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Radiother Oncol. 2015 Sep;116(3):473-9. doi: 10.1016/j.radonc.2015.03.026. Epub 2015 Apr 9.

γH2AX assay in ex vivo irradiated tumour specimens: A novel method to determine tumour radiation sensitivity in patient-derived material.

Author information

1
Department of Radiation Oncology, Medical Faculty and University Hospital, Eberhard Karls University Tübingen, Germany; German Cancer Research Center (DKFZ), Heidelberg, Germany and German Cancer Consortium (DKTK), Partner Site Tübingen, Germany.
2
German Cancer Research Center (DKFZ), Heidelberg, Germany and German Cancer Consortium (DKTK), Partner Site Dresden, Germany; Department of Radiation Oncology, Faculty of Medicine and University Hospital Carl Gustav Carus, Technische Universität Dresden, Germany; OncoRay - National Center for Radiation Research in Oncology, Faculty of Medicine and University Hospital Carl Gustav Carus, Technische Universität Dresden, Germany.
3
Department of Radiation Oncology (Maastro), GROW-School for Oncology and Developmental Biology, Maastricht University Medical Centre, The Netherlands.
4
Department of Biostatistics, The University of Texas MD Anderson Cancer Center, Houston, USA.
5
Institute for Legal Medicine, Faculty of Medicine, University Hospital Carl Gustav Carus, Technische Universität Dresden, Germany.
6
Department of Urology, Medical Faculty and University Hospital, Eberhard Karls University Tübingen, Germany.
7
Department of Pathology, Medical Faculty and University Hospital, Eberhard Karls University Tübingen, Germany.
8
Department of Neurosurgery, Medical Faculty and University Hospital, Eberhard Karls University Tübingen, Germany.
9
German Cancer Research Center (DKFZ), Heidelberg, Germany and German Cancer Consortium (DKTK), Partner Site Dresden, Germany; Department of Radiation Oncology, Faculty of Medicine and University Hospital Carl Gustav Carus, Technische Universität Dresden, Germany; OncoRay - National Center for Radiation Research in Oncology, Faculty of Medicine and University Hospital Carl Gustav Carus, Technische Universität Dresden, Germany; Institute of Radiooncology, Helmholtz-Zentrum Dresden - Rossendorf, Germany.

Abstract

PURPOSE:

To establish a clinically applicable protocol for quantification of residual γH2AX foci in ex vivo irradiated tumour samples and to apply this method in a proof-of-concept feasibility study to patient-derived tumour specimens.

MATERIAL AND METHODS:

Evaluation of γH2AX foci formation and disappearance in excised FaDu tumour specimens after (a) different incubation times in culture medium, 4Gy irradiation and fixation after 24h (cell recovery), (b) 10h medium incubation, 4Gy irradiation and fixation after various time points (double strand break repair kinetics), and (c) 10h medium incubation, irradiation with graded single radiation doses and fixation after 24h (dose-response). The optimised protocol was applied to patient-derived samples of seminoma, prostate cancer and glioblastoma multiforme.

RESULTS:

Post excision or biopsy, tumour tissues showed stable radiation-induced γH2AX foci values in oxic cells after >6h of recovery in medium. Kinetics of foci disappearance indicated a plateau of residual foci after >12h following ex vivo irradiation. Fitting the dose-response of residual γH2AX foci yielded slopes comparable with in situ irradiation of FaDu tumours. Significant differences in the slopes of ex vivo irradiated patient-derived tumour samples were found.

CONCLUSION:

A novel clinically applicable method to quantify residual γH2AX foci in ex vivo irradiated tumour samples was established. The first clinical results suggest that this method allows to distinguish between radiosensitive and radioresistant tumour types. These findings support further translational evaluation of this assay to individualise radiation therapy.

KEYWORDS:

Biomarker; DNA repair; Radiotherapy; Tumour biopsy; γH2AX foci

PMID:
25866027
DOI:
10.1016/j.radonc.2015.03.026
[Indexed for MEDLINE]
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