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Strahlentherapie und Onkologie

, Volume 189, Issue 8, pp 684–692 | Cite as

Dichloroacetate induces tumor-specific radiosensitivity in vitro but attenuates radiation-induced tumor growth delay in vivo

  • F. Zwicker
  • A. Kirsner
  • P. Peschke
  • F. Roeder
  • J. Debus
  • P.E. Huber
  • K.J. Weber
Original article

Abstract

Background

Inhibition of pyruvate dehydrogenase kinase (PDK) by dichloroacetate (DCA) can shift tumor cell metabolism from anaerobic glycolysis to glucose oxidation, with activation of mitochondrial activity and chemotherapy-dependent apoptosis. In radiotherapy, DCA could thus potentially enhance the frequently moderate apoptotic response of cancer cells that results from their mitochondrial dysfunction. The aim of this study was to investigate tumor-specific radiosensitization by DCA in vitro and in a human tumor xenograft mouse model in vivo.

Materials and methods

The interaction of DCA with photon beam radiation was investigated in the human tumor cell lines WIDR (colorectal) and LN18 (glioma), as well as in the human normal tissue cell lines HUVEC (endothelial), MRC5 (lung fibroblasts) and TK6 (lymphoblastoid). Apoptosis induction in vitro was assessed by DAPI staining and sub-G1 flow cytometry; cell survival was quantified by clonogenic assay. The effect of DCA in vivo was investigated in WIDR xenograft tumors growing subcutaneously on BALB/c-nu/nu mice, with and without fractionated irradiation. Histological examination included TUNEL and Ki67 staining for apoptosis and proliferation, respectively, as well as pinomidazole labeling for hypoxia.

Results

DCA treatment led to decreased clonogenic survival and increased specific apoptosis rates in tumor cell lines (LN18, WIDR) but not in normal tissue cells (HUVEC, MRC5, TK6). However, this significant tumor-specific radiosensitization by DCA in vitro was not reflected by the situation in vivo: The growth suppression of WIDR xenograft tumors after irradiation was reduced upon additional DCA treatment (reflected by Ki67 expression levels), although early tumor cell apoptosis rates were significantly increased by DCA. This apparently paradoxical effect was accompanied by a marked DCA-dependent induction of hypoxia in tumor-tissue.

Conclusion

DCA induced tumor-specific radiosensitization in vitro but not in vivo. DCA also induced development of hypoxia in tumor tissue in vivo. Further investigations relating to the interplay between tumor cell metabolism and tumor microenvironment are necessary to explain the limited success of metabolic targeting in radiotherapy.

Keywords

Citric acid cycle Glycolysis Irradiation Hypoxia Apoptosis 

Tumorspezifische Radiosensibilisierung durch Dichloracetat in vitro bei Verminderung der strahlungsinduzierten Tumorwachstumsverzögerung in vivo

Zusammenfassung

Hintergrund

Eine Inhibition der Pyruvatdehydrogenase (PDK) durch Dichloracetat (DCA) kann zu einer Verschiebung des Tumorzellstoffwechsels von der anaeroben Glykolyse zur Glukoseoxidation mit Aktivierung der Mitochondrienaktivität sowie der chemotherapieabhängigen Apoptose führen. In der Strahlentherapie könnte DCA potenziell die Induktion der Apoptose in Tumorzellen verstärken, die aufgrund einer mitochondrialen Dysfunktion normalerweise mäßig ist. Ziel dieser Untersuchung war, eine tumorspezifische Radiosensibilisierung durch DCA in vitro und in einem humanen Xenograft-Tumor-Maus-Modell zu untersuchen.

Material und Methoden

Die Interaktion von DCA und Photonenstrahlen wurde an den humanen Tumorzelllinien WIDR (kolorektal) und LN18 (Gliom) sowie an den Normalgewebszelllinien HUVEC (endothelial), MRC5 (Lungenfibroblasten) und TK6 (lymphoblastoid) untersucht. Die Induktion der Apoptose in vitro wurde mithilfe der DAPI-Färbung und Sub-G1-Durchflusszytometrie, das Zellüberleben mithilfe eines klonogenen Assays bestimmt. In vivo wurde der DCA-Effekt an subkutan wachsenden WIDR-Xenograft-Tumoren in Mäusen mit und ohne fraktionierte Bestrahlung untersucht. Histologische Untersuchungen schlossen die TUNEL- und Ki-67-Färbung (Apoptose und Proliferation) sowie eine Pimonidazolfärbung zum Hypoxienachweis ein.

Ergebnisse

DCA führt nur in den Tumorzelllinien (LN18, WIDR), nicht aber in den Normalgewebszelllinien (HUVEC, MRC5, TK6) zu einem verringerten klonogenen Überleben sowie einer gesteigerten spezifischen Apoptoserate und somit zu einer signifikanten tumorspezifischen Radiosensibilisierung in vitro. Umgekehrt war die Wachstumsverzögerung von WIDR-Xenograft-Tumoren nach Bestrahlung durch eine zusätzliche DCA-Behandlung, korrespondierend mit der Ki-67-Expession, geringer ausgeprägt, obwohl die frühe Tumorapoptose durch DCA signifikant gesteigert wurde. Diesen scheinbar paradoxen Effekt begleitete eine deutliche DCA-induzierte Tumorhypoxie im Tumormodell.

Schlussfolgerung

DCA bewirkt eine tumorspezifische Radiosensibilisierung in vitro, nicht aber in vivo. DCA fördert zudem die Hypoxie-Entwicklung im Tumorgewebe in vivo. Weitere Untersuchungen des komplexen Zusammenspiels zwischen dem Tumorzellstoffwechsel und der Tumormikroumwelt sind notwendig, um den begrenzten Erfolg des „metabolic targeting“ in der Strahlentherapie zu erklären.

Schlüsselwörter

Zitronensäurezyklus Glykolyse Bestrahlung Hypoxie Apoptose 

Notes

Acknowledgments

The authors wish to thank Thuy Trinh, Alexandra Tietz, Angela Funk, Marina Sczymbara and Angela Funk for their excellent technical assistance. This work was supported in part by a grant from the Kompetenzverbund Strahlenforschung (KVSF, 03NUK004A, C) of the Bundesministerien fuer Bildung, Forschung und Umwelt (BMBF/BMU).

Conflict of interest

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

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Copyright information

© Springer Heidelberg Berlin 2013

Authors and Affiliations

  • F. Zwicker
    • 1
    • 2
  • A. Kirsner
    • 1
  • P. Peschke
    • 2
  • F. Roeder
    • 1
    • 2
  • J. Debus
    • 1
    • 2
  • P.E. Huber
    • 1
    • 2
  • K.J. Weber
    • 1
  1. 1.Department of Radiation OncologyUniversity Hospital Center HeidelbergHeidelbergGermany
  2. 2.Clinical Cooperation Unit Molecular Radiation OncologyDKFZ, HeidelbergHeidelbergGermany

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