In vivo acute toxicity evaluation and in vitro molecular mechanism study of antiproliferative activity of a novel indole Schiff base β-diiminato manganeseIII complex in hormone-dependent and triple negative breast cancer cells

摘要

Breast cancer is the most frequently diagnosed cancer among women worldwide. Recently, increasing attention has been paid to the anticancer effects of transition metal complexes of indole Schiff bases. β-diiminato Manganese^III complex has shown promising cell cycle arrest and apoptosis induction against MCF-7 and MDA-MB-231 breast cancer cells. In this study, time- and dose- dependent inhibitory activity were evaluated using MTT assay after 48 h and 72 h exposure time. In addition, median effect analysis was conducted according to Chou–Talalay method to investigate whether Mn^III complex has synergistic effect in combination with chemotherapeutic drugs on inhibiting breast cancer cell growth. The molecular mechanisms underlying its potent antiproliferative effect was determined through bioluminescent caspase-3/7, -8 and -9 activity assays and quantitative expression analysis of cell cycle- and apoptosis-related genes. Furthermore, safety evaluation of Mn^III complex was assessed through the acute oral toxicity test in in vivo model. The MTT assay results revealed that it potently reduced the viability of MCF-7 (IC50 of 0.63 ± 0.07 µg/mL for 48 h and 0.39 ± 0.08 µg/mL for 72 h) and MDA-MB-231 (1.17 ± 0.06 µg/mL for 48 h, 1.03 ± 0.15 µg/mL for 72 h) cells in dose- and time-dependent manner. Combination treatment also enhanced the cytotoxic effects of doxorubicin but not tamoxifen on inhibiting breast cancer cell growth. The involvement of intrinsic and extrinsic pathway in apoptosis induction was exhibited through the increased activity of caspase-9 and caspase-8, respectively, leading to enhanced downstream executioner caspase-3/7 activity in treated MCF-7 and MDA-MB-231 cells. In addition, gene expression analysis revealed that Mn^III complex exerts its antiproliferative effect via up-and down-regulation of p21 and cyclin D1, respectively, along with increased expression of Bax/Bcl-2 ratio, TNF-α, initiator caspase-8 and -10 and effector caspase-3 in MCF-7 and MDA-MB-231 cells. However, the results did not show increased caspase-8 activity in treated MCF-7 cells. Furthermore, in vivo acute oral toxicity test revealed no signs of toxicity and mortality in treated animal models compared to the control group. Collectively, the promising inhibitory effect and molecular and mechanistic evidence of antiproliferative activity of Mn^III complex and its safety characterization have demonstrated that it may have therapeutic value in breast cancer treatment worthy of further investigation and development.