Downregulation of both mismatch repair and non-homologous end-joining pathways in hypoxic brain tumour cell lines.

Sophie Cowman,Violaine Sée,Barry Pizer

doi:10.7717/peerj.11275

Sophie Cowman, Violaine Sée + Show 1 more

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https://doi.org/10.7717/peerj.11275

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Abstract

Glioblastoma, a grade IV astrocytoma, has a poor survival rate in part due to ineffective treatment options available. These tumours are heterogeneous with areas of low oxygen levels, termed hypoxic regions. Many intra-cellular signalling pathways, including DNA repair, can be altered by hypoxia. Since DNA damage induction and subsequent activation of DNA repair mechanisms is the cornerstone of glioblastoma treatment, alterations to DNA repair mechanisms could have a direct influence on treatment success. Our aim was to elucidate the impact of chronic hypoxia on DNA repair gene expression in a range of glioblastoma cell lines. We adopted a NanoString transcriptomic approach to examine the expression of 180 DNA repair-related genes in four classical glioblastoma cell lines (U87-MG, U251-MG, D566-MG, T98G) exposed to 5 days of normoxia (21% O2), moderate (1% O2) or severe (0.1% O2) hypoxia. We observed altered gene expression in several DNA repair pathways including homologous recombination repair, non-homologous end-joining and mismatch repair, with hypoxia primarily resulting in downregulation of gene expression. The extent of gene expression changes was dependent on hypoxic severity. Some, but not all, of these downregulations were directly under the control of HIF activity. For example, the downregulation of LIG4, a key component of non-homologous end-joining, was reversed upon inhibition of the hypoxia-inducible factor (HIF). In contrast, the downregulation of the mismatch repair gene, PMS2, was not affected by HIF inhibition. This suggests that numerous molecular mechanisms lead to hypoxia-induced reprogramming of the transcriptional landscape of DNA repair. Whilst the global impact of hypoxia on DNA repair gene expression is likely to lead to genomic instability, tumorigenesis and reduced sensitivity to anti-cancer treatment, treatment re-sensitising might require additional approaches to a simple HIF inhibition.

Highlights

Glioblastoma (GBM) is a highly aggressive and infiltrative grade IV astrocytoma, the most common malignant brain tumour in adults (Dolecek et al, 2012)
We found that hypoxia affects mismatch repair (MMR), non-homologous end-joining (NHEJ) and homologous recombination repair (HRR)
HRR, NHEJ and MMR are strongly regulated by hypoxia in a range of hypoxic GBM cell lines To explore the global impact of tumour hypoxia on the expression of DNA repair genes in GBM cells, we used a NanoString assay

Summary

Introduction

Glioblastoma (GBM) is a highly aggressive and infiltrative grade IV astrocytoma, the most common malignant brain tumour in adults (Dolecek et al, 2012). Survival rates remain low partly due to the limited treatment options available. Temozolomide an alkylating agent, is the sole chemotherapeutic agent used for GBM in the U.K. Using temozolomide as an adjuvant increases mean survival rate compared to radiotherapy alone (Athanassiou et al, 2005). Despite many attempts to improve GBM outcome, survival over five years is close to zero, with numerous barriers to successful GBM treatment still remaining. One such barrier is tumour hypoxia, which can negatively influence the effectiveness of both radiotherapy and temozolomide treatment and is associated with poor patient survival for GBM and for many other solid tumours (Marampon et al, 2014; Ge et al, 2018)

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