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Ophiobolin O isolated from Aspergillus ustus induces G1 arrest of MCF-7 cells through interaction with AKT/GSK3β/cyclin D1 signaling.

Lv C, Qin W, Zhu T, Wei S, Hong K, Zhu W, Chen R, Huang C - Mar Drugs (2015)

Bottom Line: However, the anti-tumor effect and the mechanism of ophiobolin O remain unclear.In vivo, ophiobolin O suppressed tumor growth and showed little toxicity in mouse xenograft models.Overall, these findings provide theoretical basis for the therapeutic use of ophiobolin O.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, College of Basic Medical Second Military Medical University, 800 Xiangyin Road, Shanghai 200433, China. lvcuiting961021@126.com.

ABSTRACT
Ophiobolin O is a member of ophiobolin family, which has been proved to be a potent anti-tumor drug candidate for human breast cancer. However, the anti-tumor effect and the mechanism of ophiobolin O remain unclear. In this study, we further verified ophiobolin O-induced G1 phase arrest in human breast cancer MCF-7 cells, and found that ophiobolin O reduced the phosphorylation level of AKT and GSK3β, and induced down-regulation of cyclin D1. The inverse docking (INVDOCK) analysis indicated that ophiobolin O could bind to GSK3β, and GSK3β knockdown abolished cyclin D1 degradation and G1 phase arrest. Pre-treatment with phosphatase inhibitor sodium or thovanadate halted dephosphorylation of AKT and GSK3β, and blocked ophiobolin O-induced G1 phase arrest. These data suggest that ophiobolin O may induce G1 arrest in MCF-7 cells through interaction with AKT/GSK3β/cyclin D1 signaling. In vivo, ophiobolin O suppressed tumor growth and showed little toxicity in mouse xenograft models. Overall, these findings provide theoretical basis for the therapeutic use of ophiobolin O.

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Ophiobolin O induces cell cycle arrest in MCF-7 cells. (A) Ophiobolin O caused cell cycle arrest at the G1 phase. Cells were treated with vehicle and 15 μM Ophiobolin O for 12, 24, and 48 h, and then cell cycle distribution was assessed using flow cytometry; (B) The percentage of cells in different phases of the cell cycle was represented by a bar diagram. The percentage of cells in each population was shown as mean ± SD from three independent experiments. *p < 0.05, **p < 0.01; (C) Western blot analysis of G1 transition-related proteins were analyzed by Western blotting assay. GAPDH was used as an equal loading control.
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marinedrugs-13-00431-f002: Ophiobolin O induces cell cycle arrest in MCF-7 cells. (A) Ophiobolin O caused cell cycle arrest at the G1 phase. Cells were treated with vehicle and 15 μM Ophiobolin O for 12, 24, and 48 h, and then cell cycle distribution was assessed using flow cytometry; (B) The percentage of cells in different phases of the cell cycle was represented by a bar diagram. The percentage of cells in each population was shown as mean ± SD from three independent experiments. *p < 0.05, **p < 0.01; (C) Western blot analysis of G1 transition-related proteins were analyzed by Western blotting assay. GAPDH was used as an equal loading control.

Mentions: In order to test whether the inhibition effect of ophiobolin O was related to cell cycle progression, cell cycle distribution and related checkpoint factors were studied. MCF-7 cells were treated with 15 μM ophiobolin O for 12 h, 24 h and 48 h, respectively. The results showed that cells treated with ophiobolin O accumulated progressively in G1 phase (Figure 2A,B). Compared with the negative control, treatment with ophiobolin O resulted in a significant increase in the proportion of G1 phase cells (control: 40.19% ± 1.03%; 12 h: 46.09% ± 5.79%; 24 h: 56.18% ± 2.19%; 48 h: 72.56% ± 1.35%) and about a 1.8-fold increase after 48 h. Meanwhile, G2/M reduced appreciably from 32.57% ± 0.69% to 5.53% ± 0.88% during 48 h incubation. These results suggest that G1 phase arrest accounts for the anti-proliferative effect of ophiobolin O observed in MCF-7 cells. Besides, ophiobolin O treatment resulted in a time-dependent decrease in the protein expression of cyclin D1, cyclin E, CDK2 and increase of p-cyclin D1 (Thr286), p21 and p27. However, the protein levels of CDK4 and CDK6 were not significantly changed during ophiobolin O treatment (Figure 2b).


Ophiobolin O isolated from Aspergillus ustus induces G1 arrest of MCF-7 cells through interaction with AKT/GSK3β/cyclin D1 signaling.

Lv C, Qin W, Zhu T, Wei S, Hong K, Zhu W, Chen R, Huang C - Mar Drugs (2015)

Ophiobolin O induces cell cycle arrest in MCF-7 cells. (A) Ophiobolin O caused cell cycle arrest at the G1 phase. Cells were treated with vehicle and 15 μM Ophiobolin O for 12, 24, and 48 h, and then cell cycle distribution was assessed using flow cytometry; (B) The percentage of cells in different phases of the cell cycle was represented by a bar diagram. The percentage of cells in each population was shown as mean ± SD from three independent experiments. *p < 0.05, **p < 0.01; (C) Western blot analysis of G1 transition-related proteins were analyzed by Western blotting assay. GAPDH was used as an equal loading control.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4306945&req=5

marinedrugs-13-00431-f002: Ophiobolin O induces cell cycle arrest in MCF-7 cells. (A) Ophiobolin O caused cell cycle arrest at the G1 phase. Cells were treated with vehicle and 15 μM Ophiobolin O for 12, 24, and 48 h, and then cell cycle distribution was assessed using flow cytometry; (B) The percentage of cells in different phases of the cell cycle was represented by a bar diagram. The percentage of cells in each population was shown as mean ± SD from three independent experiments. *p < 0.05, **p < 0.01; (C) Western blot analysis of G1 transition-related proteins were analyzed by Western blotting assay. GAPDH was used as an equal loading control.
Mentions: In order to test whether the inhibition effect of ophiobolin O was related to cell cycle progression, cell cycle distribution and related checkpoint factors were studied. MCF-7 cells were treated with 15 μM ophiobolin O for 12 h, 24 h and 48 h, respectively. The results showed that cells treated with ophiobolin O accumulated progressively in G1 phase (Figure 2A,B). Compared with the negative control, treatment with ophiobolin O resulted in a significant increase in the proportion of G1 phase cells (control: 40.19% ± 1.03%; 12 h: 46.09% ± 5.79%; 24 h: 56.18% ± 2.19%; 48 h: 72.56% ± 1.35%) and about a 1.8-fold increase after 48 h. Meanwhile, G2/M reduced appreciably from 32.57% ± 0.69% to 5.53% ± 0.88% during 48 h incubation. These results suggest that G1 phase arrest accounts for the anti-proliferative effect of ophiobolin O observed in MCF-7 cells. Besides, ophiobolin O treatment resulted in a time-dependent decrease in the protein expression of cyclin D1, cyclin E, CDK2 and increase of p-cyclin D1 (Thr286), p21 and p27. However, the protein levels of CDK4 and CDK6 were not significantly changed during ophiobolin O treatment (Figure 2b).

Bottom Line: However, the anti-tumor effect and the mechanism of ophiobolin O remain unclear.In vivo, ophiobolin O suppressed tumor growth and showed little toxicity in mouse xenograft models.Overall, these findings provide theoretical basis for the therapeutic use of ophiobolin O.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, College of Basic Medical Second Military Medical University, 800 Xiangyin Road, Shanghai 200433, China. lvcuiting961021@126.com.

ABSTRACT
Ophiobolin O is a member of ophiobolin family, which has been proved to be a potent anti-tumor drug candidate for human breast cancer. However, the anti-tumor effect and the mechanism of ophiobolin O remain unclear. In this study, we further verified ophiobolin O-induced G1 phase arrest in human breast cancer MCF-7 cells, and found that ophiobolin O reduced the phosphorylation level of AKT and GSK3β, and induced down-regulation of cyclin D1. The inverse docking (INVDOCK) analysis indicated that ophiobolin O could bind to GSK3β, and GSK3β knockdown abolished cyclin D1 degradation and G1 phase arrest. Pre-treatment with phosphatase inhibitor sodium or thovanadate halted dephosphorylation of AKT and GSK3β, and blocked ophiobolin O-induced G1 phase arrest. These data suggest that ophiobolin O may induce G1 arrest in MCF-7 cells through interaction with AKT/GSK3β/cyclin D1 signaling. In vivo, ophiobolin O suppressed tumor growth and showed little toxicity in mouse xenograft models. Overall, these findings provide theoretical basis for the therapeutic use of ophiobolin O.

Show MeSH
Related in: MedlinePlus