Medulloblastoma (MB) is a malignant pediatric brain tumor arising in the cerebellum consisting of four distinct subgroups: WNT, SHH, Group 3 and Group 4, which exhibit different molecular phenotypes. We studied the expression of Dickkopf (DKK) 1-4 family genes, inhibitors of the Wnt signaling cascade, in MB by screening 355 expression profiles derived from four independent datasets. Upregulation of DKK1, DKK2 and DKK4 mRNA was observed in the WNT subgroup, whereas DKK3 was downregulated in 80% MBs across subgroups with respect to the normal cerebellum (p < 0.001). Since copy number aberrations targeting the DKK3 locus (11p15.3) are rare events, we hypothesized that epigenetic factors could play a role in DKK3 regulation. Accordingly, we studied 77 miRNAs predicting to repress DKK3; however, no significant inverse correlation between miRNA/mRNA expression was observed. Moreover, the low methylation levels in the DKK3 promoters (median: 3%, 5% and 5% for promoter 1, 2 and 3, respectively) excluded the downregulation of gene expression by methylation. On the other hand, the treatment of MB cells with Trichostatin A (TSA), a potent inhibitor of histone deacetylases (HDAC), was able to restore both DKK3 mRNA and protein. In conclusion, DKK3 downregulation across all MB subgroups may be due to epigenetic mechanisms, in particular, through chromatin condensation.

The insulin-like growth factor (IGF) pathway plays an important role in several brain tumor entities. However, the lack of inhibitors crossing the blood-brain barrier remains a significant obstacle for clinical translation. Here, we targeted the IGF pathway using ceritinib, an off-target inhibitor of the IGF1 receptor (IGF1R) and insulin receptor (INSR), in a pediatric patient with an unclassified brain tumor and a notch receptor 1 (NOTCH1) germline mutation. Pathway analysis of the tumor revealed activation of the sonic hedgehog (SHH), the wingless and integrated-1 (WNT), the IGF, and the Notch pathway. The proliferation of the patient tumor cells (225ZL) was inhibited by arsenic trioxide (ATO), which is an inhibitor of the SHH pathway, by linsitinib, which is an inhibitor of IGF1R and INSR, and by ceritinib. 225ZL expressed INSR but not IGF1R at the protein level, and ceritinib blocked the phosphorylation of INSR. Our first personalized treatment included ATO, but because of side effects, we switched to ceritinib. After 46 days, we achieved a concentration of 1.70 µM of ceritinib in the plasma, and after 58 days, MRI confirmed that there was a response to the treatment. Ceritinib accumulated in the tumor at a concentration of 2.72 µM. Our data suggest ceritinib as a promising drug for the treatment of IGF-driven brain tumors.

Children with high-risk SHH/TP53-mut and Group 3 medulloblastoma (MB) have a 5-year overall survival of only 40%. Innovative approaches to enhance survival while preventing adverse effects are urgently needed. We investigated an innovative therapy approach combining irradiation (RT), decitabine (DEC), and abacavir (ABC) in a patient-derived orthotopic SHH/TP53-mut and Group 3 MB mouse model. MB-bearing mice were treated with DEC, ABC and RT. Mouse survival, tumor growth (BLI, MRT) tumor histology (H/E), proliferation (Ki-67), and endothelial (CD31) staining were analyzed. Gene expression was examined by microarray and RT-PCR (Ki-67, VEGF, CD31, CD15, CD133, nestin, CD68, IBA). The RT/DEC/ABC therapy inhibited tumor growth and enhanced mouse survival. Ki-67 decreased in SHH/TP53-mut MBs after RT, DEC, RT/ABC, and RT/DEC/ABC therapy. CD31 was higher in SHH/TP53-mut compared to Group 3 MBs and decreased after RT/DEC/ABC. Microarray analyses showed a therapy-induced downregulation of cell cycle genes. By RT-PCR, no therapy-induced effect on stem cell fraction or immune cell invasion/activation could be shown. We showed for the first time that RT/DEC/ABC therapy improves survival of orthotopic SHH/TP53-mut and Group 3 MB-bearing mice without inducing adverse effects suggesting the potential for an adjuvant application of this multimodal therapy approach in the human clinic.