ESTRO 38 Abstract book

S598 ESTRO 38

or 8 Gy. Cell growth of H2B-mCherry transduced cells and cytotoxicity (CellTox™ Green Cytotoxicity Assay) were measured using live cell imaging (IncuCyte). Inhibitory effects on metabolic profiles were determined using Seahorse XF96 extracellular Flux analyzer. Results Glutamine metabolism-related markers were significantly higher in AC than SCC (Figure 1A-C). Glucose transporter 1 (GLUT1) protein was higher in solid than lepidic AC (p<0.01), but was highest in SCC (p<0.05) (Figure 1D). In AC, GLUT1 protein correlated with poor differentiation grade (p<0.001), lymph node metastases (p=0.003) and worse disease-free survival (p=0.008). Glutamine transporter (SLC1A5) protein was higher in AC with worse pTNM stage (r(s)=0.39, p=0.009). TP53 and PIK3CA mutations were more often found in SCC than AC (97.4% versus 71.2%, p=0.001; 10.5% versus 0%, p=0.011 respectively), while EGFR and KRAS mutations were more frequently observed in AC (10.2% versus 0%, p=0.042; 35.6% versus 5.3%, p=0.001 respectively). NSCLC cell lines responded differently to lonidamine and/or 968, largely corresponding with changes in glycolytic and mitochondrial metabolism upon treatment. 968 was cytotoxic in cell lines with high glutaminase C expression (H1975 and H520), whereas combination treatment was cytotoxic in KRAS mutated cell lines (SW900 and H23). H292 and HCC827 were resistant to combination treatment. Treatment with 968 and especially lonidamine resulted in radiosensitization of H292 and HCC827 in terms of decreased relative cell growth and increased cytotoxicity (Figure 2).

assay. Cell cycle analysis was performed by flow cytometry after incorporation of BrdU. HER2 cells were irradiated (GSR 137 Cs 662 keV) at different dose levels (from 1 to 8 Gy) after exposure to T-DM1. Survival fractions were determined based on clonogenic assays or on cell survival after 5 doubling times. Radiosensitivity parameter (D 10 and D 37 ) were calculated using the mean values of a and b determined from the curves drawn for best fit to the experimental data. Results The action of T-DM1 on HER2 cells showed significant lethality by deprivation of the HER2 signaling pathway and intracellular DM1 action on the cell cycle with significant G2/M phase blocking. After 72h treatment with T-DM1 alone, the median effective dose (ED 50 ) determined for the six cell lines used is function of the expression of the receptor HER2 except for one cell line (BT474). Using sub- toxic doses of T-DM1 and short-term drug exposure (6 or 12h), the cell viability still decreases dramatically after several days. After irradiation alone, D 37 and D 10 were significantly higher for the three high-HER2 expressing cells lines (HCC1954, BT474 and SKBr3) compared to the low-HER2 expressing cells (MDA-453 and ZR-75-1), with a linear increasing relationship between the radioresistance and the level of HER2 expression (D 37 : r 2 =0.9; p<0.0001 and D 10 : r 2 =0.7; p<0.0001). In combination with radiation, T-DM1 elicited strictly additive interaction. There was no significant difference on D 10 compared to control for all cell lines. HCC1954 : D 10 T-DM1 = 6.15±0.64 vs 6.0±0.24 for control; BT-474: D 10 T- DM1 = 7.86±0.22 vs 7.6.0±0.43 for control; SKBr3 : D 10 T- DM1 = 5.68±0.29 vs 5.43±0.14 for control; MDA-MB-453 : D 10 T-DM1 = 4.82±0.07 vs 4.02±0.17 for control and ZR-75- 1 = D 10 T-DM1 = 3.3±0.3 vs 3.02±0.3 for control. Conclusion Although HER2 expression is a radioresistant factor, T-DM1 did not have any radiosensitizing impact on HER2 positive breast cancer cells in this in vitro study. The major toxicity of T-DM1 alone could explain this phenomenon but further explorations are needed. PO-1077 The heterogeneous metabolic and mutational landscape of non-small cell lung carcinomas T. Van Zon - Meijer 1 , W. Peeters 1 , M. Looijen-Salamon 2 , R. Biemans 3 , L. Dubois 3 , P. Span 1 , J. Bussink 1 1 Radboud university medical center, Radiotherapy and OncoImmunology Laboratory- Department of Radiation Oncology, Nijmegen, The Netherlands ; 2 Radboud university medical center, Pathology, Nijmegen, The Netherlands ; 3 Maastricht Comprehensive Cancer Centre- Maastricht University Medical Centre, Department of Precision Medicine- The M-Lab- GROW - School for Oncology and Developmental Biology, Maastricht, The Netherlands Purpose or Objective Both hypoxia and oncogenic mutations rewire tumor metabolism causing treatment resistance. Metabolic inhibition might therefore sensitize tumors to radiation. We explored glucose and glutamine metabolism-related markers and mutation status in stage I-IIIA NSCLC in relation to histology and tumor aggressiveness characteristics. Furthermore, we examined the effect of lonidamine (hexokinase inhibitor) and/or 968 (glutaminase inhibitor) on metabolism, cell growth, cytotoxicity and radiosensitivity in NSCLC cell lines in vitro . Material and Methods Mutation analysis was performed for 97 tumors. Metabolic marker expression was measured by immunofluorescent staining (protein) and qPCR (mRNA) (n=81). Adeno- (H23, HCC827, H1975) (AC) and squamous cell carcinoma (H520, H292, SW900) (SCC) NSCLC cells were treated with lonidamine and/or 968 for 72 hours under physiological levels of glucose (1.5 mM). Cells were irradiated with 0, 4