Combination Effect of Epigenetic Regulation and Ionizing Radiation in Colorectal Cancer Cells

Abstract

Exposure of cells to ionizing radiation (IR) induces, not only, activation of multiple signaling pathways that play critical roles in cell fate determination, but also alteration of molecular pathways involved in cell death or survival. Recently, DNA methylation has been established as a critical epigenetic process involved in the regulation of gene expression in cancer cells, suggesting that DNA methylation inhibition may be an effective cancer treatment strategy. Because alterations of gene expression by DNA methylation have been considered to influence radioresponsiveness, we investigated the effect of a DNA methyltransferase inhibitor, 5-aza-2′-deoxycytidine (5-aza-dC), on radiosensitivity. In addition, we investigated the underlying cellular mechanisms of combination treatments of ionizing irradiation (IR) and 5-aza-dC in human colon cancer cells. Colon cancer cell lines were initially tested for radiation sensitivity by IR in vitro and were treated with two different doses of 5-aza-dC. Survival of these cell lines was measured using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and clonogenic assays. The effects of 5-aza-dC along with irradiation on cell growth, cell cycle distribution, apoptosis, and apoptosis-related gene expression were examined. Combination irradiation treatment with 5-aza-dC significantly decreased growth activity compared with irradiation treatment alone or with 5-aza-dC treatment alone. The percentage of HCT116 cells in the sub-G1 phase and their apoptotic rate was increased when cells were treated with irradiation in combination with 5-aza-dC compared with either treatment alone. These observations were strongly supported by increased caspase activity, increased comet tails using comet assays, and increased protein levels of apoptosis-associated molecules (caspase 3/9, cleaved PARP). Our data demonstrated that 5-aza-dC enhanced radiosensitivity in colon cancer cells, and the combination effects of 5-aza-dC with radiation showed greater cellular effects than that of single treatment, suggesting that the combination of 5-aza-dC and radiation has the potential to become a clinical strategy for the treatment of cancer.

Decoupling of DNA methylation and activity of intergenic LINE-1 promoters in colorectal cancer

“…To further investigate the relationship between methylation and expression of LCT13 we focused on HCT116 and RKO cells, established cell lines that have been widely used to study DNA methylation using inhibitors 41,42 . We treated these cells with the DNA methylation inhibitor 5-azacytidine (5-aza) and confirmed that the treatment did not affect expression of the 3 reference genes used for quantification of RT-PCR (Fig.…”

microRNA-451a regulates colorectal cancer proliferation in response to radiation

“…To identify miRs that are regulated by radiation in CRC, we implanted either HCT-116 or SW480 xenografts into nude mice. We chose these two cell lines because they have been well-characterized and grow well in vitro and in xenograft studies [ 13 , 14 ]. In addition, they provide a contrast of sorts with HCT-116 being microsatellite unstable (MSI), PIK3CA H1047R mutated and p53 WT whereas SW480 is microsatellite stable (MSS), PIK3CA WT but p53 R237H mutated.…”

Long-Lasting Remission in De Novo Breast Myeloid Sarcoma Treated with Decitabine and Radiotherapy

“…Radiation therapy is commonly employed to treat MS and in our patient it was administered bilaterally and defining a peculiar CTV which minimizes the risk of local recurrence but also the toxicity to lung and heart. Some experiences in solid tumors demonstrate an increased radiosensitivity in vitro after exposure to hypomethylating agent, mainly 5-azacytidine [9]. On the other side, in vitro models of breast cancer showed different level of methylation of DNA repair, cell cycle, and apoptosis pathway genes in response to radiation at various doses and time points [10].…”