ZME (thymol and carvacrol), cytarabine, and the MDM2 protein. RESULTS: Single-agent treatments (100 µg/mL ZME or 0.5 µM cytarabine) yielded 70-80% cell viability after 48 h. However, the combination therapy significantly enhanced cytotoxicity, inducing 47.54% apoptosis. Gene expression analysis revealed an elevated BAX/BCL2 ratio and a significant downregulation of hTERT, indicating activation of the intrinsic apoptotic pathway and inhibition of telomerase activity. In silico docking demonstrated strong binding affinities between thymol/carvacrol and the MDM2 active site, suggesting a hypothetical disruption of the MDM2-p53 interaction. CONCLUSION: The combination of ZME and cytarabine synergistically enhances apoptosis in NALM-6 cells by concurrently modulating apoptotic signaling and telomerase-related gene expression. These findings underscore the potential of ZME as a chemosensitizing agent in ALL and provide a rationale for further in vivo validation.