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Oleanane triterpenoid CDDO-Me induces apoptosis in multidrug resistant osteosarcoma cells through inhibition of Stat3 pathway.

Ryu K, Susa M, Choy E, Yang C, Hornicek FJ, Mankin HJ, Duan Z - BMC Cancer (2010)

Bottom Line: Treatment with CDDO-Me significantly decreased the level of nuclear translocation and phosphorylation of Stat3.Furthermore, CDDO-Me increased the cytotoxic effects of doxorubicin in the MDR osteosarcoma cell lines.This study provides the framework for the clinical evaluation of CDDO-Me, either as monotherapy or perhaps even more effectively in combination with doxorubicin to treat osteosarcoma and overcome drug resistance.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Orthopaedic Surgery, Massachusetts General Hospital, Boston, MA 02114, USA.

ABSTRACT

Background: The activation of signal transducer and activator of transcription 3 (Stat3) pathway correlates with tumor growth, survival, drug resistance and poor prognosis in osteosarcoma. To explore the potential therapeutic values of this pathway, we assessed both the expression and the activation of Stat3 pathway in several pairs of multidrug resistant (MDR) osteosarcoma cell lines, and tissues. To explore the potential therapeutic values of this pathway, we analyzed the ability of the synthetic oleanane triterpenoid, C-28 methyl ester of 2-cyano-3,12-dioxoolen-1,9-dien-28-oic acid (CDDO-Me), to inhibit Stat3 expression and activation as well as its effects on doxorubicin sensitivity in osteosarcoma cells.

Methods: Expression of Stat3, phosphorylated Stat3 (pStat3) and Stat3 targeted proteins, including Bcl-XL, Survivin and MCL-1 were determined in drug sensitive and MDR osteosarcoma cell lines and tissues by Western blot analysis. The effect of CDDO-Me on osteosarcoma cell growth was evaluated by MTT and apoptosis by PARP cleavage assay and caspase-3/7 activity.

Results: Stat3 pathway was activated in osteosarcoma tissues and in MDR cell lines. CDDO-Me inhibited growth and induced apoptosis in osteosarcoma cell lines. Treatment with CDDO-Me significantly decreased the level of nuclear translocation and phosphorylation of Stat3. The inhibition of Stat3 pathway correlated with the suppression of the anti-apoptotic Stat3 targeted genes Bcl-XL, survivin, and MCL-1. Furthermore, CDDO-Me increased the cytotoxic effects of doxorubicin in the MDR osteosarcoma cell lines.

Conclusions: Stat3 pathway is overexpressed in MDR osteosarcoma cells. CDDO-Me significantly inhibited Stat3 phosphorylation, Stat3 nuclear translocation and induced apoptosis in osteosarcoma. This study provides the framework for the clinical evaluation of CDDO-Me, either as monotherapy or perhaps even more effectively in combination with doxorubicin to treat osteosarcoma and overcome drug resistance.

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Caspase-3/7 activity was measured as a second parameter of apoptotic cell death. KHOS and KHOSR2 showed significant increase in apoptosis when they were treated with increasing concentrations of CDDO-Me. The experiment was repeated three times in triplicate.
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Figure 3: Caspase-3/7 activity was measured as a second parameter of apoptotic cell death. KHOS and KHOSR2 showed significant increase in apoptosis when they were treated with increasing concentrations of CDDO-Me. The experiment was repeated three times in triplicate.

Mentions: To confirm that CDDO-Me inhibits cell growth, KHOS, U-2OS, KHOSR2, U-2OSTR, and HOB-c were evaluated using MTT assay. The results showed that the growth of all cell lines was inhibited after treatment with CDDO-Me (Fig. 2A). CDDO-Me showed significantly higher antiproliferative activity in osteosarcoma cells than in osteoblast cells (p < 0.0001). The IC50 of each cells were HOB-c: 0.8 μM, KHOS: 0.15 μM, KHOSR2: 0.33 μM, U-2OS: 0.17, U-2OSTR: 0.39 μM. The effect of CDDO-Me on the induction of apoptosis was assessed by evaluating PARP cleavage and caspase assay for both drug sensitive and MDR osteosarcoma cell lines. PARP cleavage was detected in all cells after 24 hours of treatment with CDDO-Me. A dose-response analysis revealed the appearance of PARP cleavage products in the presence of 0.5 μmol/L of CDDO-Me for KHOS, KHOSR2 and 1.0 μmol/L of CDDO-Me for U-2OS, U-2OSTR (Fig. 2B). In addition, caspase-3/7 activity was significantly increased when KHOS and KHOSR2 were treated with increasing concentration of CDDO-Me (Fig. 3).


Oleanane triterpenoid CDDO-Me induces apoptosis in multidrug resistant osteosarcoma cells through inhibition of Stat3 pathway.

Ryu K, Susa M, Choy E, Yang C, Hornicek FJ, Mankin HJ, Duan Z - BMC Cancer (2010)

Caspase-3/7 activity was measured as a second parameter of apoptotic cell death. KHOS and KHOSR2 showed significant increase in apoptosis when they were treated with increasing concentrations of CDDO-Me. The experiment was repeated three times in triplicate.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2874784&req=5

Figure 3: Caspase-3/7 activity was measured as a second parameter of apoptotic cell death. KHOS and KHOSR2 showed significant increase in apoptosis when they were treated with increasing concentrations of CDDO-Me. The experiment was repeated three times in triplicate.
Mentions: To confirm that CDDO-Me inhibits cell growth, KHOS, U-2OS, KHOSR2, U-2OSTR, and HOB-c were evaluated using MTT assay. The results showed that the growth of all cell lines was inhibited after treatment with CDDO-Me (Fig. 2A). CDDO-Me showed significantly higher antiproliferative activity in osteosarcoma cells than in osteoblast cells (p < 0.0001). The IC50 of each cells were HOB-c: 0.8 μM, KHOS: 0.15 μM, KHOSR2: 0.33 μM, U-2OS: 0.17, U-2OSTR: 0.39 μM. The effect of CDDO-Me on the induction of apoptosis was assessed by evaluating PARP cleavage and caspase assay for both drug sensitive and MDR osteosarcoma cell lines. PARP cleavage was detected in all cells after 24 hours of treatment with CDDO-Me. A dose-response analysis revealed the appearance of PARP cleavage products in the presence of 0.5 μmol/L of CDDO-Me for KHOS, KHOSR2 and 1.0 μmol/L of CDDO-Me for U-2OS, U-2OSTR (Fig. 2B). In addition, caspase-3/7 activity was significantly increased when KHOS and KHOSR2 were treated with increasing concentration of CDDO-Me (Fig. 3).

Bottom Line: Treatment with CDDO-Me significantly decreased the level of nuclear translocation and phosphorylation of Stat3.Furthermore, CDDO-Me increased the cytotoxic effects of doxorubicin in the MDR osteosarcoma cell lines.This study provides the framework for the clinical evaluation of CDDO-Me, either as monotherapy or perhaps even more effectively in combination with doxorubicin to treat osteosarcoma and overcome drug resistance.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Orthopaedic Surgery, Massachusetts General Hospital, Boston, MA 02114, USA.

ABSTRACT

Background: The activation of signal transducer and activator of transcription 3 (Stat3) pathway correlates with tumor growth, survival, drug resistance and poor prognosis in osteosarcoma. To explore the potential therapeutic values of this pathway, we assessed both the expression and the activation of Stat3 pathway in several pairs of multidrug resistant (MDR) osteosarcoma cell lines, and tissues. To explore the potential therapeutic values of this pathway, we analyzed the ability of the synthetic oleanane triterpenoid, C-28 methyl ester of 2-cyano-3,12-dioxoolen-1,9-dien-28-oic acid (CDDO-Me), to inhibit Stat3 expression and activation as well as its effects on doxorubicin sensitivity in osteosarcoma cells.

Methods: Expression of Stat3, phosphorylated Stat3 (pStat3) and Stat3 targeted proteins, including Bcl-XL, Survivin and MCL-1 were determined in drug sensitive and MDR osteosarcoma cell lines and tissues by Western blot analysis. The effect of CDDO-Me on osteosarcoma cell growth was evaluated by MTT and apoptosis by PARP cleavage assay and caspase-3/7 activity.

Results: Stat3 pathway was activated in osteosarcoma tissues and in MDR cell lines. CDDO-Me inhibited growth and induced apoptosis in osteosarcoma cell lines. Treatment with CDDO-Me significantly decreased the level of nuclear translocation and phosphorylation of Stat3. The inhibition of Stat3 pathway correlated with the suppression of the anti-apoptotic Stat3 targeted genes Bcl-XL, survivin, and MCL-1. Furthermore, CDDO-Me increased the cytotoxic effects of doxorubicin in the MDR osteosarcoma cell lines.

Conclusions: Stat3 pathway is overexpressed in MDR osteosarcoma cells. CDDO-Me significantly inhibited Stat3 phosphorylation, Stat3 nuclear translocation and induced apoptosis in osteosarcoma. This study provides the framework for the clinical evaluation of CDDO-Me, either as monotherapy or perhaps even more effectively in combination with doxorubicin to treat osteosarcoma and overcome drug resistance.

Show MeSH
Related in: MedlinePlus