Study on Inhibitory Effects of Silibinin Combined with Afatinib on Proliferation and Invasion of Ovarian Cancer Cells and Its Mechanism

OBJECTIVE To explore the inhibitory effect and possible mechanism of silibinin combined with afatinib against ovarian cancer. METHODS Different concentrations of afatinib and silibinin were treated on ovarian cancer cell lines, SKOV3 or CP70 cells, alone or in combination for 48 h. The inhibitory effects of silibinin and afatinib alone or in combination on cell proliferation were detected by CCK-8 assay. The half inhibitory concentration(IC₅₀) was calculated. Transwell test was applied to detect the effect of drugs on cell proliferation. Transwell test was used to detect the effect of drugs on cell invasion. Immunofluorescence and phalloidin staining were applied to detect focal adhesion kinase(FAK) and cytoskeleton. Western blotting was used to evaluate the protein levels and phosphorylation of FAK. Finally, the expression of FAK in ovarian cancer and the correlation between expression and prognosis were analyzed using database. RESULTS Both silibinin and afatinib could inhibit the growth of SKOV3 and CP70 cells via a dose-dependent manner. The IC₅₀ values of silibinin and afatinib against SKOV3 were 81.2, 1.9 μmol·L^-1, respectively; the IC₅₀ values of silibinin and afatinib against CP70 was 154.1, 4.1 μmol·L^-1, respectively. The combination of the two drugs could significantly increase the survival inhibitory rate. In addition, both drugs inhibited the invasion of SKOV3 cells, and the combination of the two drugs significantly increased the inhibitory rate of invasion in SKOV3 cells. The combination of the two drugs inhibited the expression and phosphorylation of FAK. Furthermore, silibinin and afatinib significantly reduced the integrity of cytoskeleton, and the destruction of cytoskeleton was stronger when silibinin and afatinib were combined. Finally, FAK was highly expressed in ovarian cancer through the database, which was negatively correlated with the ovarian cancer prognosis. CONCLUSION It is preliminarily proved that the combination of silibinin and afatinib can significantly inhibit the proliferation of SKOV3 and CP70 cells, and may inhibit the invasion of SKOV3 cells by reducing the formation of cytoskeleton and suppressing the FAK signaling pathway.