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Pro-apoptotic and pro-autophagic effects of the Aurora kinase A inhibitor alisertib (MLN8237) on human osteosarcoma U-2 OS and MG-63 cells through the activation of mitochondria-mediated pathway and inhibition of p38 MAPK/PI3K/Akt/mTOR signaling pathway.

Niu NK, Wang ZL, Pan ST, Ding HQ, Au GH, He ZX, Zhou ZW, Xiao G, Yang YX, Zhang X, Yang T, Chen XW, Qiu JX, Zhou SF - Drug Des Devel Ther (2015)

Bottom Line: The results showed that ALS had potent growth inhibitory, pro-apoptotic, pro-autophagic, and EMT inhibitory effects on U-2 OS and MG-63 cells.ALS markedly induced mitochondria-mediated apoptosis with a significant increase in the expression of key pro-apoptotic proteins and a decrease in main anti-apoptotic proteins.ALS is a promising anticancer agent in OS treatment and further studies are needed to confirm its efficacy and safety in OS chemotherapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Spinal Surgery, General Hospital of Ningxia Medical University, Yinchuan, People's Republic of China ; Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA ; Department of Orthopedics, General Hospital of Tianjin Medical University, Tianjin, People's Republic of China.

ABSTRACT
Osteosarcoma (OS) is the most common malignant bone tumor occurring mostly in children and adolescents between 10 and 20 years of age with poor response to current therapeutics. Alisertib (ALS, MLN8237) is a selective Aurora kinase A inhibitor that displays anticancer effects on several types of cancer. However, the role of ALS in the treatment of OS remains unknown. This study aimed to investigate the effects of ALS on the cell growth, apoptosis, autophagy, and epithelial to mesenchymal transition (EMT) and the underlying mechanisms in two human OS cell lines U-2 OS and MG-63. The results showed that ALS had potent growth inhibitory, pro-apoptotic, pro-autophagic, and EMT inhibitory effects on U-2 OS and MG-63 cells. ALS remarkably induced G2/M arrest and down-regulated the expression levels of cyclin-dependent kinases 1 and 2 and cyclin B1 in both U-2 OS and MG-63 cells. ALS markedly induced mitochondria-mediated apoptosis with a significant increase in the expression of key pro-apoptotic proteins and a decrease in main anti-apoptotic proteins. Furthermore, ALS promoted autophagic cell death via the inhibition of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) and p38 mitogen-activated protein kinase (p38 MAPK) signaling pathways, and activation of 5'-AMP-dependent kinase (AMPK) signaling pathway. Inducers or inhibitors of apoptosis or autophagy simultaneously altered ALS-induced apoptotic and autophagic death in both U-2 OS and MG-63 cells, suggesting a crosstalk between these two primary modes of programmed cell death. Moreover, ALS suppressed EMT-like phenotypes with a marked increase in the expression of E-cadherin but a decrease in N-cadherin in U-2 OS and MG-63 cells. ALS treatment also induced reactive oxygen species (ROS) generation but inhibited the expression levels of sirtuin 1 and nuclear factor-erythroid-2-related factor 2 (Nrf2) in both cell lines. Taken together, these findings show that ALS promotes apoptosis and autophagy but inhibits EMT via PI3K/Akt/mTOR, p38 MAPK, and AMPK signaling pathways with involvement of ROS- and sirtuin 1-associated pathways in U-2 OS and MG-63 cells. ALS is a promising anticancer agent in OS treatment and further studies are needed to confirm its efficacy and safety in OS chemotherapy.

No MeSH data available.


Related in: MedlinePlus

ALS modulates the expression levels of the key EMT markers and inhibits Sirt1 expression in U-2 OS and MG-63 cells.Notes: U-2 OS and MG-63 cells were treated with ALS at 0.1, 1, and 5 μM for 24 hours and then subjected to Western blotting assay. (A) Representative blots of E-cadherin, N-cadherin, snail, slug, TCF-8/ZEB1, vimentin, β-catenin, and ZO-1 in U-2 OS and MG-63 cells, (B) quantitative expression levels of E-cadherin, N-cadherin, snail, slug, TCF-8/ZEB1, vimentin, β-catenin, and ZO-1 in U-2 OS and MG-63 cells, (C) representative blots of Sirt1 in U-2 OS and MG-63 cells and (D) quantitative expression levels of Sirt1 in U-2 OS and MG-63 cells. β-actin was used as the internal control. Data represent the mean ± SD of three independent experiments. *P<0.05; **P<0.01; and ***P<0.001 by one-way ANOVA.Abbreviations: ALS, alisertib; SD, standard deviation; ANOVA, analysis of variance; EMT, epithelial to mesenchymal transition; Sirt1, sirtuin 1; ZO-1, zona occludens protein 1; TCF-8/ZEB1, zinc finger E-box-binding homeobox 1; OS, osteosarcoma.
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f12-dddt-9-1555: ALS modulates the expression levels of the key EMT markers and inhibits Sirt1 expression in U-2 OS and MG-63 cells.Notes: U-2 OS and MG-63 cells were treated with ALS at 0.1, 1, and 5 μM for 24 hours and then subjected to Western blotting assay. (A) Representative blots of E-cadherin, N-cadherin, snail, slug, TCF-8/ZEB1, vimentin, β-catenin, and ZO-1 in U-2 OS and MG-63 cells, (B) quantitative expression levels of E-cadherin, N-cadherin, snail, slug, TCF-8/ZEB1, vimentin, β-catenin, and ZO-1 in U-2 OS and MG-63 cells, (C) representative blots of Sirt1 in U-2 OS and MG-63 cells and (D) quantitative expression levels of Sirt1 in U-2 OS and MG-63 cells. β-actin was used as the internal control. Data represent the mean ± SD of three independent experiments. *P<0.05; **P<0.01; and ***P<0.001 by one-way ANOVA.Abbreviations: ALS, alisertib; SD, standard deviation; ANOVA, analysis of variance; EMT, epithelial to mesenchymal transition; Sirt1, sirtuin 1; ZO-1, zona occludens protein 1; TCF-8/ZEB1, zinc finger E-box-binding homeobox 1; OS, osteosarcoma.

Mentions: EMT is a critical process involving the initiation, growth, invasion, and metastasis of cancer.40 EMT depends on a reduction in expression of cell adhesion molecules. Herein, we examined the effect of ALS treatment on the expression of key EMT-associated markers in U-2 OS and MG-63 cells using Western blotting assay. As shown in Figure 12, the E-cadherin expression level was increased and N-cadherin expression level was decreased after ALS treatment in both cell lines. In U-2 OS cells, there was a 1.2- and 1.3-fold increase in the expression of E-cadherin, whereas N-cadherin expression level was suppressed by 19.9% and 20.6%, when treated with 1 and 5 μM ALS for 24 hours, respectively; (P<0.05; Figure 12A and B). In MG-63 cells, there was a 1.2-fold increase in the expression of E-cadherin when cells were treated with 5 μM ALS (P<0.05; Figure 12A and B). Meanwhile, treatment of MG-63 cells with ALS at 0.1, 1, and 5 μM for 24 hours decreased the expression of N-cadherin by 39.4%, 61.7%, and 58.6%, respectively (P<0.01 or 0.001; Figure 12A and B).


Pro-apoptotic and pro-autophagic effects of the Aurora kinase A inhibitor alisertib (MLN8237) on human osteosarcoma U-2 OS and MG-63 cells through the activation of mitochondria-mediated pathway and inhibition of p38 MAPK/PI3K/Akt/mTOR signaling pathway.

Niu NK, Wang ZL, Pan ST, Ding HQ, Au GH, He ZX, Zhou ZW, Xiao G, Yang YX, Zhang X, Yang T, Chen XW, Qiu JX, Zhou SF - Drug Des Devel Ther (2015)

ALS modulates the expression levels of the key EMT markers and inhibits Sirt1 expression in U-2 OS and MG-63 cells.Notes: U-2 OS and MG-63 cells were treated with ALS at 0.1, 1, and 5 μM for 24 hours and then subjected to Western blotting assay. (A) Representative blots of E-cadherin, N-cadherin, snail, slug, TCF-8/ZEB1, vimentin, β-catenin, and ZO-1 in U-2 OS and MG-63 cells, (B) quantitative expression levels of E-cadherin, N-cadherin, snail, slug, TCF-8/ZEB1, vimentin, β-catenin, and ZO-1 in U-2 OS and MG-63 cells, (C) representative blots of Sirt1 in U-2 OS and MG-63 cells and (D) quantitative expression levels of Sirt1 in U-2 OS and MG-63 cells. β-actin was used as the internal control. Data represent the mean ± SD of three independent experiments. *P<0.05; **P<0.01; and ***P<0.001 by one-way ANOVA.Abbreviations: ALS, alisertib; SD, standard deviation; ANOVA, analysis of variance; EMT, epithelial to mesenchymal transition; Sirt1, sirtuin 1; ZO-1, zona occludens protein 1; TCF-8/ZEB1, zinc finger E-box-binding homeobox 1; OS, osteosarcoma.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4362906&req=5

f12-dddt-9-1555: ALS modulates the expression levels of the key EMT markers and inhibits Sirt1 expression in U-2 OS and MG-63 cells.Notes: U-2 OS and MG-63 cells were treated with ALS at 0.1, 1, and 5 μM for 24 hours and then subjected to Western blotting assay. (A) Representative blots of E-cadherin, N-cadherin, snail, slug, TCF-8/ZEB1, vimentin, β-catenin, and ZO-1 in U-2 OS and MG-63 cells, (B) quantitative expression levels of E-cadherin, N-cadherin, snail, slug, TCF-8/ZEB1, vimentin, β-catenin, and ZO-1 in U-2 OS and MG-63 cells, (C) representative blots of Sirt1 in U-2 OS and MG-63 cells and (D) quantitative expression levels of Sirt1 in U-2 OS and MG-63 cells. β-actin was used as the internal control. Data represent the mean ± SD of three independent experiments. *P<0.05; **P<0.01; and ***P<0.001 by one-way ANOVA.Abbreviations: ALS, alisertib; SD, standard deviation; ANOVA, analysis of variance; EMT, epithelial to mesenchymal transition; Sirt1, sirtuin 1; ZO-1, zona occludens protein 1; TCF-8/ZEB1, zinc finger E-box-binding homeobox 1; OS, osteosarcoma.
Mentions: EMT is a critical process involving the initiation, growth, invasion, and metastasis of cancer.40 EMT depends on a reduction in expression of cell adhesion molecules. Herein, we examined the effect of ALS treatment on the expression of key EMT-associated markers in U-2 OS and MG-63 cells using Western blotting assay. As shown in Figure 12, the E-cadherin expression level was increased and N-cadherin expression level was decreased after ALS treatment in both cell lines. In U-2 OS cells, there was a 1.2- and 1.3-fold increase in the expression of E-cadherin, whereas N-cadherin expression level was suppressed by 19.9% and 20.6%, when treated with 1 and 5 μM ALS for 24 hours, respectively; (P<0.05; Figure 12A and B). In MG-63 cells, there was a 1.2-fold increase in the expression of E-cadherin when cells were treated with 5 μM ALS (P<0.05; Figure 12A and B). Meanwhile, treatment of MG-63 cells with ALS at 0.1, 1, and 5 μM for 24 hours decreased the expression of N-cadherin by 39.4%, 61.7%, and 58.6%, respectively (P<0.01 or 0.001; Figure 12A and B).

Bottom Line: The results showed that ALS had potent growth inhibitory, pro-apoptotic, pro-autophagic, and EMT inhibitory effects on U-2 OS and MG-63 cells.ALS markedly induced mitochondria-mediated apoptosis with a significant increase in the expression of key pro-apoptotic proteins and a decrease in main anti-apoptotic proteins.ALS is a promising anticancer agent in OS treatment and further studies are needed to confirm its efficacy and safety in OS chemotherapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Spinal Surgery, General Hospital of Ningxia Medical University, Yinchuan, People's Republic of China ; Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA ; Department of Orthopedics, General Hospital of Tianjin Medical University, Tianjin, People's Republic of China.

ABSTRACT
Osteosarcoma (OS) is the most common malignant bone tumor occurring mostly in children and adolescents between 10 and 20 years of age with poor response to current therapeutics. Alisertib (ALS, MLN8237) is a selective Aurora kinase A inhibitor that displays anticancer effects on several types of cancer. However, the role of ALS in the treatment of OS remains unknown. This study aimed to investigate the effects of ALS on the cell growth, apoptosis, autophagy, and epithelial to mesenchymal transition (EMT) and the underlying mechanisms in two human OS cell lines U-2 OS and MG-63. The results showed that ALS had potent growth inhibitory, pro-apoptotic, pro-autophagic, and EMT inhibitory effects on U-2 OS and MG-63 cells. ALS remarkably induced G2/M arrest and down-regulated the expression levels of cyclin-dependent kinases 1 and 2 and cyclin B1 in both U-2 OS and MG-63 cells. ALS markedly induced mitochondria-mediated apoptosis with a significant increase in the expression of key pro-apoptotic proteins and a decrease in main anti-apoptotic proteins. Furthermore, ALS promoted autophagic cell death via the inhibition of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) and p38 mitogen-activated protein kinase (p38 MAPK) signaling pathways, and activation of 5'-AMP-dependent kinase (AMPK) signaling pathway. Inducers or inhibitors of apoptosis or autophagy simultaneously altered ALS-induced apoptotic and autophagic death in both U-2 OS and MG-63 cells, suggesting a crosstalk between these two primary modes of programmed cell death. Moreover, ALS suppressed EMT-like phenotypes with a marked increase in the expression of E-cadherin but a decrease in N-cadherin in U-2 OS and MG-63 cells. ALS treatment also induced reactive oxygen species (ROS) generation but inhibited the expression levels of sirtuin 1 and nuclear factor-erythroid-2-related factor 2 (Nrf2) in both cell lines. Taken together, these findings show that ALS promotes apoptosis and autophagy but inhibits EMT via PI3K/Akt/mTOR, p38 MAPK, and AMPK signaling pathways with involvement of ROS- and sirtuin 1-associated pathways in U-2 OS and MG-63 cells. ALS is a promising anticancer agent in OS treatment and further studies are needed to confirm its efficacy and safety in OS chemotherapy.

No MeSH data available.


Related in: MedlinePlus