Silibinin, Synergistically Enhances Vinblastine-Mediated Apoptosis in Triple Negative Breast Cancer Cell Line: Involvement of Bcl2/Bax and Caspase-3 Pathway

Abstract

Background: Triple-negative breast cancer (TNBC) with a poor prognosis and survival is the most invasive subtype of breast cancer.  Usually, TNBC requires a chemotherapy regimen at all stages, but chemotherapy drugs have shown many side effects. We assumed that combination therapy of vinblastine and silibinin might reduce the vinblastine toxicity and dose of vinblastine. Materials and Methods: The MDA-MB-231 were cells subjected to MTT assay for IC50 determination and combination effects, which were measured based on Chou-Talalay's method. The type of cell death was determined by using a Flow-cytometric assay. Cell death pathway markers, including Bcl-2, Bax, and caspase-3 were analyzed by western blot and Real-Time PCR. Results: The treatment of MDA-MB-231 cells exhibited IC50 and synergism at the combination of 30 µM of silibinin and 4 µm of vinblastine in cell viability assay (CI=0.69). YO-PRO-1/PI double staining results showed a significant induction of apoptosis when MDA-MB-231 cells were treated with a silibinin and vinblastine combination (p<0.01). Protein levels of Bax and cleaved caspase-3 were significantly upregulated, and Bcl-2 downregulated significantly. Significant upregulation of Bax (2.96-fold) and caspase-3 (3.46-fold) while Bcl-2 was downregulated by 2-fold. Conclusion: Findings established a preclinical rationale for the combination of silibinin and vinblastine. This combination produces synergistic effects in MDA-MB-231 cells by altering pro- and anti-apoptotic genes, which may reduce the toxicity and side effects of vinblastine.