In Vitro Evaluation of the Potential Synergistic Interactions Between Apigenin and Bortezomib in U266 Multiple Myeloma Cells

Purpose This study aimed to evaluate the cytotoxic, apoptotic, and cell cycle regulatory effects of the combination of Apigenin, a natural flavonoid, and the proteasome inhibitor Bortezomib on U266 multiple myeloma cells, and to quantify the potential synergistic interaction. Methods U266 cells were treated with different doses of Bortezomib and Apigenin at 24, 48, and 72 hours of incubation. Cell viability was assessed using the CCK-8 assay, and the Combination Index (CI) was calculated using the Chou–Talalay method. Additionally, apoptotic response and changes in PI3K/MAPK/Bcl-2 protein levels were examined using Annexin V/Dead Cell, cell cycle (Muse™ Cell Cycle), and ELISA analyses. Results Combination therapy was characterized by a significant decrease in cell viability compared to single-agent treatments, an increase in early and late apoptosis rates, and suppression of the PI3K/MAPK/Bcl-2 pathways. CI < 1 was detected in all dose groups, with the strongest synergy observed in the 50 nM Bortezomib + 5 µM Apigenin combination (CI = 0.16996). Apigenin treatment caused S-phase accumulation in the cell cycle, while Bortezomib induced G₂/M phase arrest; the combination balanced these effects and significantly enhanced apoptosis. Conclusions The findings demonstrate that Apigenin is not only a supportive phytochemical but also a multi-targeted chemotherapeutic modulator capable of enhancing the therapeutic efficacy of Bortezomib. This combination has the potential to reduce treatment-related neurotoxicity and resistance development by providing similar antitumor activity at lower doses. In conclusion, the Apigenin–Bortezomib combination emerges as a strong and clinically promising candidate for next-generation adjuvant strategies in multiple myeloma treatment, based on biological synergy.