Abstract Book

S77

ESTRO 37

J. Gordito Soler 4 , J. Praena Fernández 5 , M. Ortiz Gordillo 4 , J. López Guerra 4 1 Institute of Biomedicine of Seville, Radiation Oncology, Sevilla, Spain 2 University Hospital of Cruces, Radiation Oncology, Bilbao, Spain 3 University Hospital Virgen del Rocío, Medical Oncology, Seville, Spain 4 University Hospital Virgen del Rocío, Radiation Oncology, Seville, Spain 5 University Hospital Virgen del Rocío, Methodology Unit, Seville, Spain Purpose/Objective Retrospective unicenter studies have shown that HSPB1 rs2868371 single nucleotide polymorphism (SNP) is associated with survival in non-small cell lung cancer (NSCLC) patients related to a very different response to radiation therapy (RT). Therefore, the aim of the current study is to validate this finding in a prospective multicenter (2 institutions) setting. Material/methods: DNA samples from 181 NSCLC patients have been collected after consultation at the Radiation Oncology Department from May 2012 to August 2016 in order to perform a genotyping analysis seeking associations between survival and HSPB1 rs2868371 SNP . From these patients, 155 (86%) received concurrent radio(chemo)therapy (platinum based), and 27 patients (15%) underwent surgery. All the patients were caucasians, 163 males (90%) and 18 females (10%), the median age was 66 years old. Median follow-up time for all patients was 13 months (range, 0.1-51 months). According to the clinical stage, 10 patients (5.5%) were stage I-II, 82 (45.4%) were stage IIIA, 64 (35.3%) were stage IIIB and 25 (13.8%) were stage IV. The histologies were 33.7% (N=61) adenocarcinoma, 57% (N=103) squamous cell carcinoma, 3.3% (N=6) large cell carcinoma, and 6% (N=11) not specified. Genotype analysis was performed from DNA isolated from blood samples, by real-time polymerase chain reaction (PCR) using TaqMan® SNP Genotyping Assays. Additionally, genotypes were verified by direct sequencing in paired control/tumor paraffin tissue sections in 27 patients. For statistical analysis, Kaplan-Meier cumulative probability and Cox proportional hazards analysis were performed in order to evaluate the effect of different genotypes on survival. Results Multivariate analysis showed that early stage (HR: 0.41; P=0.049), surgery (HR: 0.33; P=0.003), and concurrent radio(chemo)therapy (HR:0.51;P=0.009) were significantly associated with better overall survival (OS). The HSPB1 rs2868371 CC genotype was associated with better survival compared with the rs2868371 GG/GC genotypes (HR=0.349; 95% CI=0.127-0.964; P=0.042). Two years OS for 72% of patients carrying the rs2868371 CC genotype and 36% for patients with rs2868371 GG/GC genotypes (log-rank test: P=0.013). In addition, the rs2868371 CC genotype was also found to be associated with better disease-free survival (HR: 0.357; 95% CI: 0.128-0.991; P=0.048). Conclusion Our findings validate a strong relationship between the functional HSPB1 promoter variant rs2868371 and OS among NSCLC patients. High expression of HSPB1 can mediate cancer progression through inhibition of the programmed cell death and senescence in lung cancer. A plausible explanation for a better OS in patients carrying the C allele may rely on the presence of a functional binding site in this region where the C allele prompt to a lower transcription factor-DNA binding affinity responsible of a cascade of downstream events able to alter endogenous HSPB1 expression levels.

Helmholtz-Zentrum Dresden-Rossendorf- Institute of Radiooncology – OncoRay- Dresden- German, 12 OncoRay - National Center for Radiation Research in Oncology- Faculty of Medicine and University Hospital Carl Gustav Carus- Technische Universität Dresden and Helmholtz-Zentrum Dresden-Rossendorf- Institute of Radiooncology – OncoRay- Dresden- German, Purpose or Objective c-MYC is a broad transcription regulator, which is upregulated in prostate cancer stem cells (CSC) and activates expression of genes regulating glutaminolysis. Glutaminolysis is important for biosynthesis and energy production, and also contributes to the ROS scavenging and activation of the pro-survival signaling pathways. Tumors with enhanced c-MYC expression, such as prostate cancers have a particularly high demand for glutamine. Herein, we investigated the role of glutamine metabolism pathways for prostate cancer (PCa) radioresistance. Material and Methods PCa cell lines and their radioresistant (RR) sublines were analyzed by metabolomics and gene expression profiling. The relative cell sensitivity to the inhibition of glutaminolysis was measured by analysis of cell viability (MTT), apoptosis and necrosis (flow cytometry and Western blotting), levels of ROS and glutathione (flow cytometry), radiosensitivity (colony formation assay), DNA repair (γH2A.X foci) and by in vivo tumor growth in NMRI (nu/nu) mice. Primary cell cultures from 12 tumor biopsies and matched benign tissues from PCa patients were characterized by radiobiological 3D CFA and by gene expression profiling, and relative radioresistance was correlated with expression levels of the genes regulating glutaminolysis. Analysis of the Cancer Genome Atlas (TCGA) datasets was performed using Statistical Utilities for Microarray and Omics data (SUMO) software package. MYC gene expression in PCa patients treated with radiotherapy (RT) was analyzed by nanoString technology using the formalin-fixed paraffin-embedded (FFPE) material. Results Glutaminolysis is upregulated in RR cells, where glutamine is mostly used for production of α- ketoglutarate which is involved in the regulation of ROS scavenging, epigenetic resetting and CSC maintenance. Deprivation of glutamine or siRNA-mediated inhibition of glutaminolysis leads to the induction of endoplasmic reticulum (ER) stress and cell death, and to the inhibition of the mTOR signaling pathway, DNA repair, clonogenicity, spherogenicity and in vivo tumorigenicity after irradiation with a more pronounced effect for RR cells. A high expression of the genes regulating PCa glutaminolysis correlates with clonogenic survival of primary cell cultures after irradiation and is significantly associated with a decrease in relapse free survival after RT based on the analysis of TCGA datasets. NanoString analysis of the FFPE tissues of PCa patients (n=75) treated with RT at the Department of Radiotherapy and Radiation Oncology, University Hospital Carl Gustav Carus and Faculty of Medicine showed a significant correlation of low MYC expression to better clinical outcome (PSA- free survival) (p = 0,004). Conclusion Inhibition of glutaminolysis increases the cytotoxic effects of radiation in prostate tumor cells. Expression of the proteins involved in glutaminolysis can be potentially used to predict clinical outcome after RT in prostate cancer patients. OC-0153 Multicenter prospective validation of HSPB1 rs2868371 SNP and survival in non-small cell lung cancer M. Enguix-Riego 1 , J. Cacicedo Fernández de Bobadilla 2 , D. Herrero Rivera 3 , D. Delgado 4 , J. Nieto-Guerrero Gómez 4 ,