ESTRO 36 Abstract Book

S538 ESTRO 36 2017 _______________________________________________________________________________________________

rhabdospheres

derived

from

the

embryonal

effectively induced ubiquitination and degradation of Nrf2, and inhibited translocation of Nrf2 to the nucleus. Consequently, expression of Nrf2 downstream genes was reduced, and the Nrf2-dependent antioxidant system was suppressed. Endogenous ROS was higher than before ISL treatment, causing redox imbalance and oxidative stress in HepG2 cells. Moreover, pretreatment with ISL for 6 h followed by X-ray irradiation significantly increased γ- H2AX foci and cell apoptosis, and reduced clonogenic potential compared with cells irradiated with X-rays alone. In addition, HepG2 xenografts, ISL, and X-ray cotreatments induced greater apoptosis and tumor growth inhibition, when compared with X-ray treatments alone. Additionally, HepG2 xenografts, in which Nrf2 was expressed at very low levels due to ectopic expression of Keap1, showed that ISL-mediated radiosensitization was Keap1 dependent. Conclusion ISL inhibited the Nrf2-antioxidant pathway by increasing the levels of Keap1 and ultimately inducing oxidative stress via disturbance of the redox status. The antioxidant ISL possessed pro-oxidative properties, and enhanced the radiosensitivity of liver cancer cells, both in vivo and in vitro . Taken together, these results demonstrated the effectiveness of using ISL to decrease radioresistance, suggesting that ISL could be developed as an adjuvant radiosensitization drug. Disturbance of redox status could be a potential target for radiosensitization. PO-0982 Fused Toes Homolog (FTS) is a potential target for Notch-mediated radioresistance in cervical cancer W.Y. Park 1 , P.D. Subramania 1 , J.R. Yu 2 1 Chungbuk National University Hospital, Dept of Radiation Oncology, Cheongju, Korea Republic of 2 Konkuk University, Department of Environmental and Tropical Medicine, Chungju, Korea Republic of Purpose or Objective Radiation therapy is one of the major treatment modalities for cervical cancer. Increasing evidences suggest that cancer stem cells (CSC) in tumours contribute to radioresistance and recurrence. Notch pathway plays a vital role in maintenance of cancer stemness and its activation leads to disease progression and metastasis. FTS gene was initially identified as one of six genes deleted in a mouse mutant called Fused Toes, due to defects in limb development, and referred as FT1/FTS. However, the function of FTS has not been elucidated well in human. We previously reported that FTS plays an essential role in nuclear phosphorylation of EGFR and repair of DNA damage, and epithelial-mesenchymal transition. In this study, we evaluated the role of FTS in Notch signaling and CSCs. Material and Methods A human cervical cancer cell line (ME180) was used. Silencing of FTS was obtained using siRNA. Western blot and immunofluorescence was done to analyze the expression and localization of the proteins. Results Protein expression of Notch 1, cleaved Notch1, Notch 3, γ-secretase complex and its downstream Hes-1was increased by ionizing radiation and it was reduced by FTS- silencing. Spheroid formation ability and cancer stem cell markers Nanog, Oct-4A, Sox2 were reduced by FTS- silencing. Cell survival was decreased by FTS-silencing. Conclusion FTS is involved in the regulation of Notch signaling and CSC maintenance. FTS can be a target to overcome Notch– mediated radioresistance in cervical cancer. PO-0983 Antrodia cinnamomea Regulates DNA Repair and Enhances Radiosensitivity of Esophageal Cancer Cells

rhabdomyosarcoma cell lines. Material and Methods

Rhabdospheres enriched in cancer stem like cells were obtained growing ERMS cells in non adherent condition in stem cell medium. Stem cell markers were evaluated by FACS analysis and immunoblotting. ERK1/2, myogenic markers, proteins of DNA repair and bone marrow X-linked kinase (BMX) expression were evaluated by immunoblotting analysis. Radiation was delivered using an x-6 MV photon linear accelerator. Xenografts were obtained in NOD/SCID mice by subcutaneously injection of rhabdosphere cells or cells pretreated with U0126 in stem cell medium. Results MEK/ERK inhibitor U0126 dramatically prevented rhabdosphere formation and down-regulated stem cell markers CD133, CXCR4 and Nanog expression, but enhanced ALDH, MAPK phospho-active p38 and differentiative myogenic markers. By contrast, MAPK p38 inhibition accelerated rhabdosphere formation and enhanced phospho-active ERK1/2 and Nanog expression. ERMS cells, chronically treated with U0126 and then xeno- transplanted in NOD/SCID mice, delayed tumor development and reduced tumor mass when compared with tumor induced by rhabdosphere cells. U0126 intraperitoneal administration to mice bearing rhabdosphere-derived tumors inhibited tumor growth . The MEK/ERK pathway role in rhabdosphere radiosensitivity was investigated in vitro. Disassembly of rhabdospheres was induced by both radiation or U0126, and further enhanced by combined treatment. In U0126- treated rhabdospheres, the expression of the stem cell markers CD133 and CXCR4 decreased and dropped even more markedly following combined treatment. The expression of BMX, a negative regulator of apoptosis, also decreased following combined treatment, which suggests an increase in radiosensitivity of rhabdosphere cells. Conclusion Our results indicate that the MEK/ERK pathway plays a prominent role in maintaining the stem-like phenotype of ERMS cells, their survival and their innate radioresistance. Thus, therapeutic strategies that target cancer stem cells, which are resistant to traditional cancer therapies, may benefit from MEK/ERK inhibition combined with traditional radiotherapy, thereby providing a promising therapy for embryonal rhabdomyosarcoma. PO-0981 Disturbance of redox status enhances radiosensitivity of hepatocellular carcinoma H. Zhang 1 , C. Sun 1 1 Institute of Modern Physics- Chinese Academy of Sciences, Department of Heavy Ion Radiation Biology and Medicine, Lanzhou, China Purpose or Objective High constitutive expression of Nrf2 has been found in many types of cancers, and this high level of Nrf2 also favors resistance to drugs and radiation. Here we investigate how isoliquiritigenin (ISL), a natural antioxidant, inhibits the Nrf2-dependent antioxidant pathway and enhances the radiosensitivity of HepG2 cells Treatment of HepG2 cells with ISL for 6 h, Keap1 and ubiquitination of Nrf2 were measured by RT-PCR and Western blot. Pretreatment with ISL for 6 h followed by X- ray irradiation, confocal microscopy was used to visualize Nrf2 translocation to the nucleus and γ-H2AX foci. To investigate the radiosensitization effect of ISL, apoptosis, clonogenic potential and HepG2 xenografts were examined. Results Treatment of HepG2 cells with ISL for 6 h selectively enhanced transcription and expression of Keap1. Keap1 and HepG2 xenografts. Material and Methods