Effect and mechanism of silibinin on the inhibition of ALK positive NSCLC cells by sensitizing crizotinib
10.3760/cma.j.issn.0253-3766.2017.09.003
- VernacularTitle: 水飞蓟宾增敏克唑替尼抑制间变性淋巴瘤激酶阳性非小细胞肺癌的作用及机制
- Author:
Caiyu LIN
1
;
Conghua LU
1
;
Yonghong PAN
1
;
Lin JIAO
1
;
Hengyi CHEN
1
;
Li LI
1
;
Yong HE
1
Author Information
1. Department of Respiratory, Daping Hospital, the Third Military Medical University, Chongqing 400042, China
- Publication Type:Journal Article
- Keywords:
Crizotinib;
Silibinin;
Anaplastic lymphoma kinase;
Carcinoma, non-small-cell lung
- From:
Chinese Journal of Oncology
2017;39(9):650-656
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To explore the synergistic effect of silibinin combined with crizotinib on anaplastic lymphoma kinase positive (ALK+ ) non-small cell lung cancer (NSCLC) cells and its mechanism.
Methods:H2228 and H3122 cells were treated with silibinin, crizotinib alone or in combination. Cell proliferation was measured by 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and colony formation assay. Migration or invasion ability was tested by wound healing assay or transwell assay, respectively. Expressions of E-Cadherin and vimentin protein were examined by immunofluorescence staining. The protein expressions of ALK, p-ALK, E-Cadherin and Vimentin were detected by western blotting.The anti-cancer effect of silibinin combined with crizotinib in vivo was determined by subcutaneously injecting 2×106 H2228 cells into immunodeficient nude mice.
Results:The result of MTT assay showed that the cell viability of H2228 or H3122 treated with 100 μmol/L silibinin was (88.38±4.10)% or (72.27±3.62)%, respectively, marginally decreased compared with that of the control. The 50% inhibitory concentration (IC50) of H2228 cells treated with crizotinib alone or combined with 100 μmol/L silibinin was (917.10±7.75) nmol/L or (238.73±7.67) nmol/L, respectively. The IC50 of H3122 cells treated with crizotinib alone or combined with 100 μmol/L silibinin was (472.50±15.70) nmol/L or (206.10±12.01) nmol/L, respectively. The IC50s of H2228 and H3122 cells were significantly decreased by combined treatment of crizotinib and silibinin compared to crizotinib treatment alone (P<0.01). When compared with the control group, colony forming ratios of H2228 cells were (83.34±2.72)% in 100 μmol/L silibinin treatment group, (69.42±3.06)% in 400 nmol/L crizotinib treatment group and (27.32±1.42)% in combined treatment group. When compared with the control group, colony forming ratios of H3122 cells were (84.45±5.67)% in 100 μmol/L silibinin treatment group, (45.02±5.83)% in 400 nmol/L crizotinib treatment group and (17.43±3.83)% in combined treatment group. Silibinin combined with crizotinib treatment significantly inhibited the colony formation ability of H2228 and H3122 cells (P<0.01). Migration and invasion results showed that combined treatment of crizotinib and silibinin markedly inhibited the migration and invasion ability of H2228 cells (P<0.01). Western blot results indicated that treated with silibinin alone or in combination of crozitinib for 48 hours, the protein level of E-cadherin in H2228 cells was upregulated, while the expressions of p-ALK and vimentin were downregulated, without obvious alteration of ALK protein expression. In the xenograft model, the mean tumor weight was (9.40±2.58)g in crizotinib treatment group and (4.58±1.07)g in the combined treatment group. The inhibitory effect of tumor growth in vivo of combined treatment was significantly superior to that of crizotinib treatment alone (P<0.05).
Conclusion:Silibinin enhances the inhibitory effect of crizotinib on ALK positive NSCLC cells, which may be associated with suppression of ALK activity and mesenchymal-epithelial transition.
