Hypoxia down-regulates DNA double strand break repair gene expression in prostate cancer cells

Meng, Alice X.; Jalali, Farid; Cuddihy, Andrew; Chan, Norman; Bindra, Ranjit S.; Glazer, Peter M.; Bristow, Robert G.

doi:10.1016/j.radonc.2005.06.025

« Previous Next »Radiotherapy & Oncology
Volume 76, Issue 2 , Pages 168-176, August 2005

Hypoxia down-regulates DNA double strand break repair gene expression in prostate cancer cells

Alice X. Meng
- Ontario Cancer Institute and Princess Margaret Hospital (University Health Network), Toronto, Ont., Canada
Farid Jalali
- Ontario Cancer Institute and Princess Margaret Hospital (University Health Network), Toronto, Ont., Canada
Andrew Cuddihy
- Ontario Cancer Institute and Princess Margaret Hospital (University Health Network), Toronto, Ont., Canada
Norman Chan
- Ontario Cancer Institute and Princess Margaret Hospital (University Health Network), Toronto, Ont., Canada
Ranjit S. Bindra
- Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, CT, USA
Peter M. Glazer
- Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, CT, USA
Robert G. Bristow
- Ontario Cancer Institute and Princess Margaret Hospital (University Health Network), Toronto, Ont., Canada
- Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, CT, USA
- Corresponding author. Robert G. Bristow, Department of Radiation Oncology, Ontario Cancer Research Network, Princess Margaret Hospital, 610 University Avenue, Rm 5-923, Toronto, Ont., Canada M5G2M9.

Received 10 May 2005; received in revised form 13 May 2005; accepted 19 June 2005. published online 18 July 2005.

Abstract

Background and purpose

Intratumoral hypoxia has been correlated with poor clinical outcome in prostate cancer. Prostate cancer cells can be genetically unstable and have altered DNA repair. We, therefore, hypothesized that the expression of DNA double-strand break (DNA-dsb) repair genes in normal and malignant prostate cultures can be altered under hypoxic conditions.

Methods and materials

The expression of homologous recombination (HR) and non-homologous recombination (NHEJ) genes following gas hypoxia (0.2%) or exposure to HIF1α-inducing agent, CoCl₂ (100μM), was determined for normal diploid fibroblasts (GM05757) and the pre-malignant and malignant prostate cell lines, BPH-1, 22RV-1, DU145 and PC3. RNA and protein levels were determined using RT-PCR and Western blotting. Additionally, p53 genotype and function, the level of hypoxia-induced apoptosis, and cell cycle distribution, were determined to correlate to changes in DNA-dsb gene expression.

Results

Induction of hypoxia was confirmed using HIF1α and VEGF expression in gas- and CoCl₂-treated cultures. Hypoxia (48–72h of 0.2% O₂) decreased RNA expression of a number of HR-related genes (e.g. Rad51, Rad52, Rad54, BRCA1, BRCA2) in both normal and malignant cultures. Similar decreases in RNA pertaining to the NHEJ-related genes (e.g. Ku70, DNA-PKcs, DNA Ligase IV, Xrcc4) were observed. In selected cases, hypoxia-mediated decreases in RNA expression led to decreased DNA-dsb protein expression. CoCl₂-treated cultures did not show decreased DNA-dsb protein expression. The ability of hypoxia to down-regulate Rad51 and other HR-associated genes under hypoxia was not correlated to c-Abl or c-Myc gene expression, p53 genotype or function, propensity for hypoxia-mediated apoptosis, or specific changes in cell cycle distribution.

Conclusions

Hypoxia can down-regulate expression of DNA-dsb repair genes in both normal and cancer cells. If associated with a functional decrease in DNA-dsb repair, this observation could provide a potential basis for the observed genetic instability within tumor cells exposed to hypoxia.

Keywords: Hypoxia, DNA repair, Rad51, Prostate cancer, Genetic instability