PO-481 Patient-derived sarcoma primary cell lines with preserved cancer stem cell properties to screen anticancer compounds

Abstract

IntroductionSarcomas often show a limited response to current treatments. A hypothesis to explain this resistance relies on the existence of subpopulations of drug-resistant cancer stem cells (CSCs) responsible for relapses and metastasis. Therefore, there is a need for patient-close disease models amenable for testing the effect of anti-cancer therapies on CSCs. As a proof-of principle of their possible utility in future personalise medicine strategies, we have established a panel of patient-derived sarcoma primary cell lines, characterised their CSC subpopulations and tested the anti-tumour activity of the mithramycin analogue EC-8042 on these cell lines.Material and methodsFresh surgical samples of sarcomas were disaggregated and put in culture. In those cell lines adapted to grow in vitro, their proliferation ability and their potential to growth tumour xenografts were evaluated. CSCs subpopulations were characterised according to their ability to grow as tumorspheres, and the presence of cells presenting ALDH1 activity (Aldefluor© activity) or transcriptional activity due to SOX2/OCT4 in cells expressing the reporter system Sore6 (Sore6 activity). Cell survival, apoptotic induction and modulation of Sore6 activity was analysed after drug treatments. Finally, the expression/phosphorylation of signalling proteins was analysed by Western blotting.Results and discussionsWe have established a panel of patient-derived primary cell lines able to reproduce in vivo the histopathological features of their tumours of origin. Most of these cell lines were able to grow as serially passaged tumorspheres and showed between 1% and 15% of Aldefluor© positive cells. In addition, we detected Sore6 activity in 25% of cells in a cell line expressing the Sore6 reporter. Finally, these cell lines showed a distinct sensitivity to EC8042, with IC50 values ranging from 0.1 to 1 µM. EC-8042 was able to inhibit the expression of CSC-related factors such as SOX2, NOTCH1 and C-MYC in a dose-dependent fashion and moreover, this drug efficiently targeted Sore6 positive cells. Importantly, we found a strong correlation between the presence of intra-cellular NOTCH1 and the cytotoxic response to EC-8042.ConclusionThis panel of sarcoma primary cell lines with preserved CSC properties constitutes a valuable patient-close platform to test anti-cancer drugs. We found that EC-8042 could act as an anti-pluripotency agent in sarcomas and we identified intra-cellular NOTCH1 as a possible marker of response for this drug.