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Inhibitory effects of metformin at low concentration on epithelial-mesenchymal transition of CD44(+)CD117(+) ovarian cancer stem cells.

Zhang R, Zhang P, Wang H, Hou D, Li W, Xiao G, Li C - Stem Cell Res Ther (2015)

Bottom Line: Low dose of metformin reduced survivin expression in CSCs.Metformin was also associated with a reduction of snail2, twist, and vimentin in CD44(+)CD117(+) ovarian CSCs in vivo.Our results implicate that metformin at low dose inhibits selectively CD44(+)CD117(+) ovarian CSCs through inhibition of EMT and potentiates the effect of cisplatin.

View Article: PubMed Central - PubMed

Affiliation: Department of Gynecology, Xinhua Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200092, China. rrzhangcn@126.com.

ABSTRACT

Background: Although metformin, a first-line drug for treating diabetes, may play an important role in inhibition of epithelial ovarian cancer cell growth and cancer stem cells (CSCs), metformin at low dose showed less effect on the proliferation of ovarian cancer cells. In this study, we evaluated the effect of metformin at low dose on ovarian CSCs in order to understand the molecular mechanisms underlying.

Methods: The inhibitory effects of metformin at los dose on proliferation and population of ovarian cancer cells including SKOV3 and A2780 were assessed by cell proliferation assay and flow cytometry. Quantitative real-time PCR assay on expression of Bcl-2, Survivin and Bax was performed to determine the effect of metformin at low dose on epithelial-mesenchymal transition (EMT) of cancer cells and CSCs. Tumor sphere formation assay was also performed to evaluate the effect of metformin on spheres forming ability of CSCs. The therapeutic efficacy and the anti-CSC effects of metformin at low dose were investigated by using both SKOV3 cells and primary tumor xenografts. In addition, the CSC frequency and EMT in tumor xenograft models were also assessed by flow cytometry and quantitative real-time PCR.

Results: Metformin at low dose did not affect the proliferation of ovarian cancer cells. However, it inhibited population of CD44(+)CD117(+) selectively, neither CD133(+) nor ALDH(+) cells. It suppressed expression of snail2, twist and vimentin significantly in cancer cells and CD44(+)CD117(+) CSCs in vitro. Low dose of metformin reduced survivin expression in CSCs. Low concentrations of metformin inhibited the secondary and the tertiary tumor sphere formation, decreased SKOV3 and primary ovarian tumor xenograft growth, enhanced the anticancer effect of cisplatin, and lowered the proportion of CD44(+)CD117(+) CSCs in the xenograft tissue. Metformin was also associated with a reduction of snail2, twist, and vimentin in CD44(+)CD117(+) ovarian CSCs in vivo.

Conclusions: Our results implicate that metformin at low dose inhibits selectively CD44(+)CD117(+) ovarian CSCs through inhibition of EMT and potentiates the effect of cisplatin.

No MeSH data available.


Related in: MedlinePlus

Metformin and cisplatin inhibit tumor growth in patient-derived xenograft mice. Six to 8 weeks after subcutaneous injection of ovarian patient ovarian cancer cells, the treatments were started when the tumor size reached approximately 150 mm3. Four groups of mice (n = 6) were treated with saline, metformin, cisplatin, and the combination of metformin and cisplatin for 4 weeks. Tumor size and body weight were measured weekly throughout the experiment period. Three mice from each group were sacrificed after 4 weeks of treatment for analysis and the other three mice were kept to observe for an additional 5 weeks. Summary results from three separate experiments are presented (a). Flow cytometry was performed to isolate CD44+CD117+ CSCs of each treatment group. Metformin treatment significantly reduced the CD44+CD117+ CSC population (b). Metformin significantly reduced snail2, twist, and vimentin, but not Snail1expression, and upregulated E-cadherin expression (c). * Significant difference compared with control (P < 0.05)
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Fig6: Metformin and cisplatin inhibit tumor growth in patient-derived xenograft mice. Six to 8 weeks after subcutaneous injection of ovarian patient ovarian cancer cells, the treatments were started when the tumor size reached approximately 150 mm3. Four groups of mice (n = 6) were treated with saline, metformin, cisplatin, and the combination of metformin and cisplatin for 4 weeks. Tumor size and body weight were measured weekly throughout the experiment period. Three mice from each group were sacrificed after 4 weeks of treatment for analysis and the other three mice were kept to observe for an additional 5 weeks. Summary results from three separate experiments are presented (a). Flow cytometry was performed to isolate CD44+CD117+ CSCs of each treatment group. Metformin treatment significantly reduced the CD44+CD117+ CSC population (b). Metformin significantly reduced snail2, twist, and vimentin, but not Snail1expression, and upregulated E-cadherin expression (c). * Significant difference compared with control (P < 0.05)

Mentions: Data indicated that treatment with metformin or cisplatin alone resulted in a delay of tumor growth. However, in the group treated with metformin and cisplatin, tumor growth was extremely slow, even near the cessation of treatment, showing a significant enhanced anti-tumor effect (Fig. 6a). The effect of metformin on expression of EMT marker CD44+CD117+ was analyzed by real-time RT-PCR. We obtained similar results when measuring the effect of metformin, used alone or in combination with cisplatin, on the CD44+CD117+ CSC population (Fig. 6b). Treatment with metformin resulted in a 2.8-fold decrease in the CD44+CD117+ population, and an increase of 2.2-fold for cisplatin, and combined treatment with metformin and cisplatin resulted in a 4.6-fold decrease in the CD44+CD117+ population compared with cisplatin alone, and even a 2.1-fold decrease in the CD44+CD117+ population compared with control, further demonstrating that metformin is capable of targeting the CSC population (Fig. 6b). Real-time RT-PCR experiment demonstrated that metformin had no effect on Snail 1 expression, but led to a 11.1-fold decrease in Snail 2, a 10-fold decrease in Twist, a 11.1-fold decrease in Vimentin expression, and a 2.9-fold increase in E-cadherin expression (Fig. 6c).Fig. 6


Inhibitory effects of metformin at low concentration on epithelial-mesenchymal transition of CD44(+)CD117(+) ovarian cancer stem cells.

Zhang R, Zhang P, Wang H, Hou D, Li W, Xiao G, Li C - Stem Cell Res Ther (2015)

Metformin and cisplatin inhibit tumor growth in patient-derived xenograft mice. Six to 8 weeks after subcutaneous injection of ovarian patient ovarian cancer cells, the treatments were started when the tumor size reached approximately 150 mm3. Four groups of mice (n = 6) were treated with saline, metformin, cisplatin, and the combination of metformin and cisplatin for 4 weeks. Tumor size and body weight were measured weekly throughout the experiment period. Three mice from each group were sacrificed after 4 weeks of treatment for analysis and the other three mice were kept to observe for an additional 5 weeks. Summary results from three separate experiments are presented (a). Flow cytometry was performed to isolate CD44+CD117+ CSCs of each treatment group. Metformin treatment significantly reduced the CD44+CD117+ CSC population (b). Metformin significantly reduced snail2, twist, and vimentin, but not Snail1expression, and upregulated E-cadherin expression (c). * Significant difference compared with control (P < 0.05)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4697317&req=5

Fig6: Metformin and cisplatin inhibit tumor growth in patient-derived xenograft mice. Six to 8 weeks after subcutaneous injection of ovarian patient ovarian cancer cells, the treatments were started when the tumor size reached approximately 150 mm3. Four groups of mice (n = 6) were treated with saline, metformin, cisplatin, and the combination of metformin and cisplatin for 4 weeks. Tumor size and body weight were measured weekly throughout the experiment period. Three mice from each group were sacrificed after 4 weeks of treatment for analysis and the other three mice were kept to observe for an additional 5 weeks. Summary results from three separate experiments are presented (a). Flow cytometry was performed to isolate CD44+CD117+ CSCs of each treatment group. Metformin treatment significantly reduced the CD44+CD117+ CSC population (b). Metformin significantly reduced snail2, twist, and vimentin, but not Snail1expression, and upregulated E-cadherin expression (c). * Significant difference compared with control (P < 0.05)
Mentions: Data indicated that treatment with metformin or cisplatin alone resulted in a delay of tumor growth. However, in the group treated with metformin and cisplatin, tumor growth was extremely slow, even near the cessation of treatment, showing a significant enhanced anti-tumor effect (Fig. 6a). The effect of metformin on expression of EMT marker CD44+CD117+ was analyzed by real-time RT-PCR. We obtained similar results when measuring the effect of metformin, used alone or in combination with cisplatin, on the CD44+CD117+ CSC population (Fig. 6b). Treatment with metformin resulted in a 2.8-fold decrease in the CD44+CD117+ population, and an increase of 2.2-fold for cisplatin, and combined treatment with metformin and cisplatin resulted in a 4.6-fold decrease in the CD44+CD117+ population compared with cisplatin alone, and even a 2.1-fold decrease in the CD44+CD117+ population compared with control, further demonstrating that metformin is capable of targeting the CSC population (Fig. 6b). Real-time RT-PCR experiment demonstrated that metformin had no effect on Snail 1 expression, but led to a 11.1-fold decrease in Snail 2, a 10-fold decrease in Twist, a 11.1-fold decrease in Vimentin expression, and a 2.9-fold increase in E-cadherin expression (Fig. 6c).Fig. 6

Bottom Line: Low dose of metformin reduced survivin expression in CSCs.Metformin was also associated with a reduction of snail2, twist, and vimentin in CD44(+)CD117(+) ovarian CSCs in vivo.Our results implicate that metformin at low dose inhibits selectively CD44(+)CD117(+) ovarian CSCs through inhibition of EMT and potentiates the effect of cisplatin.

View Article: PubMed Central - PubMed

Affiliation: Department of Gynecology, Xinhua Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200092, China. rrzhangcn@126.com.

ABSTRACT

Background: Although metformin, a first-line drug for treating diabetes, may play an important role in inhibition of epithelial ovarian cancer cell growth and cancer stem cells (CSCs), metformin at low dose showed less effect on the proliferation of ovarian cancer cells. In this study, we evaluated the effect of metformin at low dose on ovarian CSCs in order to understand the molecular mechanisms underlying.

Methods: The inhibitory effects of metformin at los dose on proliferation and population of ovarian cancer cells including SKOV3 and A2780 were assessed by cell proliferation assay and flow cytometry. Quantitative real-time PCR assay on expression of Bcl-2, Survivin and Bax was performed to determine the effect of metformin at low dose on epithelial-mesenchymal transition (EMT) of cancer cells and CSCs. Tumor sphere formation assay was also performed to evaluate the effect of metformin on spheres forming ability of CSCs. The therapeutic efficacy and the anti-CSC effects of metformin at low dose were investigated by using both SKOV3 cells and primary tumor xenografts. In addition, the CSC frequency and EMT in tumor xenograft models were also assessed by flow cytometry and quantitative real-time PCR.

Results: Metformin at low dose did not affect the proliferation of ovarian cancer cells. However, it inhibited population of CD44(+)CD117(+) selectively, neither CD133(+) nor ALDH(+) cells. It suppressed expression of snail2, twist and vimentin significantly in cancer cells and CD44(+)CD117(+) CSCs in vitro. Low dose of metformin reduced survivin expression in CSCs. Low concentrations of metformin inhibited the secondary and the tertiary tumor sphere formation, decreased SKOV3 and primary ovarian tumor xenograft growth, enhanced the anticancer effect of cisplatin, and lowered the proportion of CD44(+)CD117(+) CSCs in the xenograft tissue. Metformin was also associated with a reduction of snail2, twist, and vimentin in CD44(+)CD117(+) ovarian CSCs in vivo.

Conclusions: Our results implicate that metformin at low dose inhibits selectively CD44(+)CD117(+) ovarian CSCs through inhibition of EMT and potentiates the effect of cisplatin.

No MeSH data available.


Related in: MedlinePlus