Inhibition of DNA-PKcs enhances radiosensitivity and increases the levels of ATM and ATR in NSCLC cells exposed to carbon ion irradiation

Abstract

Non-small cell lung cancer (NSCLC) exhibits radioresistance to conventional rays, due to its DNA damage repair systems. NSCLC may potentially be sensitized to radiation treatment by reducing those factors that continuously enhance the repair of damaged DNA. In the present study, normal lung fibroblast MRC-5 and lung cancer A549 cells were treated with NU7026 and CGK733, which are inhibitors of the DNA-dependent protein kinase catalytic subunit (PKcs) and ataxia telangiectasia mutated (ATM) and ataxia telangiectasia and Rad3-related (ATR), respectively, followed by exposure to X-rays and carbon ion irradiation. The cytotoxic activity, cell survival rate, DNA damage repair ability, cell cycle arrest and apoptosis rate of the treated cells were analyzed with MTT assay, colony formation assay, immunofluorescence and flow cytometry, respectively. The transcription and translation levels of the ATM, ATR and DNA-PKcs genes were detected by reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. The results indicated that the radiosensitivity and DNA repair ability of A549 cells were reduced, and the percentages of apoptotic cells and those arrested at the G2/M phase of the cell cycle were significantly increased, following ionizing radiation with inhibitor-pretreatment. The expression levels of ATM, ATR, DNA-PKcs and phosphorylated histone H2AX, a biomarker for DNA double-strand breaks, were all upregulated at the transcriptional or translational level in A549 cells treated with carbon ion irradiation, compared with the control and X-rays-treated cells. In addition, the treatment with 5–50 µM NU7026 or CGK733 did not produce any obvious cytotoxicity in MRC-5 cells, and the effect of the DNA-PKcs-inhibitor on enhancing the radiosensitivity of A549 cells was stronger than that observed for the ATM and ATR-inhibitor. These findings demonstrated a minor role for ATM and ATR in radiation-induced cell death, since the upregulation of ATM and ATR did not rescue the A549 cells subjected to ionizing irradiation. Therefore, future studies on DNA-PKcs, ATM and ATR may lead to novel specific therapies that supplement general radiotherapy for the treatment of lung cancer.

Radioresistance of chordoma cells is associated with the ATM/ATR pathway, in which RAD51 serves as an important downstream effector

“…Several lines of evidence have demonstrated that DNA repair mechanisms are employed by cancer cells to resist ionizing radiation, resulting in cancer recurrence and final radiotherapy failure ( 4 – 6 ). It has been reported that the protein kinases, ataxia telangiectasia mutated (ATM) and ATM and Rad3 related (ATR), are major regulators in the DNA damage response to radiation ( 4 , 5 ).…”

<p>NVP-BSK805, an Inhibitor of JAK2 Kinase, Significantly Enhances the Radiosensitivity of Esophageal Squamous Cell Carcinoma in vitro and in vivo</p>

“…[14][15][16] Several kinase inhibitors such as wee1 kinase inhibitor AZD1775, inhibitor of DNA-PKcs kinase and ATR inhibitor were proven to enhance the radiosensitivity of human cancers. [17][18][19] In our study, by screening of 93 kinase inhibitors, we found NVP-BSK805, a specific inhibitor of JAK2 kinase, 20 significantly improved the radiosensitivity of esophageal cancer cells in vitro and in vivo. As found in other models, 21 JAK2 inhibition by NVP-BSK805 efficiently blocked the phosphorylation of STAT5.…”

Novel recombinant protein FlaA N/C increases tumor radiosensitivity via NF-κB signaling in murine breast cancer cells

“…A large number of studies have shown that apoptosis is important in tumor radiotherapy ( 30 , 31 ). Therefore, the present study further tested whether FlaA N/C affects cell apoptosis after radiation.…”