Modulation of Drug Resistance and Apoptotic Pathways Underlies the Enhanced Antitumor Effect of Ellagic Acid–Irinotecan Combination in Glioma

Gliomas account for over half of all primary malignant tumors of the central nervous system and remain associated with poor prognosis. Although irinotecan is an effective chemotherapeutic agent, its clinical utility is limited by systemic toxicity, prompting interest in phytochemicals such as ellagic acid (EA) as potential sensitizers. This study aimed to investigate whether EA enhances the antiproliferative and pro-apoptotic effects of irinotecan in C6 glioma cells. C6 glioma cells were treated with EA (100 µM), irinotecan (100 µM), or their combination for 24, 48, and 72 h. Cell proliferation was assessed by BrdU assay, p53 and caspase-3 protein expression by immunocytochemistry (H-SCORE), and multidrug resistance gene 1 (MDR1), MGMT, p53, and caspase-3 mRNA levels by RT-qPCR. EA significantly enhanced irinotecan-mediated suppression of proliferation at 24 h (p < 0.001), 48 h (p < 0.001), and 72 h (p < 0.001), with the combination producing the strongest inhibition across all time points. Immunocytochemical p53 expression increased significantly in all treatment groups at 24 h and 48 h (EA: p < 0.01; irinotecan: p < 0.01; EA + irinotecan: p < 0.01) and remained elevated at 72 h (p < 0.05). Caspase-3 immunoreactivity showed robust early activation at 24 h (Ir: p < 0.05; EA: p < 0.01), persisted at 48 h (p < 0.01), and remained significantly elevated in the EA group at 72 h (p < 0.001). At the mRNA level, irinotecan induced the highest p53 expression at 24 h (p < 0.001), with sustained elevation at 48 h and 72 h (p < 0.001 and p < 0.05, respectively). Caspase-3 mRNA peaked at 24 h only in the irinotecan group (p < 0.001). EA significantly increased MDR1 and MGMT transcription at 24 h and 48 h (p < 0.001), whereas the EA + irinotecan combination attenuated this increase and remained close to control levels at early time points. MGMT remained significantly elevated in EA and EA + irinotecan groups through 72 h (p < 0.001). EA cooperatively enhanced the antitumor activity of irinotecan primarily by enhancing proliferation inhibition and modulating drug-resistance gene expression, while maintaining, rather than further augmenting, apoptotic protein markers comparable to those induced by single-agent treatments. These findings support EA as a promising adjunct to irinotecan-based glioma therapy.