ESTRO 2023 - Abstract Book

S591

Monday 15 May 2023

ESTRO 2023

Glioblastoma (GBM) is the most common primary brain tumor with almost 100% recurrence rate after surgery and radiochemotherapy. Previously, Liu laboratory at DKFZ found that glycerol-3-phosphate dehydrogenase 1 (GPD1) was expressed specifically in GBM stem cells (GSCs), and that GPD1 expression drove tumor relapse after chemotherapy. So far the role of GPD1 expression in GBM receiving radiation therapy remains unclear. We therefore investigated the role of GPD1 expression in GBM receiving radiation therapy. Materials and Methods We built GPD1 overexpression model and GPD1 Crispr/cas 9 knockout (KO) model in vitro. Furthermore, we induced organoids with human induced pluripotent stem cells (iPSC), and accordingly conducted PTEN & P53 KO organoids (C1) and PTEN & P53 & GPD1 KO organoids (G1). The Cancer Genome Atlas (TCGA) database was queried for the impact of GPD1 gene expression on survival of GBM patients. R 4.0.3 was used to determine the optimal cutoff value of GPD1. Results 3D-clonogenic survival assay was conducted with GPD1 overexpression NCH601 cells, and showed that they became more resistant to radiation after GPD1 overexpression (P = 0.049). Meanwhile, 3D-clonogenic survival assay was also conducted with GPD1 KO NCH601 cells, and showed that they became more sensitive to radiation after KD GPD1 (P = 0.007). In addition, we co-stained GPD1 and SOX-2 (a common biomarker for GSCs) in C1 organoids with immunofluorescence (IF) staining, and found that almost all GPD1 cells were SOX-2 positive, and GPD1 cells located close to the hypoxic core. Meanwhile, we stained rH2AX in C1 and G1 organoids 1h and 24h after radiation, and determined that G1 organoids had much more rH2AX foci number per nuclei than C1 organoids 1h after radiation (11 vs. 6), and recovered worse 24h after radiation (P < 0.05). Using the TCGA database, we found that GPD1 was highly expressed in GBM tissues, compared with normal brain tissues. For 501 GBM patients both clinical and GPD1 gene expression data were available in the TCGA database. 275 of these patients underwent radiation therapy with or without chemotherapy. The cutoff value of GPD1 expression was 4.449 log2(affy RMA) (range, 3.973-8.612), 183 tumors showed high GPD1 expression and 92 low expression. Patients with high GPD1 expressing GMB had significantly inferior OS than those with low GPD1 expression (P = 0.012), with median survival times of 15.9 months vs. 20.9 months. Conclusion GPD1 expression in GSCs plays a (significant) role in radiation resistance of GBM in preclinical models in vitro and is associated with impaired overall survival of patients receiving radiation therapy with or without chemotherapy. Further preclinical research is currently ongoing to determine the mechanisms by which GPD1 expression confers radiation resistance. Reference: Patricia Rusu, et al. GPD1 Specifically Marks Dormant Glioma Stem Cells with a Distinct Metabolic Profile. Cell Stem Cell. 2019;25(2):241-257. MO-0723 Previous Irradiation Impairs the DNA DSB Repair System in Sheep Lung-Derived Epithelial Cells. L. Bodgi 1,2 , J. Al-Choboq 1 , J. Azzi 1 , J. Bou Gharios 1 , C. Feghaly 1 , R. Challita 1 , H. Bou Hadir 1 , F. Abi Antoun 1 , L. Dib 1 , M. Jammal 1 , F. Olleik 1 , T. Araji 3 , P. Taddei 4 , F. Geara 1 , L. Dosch 5 , P. Sfeir 6 , A. Jurjus 2 , W. Abou Kheir 2 , B. Youssef 1 1 American University of Beirut Medical Center, Radiation Oncology, Beirut, Lebanon; 2 American University of Beirut, Anatomy, Cell Biology and Physiological Sciences, Beirut, Lebanon; 3 American University of Beirut, Anatomy, Cell biology and Physiological Sciences, Beirut, Lebanon; 4 Mayo Clinic, Radiation Oncology, Rochester, USA; 5 American University of Beirut Medical Center, Faculty of Medicine, Beirut, Lebanon; 6 American University of Beirut Medical Center, Surgery, Beirut, Lebanon Purpose or Objective Childhood cancer survivors who receive early radiotherapy (RT) treatments face the likelihood of long-term effects from their treatment that can be debilitating, chronic, and even fatal. One of these late effects that bear the greatest concern is the development of a second radiogenic cancer. In fact, irradiating normal tissues can cause mutations that might affect the DNA damage repair pathways, proto-oncogenes, and tumor suppressor genes, which enhances the risk of secondary cancer. In order to mimic pediatric craniospinal irradiation, a sheep regiment (5 treated, 4 controls) was treated with radiation administered to the thecal sac. However, incidental irradiation was also delivered to nearby tissues. The purpose of this project is to study the effect of previous RT on the DNA double-strand breaks repair kinetics. Three years after receiving the treatment, biopsies from lung tissues exposed to low and high doses of X-rays were tested and compared for treated and untreated sheep. Materials and Methods A total of 9 sheep, 5 treated and 4 control, were included in this study. The treated sheep received a total dose of 28 Gy, administered in 8 sessions, targeting their spinal thecal sac. Treatment areas were divided into two regions, depending on the received dose: high dose (HD, dose> 20 Gy) and low dose (LD, dose <2 Gy). Three years after their treatment, lung biopsies were taken from the HD, LD areas and from the control untreated sheep. Lung epithelial cells were derived, cultured and amplified. This study included 5 control, 4 LD, and 4 HD primary lung epithelial cells. After amplification, cells were irradiated with a 2 Gy dose, and the kinetics of DNA DSB repair was assessed through anti γ H2AX and anti-pATM immunofluorescence. The number of foci was assessed without re-irradiation, and 10 min, 1h, 4hrs, and 24hrs after irradiation. A non-parametric Kruskal-Wallis test was performed to compare the values between the control, LD, and HD cell lines. Results Without any irradiation, both LD (1.8 ±0.5 foci, p<0.05) and HD (1.55±36 foci, p<0.01) cells had a significantly higher number of spontaneous γ H2AX foci when compared with the control (0.7±0.1 foci). 10 mins after a 2 Gy re-irradiation, only LD cells had a significantly lower number of foci (49 ± 1.6 foci) than the control (60.7±1.8 foci). Interestingly, the number of residual foci 24 hrs after irradiation was higher for both LD (5.7±0.9 foci, p<0.01) and HD (7.25±1 foci, p<0.01) when compared with