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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.

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Clotrimazole inhibits colony formation of OSCC cells.OSCC cells (CAL27, SCC25, and UM1) grown in 6-well plates (1000 cells/well) were incubated with various concentrations of clotrimazole (0, 10, 20 and 30 µM) for two weeks. Cell colonies were stained and counted as described in the Methods section. The results presented as mean ± standard deviation values for three independent experiments. *P<0.05; **P<0.01 compared with solvent control.
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pone-0098885-g002: Clotrimazole inhibits colony formation of OSCC cells.OSCC cells (CAL27, SCC25, and UM1) grown in 6-well plates (1000 cells/well) were incubated with various concentrations of clotrimazole (0, 10, 20 and 30 µM) for two weeks. Cell colonies were stained and counted as described in the Methods section. The results presented as mean ± standard deviation values for three independent experiments. *P<0.05; **P<0.01 compared with solvent control.

Mentions: To investigate the effects of clotrimazole on human OSCC cells growth in vitro, three OSCC cell lines CAL27, SCC25 and UM1 were used in our study. Treatment with clotrimazole led to significant inhibition of the cell viability in a dose-and time-dependent manner at each concentration (20, 30, 40 and 50 µM) compared with control cells (P<0.05, respectively) in these OSCC cell lines (Fig.1). After 40 µM clotrimazole treatment for 48 h, the inhibition rate reached 53.1% in CAL27 cells, 43.8% in SCC25 cells, and 61.5% in UM1 cells. The IC50 values for 48 h of clotrimazole treatment were 35.9 µM, 35.6 µM, and 31.4 µM in CAL27, SCC25, and UM1 cells, respectively. Therefore, we used 30 µM and 40 µM clotrimazole that encompassed concentrations above and below IC50 values for further experiments. Next, the ability of these three cell lines to form colonies with or without clotrimazole treatment was examined for a period of two weeks. Clotrimazole markedly decreased cell colony formation in a dose-dependent manner (Fig. 2A). At the low concentration of 10 µM, clotrimazole inhibited cell colony formation of CAL27, SCC25 and UM1 cells by 54.2%, 54.1% and 54.4%, respectively (P<0.05). Additionally, at the concentration of 30 µM clotrimazole, cell colony formation was strongly reduced by 95.0%, 95.5% and 93.0%, respectively (P<0.001), as compared with the respective control groups (Fig.2B). Collectively, our results indicated that clotrimazole inhibited the growth of OSCC cells in vitro.


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 inhibits colony formation of OSCC cells.OSCC cells (CAL27, SCC25, and UM1) grown in 6-well plates (1000 cells/well) were incubated with various concentrations of clotrimazole (0, 10, 20 and 30 µM) for two weeks. Cell colonies were stained and counted as described in the Methods section. The results presented as mean ± standard deviation values for three independent 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-g002: Clotrimazole inhibits colony formation of OSCC cells.OSCC cells (CAL27, SCC25, and UM1) grown in 6-well plates (1000 cells/well) were incubated with various concentrations of clotrimazole (0, 10, 20 and 30 µM) for two weeks. Cell colonies were stained and counted as described in the Methods section. The results presented as mean ± standard deviation values for three independent experiments. *P<0.05; **P<0.01 compared with solvent control.
Mentions: To investigate the effects of clotrimazole on human OSCC cells growth in vitro, three OSCC cell lines CAL27, SCC25 and UM1 were used in our study. Treatment with clotrimazole led to significant inhibition of the cell viability in a dose-and time-dependent manner at each concentration (20, 30, 40 and 50 µM) compared with control cells (P<0.05, respectively) in these OSCC cell lines (Fig.1). After 40 µM clotrimazole treatment for 48 h, the inhibition rate reached 53.1% in CAL27 cells, 43.8% in SCC25 cells, and 61.5% in UM1 cells. The IC50 values for 48 h of clotrimazole treatment were 35.9 µM, 35.6 µM, and 31.4 µM in CAL27, SCC25, and UM1 cells, respectively. Therefore, we used 30 µM and 40 µM clotrimazole that encompassed concentrations above and below IC50 values for further experiments. Next, the ability of these three cell lines to form colonies with or without clotrimazole treatment was examined for a period of two weeks. Clotrimazole markedly decreased cell colony formation in a dose-dependent manner (Fig. 2A). At the low concentration of 10 µM, clotrimazole inhibited cell colony formation of CAL27, SCC25 and UM1 cells by 54.2%, 54.1% and 54.4%, respectively (P<0.05). Additionally, at the concentration of 30 µM clotrimazole, cell colony formation was strongly reduced by 95.0%, 95.5% and 93.0%, respectively (P<0.001), as compared with the respective control groups (Fig.2B). Collectively, our results indicated that clotrimazole inhibited the growth of OSCC cells in vitro.

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