Anticancer effect of silibinin on the xenograft model using MDA-MB-468 breast cancer cells

Abstract

Purpose: The aim of this study is to know whether silibinin has an anticancer effect on triple negative breast cancer xenograft model using MDA-MB-468 cells.

Methods: To establish the xenograft model, we injected the MDA-MB-468 cells into female Balb/c-nude mice. After establishing a xenograft model, oral silibinin was administered to the tested mice in the way of 200 mg/kg for 45 days. The difference of mean tumor volume between silibinin fed mice and control mice was analyzed. The epidermal growth factor receptor (EGFR) phosphorylation in MDA-MB-468 cells was analyzed by Western blotting. The expression of VEGF, COX-2, and MMP-9 genes in tumor tissue was analyzed by real-time polymerase chain reaction (PCR).

Results: In the xenograft model using MDA-MB-468 cells, we found that oral administration of silibinin significantly suppressed the tumor volume (silibinin treated mice vs. control mice; 230.3 ± 61.6 mm(3) vs. 435.7 ± 93.5 mm(3), P < 0.001). The phosphorylation of EGFR in MDA-MB-468 cells was inhibited by treatment with 50 µg/mL of silibinin. In real time-PCR analysis of tumor tissue obtained from sacrificed mice, the gene expression of MMP-9, VEGF, and COX-2 was 51.8%-80% smaller in silibinin group than that of control group and we can also verify the similar result using Western blotting analysis.

Conclusion: We verified that silibinin had anticancer effect on xenograft model of MDA-MB-468 cells in the way of preventing the phosphorylation of EGFR and eventually suppressed the production of COX-2, VEGF, and MMP-9 expression. Finally, the tumor volume of xenograft models was decreased after administration of Silibinin.

Keywords: Silibinin; Triple negative breast neoplasms; Xenograft.

Fig. 1

Establishment of triple negative breast cancer xenograft model using MDA-MB-468 cells.

Fig. 2

Epidermal growth factor receptor (EGFR) family expression of MDA-MB-231 and MDA-MB-468 cell line using Western blotting.

Fig. 3

Time-dependent change in tumor size of the control mice and the silibinin-treated mice. (A) Tumor size of the control mice and the silibinin-treated mice on the 45th day of silibinin administration; (B) tumor size of the control mice and the silibinin-treated mice extracted after mouse euthanasia; (C) difference of the mean tumor volume for each group on the 3rd day, the 10th day, the 17th day, the 24th day, the 31st day, the 38th day, and the 45th day for the silibinin-treated mice and the control mice. ^*P < 0.05; ^**P < 0.01.

Fig. 4

Epidermal growth factor receptor (EGFR) phosphorylation of MDA-MB-468 breast cancer cell treated with silibinin using Western blotting. Con, control.

Fig. 5

VEGF, COX-2, and MMP-9 gene expression using tumor tissue extracted from the triple negative breast cancer animal model. (A) According to real-time polymerase chain reaction analysis for tumor tissue, COX-2, MMP-9, and VEGF gene expression was reduced by 50%-80% in the silibinin-treated mice compared to the control mice; (B) Western blotting analysis also showed that cyclooxygenase-2 (COX-2) and vascular endothelial growth factor (VEGF) protein expression was reduced in the silibinin-treated mice.