Background: Breast cancer remains a leading cause of mortality among women, prompting the development of novel therapeutics. Nanoparticle-based agents and probiotic derivatives have emerged as promising candidates. This study evaluates the synergistic cytotoxic and apoptotic effects of zinc oxide (ZnO) nanoparticles and Lactococcus lactis lysate in MCF-7 breast cancer cells. Materials and Methods: Lactococcus lactis (NCTC 6681) was cultured in MRS broth, and its lysate was prepared by repeated freeze-thaw cycles. MCF-7 cells were treated with ZnO nanoparticles (3.125–1600 µg/mL), L. lactis lysate, the combination of ZnO nanoparticles and L. lactis lysate, and tamoxifen as a positive control. Cytotoxicity was assessed via MTT assay at 24, 48, and 72 hours. Expression levels of bax, bcl2, p21, casp9, and cdk4 were measured by Real-Time PCR, using GAPDH as the reference gene. Results: IC50 values at 24 hours for ZnO, L. lactis lysate, combination of ZnO and L. lactis lysate, and tamoxifen were 49.68 µg/mL, 61.53 µg/mL, 31.05 µg/mL, and 11.68 µg/mL respectively while at 48 and 72 hours, the combination showed enhanced cytotoxicity at IC50 = 26.38 and 24.79 µg/mL. Gene expression analysis revealed that the combination treatment significantly upregulated pro-apoptotic markers BAX (1.723-fold), CASP9 (1.75321-fold), and P21 (1.72914-fold), while downregulating the anti-apoptotic BCL2 (0.55287-fold) and cell cycle promoter CDK4 (0.544-fold). These effects closely paralleled those of tamoxifen, which induced comparable changes. (BAX: 1.8066-fold; CASP9: 1.82977-fold; P21: 1.79419-fold; BCL2: 0.50991-fold; CDK4: 0.49655-fold). Conclusion: Co-treatment with ZnO nanoparticles and L. lactis lysate exerts a synergistic pro-apoptotic effect on MCF-7 cells. These findings support their potential as an adjunct therapeutic strategy in breast cancer management.
