Limits...
The in vitro and in vivo antitumor effects of clotrimazole on oral squamous cell carcinoma.

Wang J, Jia L, Kuang Z, Wu T, Hong Y, Chen X, Leung WK, Xia J, Cheng B - PLoS ONE (2014)

Bottom Line: Clotrimazole inhibited proliferation in all three OSCC cell lines in a dose-and time-dependent manner, and significantly reduced the colony formation of OSCC cells in vitro.In addition, clotrimazole induced apoptosis in OSCC cells, and significantly down-regulated the anti-apoptotic protein Bcl-2 and up-regulated the pro-apoptotic protein Bax.Our findings demonstrated a potent anticancer effect of clotrimazole by inducing cell cycle arrest and cellular apoptosis in OSCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Medicine, Guanghua School of Stomatology, Guangdong Provincial Key Laboratory of Stomatology, Sun Yat-sen University, Guangzhou, Guangdong, China.

ABSTRACT

Background: Clotrimazole is an antifungal imidazole derivative showing anti- neoplastic effect in some tumors, but its anticancer potential is still unclear in oral squamous cell carcinoma (OSCC). The aim of this study was to evaluate the antitumor effect of clotrimazole, and to investigate the possible mechanism of clotrimazole-mediated antitumor activity in OSCC.

Methodology: In vitro experiments, the cell viability and clonogenic ability of three human OSCC cell lines CAL27, SCC25 and UM1 were detected after clotrimazole treatment by CCK8 assay and colony formation assay. Cell cycle progression and apoptosis were assessed by flow cytometry, and the involvement of several mediators of apoptosis was examined by western blot analysis. Then, the in vivo antitumor effect of clotrimazole was investigated in CAL27 xenograft model. Immunohistochemistry and western blot analysis were performed to determine the presence of apoptotic cells and the expression of Bcl-2 and Bax in tumors from mice treated with or without clotrimazole.

Results: Clotrimazole inhibited proliferation in all three OSCC cell lines in a dose-and time-dependent manner, and significantly reduced the colony formation of OSCC cells in vitro. Clotrimazole caused cell cycle arrest at the G0/G1 phase. In addition, clotrimazole induced apoptosis in OSCC cells, and significantly down-regulated the anti-apoptotic protein Bcl-2 and up-regulated the pro-apoptotic protein Bax. Notably, clotrimazole treatment inhibited OSCC tumor growth and cell proliferation in CAL27 xenograft model. Clotrimazole also markedly reduced Bcl-2 expression and increased the protein level of Bax in tumor tissues of xenograft model.

Conclusion: Our findings demonstrated a potent anticancer effect of clotrimazole by inducing cell cycle arrest and cellular apoptosis in OSCC.

Show MeSH

Related in: MedlinePlus

Clotrimazole induces apoptosis of OSCC cells.CAL27, SCC25 and UM1 cells were incubated with various concentrations of clotrimazole (0, 30 and 40 µM) for 24 h and labeled with Annexin V and propidium iodide (PI). (A) Apoptosis of OSCC cells was analyzed by flow cytometry. The bottom right quadrant represents the percentage of early apoptotic cells (Annexin V+/PI−), whereas the top right quadrant is the percentage of late apoptotic cells (Annexin V+/PI+). (B) Percentages of cells in apoptosis at each clotrimazole concentration. The results are presented as the mean of three similar experiments. *P<0.05; **P<0.01 compared with solvent control.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4043897&req=5

pone-0098885-g004: Clotrimazole induces apoptosis of OSCC cells.CAL27, SCC25 and UM1 cells were incubated with various concentrations of clotrimazole (0, 30 and 40 µM) for 24 h and labeled with Annexin V and propidium iodide (PI). (A) Apoptosis of OSCC cells was analyzed by flow cytometry. The bottom right quadrant represents the percentage of early apoptotic cells (Annexin V+/PI−), whereas the top right quadrant is the percentage of late apoptotic cells (Annexin V+/PI+). (B) Percentages of cells in apoptosis at each clotrimazole concentration. The results are presented as the mean of three similar experiments. *P<0.05; **P<0.01 compared with solvent control.

Mentions: To further investigate whether clotrimazole initiates cellular apoptosis, OSCC cells were incubated with clotrimazole (0, 30, and 40 µM) for 24 h and analyzed by flow cytometry. The percentages of OSCC cells undergoing early apoptosis and late apoptosis were increased by clotrimazole treatment in a concentration-dependent manner (P<0.05) (Fig.4). The percentage of cells undergoing apoptosis was determined by the sum of cells in early and late apoptosis. Clotrimazole (40 µM) induced a significant increase in the proportion of apoptotic cells in CAL27, SCC25 and UM1 cells (12.3±0.9% vs. 2.0±0.1%, 12.6±1.1% vs. 1.9±0.1%, and 13.0±1.9% vs. 1.7±0.3%, respectively). The mechanisms underlying the apoptosis-inducing effect of clotrimazole in OSCC cells were assessed by western blot analysis. The levels of apoptosis-related proteins such as anti-apoptotic protein Bcl-2 and pro-apoptotic protein Bax were examined in total protein from tumor cells treated with DMSO or clotrimazole(40 µM) for 12 h, 24 h and 48 h. Our results showed that treatment with clotrimazole for 48 h induced a 3.3-fold and 1.6-fold decrease of Bcl2 protein expression in CAL27 and UM1 cells, respectively, as compared with respective control cells. Conversely, clotrimazole treatment induced a 2.3-fold and 1.6-fold increase of Bax protein expression in CAL27 and UM1 cells, respectively (Fig.5). These results indicate that an apoptotic mechanism is implicated in the clotrimazole-induced growth-inhibitory effects in OSCC cells.


The in vitro and in vivo antitumor effects of clotrimazole on oral squamous cell carcinoma.

Wang J, Jia L, Kuang Z, Wu T, Hong Y, Chen X, Leung WK, Xia J, Cheng B - PLoS ONE (2014)

Clotrimazole induces apoptosis of OSCC cells.CAL27, SCC25 and UM1 cells were incubated with various concentrations of clotrimazole (0, 30 and 40 µM) for 24 h and labeled with Annexin V and propidium iodide (PI). (A) Apoptosis of OSCC cells was analyzed by flow cytometry. The bottom right quadrant represents the percentage of early apoptotic cells (Annexin V+/PI−), whereas the top right quadrant is the percentage of late apoptotic cells (Annexin V+/PI+). (B) Percentages of cells in apoptosis at each clotrimazole concentration. The results are presented as the mean of three similar experiments. *P<0.05; **P<0.01 compared with solvent control.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4043897&req=5

pone-0098885-g004: Clotrimazole induces apoptosis of OSCC cells.CAL27, SCC25 and UM1 cells were incubated with various concentrations of clotrimazole (0, 30 and 40 µM) for 24 h and labeled with Annexin V and propidium iodide (PI). (A) Apoptosis of OSCC cells was analyzed by flow cytometry. The bottom right quadrant represents the percentage of early apoptotic cells (Annexin V+/PI−), whereas the top right quadrant is the percentage of late apoptotic cells (Annexin V+/PI+). (B) Percentages of cells in apoptosis at each clotrimazole concentration. The results are presented as the mean of three similar experiments. *P<0.05; **P<0.01 compared with solvent control.
Mentions: To further investigate whether clotrimazole initiates cellular apoptosis, OSCC cells were incubated with clotrimazole (0, 30, and 40 µM) for 24 h and analyzed by flow cytometry. The percentages of OSCC cells undergoing early apoptosis and late apoptosis were increased by clotrimazole treatment in a concentration-dependent manner (P<0.05) (Fig.4). The percentage of cells undergoing apoptosis was determined by the sum of cells in early and late apoptosis. Clotrimazole (40 µM) induced a significant increase in the proportion of apoptotic cells in CAL27, SCC25 and UM1 cells (12.3±0.9% vs. 2.0±0.1%, 12.6±1.1% vs. 1.9±0.1%, and 13.0±1.9% vs. 1.7±0.3%, respectively). The mechanisms underlying the apoptosis-inducing effect of clotrimazole in OSCC cells were assessed by western blot analysis. The levels of apoptosis-related proteins such as anti-apoptotic protein Bcl-2 and pro-apoptotic protein Bax were examined in total protein from tumor cells treated with DMSO or clotrimazole(40 µM) for 12 h, 24 h and 48 h. Our results showed that treatment with clotrimazole for 48 h induced a 3.3-fold and 1.6-fold decrease of Bcl2 protein expression in CAL27 and UM1 cells, respectively, as compared with respective control cells. Conversely, clotrimazole treatment induced a 2.3-fold and 1.6-fold increase of Bax protein expression in CAL27 and UM1 cells, respectively (Fig.5). These results indicate that an apoptotic mechanism is implicated in the clotrimazole-induced growth-inhibitory effects in OSCC cells.

Bottom Line: Clotrimazole inhibited proliferation in all three OSCC cell lines in a dose-and time-dependent manner, and significantly reduced the colony formation of OSCC cells in vitro.In addition, clotrimazole induced apoptosis in OSCC cells, and significantly down-regulated the anti-apoptotic protein Bcl-2 and up-regulated the pro-apoptotic protein Bax.Our findings demonstrated a potent anticancer effect of clotrimazole by inducing cell cycle arrest and cellular apoptosis in OSCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Medicine, Guanghua School of Stomatology, Guangdong Provincial Key Laboratory of Stomatology, Sun Yat-sen University, Guangzhou, Guangdong, China.

ABSTRACT

Background: Clotrimazole is an antifungal imidazole derivative showing anti- neoplastic effect in some tumors, but its anticancer potential is still unclear in oral squamous cell carcinoma (OSCC). The aim of this study was to evaluate the antitumor effect of clotrimazole, and to investigate the possible mechanism of clotrimazole-mediated antitumor activity in OSCC.

Methodology: In vitro experiments, the cell viability and clonogenic ability of three human OSCC cell lines CAL27, SCC25 and UM1 were detected after clotrimazole treatment by CCK8 assay and colony formation assay. Cell cycle progression and apoptosis were assessed by flow cytometry, and the involvement of several mediators of apoptosis was examined by western blot analysis. Then, the in vivo antitumor effect of clotrimazole was investigated in CAL27 xenograft model. Immunohistochemistry and western blot analysis were performed to determine the presence of apoptotic cells and the expression of Bcl-2 and Bax in tumors from mice treated with or without clotrimazole.

Results: Clotrimazole inhibited proliferation in all three OSCC cell lines in a dose-and time-dependent manner, and significantly reduced the colony formation of OSCC cells in vitro. Clotrimazole caused cell cycle arrest at the G0/G1 phase. In addition, clotrimazole induced apoptosis in OSCC cells, and significantly down-regulated the anti-apoptotic protein Bcl-2 and up-regulated the pro-apoptotic protein Bax. Notably, clotrimazole treatment inhibited OSCC tumor growth and cell proliferation in CAL27 xenograft model. Clotrimazole also markedly reduced Bcl-2 expression and increased the protein level of Bax in tumor tissues of xenograft model.

Conclusion: Our findings demonstrated a potent anticancer effect of clotrimazole by inducing cell cycle arrest and cellular apoptosis in OSCC.

Show MeSH
Related in: MedlinePlus