Histone deacetylase inhibitor MS-275 alone or combined with bortezomib or sorafenib exhibits strong antiproliferative action in human cholangiocarcinoma cells

Abstract

Aim: To investigate the antiproliferative effect of the histone deacetylase (HDAC) inhibitor MS-275 on cholangiocarcinoma cells alone and in combination with conventional cytostatic drugs (gemcitabine or doxorubicin) or the novel anticancer agents sorafenib or bortezomib.

Methods: Two human bile duct adenocarcinoma cell lines (EGI-1 and TFK-1) were studied. Crystal violet staining was used for detection of cell number changes. Cytotoxicity was determined by measuring the release of the cytoplasmic enzyme lactate dehydrogenase (LDH). Apoptosis was determined by measuring the enzyme activity of caspase-3. Cell cycle status reflected by the DNA content was detected by flow cytometry.

Results: MS-275 treatment potently inhibited the proliferation of EGI-1 and TFK-1 cholangiocarcinoma cells by inducing apoptosis and cell cycle arrest. MS-275-induced apoptosis was characterized by activation of caspase-3, up-regulation of Bax and down-regulation of Bcl-2. Cell cycle was predominantly arrested at the G(1)/S checkpoint, which was associated with induction of the cyclin-dependent kinase inhibitor p21(Waf/CIP1). Furthermore, additive anti-neoplastic effects were observed when MS-275 treatment was combined with gemcitabine or doxorubicin, while combination with the multi-kinase inhibitor sorafenib or the proteasome inhibitor bortezomib resulted in overadditive anti-neoplastic effects.

Conclusion: The growth of human cholangiocarcinoma cells can be potently inhibited by MS-275 alone or in combination with conventional cytostatic drugs or new, targeted anticancer agents.

Figure 1

Anti-proliferative effects of MS-275 on human EGI-1 and TFK-1 cholangiocarcinoma cells. mean ± SEM, n > 5, ^aP < 0.05 vs controls.

Figure 2

MS-275-induced LDH release into the supernatant of EGI-1 and TFK-1 cells. mean ± SEM, n > 3 independent experiments; No significant increase in LDH release was observed.

Figure 3

MS-275-induced apoptosis-specific caspase-3 activity increase in EGI-1 and TFK-1 cells. mean ± SEM, n > 3 independent experiments, ^aP < 0.05 vs controls.

Figure 4

Induction of cell cycle arrest by MS-275 on EGI-1 and TFK-1 cells. means of > 3 independent experiments, ^aP < 0.05 vs controls.

Figure 5

MS-275-induced modulation of apoptosis- and cell cycle-relevant proteins (n = 3 independent experiments).

Figure 6

Anti-proliferative effects of MS-275 plus cytostatics on EGI-1 and TFK-1 cells. mean ± SEM, n = 3-4 independent experiments.

Figure 7

Anti-proliferative effects of MS-275 plus sorafenib or bortezomib on EGI-1 and TFK-1. mean ± SEM, n = 3-4 independent experiments.