Lenalidomide improvement of cisplatin antitumor efficacy on triple-negative breast cancer cells in vitro

Abstract

Lenalidomide is an immunomodulatory drug and possesses anti-angiogenic and immunomodulatory activities against multiple myeloma. The present study assessed the in vitro effect of lenalidomide combined with cisplatin on MDA-MB-231, a triple-negative breast cancer (TNBC) cell line and explored the underlying molecular mechanism of this combination. Cell viability, apoptosis and the protein expression of phosphorylated (p) and total extracellular signal-regulated kinase (ERK), B-cell lymphoma-2 (Bcl-2), caspase-3, cleaved poly-adenosine diphosphate-ribose polymerase (cPARP), vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) were measured in MDA-MB-231 cells treated with different concentrations of lenalidomide, cisplatin and their combination using different biochemical assays. Lenalidomide demonstrated no significant effect on the cell viability of MDA-MB-231 cells, even at high concentrations, whereas lenalidomide in combination with cisplatin, significantly reduced cisplatin IC₅₀ from 7.8 to 3.0 µM in MDA-MB-231 cells. In addition, lenalidomide and cisplatin in combination significantly induced cell apoptosis by 1.6- and 1.38-fold, respectively compared with lenalidomide and cisplatin alone (P<0.05). The expression levels of VEGF, bFGF and Bcl-2 proteins were significantly reduced (P<0.01), whereas caspase-3 and cleaved PARP expression were significantly increased in MDA-MB-231 cells treated with the combination compared to those treated with single agents (P<0.01). Lenalidomide treatment alone significantly reduced the p-ERK level compared with the control (P<0.05) and cisplatin treatment alone significantly increased it (P<0.01), however treatment with them in combination significantly reduced the p-ERK level in MDA-MB-231 cells compared with cisplatin treatment alone (P<0.05). In conclusion, the present study provides the basis for using lenalidomide in combination with cisplatin in TNBC therapy.

Keywords: apoptosis; cisplatin; lenalidomide; triple-negative breast cancer.

Figure 1.

Effect of lenalidomide and cisplatin and their combination on MDA-MB-231 cell viability in vitro. MDA-MB-231 cells were grown and treated with (A) cisplatin at varying concentrations for 72 h. *P<0.05, **P<0.01 and ***P<0.001 vs. 0.0 µM cisplatin group; (B) cisplatin at 24, 48 and 72 h; (C) lenalidomide; or (D) their combination up to 72 h and then subjected to cell viability MTT assay. *P<0.05, **P<0.01 and ***P<0.001 vs. corresponding cisplatin + lenalidomide group. NS, no significant difference.

Figure 2.

Effect of lenalidomide and cisplatin on MDA-MB-231 cell apoptosis in vitro. MDA-MB-231 cells were grown and treated with lenalidomide, cisplatin, or their combination for 72 h and apoptotic cells were detected using FITC Annexin V-FITC/PI apoptosis detection kit. The bar graph is a quantitative presentation of the flow cytometric data. **P<0.01 and ***P<0.001. NS, no significant difference; NC, negative control; Len, lenalidomide; Cis, cisplatin; FITC, fluorescein isothiocyanate; PI, propidium iodide; NC, negative control.

Figure 3.

Effect of lenalidomide and cisplatin on modulating the different protein expression in MDA-MB-231 cells. MDA-MB-231 cells were grown and treated with lenalidomide, cisplatin, or their combination for 72 h and then subjected to western blot analysis. The bar graphs are a quantitative presentation of the western blot analyses. *P<0.05 and **P<0.01. ERK, extracellular signal-regulated kinase; p-, phosphorylated; Bcl-2, B-cell lymphoma-2; cPARP, cleaved poly-adenosine diphosphate-ribose polymerase; VEGF, vascular endothelial growth factor; bFGF, basic fibroblast growth factor; NC, negative control.