Augmenting the cytotoxicity and apoptosis induction of green synthesized copper oxide nanoparticles from Artemisia sieberi by combining with tamoxifen: Mechanism involving Bax/Bcl-2 modulation.

Treatment of breast cancer is severely hindered by dose-limiting toxicity and acquired Tamoxifen (Tam) resistance. This study investigated the synergistic anti-cancer effect and molecular mechanism of green-synthesized Copper Oxide Nanoparticles (CuO NPs) when combined with Tam against the MCF-7 human breast cancer cell line. CuO NPs were successfully produced using an aqueous extract of Artemisia sieberi as a dual reducing/capping agent. Characterization confirmed the formation of stable nanoparticles, evidenced by a Surface Plasmon Resonance (SPR) peak at 552 nm, an average size of ∼36 nm (TEM), and excellent colloidal stability with a highly negative Zeta Potential (-33.5 mV). The MTT assay demonstrated that the combination regimen exhibited a strong synergistic cytotoxic effect (CI<1) toward MCF-7 cells. Synergy was achieved even at low concentrations, such as 15.0 μg/mL CuO NPs:1.0 μM Tam (CI=0.99), with the most potent synergy (CI=0.69) observed at the maximum dose. Quantitative analysis via Annexin V/PI flow cytometry confirmed this enhancement, showing the combination reduced the viable cell population to a minimal 4.87 %. Mechanistically, RT-qPCR analysis revealed a maximal perturbation of the mitochondrial apoptotic pathway. The combined treatment simultaneously maximized the up-regulation of the pro-apoptotic gene BAX and the down-regulation of the anti-apoptotic gene BCL-2. This synergistic action resulted in a 16.74-fold increase in the critical BAX/BCL-2 ratio, approximately two times greater than either monotherapy. These findings validate CuO NPs co-administration with Tamoxifen as a promising strategy for overcoming acquired Tamoxifen resistance.