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The synergistic in vitro and in vivo antitumor effect of combination therapy with salinomycin and 5-fluorouracil against hepatocellular carcinoma.

Wang F, Dai W, Wang Y, Shen M, Chen K, Cheng P, Zhang Y, Wang C, Li J, Zheng Y, Lu J, Yang J, Zhu R, Zhang H, Zhou Y, Xu L, Guo C - PLoS ONE (2014)

Bottom Line: The combination of Sal and 5-FU resulted in a synergistic antitumor effect against liver tumors both in vitro and in vivo.Sal reversed the 5-FU-induced increase in CD133(+) EPCAM(+) cells, epithelial-mesenchymal transition and activation of the Wnt/β-catenin signaling pathway.The combination of Sal and 5-FU may provide us with a new approach to reverse drug resistant for the treatment of patients with HCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, Shanghai Tenth People's Hospital, Tongji University of Medicine, Shanghai, PR China.

ABSTRACT
Hepatocellular carcinoma (HCC) is one of the few cancers in which a continuous increase in incidence has been observed over several years. Drug resistance is a major problem in the treatment of HCC. In the present study, we used salinomycin (Sal) and 5-fluorouracil (5-FU) combination therapy on HCC cell lines Huh7, LM3 and SMMC-7721 and nude mice subcutaneously tumor model to study whether Sal could increase the sensitivity of hepatoma cells to the traditional chemotherapeutic agent such as 5-FU. The combination of Sal and 5-FU resulted in a synergistic antitumor effect against liver tumors both in vitro and in vivo. Sal reversed the 5-FU-induced increase in CD133(+) EPCAM(+) cells, epithelial-mesenchymal transition and activation of the Wnt/β-catenin signaling pathway. The combination of Sal and 5-FU may provide us with a new approach to reverse drug resistant for the treatment of patients with HCC.

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Effects of 5-FU, Sal and their combination on cancer stem cell properties in HCC cells.(A) Flow cytometry assays showed that treatment with 5-FU increased the proportion of the CD133+ EPCAM+ Huh7 cell subpopulation compared with the control group. In contrast, treatment with Sal reduced this proportion compared with the control group. 5-FU combined with Sal reduced this proportion compared with the 5-FU group (*p<0.05). (B) Colony-forming assays were performed to measure the proliferative ability of single cancer cells. The number of colonies increased in the 5-FU treatment group compared with the control group, decreased in the Sal treatment group compared with the control group. The number of colonies was significantly lower in the Sal plus 5-FU combination group compared with the 5-FU treatment group (*p<0.05). (C) Immunohistochemistry indicates CD133+ and EPCAM+ expression in the tumors of mouse xenograft models. (Magnification is 200×).
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pone-0097414-g003: Effects of 5-FU, Sal and their combination on cancer stem cell properties in HCC cells.(A) Flow cytometry assays showed that treatment with 5-FU increased the proportion of the CD133+ EPCAM+ Huh7 cell subpopulation compared with the control group. In contrast, treatment with Sal reduced this proportion compared with the control group. 5-FU combined with Sal reduced this proportion compared with the 5-FU group (*p<0.05). (B) Colony-forming assays were performed to measure the proliferative ability of single cancer cells. The number of colonies increased in the 5-FU treatment group compared with the control group, decreased in the Sal treatment group compared with the control group. The number of colonies was significantly lower in the Sal plus 5-FU combination group compared with the 5-FU treatment group (*p<0.05). (C) Immunohistochemistry indicates CD133+ and EPCAM+ expression in the tumors of mouse xenograft models. (Magnification is 200×).

Mentions: EpCAM and CD133 have been used as cancer stem cells (CSCs) markers in HCC. Research has shown that both EpCAM and CD133 surface markers were more representative for CSC s in HCC Huh7 cells [24]. We performed flow cytometry to determine the effects of 5-FU and Sal on the proportion of HCC cells with the CD133(+) EPCAM(+) antigenic phenotype (Fig. 3A). Treatment with 5-FU increased the proportion of the CD133(+) EPCAM(+) cell subpopulation from 27.77±4.72% (vehicle-treated controls) to 53.5±3.17% (*p<0.05). In contrast, treatment with Sal reduced this proportion from 27.77±4.72% (vehicle-treated controls) to 6±1.70% (*p<0.05). There was a significant decrease in the CD133(+) EPCAM(+) cell subpopulation in the 5-FU plus Sal combination therapy group compared with 5-FU monotherapy (26.73±8.27% vs 53.57±3.17, *p<0.05). We know that cancer stem cells have a strong proliferative ability, thus, colony-forming assays (Fig. 3B) were performed to measure the proliferative ability of single cancer cells. Huh7 cells were treated with DMSO vehicle, 5-FU (44 ug/ml), Sal (2 µM) and Sal plus 5-FU for 96 h. In all cases, colonies were visible after 10 days. The number of colonies increased in the 5-FU treatment group (8.25±0.25 colonies/high power field (HPF)), and decreased in the Sal treatment group (1.83±0.29 colonies/HPF), relative to vehicle-treated controls (4.75±0.05 colonies/HPF) (*p<0.05). The number of colonies was significantly lower in the Sal plus 5-FU combination group (4.42±0.29 colonies/HPF) compared with the 5-FU treatment group (8.25±0.25 colonies/HPF) (*p<0.05).


The synergistic in vitro and in vivo antitumor effect of combination therapy with salinomycin and 5-fluorouracil against hepatocellular carcinoma.

Wang F, Dai W, Wang Y, Shen M, Chen K, Cheng P, Zhang Y, Wang C, Li J, Zheng Y, Lu J, Yang J, Zhu R, Zhang H, Zhou Y, Xu L, Guo C - PLoS ONE (2014)

Effects of 5-FU, Sal and their combination on cancer stem cell properties in HCC cells.(A) Flow cytometry assays showed that treatment with 5-FU increased the proportion of the CD133+ EPCAM+ Huh7 cell subpopulation compared with the control group. In contrast, treatment with Sal reduced this proportion compared with the control group. 5-FU combined with Sal reduced this proportion compared with the 5-FU group (*p<0.05). (B) Colony-forming assays were performed to measure the proliferative ability of single cancer cells. The number of colonies increased in the 5-FU treatment group compared with the control group, decreased in the Sal treatment group compared with the control group. The number of colonies was significantly lower in the Sal plus 5-FU combination group compared with the 5-FU treatment group (*p<0.05). (C) Immunohistochemistry indicates CD133+ and EPCAM+ expression in the tumors of mouse xenograft models. (Magnification is 200×).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4016361&req=5

pone-0097414-g003: Effects of 5-FU, Sal and their combination on cancer stem cell properties in HCC cells.(A) Flow cytometry assays showed that treatment with 5-FU increased the proportion of the CD133+ EPCAM+ Huh7 cell subpopulation compared with the control group. In contrast, treatment with Sal reduced this proportion compared with the control group. 5-FU combined with Sal reduced this proportion compared with the 5-FU group (*p<0.05). (B) Colony-forming assays were performed to measure the proliferative ability of single cancer cells. The number of colonies increased in the 5-FU treatment group compared with the control group, decreased in the Sal treatment group compared with the control group. The number of colonies was significantly lower in the Sal plus 5-FU combination group compared with the 5-FU treatment group (*p<0.05). (C) Immunohistochemistry indicates CD133+ and EPCAM+ expression in the tumors of mouse xenograft models. (Magnification is 200×).
Mentions: EpCAM and CD133 have been used as cancer stem cells (CSCs) markers in HCC. Research has shown that both EpCAM and CD133 surface markers were more representative for CSC s in HCC Huh7 cells [24]. We performed flow cytometry to determine the effects of 5-FU and Sal on the proportion of HCC cells with the CD133(+) EPCAM(+) antigenic phenotype (Fig. 3A). Treatment with 5-FU increased the proportion of the CD133(+) EPCAM(+) cell subpopulation from 27.77±4.72% (vehicle-treated controls) to 53.5±3.17% (*p<0.05). In contrast, treatment with Sal reduced this proportion from 27.77±4.72% (vehicle-treated controls) to 6±1.70% (*p<0.05). There was a significant decrease in the CD133(+) EPCAM(+) cell subpopulation in the 5-FU plus Sal combination therapy group compared with 5-FU monotherapy (26.73±8.27% vs 53.57±3.17, *p<0.05). We know that cancer stem cells have a strong proliferative ability, thus, colony-forming assays (Fig. 3B) were performed to measure the proliferative ability of single cancer cells. Huh7 cells were treated with DMSO vehicle, 5-FU (44 ug/ml), Sal (2 µM) and Sal plus 5-FU for 96 h. In all cases, colonies were visible after 10 days. The number of colonies increased in the 5-FU treatment group (8.25±0.25 colonies/high power field (HPF)), and decreased in the Sal treatment group (1.83±0.29 colonies/HPF), relative to vehicle-treated controls (4.75±0.05 colonies/HPF) (*p<0.05). The number of colonies was significantly lower in the Sal plus 5-FU combination group (4.42±0.29 colonies/HPF) compared with the 5-FU treatment group (8.25±0.25 colonies/HPF) (*p<0.05).

Bottom Line: The combination of Sal and 5-FU resulted in a synergistic antitumor effect against liver tumors both in vitro and in vivo.Sal reversed the 5-FU-induced increase in CD133(+) EPCAM(+) cells, epithelial-mesenchymal transition and activation of the Wnt/β-catenin signaling pathway.The combination of Sal and 5-FU may provide us with a new approach to reverse drug resistant for the treatment of patients with HCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, Shanghai Tenth People's Hospital, Tongji University of Medicine, Shanghai, PR China.

ABSTRACT
Hepatocellular carcinoma (HCC) is one of the few cancers in which a continuous increase in incidence has been observed over several years. Drug resistance is a major problem in the treatment of HCC. In the present study, we used salinomycin (Sal) and 5-fluorouracil (5-FU) combination therapy on HCC cell lines Huh7, LM3 and SMMC-7721 and nude mice subcutaneously tumor model to study whether Sal could increase the sensitivity of hepatoma cells to the traditional chemotherapeutic agent such as 5-FU. The combination of Sal and 5-FU resulted in a synergistic antitumor effect against liver tumors both in vitro and in vivo. Sal reversed the 5-FU-induced increase in CD133(+) EPCAM(+) cells, epithelial-mesenchymal transition and activation of the Wnt/β-catenin signaling pathway. The combination of Sal and 5-FU may provide us with a new approach to reverse drug resistant for the treatment of patients with HCC.

Show MeSH
Related in: MedlinePlus