PO-442 In vitroassessment of sonic hedgehog (shh) pathway inhibitior, lde225, in paediatric ependymoma rela subgroup shows antiproliferative and proapoptotic effects

T Magalhaes, GAlencastro Veiga Cruzeiro, K Silva Borges, K Rodrigues da Silva, G Ribeiro de Souza, RGomes de Paula Queiroz, C Alberto Scridelli, L Gonzaga Tone
2018 Novel Targets, Delivery Systems   unpublished
SHH TP53 mutated cell lines regarding proliferation and apoptosis. Material and methods Expression of YAP in MB was assessed in 763 medulloblastoma patients trough in silico analysis utilising R2 genomic data base. It was utilised the cell lines DAOY and UW228 as TP53 mutated and ONS-76 as TP53 Wild Type. Cytoplasmatic and Nuclear detection of YAP was perfomed trough Immunofluorescence. Verteporfin, a YAP inhibitor was purchased from Sigma Aldrich. CCK8 Proliferation assay was perfomed in 48
more » ... rs incubating cells with concentrations ranging from 2,5 mM to 15 mM. Apoptosis and necrosis was assessed trough High content screening assay (HCS) trough Propidium Iodide and Anexin-FITC. Statistical analysis was perfomed trough One-way ANOVA followed by Bonferroni. All procedures performed were in accordance with the ethical standards of the institutional and/or national research committee. Results and discussions We have found an overexpression of YAP in MB SHH compared to others molecular subgroups and predominantly in SHH TP53 mutated subtype (alpha subtype). Cytoplasmatic and Nuclear YAP was identified in all cell lines. It was found a decrease in cell proliferation when incubated with Vertporfin. Interestingly, in 10 mM concentration, DAOY showed lower proliferation ratio (0.37) (p<0.05) compared to ONS-76 (0.47)(p<0.05). However, in UW228 was identified a proliferation ratio of (0.88)(p<0.05). DAOY and UW228 bears distinct TP53 mutation site, which might explain its different behaviour when exposed to compound. HCS showed an increase in apoptosis and necrosis levels in DAOY and ONS-76 cell line when incubated with Verteporfin. Conclusion This initial screening with Verteporfin lead us to primary conclusion such as the role of YAP promoting cell proliferation and cease apoptosis. However, the biological role of YAP still remains to be explored in MB SHH TP53.
doi:10.1136/esmoopen-2018-eacr25.467 fatcat:dl7pguwrkjagjpxgtzbtceswh4