Open Access Research article

The radiosensitising effect of gemcitabine and its main metabolite dFdU under low oxygen conditions is in vitro not dependent on functional HIF-1 protein

An Wouters1*, Bea Pauwels1, Natalie Burrows23, Marc Baay1, Vanessa Deschoolmeester1, Trung Nghia Vu4, Kris Laukens4, Paul Meijnders5, Dirk Van Gestel5, Kaye J Williams2, Danielle Van den Weyngaert5, Jan B Vermorken16, Patrick Pauwels17, Marc Peeters16 and Filip Lardon1

Author Affiliations

1 Center for Oncological Research Antwerp, University of Antwerp, Universiteitsplein 1, 2610 Wilrijk, Belgium

2 Hypoxia and Therapeutics Group, University of Manchester, Oxford Road, Manchester M13 9PT, UK

3 Cambridge Institute for Medical Research, University of Cambridge, 4.17 Wellcome Trust/MRC Building, Addenbrooke’s Hospital, Hills Road, Cambridge CB2 0XY, UK

4 Biomedical Informatics Research Center Antwerp (Biomina), University of Antwerp, Middelheimlaan 1, 2020 Antwerpen, Belgium

5 Department of Radiotherapy, University Radiotherapy Antwerp (URA), Lindendreef 1, 2020 Antwerp, Belgium

6 Department of Oncology, Antwerp University Hospital, Wilrijkstraat 10, 2650 Edegem, Belgium

7 Department of Pathology, Antwerp University Hospital, Wilrijkstraat 10, 2650 Edegem, Belgium

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BMC Cancer 2014, 14:594  doi:10.1186/1471-2407-14-594

Published: 16 August 2014

Abstract

Background

Regions within solid tumours often experience oxygen deprivation, which is associated with resistance to chemotherapy and irradiation. The aim of this study was to evaluate the radiosensitising effect of gemcitabine and its main metabolite dFdU under normoxia versus hypoxia and to determine whether hypoxia-inducible factor 1 (HIF-1) is involved in the radiosensitising mechanism.

Methods

Stable expression of dominant negative HIF-1α (dnHIF) in MDA-MB-231 breast cancer cells, that ablated endogenous HIF-1 transcriptional activity, was validated by western blot and functionality was assessed by HIF-1α activity assay. Cells were exposed to varying oxygen environments and treated with gemcitabine or dFdU for 24 h, followed by irradiation. Clonogenicity was then assessed. Using radiosensitising conditions, cells were collected for cell cycle analysis.

Results

HIF-1 activity was significantly inhibited in cells stably expressing dnHIF. A clear radiosensitising effect under normoxia and hypoxia was observed for both gemcitabine and dFdU. No significant difference in radiobiological parameters between HIF-1 proficient and HIF-1 deficient MDA-MB-231 cells was demonstrated.

Conclusions

For the first time, radiosensitisation by dFdU, the main metabolite of gemcitabine, was demonstrated under low oxygen conditions. No major role for functional HIF-1 protein in radiosensitisation by gemcitabine or dFdU could be shown.

Keywords:
Hypoxia; Radiosensitisation; Gemcitabine; dFdU; HIF-1