MEDB-09. Unraveling the role of unfolded protein response in medulloblastoma cancer stem cells

Medulloblastoma (MB) is the most common malignant childhood brain tumor. The current clinical approach consists of multimodal strategies with debilitating long-term effects and risk of tumor recurrence. Medulloblastoma stem cells (MBSCs) are a fraction of tumor cells with high proliferation potential and the capability to adapt to adverse/restrictive conditions in tumor milieu thus driving the refractoriness to conventional therapies. Recently, high basal levels of Unfolded Protein Response (UPR) molecules have been found in tumors of different tissue-origin and are correlated with poor prognosis and low patient survival. However, little is known about the role of UPR in MB. We investigated the expression and activation of UPR players in MBSCs. Human group 3 MB (G3MB) cell lines, specifically CHLA-01, D283- and D341-Med, were grown in Vitamin A and/or FBS or in stem selective medium (B27™) for 72 h before collection. Cells were fixed, stained with proper primary antibodies and images were acquired by confocal microscopy. The analysis of the transcription factors ATF-4 and CHOP revealed their elevated nuclear expression and co-localization, which resulted to be more marked in G3MB stem-like cells than in the differentiated ones. Also the ATF-6 branch was investigated, in differentiating conditions D283 and D341-Med showed a greater activation of ATF-6, represented by its nuclear localization, in respect to stem cells, while CHLA-01 did not show differences. Conversely XBP1, the transcription factor downstream IRE1 signaling, was not expressed in the three cell lines. Lastly, a Kaplan-Meier analysis on MB patients showed a worse prognosis with a shorter survival rate of patients expressing high ATF4 transcript levels. Our results reveal, even in resting conditions, preferential activation of the PERK branch in G3MB cells grown in stem-like condition suggesting that ATF-4 might be a promising therapeutic and prognostic factor to specifically target the stem compartment in aggressive MB.