Pharmacological genome demethylation increases radiosensitivity of head and neck squamous carcinoma cells

Abstract

Abstract. Aberrant inactivation of tumor suppressor genes by promoter hypermethylation has been recognized as a crucial step of tumor development and is related to aggressiveness and therapy resistance. To identify potential novel treatment strategies, we evaluated pharmacological genome demethylation for the increase of irradiation treatment effectiveness in head and neck squamous cell carcinoma (HNSCC) in this in vitro study. HNSCC cells were cultured with 2 different concentrations of 5-azacytidine (5-Aza) for 72 h, followed by a single fraction irradiation with 4 or 50 Gy, respectively. To show successful genome demethylation, the methylation status of the tumor suppressor gene hic1 (hypermethylated in cancer) promoter was analyzed by methylation specific PCR (MSP) as well as hic1 transcription by quantitative RT-PCR. Survival, apoptosis, viability, and migration of the tumor cells were analyzed as functional parameters of combined treatment response. After 5-Aza treatment the hic1 promoter was demethylated and gene transcription restored demonstrating genome demethylation. 5-Aza treated cells tended to be less viable and showed decreased survival indicated by lower colony numbers. Apoptosis and migration were not affected. The combined application of irradiation and 5-Aza significantly reduced survival compared to the single treatments. Accordingly, apoptosis was strongly increased after combined 4 Gy/5-Aza treatment. Viability was not additionally affected by combined treatment. Migration was affected weakly by combined high dosage irradiation/5-Aza treatment. Our data show that the combined application of 5-Aza and irradiation is effective in vitro. A demethylating concept prior to irradiation should be further evaluated for its potential to reduce irradiation resistance.

DNA methylome analysis identifies epigenetic silencing of FHIT as a determining factor for radiosensitivity in oral cancer: an outcome-predicting and treatment-implicating study

“…Remarkably, epigenetic interventions using DNMT inhibitors significantly enhanced the radiosensitivity of resistant oral cancer cells, in vivo. Our results together with previous observations [30-31] suggest that concomitant treatment of DNA-demethylating agents and radiotherapy is a new treatment direction for this aggressive malignancy.…”

“…For examples, decitabine (5-Aza) has been reported to enhance radiosensitivity through activating G2/M checkpoint responses, inducing apoptosis in osteosarcoma [61], breast cancer [62], colorectal cancer [63], medulloblastoma [64], and head-and-neck cancers [30-31]. Our results may provide an explanation for the phenomenon that inhibitors of DNMT or EZH2 reverse repressive epigenomes to resensitize radioresistant cancer cells to ionizing radiation.…”

Radiation-induced epigenetic DNA methylation modification of radiation-response pathways

“…Epigenetic therapies targeting DNA methylation are under study and have been shown to sensitize cells to radiation. 4,[15][16][17] Treatment of cells with the DNA methyltransferase inhibitors psammaplin A or zebularine partially abrogated the G 2 phase arrest, increased apoptosis, and reduced the ability to repair DNA double strand breaks in cells exposed to 6 Gy radiation. 4 The DNA damage response is complex and involves changes to the cell cycle, activation of DNA repair pathways, and induction of apoptosis or, alternatively, senescence.…”

Effects of ionizing radiation on DNA methylation: from experimental biology to clinical applications

“…The results from recent in vitro clonogenic studies suggest that DNMT inhibitors may also modulate the cancer cell response to IR. For instance, nucleoside analogs increased the sensitivity to radiotherapy in glioblastoma (Dote et al, 2005), gastric (Qiu et al, 2009), colorectal (Hofstetter et al, 2010), head and neck (Brieger et al, 2012), and nasopharyngeal (Jiang et al, 2014) cancer cell lines. At the same time, the effect of DNMT inhibitors-induced radiosensitization was not uniform, and in some cell lines it was little to negligible (Qiu et al, 2009; Kim et al, 2012).…”