ESTRO 38 Abstract book

S593 ESTRO 38

Purpose or Objective Glioblastomas (GBM) are highly radio- and chemoresistant tumors associated with poor outcome. Cyclin D2 (CCND2) is known to play a critical role in cell cycle progression, tumorigenicity and radioresistance in GBM, particularly in the subpopulation of glioma stem cells. The aim of our study is to better understand the evolution of CCND2 expression during treatments and to investigate its clinical significance in GBM. Material and Methods Immunohistochemical expression analysis of CCND2 was carried out on paired GBM samples of 72 patients, comparing initial resections with recurrent tumors after radiation therapy alone (RT) (n=36) or radio- chemotherapy with temozolomide according to the Stupp regimen (RT-TMZ) (n=36). Semi-quantitative analysis of CCND2 nuclear staining was performed by two independent observers. Every tumor was scored according to the number of stained cells (low nuclear expression < 30% and high nuclear expression ≥ 30%). (Figure 1) Multivariate logistic regression analysis was conducted to investigate clinical and demographic risk factors of early mortality in GBM patients.

Purpose or Objective Human induced pluripotent stem cells (hiPSCs) constitute a real breakthrough in regenerative medicine. We differentiated hiPSCs towards chondrocytes (hiPSC- derived chondrocytes, hiPSC-DCHs) as a real hope for patients diagnosed with osteoarthritis. However, it is still a research area that must be more precisely investigated to ensure safety of patients. Thus, we investigate the DDR mechanisms of these cells exposed to IR. Previously, we demonstrated results concerning γH2AX, DNA repair, apoptosis and senescence. In this study, we aim to show the analyses of cell cycle, ROS level and changes in mitochondrial membrane potential of irradiated cells. Material and Methods We differentiated of hiPSCs into chondrocyte-like cells. The hiPSC-derived chondrocytes were treated with IR (0; 1; 2 and 5 [Gy]). Then, we performed analysis of cell cycle using propidium iodide (9h after IR as a most sensitive time point that we previously selected). Immediately after IR, we investigated level of generated ROS taking advantage of CellROX Green Flow Cytometry Assay Kit (TBHP served as a positive control). Finally, the analysis of mitochondrial membrane potential was based on JC-1 staining (CCCP served as a positive control). Results The irradiated hiPSCs demonstrated accumulation of cells in S phase. On the contrary, both mature chondrocytes and hiPSC-DCHs revealed arrest of cell cycle in G2 phase. Furthermore, chondrocyte-like cells obtained from hiPSCs revealed characterization of cell cycle similar to fully differentiated cells (hiPSC-DCHs vs chondrocytes). In that case the majority of cells were noticeable in G1 phase. In contrast, non-irradiated hiPSCs as highly proliferative type of cells demonstrate the highest percentage of S phase. The IR did not cause a dramatic differences in the ROS level in all investigated types of cells (hiPSCs, hiPSC-DCHs and mature chondrocytes). However, there was a significant dissimilarity between overall profile of hiPSCs and differentiated cells (both hiPSC-DCHs and chondrocytes). The non- and irradiated hiPSCs showed the relatively high level of ROS- that was shifted to the positive control- in contrast to differentiated cells. In turn, the most notable changes of mitochondrial membrane potential was observed as hiPSC-DCHs (above 30% of stained cells) revealing - in this aspect - their high radiosensitivity. Conclusion We provided a new knowledge concerning hiPSC-derived cells. We can observe that radioresistance of hiPSC-DCHs significantly differ from both “parental” hiPSC and mature chondrocytes generating a unique, individual and specific DDR of these cells. It may have an influence on their genetic stability. In further part of research we plan to focus on testing hiPSC-DCHs in in vivo model. Funding: National Science Centre (2012/E/NZ3/01819) and (2016/23/N/NZ7/01892) PO-1067 High expression of CCND2 in glioblastoma is associated with an increased risk of early mortality. C. Bouchart 1 , A. Trépant 2 , M. Hein 3 , D. Van Gestel 1 , P. Demetter 4 1 Institut Jules Bordet - Université Libre de Bruxelles, Department of Radiation-Oncology, Brussels, Belgium ; 2 Erasme University Hospital - Université Libre de Bruxelles, Department of Pathology and DIAPATH- Center of Microscopy and Molecular Imaging CMMI, Brussels, Belgium ; 3 Erasme University Hospital - Université Libre de Bruxelles, Department of Psychiatry and Sleep Laboratory, Brussels, Belgium ; 4 Institut Jules Bordet - Université Libre de Bruxelles, Department of Pathology, Brussels, Belgium

Results Compared to the primary tumor, CCND2 expression was decreased in GBM recurrences after treatment with RT (32.00 ± 19.00 vs 20.00 ± 19.00%, p = 0.013) or RT-TMZ (31.00 ± 20.00 vs 19.00 ± 18.00%, p = 0.001). The decrease in CCND2 expression did not differ significantly according to the treatment used (p = 0.731). Multivariate logistic regression analysis revealed that significant risk factors predicting early mortality (<12 months) in GBM patients are: time to recurrence ≤6 months [OR 23.88 (CI 95% 4.36 to 130.68)], subtotal surgery at recurrence [OR 10.58 (CI 95% 1.81 to 62.01)] and high nuclear expression of CCND2 (≥ 30%) at initial surgery [OR 7.97 (CI 95% 1.35 to 46.98)]. The following variables were not included in the model because they were not significant: gender, age, preoperative use of corticosteroids, type of initial surgery,