An impaired apoptotic signal pathway plays an important role in human cancer development, and tumor-cell apoptosis is the key to many anticancer drug treatments.5
The two main signal-transduction pathways of apoptosis are endogenous and exogenous pathways, ie, mitochondrial pathways and death-receptor pathways. Cell-death signals can activate Bax and promote Bax to translocate from cytoplasm to mitochondrial membrane. Bax soon forms homodimers after translocation to the mitochondrial membrane with the change in permeability of the mitochondrial membrane, the interaction with the spilled cytochrome C and apoptosis-related molecules of Apaf-1, followed by the activation of caspase signal-transduction pathway from the initial activation of caspase-2, caspase-8, caspase-9, and caspase-10 downstream of caspase-3, caspase-6, and caspase-7, ultimately leading to cell apoptosis. Bcl-2 was mainly localized in the mitochondrial membrane and formed a heterodimer with Bax, restricting Bax apoptosis. The ratio of Bcl-2 and Bax determines whether the cells will be apoptotic.5
Many chemotherapeutic drugs or sex hormone-receptor blockers may affect endogenous signaling and promote apoptosis of tumor cells.2
Mifepristone is a PR antagonist through its competitive binding PR playing the role of antiprogesterone, and has been widely used for the treatment of gynecological sex hormone-dependent tumors.7
By MTT assay, this study found that mifepristone significantly inhibited the proliferation of cholangiocarcinoma cells’ in vitro activity in a dose-/time-dependent manner. Li et al’s findings indicated that mifepristone had an inhibitory effect on ovarian cancer cell lines’ in vitro proliferation, which increased apoptosis rate in a certain concentration range in a dose-/time-dependent manner.19
Li’s experimental results showed that mifepristone allowed PR-positive gastric cancer cell lines to reduce in the S/G2
/M phase and increase in the G0
phase, and the proliferation index was significantly reduced, helping to induce gastric cancer cells’ apoptosis.11
The literature has reported that the Bcl-2 protein-positive expression rate of cholangiocarcinoma cells was up to 31.7%–72.5%, and regarded Bcl-2 protein expression as characteristic signs of cholangiocarcinoma cells.20
Bcl-2 positive expression and PR-positive expression were significantly correlated. This suggested that Bcl-2–positive tumors were progesterone-dependent.12
The results of this study showed that mifepristone could promote apoptosis of cholangiocarcinoma cells through regulation of Bax gene expression and lower expression of the Bcl-2 gene. Its mechanism might be related with the expression of Bcl-2 inhibited by a PR-blocking effect and/or the non-PR regulation enhanced Bax gene expression. Yin et al reported that under progesterone, uterine leiomyoma cells had pregnant hormone response element–like sequence in the Bcl-2 promoter region and combined with PR to adjust to a high expression of Bcl-2, thus inhibiting the Bcl-2/Bax pathway in inducing apoptosis.22
Kandouz et al’s experiments confirmed that progesterone might increase breast cancer cells’ Bcl-2 mRNA and protein expression.23
Liu et al reported that progesterone could nourish the mother-cell caspase-3 expression reduction, and Bcl-2 expression was significantly increased.24
The results of this study showed that mifepristone allowed Bcl-2 gene-expression decrease and at the same time allowed the increase of Bax gene expression, which increased cholangiocarcinoma cell apoptotic activity significantly. It was not difficult to infer that mifepristone inhibits Bcl-2 expression and promotes apoptosis of tumor cells by blocking PR-antagonist PR agonists. The exact mechanism of mifepristone’s induction on Bax gene expression needs further study.
Fas belongs to the TNF receptor family, which is an important cell membrane-death receptor. The natural ligand of Fas is Fasl. The Fas/Fasl apoptotic pathway is involved in host defense, immune surveillance, and the aging process. Abnormalities in the Fas/Fasl system are associated with tumor occurrence and the development of cancer.25
Fas expression of cholangiocarcinoma cell has heterogeneity, which can be divided into Fas-positive expression and Fas-negative expression.2
Cancer cells with an expression of Fas-negative were inoculated in nude mice, and tumor cells had fast growth and large volume; cancer cells with Fas-positive expression were inoculated in nude mice, and tumor cells had slow growth and small volume. These results were related to the fact that apoptosis was easily induced by Fas-positive expression.2
Liu reported that progesterone in PR could inhibit the expression of Fas.24
Therefore, mifepristone could upregulate Fas expression by blocking PR.
IFN-γ could induce the expression of Fas, Bax, and caspase-3, making human cholangiocarcinoma cells and rabbit placental cells significantly increase Fas expression.2
The apoptosis signal transduction mediated by Fas/Fasl requires costimulatory signals of T-cell receptors and major histocompatibility complex (MHC) class I antigens. IFN is an important biological factor regulating high expression of MHC class I antigens, in which IFN-γ has stronger expression to promote MHC I than IFN-α or IFN-β. Tumor cells may be protected without the risk of Fas-mediated apoptosis in the loss of MHC class I antigens’ costimulatory signal system.14
The results of this study showed that the expression of Fas significantly increased when it was cocultured with IFN-γ and cholangiocarcinoma cells, making the effect of mifepristone induce a significant increase of apoptosis in cholangiocarcinoma cells, and apoptosis rate was significantly higher than the simple application of the mifepristone group (P
< 0.01). Therefore, IFN-γ and mifepristone coinduced was expected to be a new method of treatment for cholangiocarcinoma.
Overall, mifepristone in vitro could inhibit the proliferation activity of cholangiocarcinoma cell to promote the occurrence of apoptosis. Its mechanism was related to mitochondrial apoptotic and death-receptor apoptotic pathways to be actively upregulated. IFN-γ had the role of inducing Fas gene expression and thus increased mifepristone-induced apoptosis of cholangiocarcinoma cells.