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Radiosensitization by histone deacetylase inhibition in an osteosarcoma mouse model

Radiosensitivierung durch Histon-Deacetylasehemmung im Osteosarkom- Mausmodell

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Abstract

Background

Osteosarcomas (OS) are highly malignant and radioresistant tumors. Histone deacetylase inhibitors (HDACi) constitute a novel class of anticancer agents. We sought to investigate the effect of combined treatment with suberoylanilide hydroxamic acid (SAHA) and radiotherapy in OS in vivo.

Methods

Clonogenic survival of human OS cell lines as well as tumor growth delay of OS xenografts were tested after treatment with either vehicle, radiotherapy (XRT), SAHA, or XRT and SAHA. Tumor proliferation, necrosis, microvascular density, apoptosis, and p53/p21 were monitored by immunohistochemistry. The CD95 pathway was performed by flow cytometry, caspase (3/7/8) activity measurements, and functional inhibition of CD95 death signaling.

Results

Combined treatment with SAHA and XRT markedly reduced the surviving fraction of OS cells as compared to XRT alone. Likewise, dual therapy significantly inhibited OS tumor growth in vivo as compared to XRT alone, reflected by reduced tumor proliferation, impaired angiogenesis, and increased apoptosis. Addition of HDACi to XRT led to elevated p53, p21, CD95, and CD95L expression. Inhibition of CD95 signaling reduced HDACi- and XRT-induced apoptosis.

Conclusion

Our data show that HDACi increases the radiosensitivity of osteosarcoma cells at least in part via ligand-induced apoptosis. HDACi thus emerge as potentially useful treatment components of OS.

Zusammenfassung

Zielsetzung

Osteosarkome (OS) sind hochmaligne und relative radioresistente Tumore. Histon-Deacetylase-Inhibitoren (HDACi) stellen eine neue Substanzklasse in der Tumortherapie dar, insbesondere in Kombination mit anderen antineoplastischen Therapiestrategien. Wir haben deshalb die Effekte der Kombinationstherapie von Suberoylanilide Hydroxamic Acid (SAHA) und konventioneller Radiotherapie im OS-Mausmodell untersucht.

Methoden

Es wurde die Tumorwachstumsverzögerung in OS-Xenografts nach Behandlung mit Radiotherapie (XRT), SAHA alleine oder XRT und SAHA untersucht. Die Tumorproliferation, Tumornekrose, Tumorvaskularisation, Apoptose und p53/p21-Expression wurden immunohistochemisch analysiert. Die Bestimmung der CD95-vermittelten Apoptose erfolgte durchflusszytometrisch sowie durch Messung der Caspase-(3-/7-/8-)Aktivität und funktionaler Inhibition des CD95-Liganden.

Ergebnisse

Die Kombinationsbehandlung führt zu einer signifikanten Reduktion des Tumorwachstums in vivo im Vergleich zur alleinigen Radiotherapie, reflektiert durch eine verminderte Tumorproliferation sowie reduzierte Angiogenese und gesteigerte Apoptoseinduktion. Die Kombination von HDACi und XRT resultiert in einer gesteigerten p53-, p21-, CD95- und CD95L-Expression. Eine Inhibition des CD95-Liganden führt zu einer reduzierten HDACi- und XRT-induzierten Apoptoserate nach Kombinationstherapie.

Schlußfolgerung

Die Ergebnisse zeigen, dass eine HDACi die Radiosensitivität von Osteosarkomzellen signifikant erhöht. Dies erfolgt – zumindest teilweise – durch die CD95-abhängige Apoptoseinduktion. Die Autoren sind daher der Meinung, dass HDACi eine potenziell wirksame Therapiestrategie beim OS darstellt.

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Conflict of interest

On behalf of all authors, the corresponding author states that there are no conflicts of interests.

Acknowledgments

This study was supported by the German Krebshilfe (Deutsche Krebshilfe, 109665 and Max-Eder 108876), the German Federal Ministry of Research and Technology (Bundesministerium für Bildung und Forschung; BMBF 03NUK004A/C) and the Dietmar Hopp Stiftung. Furthermore, we would like to thank Ludmilla Frick, Sylvia Trinh, Claudia Rittmueller, Gabriele Becker, Alexandra Tietz, Angela Funk, and Andreas Griesbach for their excellent technical assistance.

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Author notes

  1. C. Blattmann M.D.

    Present address: Pädiatrie 5, Olgahospital, Bismarckstr. 8, 70176, Stuttgart, Germany

Authors and Affiliations

  1. Department of Pediatric Oncology, Hematology and Immunology, University Children’s Hospital of Heidelberg, Heidelberg, Germany

    C. Blattmann M.D., E. Roth M.D. & A.E. Kulozik

  2. Departments of Radiotherapy, Molecular- and Translational Radiation Oncology, Institute of Radiation Oncology (HIRO), National Center for Tumor diseases (NCT), German Cancer Research Center (DKFZ) , Heidelberg, Germany

    M. Thiemann PhD, A. Christmann M.D., J. Debus, P.E. Huber, S. Oertel M.D. & A. Abdollahi M.D.

  3. Institute of Pathology, University of Heidelberg, Heidelberg, Germany

    A. Stenzinger M.D., V. Ehemann PhD & W. Weichert

  4. Center of Cancer Systems Biology, St. Elizabeth’s Medical Center, Tufts University School of Medicine, Boston, MA, USA

    A. Abdollahi M.D.

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  1. C. Blattmann M.D.
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Correspondence to C. Blattmann M.D..

Additional information

Oertel and Abdollahi share senior authorship.

Electronic supplementary material

66_2013_372_MO1_ESM.pdf

Supplemental Figures: „Determination of XRT dosage“ and „Course of weight in xenografts treated with vehicle, SAHA, XRT or XRT and SAHA“ (PDF 90kB)

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Blattmann, C., Thiemann, M., Stenzinger, A. et al. Radiosensitization by histone deacetylase inhibition in an osteosarcoma mouse model. Strahlenther Onkol 189, 957–966 (2013). https://doi.org/10.1007/s00066-013-0372-8

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  • DOI: https://doi.org/10.1007/s00066-013-0372-8

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