Reactive oxygen species and p21Waf1/Cip1 are both essential for p53-mediated senescence of head and neck cancer cells.

A L Fitzgerald,V Sandulache,A A Osman,H Skinner,J N Myers,T-X Xie,A Patel

doi:10.1038/cddis.2015.44

Abstract

Treatment of head and neck squamous cell carcinoma, HNSCC, often requires multimodal therapy, including radiation therapy. The efficacy of radiotherapy in controlling locoregional recurrence, the most frequent cause of death from HNSCC, is critically important for patient survival. One potential biomarker to determine radioresistance is TP53 whose alterations are predictive of poor radiation response. DNA-damaging reactive oxygen species (ROS) are a by-product of ionizing radiation that lead to the activation of p53, transcription of p21cip1/waf1 and, in the case of wild-type TP53 HNSCC cells, cause senescence. The expression of p21 and production of ROS have been associated with the induction of cellular senescence, but the intricate relationship between p21 and ROS and how they work together to induce senescence remains elusive. For the first time, we show that persistent exposure to low levels of the ROS, hydrogen peroxide, leads to the long-term expression of p21 in HNSCC cells with a partially functional TP53, resulting in senescence. We conclude that the level of ROS is crucial in initiating p53's transcription of p21 leading to senescence. It is p21's ability to sustain elevated levels of ROS, in turn, that allows for a long-term oxidative stress, and ensures an active p53–p21–ROS signaling loop. Our data offer a rationale to consider the use of either ROS inducing agents or therapies that increase p21 expression in combination with radiation as approaches in cancer therapy and emphasizes the importance of considering TP53 status when selecting a patient's treatment options.

Highlights

TP53 missense mutations used for this study, we have developed the Evolutionary Action scoring system.[16]
Being that senescence is the primary mechanism of radiation-induced cell death in Head and neck squamous cell carcinoma (HNSCC) cells,[30] a senescence-associated β-galactosidase (SA-β-gal) staining was performed and showed that 4 days after treatment with 4-Gy ionizing radiation, cells with wtp[53] had a significantly higher SA-βgal-positive staining (48–60%) compared with the SA-β-galpositive staining in TP53 null and all TP53 mutant cells
Given the prominent role of radiation therapy in the treatment of HNSCC, understanding the mechanisms of radiation resistance is of the utmost importance

Summary

Introduction

TP53 missense mutations used for this study, we have developed the Evolutionary Action scoring system.[16]. Expression of the p21waf1/cip[1] gene is transcriptionally regulated by the p53 protein and is an essential mediator of p53-induced cell cycle arrest.[18] p21’s role in senescence, a terminal state of differentiation in which cells are permanently and irreversibly arrested, has been established under various conditions.[19,20,21] Depending on cell type and stimuli, p21 is influential in causing cell senescence by multiple mechanisms, such as: binding to PCNA, leading to inhibition of DNA synthesis,[22] interfering with cyclin-dependent kinase complexes,[23,24] inhibiting phosphorylation of Rb and, causing the downregulation of many E2F-regulated genes, which are involved in controlling progression of the cell cycle and DNA replication.[25] A cell in a senescence state is considered to be tumor suppressive in that it can prevent transformation of cells;[26] in addition, it can be a pathway to chemotherapy- or radiotherapy-induced cell death.[27] Both p21 and ROS have been shown to be involved in senescence and the complex relationship between p21 expression and ROS production has been studied for many years, questions as to how p21 and ROS regulate one another’s expression remain. The elevated expression of p21, in turn, sustains the level of elevated ROS, causing HNSCC cells to undergo cell death via a senescence-mediated pathway

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Conclusion