Carnosic Acid Mediates Production of Reactive Oxygen Species to Regulate Mitogen-Activated Protein Kinase Pathway Phosphorylation and Induce Apoptosis in Human Breast Cancer Cells.

BACKGROUND: Reactive oxygen species (ROS) can induce cancer cell apoptosis, which plays a crucial role in breast cancer therapy. Carnosic acid (CA) exerts an anti-tumor effect via generating ROS or activating the mitochondria-related apoptosis pathway in vitro and in vivo. The deoxy-glucose derivative CN5DG labeled with (99m)Tc can be used for breast cancer imaging and evaluating the therapeutic efficacy of CA. METHODS: Inhibition of cancer cell proliferation by CA was assessed by MTT and cell colony-formation assays. The mechanism of CA-induced cancer cell apoptosis was examined by western blotting and the apoptosis rate was detected by flow cytometry. The in vivo anti-tumor effect of CA was assessed by immunohistochemistry and (99m)Tc-CN5DG imaging. Its biodistribution was determined to evaluate the accumulation of (99m)Tc-CN5DG in xenografts. RESULTS: CA promoted cancer cell apoptosis via ROS generation, which activated c-Jun N-terminal kinase (JNK) and p38 phosphorylation. The apoptosis rates of T47D and MCF7 cells treated with CA (IC50 concentration) for 24 h were 44.97% ± 1.56% and 39.74% ± 1.78%, respectively. The antioxidant N-acetyl-L-cysteine (5 μM) abolished CA-induced apoptosis. MCF-7 xenografts without CA were visualized in vivo at all time points by (99m)Tc-CN5DG imaging (tumor/muscle ratio: 2.52 ± 0.10 at 4 h), but xenografts treated with CA were not visualized (tumor/muscle ratio: 1.36 ± 0.34 at 4 h). Evaluation of the biodistribution by γ-counter also demonstrated the efficacy of (99m)Tc-CN5DG. Bcl-2 and Ki-67 expression were higher while Bax expression was lower in the control group compared with the CA treatment group. The tumor/muscle radioactivity count ratio was lower following treatment with CA compared with the control group. CONCLUSION: CA can induce breast cancer apoptosis via ROS generation and activation of JNK and p38 phosphorylation. The anti-tumor effect of CA can be assessed using (99m)Tc-CN5DG SPECT imaging.