Radiotherapy & Oncology
Volume 92, Issue 3 , Pages 334-338, September 2009

2-Methoxyestradiol-induced radiosensitization is independent of SOD but depends on inhibition of Akt and DNA-PKcs activities

  • Urszula Florczak

      Affiliations

    • Department of Radiation Oncology, University of Tuebingen, Germany
    • These authors shared the first authorship of this paper.
    • ,
  • Mahmoud Toulany

      Affiliations

    • Department of Radiation Oncology, University of Tuebingen, Germany
    • These authors shared the first authorship of this paper.
    • ,
  • Rainer Kehlbach

      Affiliations

    • Department of Diagnostic and Interventional Radiology, University of Tuebingen, Germany
    • ,
  • H. Peter Rodemann

      Affiliations

    • Department of Radiation Oncology, University of Tuebingen, Germany
    • Corresponding Author InformationCorresponding author. Address: Division of Radiobiology and Molecular Environmental Research, Department of Radiation Oncology, Eberhard-Karls University Tuebingen, Roentgenweg 11, 72076 Tuebingen, Germany.

Received 24 April 2009; received in revised form 2 June 2009; accepted 5 June 2009. published online 09 July 2009.

Abstract 

Background and purpose

2-Methoxyestradiol (2-ME) is described as an inhibitor of the superoxide dismutase (SOD) enzyme activity. However, it attenuates PI3K/Akt pathway and induces radiosensitization in human tumor cells as well. Since the activation of catalytic subunit of DNA-protein kinase (DNA-PKcs) is partially regulated by Akt activity, in the present study we investigated whether 2-ME-induced radiosensitization is dependent on inhibition of Akt and DNA-PKcs activities or on SOD targeting.

Materials and methods

This study was performed using the lung carcinoma cell line A549. Ionizing radiation-induced SOD activity was analyzed by superoxide dismutase activity assay. Applying Western blotting, the pattern of radiation-induced SOD expression and activation of Akt as well as DNA-PKcs was analyzed. Colony formation assay and γH2AX foci assay were performed to measure radiosensitization and DNA-double strand break (DNA-DSB) repair. To downregulate SOD expression small interfering RNA (siRNA) was used.

Results

Irradiation with 4Gy stimulated SOD enzyme activity as early as 1min after radiation exposure. Expression of Cu/Zn-SOD (SOD1) as well as Mn-SOD (SOD2) was increased by single doses of 1–4Gy within 24–36h. 2-ME blocked radiation-induced SOD enzyme activity but not protein expression and enhanced radiation sensitivity. Pretreatment with 2-ME blocked IR-induced Akt as well as DNA-PKcs phosphorylation and impaired the repair of DNA-DSB. SiRNA targeting of SOD1 and SOD2 affected neither DNA-PKcs phosphorylation nor post-irradiation survival while inhibition of Akt by specific inhibitor abrogated 2-ME-induced radiosensitization.

Conclusion

These results may indicate that 2-ME-induced radiosensitization is independent of SOD inhibition but mainly depends on inhibition of Akt and DNA-PKcs activities.

Keywords: Ionizing radiation, Superoxide dismutases, 2-ME, DNA-PKcs, Akt

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PII: S0167-8140(09)00317-X

doi:10.1016/j.radonc.2009.06.005

Radiotherapy & Oncology
Volume 92, Issue 3 , Pages 334-338, September 2009

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