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The AKT inhibitor MK-2206 is cytotoxic in hepatocarcinoma cells displaying hyperphosphorylated AKT-1 and synergizes with conventional chemotherapy.

Simioni C, Martelli AM, Cani A, Cetin-Atalay R, McCubrey JA, Capitani S, Neri LM - Oncotarget (2013)

Bottom Line: The inhibitor decreased cell viability and induced cell cycle arrest in the G0/G1 phase of the cell cycle, with a higher efficacy in cells with hyperphosphorylated Akt-1.MK-2206 synergized with doxorubicin, a chemotherapeutic drug widely used for HCC treatment.Our findings suggest that the use of Akt inhibitors, either alone or in combination with doxorubicin, may be considered as an attractive therapeutic regimen for the treatment of HCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy.

ABSTRACT
Hepatocellular carcinoma (HCC) is one of the most common potentially lethal human malignancies worldwide. Advanced or recurrent HCC is frequently resistant to conventional chemotherapeutic agents and radiation. Therefore, targeted agents with tolerable toxicity are mandatory to improve HCC therapy and prognosis. In this neoplasia, the PI3K/Akt signaling network has been frequently shown to be aberrantly up-regulated. To evaluate whether Akt could represent a target for treatment of HCC, we studied the effects of the allosteric Akt inhibitor, MK-2206, on a panel of HCC cell lines characterized by different levels of Akt-1 activation. The inhibitor decreased cell viability and induced cell cycle arrest in the G0/G1 phase of the cell cycle, with a higher efficacy in cells with hyperphosphorylated Akt-1. Moreover, MK-2206 induced apoptosis, as documented by Annexin V labeling, and also caused autophagy, as evidenced by increased levels of the autophagy marker LC3A/B. Autophagy was shown to be a protective mechanism against MK-2206 cytotoxicity. MK-2206 down-regulated, in a concentration-dependent manner, the phosphorylation levels of Akt-1 and its downstream targets, GSK3 α/β and FOXO3A. MK-2206 synergized with doxorubicin, a chemotherapeutic drug widely used for HCC treatment. Our findings suggest that the use of Akt inhibitors, either alone or in combination with doxorubicin, may be considered as an attractive therapeutic regimen for the treatment of HCC.

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MK-2206 induces apoptosis and autophagy in HCC cell lines(A, B) Analysis of Annexin-V positive cells after MK-2206 treatment using the Muse™ Cell Analyzer in PLC and Mahlavu cells. The analysis was performed after 24h of treatment with increasing concentrations of MK-2206. The results are the mean ± s.d. of three different experiments. Asterisks indicate significant differences (p<0.05) in comparison with control (CTRL). BK represents the unstained samples. In both cell lines the apoptotic process was further increased when the drug was administered at the concentration of 10 μM. (C) Western blot analysis documenting increased expression of the fast-migrating (lipidated) form of LC3A/B in Mahlavu and SNU475 cell lines treated with MK-2206. An antibody to β-actin documented equal lane loading. (D) xCELLigence System analysis documenting the effects of chloroquine (CQ) on growth of Mahlavu cells treated with MK-2206. One representative of three different experiments is shown.
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Figure 3: MK-2206 induces apoptosis and autophagy in HCC cell lines(A, B) Analysis of Annexin-V positive cells after MK-2206 treatment using the Muse™ Cell Analyzer in PLC and Mahlavu cells. The analysis was performed after 24h of treatment with increasing concentrations of MK-2206. The results are the mean ± s.d. of three different experiments. Asterisks indicate significant differences (p<0.05) in comparison with control (CTRL). BK represents the unstained samples. In both cell lines the apoptotic process was further increased when the drug was administered at the concentration of 10 μM. (C) Western blot analysis documenting increased expression of the fast-migrating (lipidated) form of LC3A/B in Mahlavu and SNU475 cell lines treated with MK-2206. An antibody to β-actin documented equal lane loading. (D) xCELLigence System analysis documenting the effects of chloroquine (CQ) on growth of Mahlavu cells treated with MK-2206. One representative of three different experiments is shown.

Mentions: In order to establish whether decreased cell growth was related to apoptosis in HCC cell lines, we analyzed programmed cell death by Annexin-V/7-AAD-assay using the MuseTM Cell Analyzer. PLC and Mahlavu cells were treated with increasing concentrations of MK-2206 for 24h and then analyzed for Annexin-V labeling. MK-2206 induced concentration-dependent apoptosis in both cell lines. However, at 5 μM concentration of MK-2206, apoptosis was 11.7% in PLC cells, (Fig. 3A), whereas it reached 51.6 % in Mahlavu cells (Fig. 3B).


The AKT inhibitor MK-2206 is cytotoxic in hepatocarcinoma cells displaying hyperphosphorylated AKT-1 and synergizes with conventional chemotherapy.

Simioni C, Martelli AM, Cani A, Cetin-Atalay R, McCubrey JA, Capitani S, Neri LM - Oncotarget (2013)

MK-2206 induces apoptosis and autophagy in HCC cell lines(A, B) Analysis of Annexin-V positive cells after MK-2206 treatment using the Muse™ Cell Analyzer in PLC and Mahlavu cells. The analysis was performed after 24h of treatment with increasing concentrations of MK-2206. The results are the mean ± s.d. of three different experiments. Asterisks indicate significant differences (p<0.05) in comparison with control (CTRL). BK represents the unstained samples. In both cell lines the apoptotic process was further increased when the drug was administered at the concentration of 10 μM. (C) Western blot analysis documenting increased expression of the fast-migrating (lipidated) form of LC3A/B in Mahlavu and SNU475 cell lines treated with MK-2206. An antibody to β-actin documented equal lane loading. (D) xCELLigence System analysis documenting the effects of chloroquine (CQ) on growth of Mahlavu cells treated with MK-2206. One representative of three different experiments is shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3824526&req=5

Figure 3: MK-2206 induces apoptosis and autophagy in HCC cell lines(A, B) Analysis of Annexin-V positive cells after MK-2206 treatment using the Muse™ Cell Analyzer in PLC and Mahlavu cells. The analysis was performed after 24h of treatment with increasing concentrations of MK-2206. The results are the mean ± s.d. of three different experiments. Asterisks indicate significant differences (p<0.05) in comparison with control (CTRL). BK represents the unstained samples. In both cell lines the apoptotic process was further increased when the drug was administered at the concentration of 10 μM. (C) Western blot analysis documenting increased expression of the fast-migrating (lipidated) form of LC3A/B in Mahlavu and SNU475 cell lines treated with MK-2206. An antibody to β-actin documented equal lane loading. (D) xCELLigence System analysis documenting the effects of chloroquine (CQ) on growth of Mahlavu cells treated with MK-2206. One representative of three different experiments is shown.
Mentions: In order to establish whether decreased cell growth was related to apoptosis in HCC cell lines, we analyzed programmed cell death by Annexin-V/7-AAD-assay using the MuseTM Cell Analyzer. PLC and Mahlavu cells were treated with increasing concentrations of MK-2206 for 24h and then analyzed for Annexin-V labeling. MK-2206 induced concentration-dependent apoptosis in both cell lines. However, at 5 μM concentration of MK-2206, apoptosis was 11.7% in PLC cells, (Fig. 3A), whereas it reached 51.6 % in Mahlavu cells (Fig. 3B).

Bottom Line: The inhibitor decreased cell viability and induced cell cycle arrest in the G0/G1 phase of the cell cycle, with a higher efficacy in cells with hyperphosphorylated Akt-1.MK-2206 synergized with doxorubicin, a chemotherapeutic drug widely used for HCC treatment.Our findings suggest that the use of Akt inhibitors, either alone or in combination with doxorubicin, may be considered as an attractive therapeutic regimen for the treatment of HCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy.

ABSTRACT
Hepatocellular carcinoma (HCC) is one of the most common potentially lethal human malignancies worldwide. Advanced or recurrent HCC is frequently resistant to conventional chemotherapeutic agents and radiation. Therefore, targeted agents with tolerable toxicity are mandatory to improve HCC therapy and prognosis. In this neoplasia, the PI3K/Akt signaling network has been frequently shown to be aberrantly up-regulated. To evaluate whether Akt could represent a target for treatment of HCC, we studied the effects of the allosteric Akt inhibitor, MK-2206, on a panel of HCC cell lines characterized by different levels of Akt-1 activation. The inhibitor decreased cell viability and induced cell cycle arrest in the G0/G1 phase of the cell cycle, with a higher efficacy in cells with hyperphosphorylated Akt-1. Moreover, MK-2206 induced apoptosis, as documented by Annexin V labeling, and also caused autophagy, as evidenced by increased levels of the autophagy marker LC3A/B. Autophagy was shown to be a protective mechanism against MK-2206 cytotoxicity. MK-2206 down-regulated, in a concentration-dependent manner, the phosphorylation levels of Akt-1 and its downstream targets, GSK3 α/β and FOXO3A. MK-2206 synergized with doxorubicin, a chemotherapeutic drug widely used for HCC treatment. Our findings suggest that the use of Akt inhibitors, either alone or in combination with doxorubicin, may be considered as an attractive therapeutic regimen for the treatment of HCC.

Show MeSH
Related in: MedlinePlus