Kooperativer Bibliotheksverbund

In: Neuro-Oncology, 2018, Vol. 20(suppl6), pp.vi45-vi46

Description: We have previously demonstrated that Myc-overexpressing medulloblastoma cells are sensitive to apoptosis induction in response to mitotic inhibition with an Aurora kinase B inhibitor. Profiling of differential mRNA expression by microarray and RT-PCR revealed that alpha cardiac actin 1 ( ACTC1 ) mRNA expression is upregulated in cells resistant to apoptosis triggered by Aurora kinase B inhibition and this upregulation is absent in Myc-overexpressing cells. We confirmed expression of ACTC1 protein by Western blot in SHH subgroup (DAOY, UW228, UW426) and Group 3 subgroup cell lines (D458, D425) and observed that expression is lower in Group 3 cells. These findings were further validated by analysis of ACTC1 mRNA expression among all four medulloblastoma subgroups by microarray in a set of 763 medulloblastomas which revealed increased expression of ACTC1 in SHH and WNT subgroups compared to Group 3 and Group 4. Inhibition of Aurora kinase B in SHH cells that overexpress Myc results in a reduction in ACTC1 protein level after 96 hours and this is not observed in the isotype control cells with basal Myc expression. Reduction in ACTC1 protein levels in SHH cells overexpressing Myc is associated with Caspase 3 cleavage. ACTC1 may play a role in blocking apoptosis triggered by mitotic inhibition in medulloblastoma. Further experiments to test this hypothesis are planned. These finding could potentially impact on chemotherapy choice in the treatment of SHH and WNT tumours, which demonstrate increased ACTC1 expression.

Keywords: Medicine;

ISSN: 1522-8517

E-ISSN: 1523-5866

DOI: 10.1093/neuonc/noy148.180

In: Neuro-Oncology, 2015, Vol. 17(suppl3), pp.iii28-iii28

Description: Sonic Hedgehog (SHH) signaling is active in 30% of childhood medulloblastomas. Dysregulation of p53 signaling is especially prognostic for poor survival in SHH medulloblastoma. But, p53 is mutated in 〈10% of medulloblastomas. Genomic analyses have demonstrated high-level amplification of PPM1D (WIP1) in p53 wild-type SHH medulloblastomas. WIP1 functions as an oncogene, in part by inactivating p53. We hypothesized that, in the absence of p53 mutation, WIP1 overexpression augments SHH-driven medulloblastoma tumorigenesis and is an important target in the treatment of SHH-active medulloblastomas. WIP1 overexpression in murine granule neuron precursor (GNP) cells enhanced expression of Shh target genes, increased the percentage of cells in S phase, and increased cell viability in response to Shh stimulation. Although WIP1 is known to primarily act through p53 pathways, we found that the interaction between WIP1 and Shh was partially p53-independent. To further understand WIP1 mechanisms in vivo, we developed a mouse in which WIP1 is expressed in the developing brain under control of the Neurod2 promoter (ND2:WIP1). The external granule layer in early post-natal ND2:WIP1 mice exhibited increased proliferation and expression of Shh downstream targets. When ND2:WIP1 transgenic mice were crossed with SmoA1/SmoA1 medulloblastoma-prone mice, medulloblastoma incidence increased and double transgenic SmoA1/SmoA1; ND2:WIP1+ mice demonstrated reduced survival. Conversely, Wip1 knock out significantly suppressed medulloblastoma formation in SmoA1/SmoA1 as well as Tamoxifen-treated Math1-creER; Ptc1 fl/fl mice. Treatment with a lentivirus that knocks down Wip1 or with the WIP1 inhibitor CCT007093 inhibits the effects of Shh stimulation and potentiates the effects of the SHH pathway-inhibiting drugs SANT-1, cyclopamine, or the clinically-relevant LDE225 on the growth of either SmoA1/SmoA1 or Math1-creER + ; Ptc1 fl/fl medulloblastoma cells in vitro. This suggests an important cross-talk between SHH and WIP1 that accelerates tumorigenesis. Our findings also suggest an important role for WIP1 inhibition in the treatment of SHH-active medulloblastomas.

Keywords: Medicine;

ISSN: 1522-8517

E-ISSN: 1523-5866

DOI: 10.1093/neuonc/nov061.112

In: Neuro-Oncology, 2014, Vol. 16(suppl5), pp.v23-v23

Description: Major research efforts have focused on the isolation and characterization of brain tumor stem cells, or propagating cells (BTPC) in a variety of brain cancers. Elucidating cell surface marker profiles that can be used to selectively isolate this cellular population is an imperative first step in the development of targeted therapies. Medulloblastoma (MB) is the most common form of pediatric brain cancer. MB is divided into 4 molecular subgroups; Wnt, Sonic Hedgehog (SHH), Group 3 and Group 4. Given the variable results obtained for currently utilized markers, as well as the cellular heterogeneity within and between MB sub-groups, it is likely there are additional surface marker profiles capable of selecting for sub-type specific MB BTPCs. We set out to identify novel surface marker combinations capable of selecting for TPCs in SHH MB. We employed the new BD Bioscience Lyoplate screening platform to compare 242 human cell surface marker levels across high and low self-renewing SHH MB sub-clones. The top 25 markers were refined by evaluating expression levels in Shh vs Group 3,4 and Wnt variants in transcriptome datasets representing 548 patient samples. Four markers, CD271, CD106/VCAM1, EGFR and CD171/NCAM-L1 showed consistent differential expression in the SHH subtype relative to the other variants. Flow cytometry validation in additional cell lines confirmed these findings. As a proof of principle, functional characterization of CD271 in SHH MB in vitro and in vivo was performed. Using fluorescence activated cell sorting and gain/loss of function studies, our results suggest that CD271 selects for MB progenitor cells. This work highlights a new approach to screening for differentially expressed surface markers across matched samples. We delineated a cell surface fingerprint for BTPC populations from MB molecular variants, however the utility can be seen in normal stem cell biology and across all forms of cancer.

Keywords: Medicine;

ISSN: 1522-8517

E-ISSN: 1523-5866

DOI: 10.1093/neuonc/nou239.2

In: Neuro-Oncology, 2016, Vol. 18(suppl3), pp.iii116-iii117

Keywords: Medicine;

ISSN: 1522-8517

E-ISSN: 1523-5866

DOI: 10.1093/neuonc/now076.83

In: Neuro-Oncology, 2014, Vol. 16(suppl5), pp.v102-v102

Description: OBJECTIVES: The advent of integrated genomics has fundamentally changed our understanding of medulloblastoma. Although survival differences have been shown to exist between the four principle subgroups, treatment related differences have yet to be elucidated. We sought to delineate these differences at a large referral centre. METHODS: All patients over age 3 treated with surgery, craniospinal irradiation and adjuvant chemotherapy were identified at the Hospital for Sick Children in Toronto from 1998-2012. RESULTS: Ninety-four patients were identified who met our inclusion criteria. Two periods were identified, those patients treated prior to 2006 as per the open protocols of the Children's Oncology Group (CCG9961, POG9631), and patients treated after 2006 treated as per the SJMB03 protocol. Five-year progression free survival over the entire cohort was 78%. When stratified by treatment, 5 year survival pre and post 2006 were identical (76.8% pre-2006 and 79.3% post-2006). When re-analysed in a subgroup specific manner, we find no significant differences in progression-free survival pre and post 2006. Strikingly, we found that Group 3 and 4 patients have excellent survivals compared to those previously reported, with 5 year progression-free survival in average risk Group 4 patients of over 90% and over 75% in average risk Group 3 patients regardless of treatment protocol. SHH patients did relatively poorly across both treatment protocols with 5 year progression free survival of 60% likely owing to a high proportion of TP53 mutated patients. CONCLUSIONS: In a cohort of homogenously treated patients over age 3, progression free survival appears to be improved compared to initial reports based on retrospective cohorts. The impact of subgroup affiliation in children over age 3 needs to assessed in large prospectively treated cooperative protocols to determine the subgroup specificity of treatment regiments.

Keywords: Medicine;

ISSN: 1522-8517

E-ISSN: 1523-5866

DOI: 10.1093/neuonc/nou256.26

In: Brain, 2018, Vol. 141(5), pp.1300-1319

Description: The molecular events underlying dissemination of group 3 medulloblastoma remain elusive. Ferrucci et al. show that PRUNE1 overexpression enhances the canonical TGF-β cascade, upregulates OTX2 and SNAIL, and inhibits the tumour suppressor PTEN. They describe anti-metastatic properties of an anti-PRUNE1 drug, and identify further deleterious gene variants as therapeutic targets. Genetic modifications during development of paediatric groups 3 and 4 medulloblastoma are responsible for their highly metastatic properties and poor patient survival rates. PRUNE1 is highly expressed in metastatic medulloblastoma group 3, which is characterized by TGF-β signalling activation, c-MYC amplification, and OTX2 expression. We describe the process of activation of the PRUNE1 signalling pathway that includes its binding to NME1, TGF-β activation, OTX2 upregulation, SNAIL ( SNAI1 ) upregulation, and PTEN inhibition. The newly identified small molecule pyrimido-pyrimidine derivative AA7.1 enhances PRUNE1 degradation, inhibits this activation network, and augments PTEN expression. Both AA7.1 and a competitive permeable peptide that impairs PRUNE1/NME1 complex formation, impair tumour growth and metastatic dissemination in orthotopic xenograft models with a metastatic medulloblastoma group 3 cell line (D425-Med cells). Using whole exome sequencing technology in metastatic medulloblastoma primary tumour cells, we also define 23 common ‘non-synonymous homozygous’ deleterious gene variants as part of the protein molecular network of relevance for metastatic processes. This PRUNE1/TGF-β/OTX2/PTEN axis, together with the medulloblastoma-driver mutations, is of relevance for future rational and targeted therapies for metastatic medulloblastoma group 3. 10.1093/brain/awy039_video1 awy039media1 5742053534001

Keywords: Medulloblastoma ; Metastatic Cns Tumour ; Molecular Genetics ; Genetic Network ; Oncology

ISSN: 0006-8950

E-ISSN: 1460-2156

DOI: 10.1093/brain/awy039

In: Neuro-Oncology, 2018, Vol. 20(suppl6), pp.vi33-vi33

Description: BACKGROUND/OBJECTIVES: MYC amplification in medulloblastoma (MB) determines highly aggressive disease, underscoring an urgent need for novel therapies. Let-7 microRNAs (miRNAs) inhibit tumor progression and regulate metabolism by degrading several mRNAs, including MYC. Let-7 miRNAs are frequently repressed in cancer, including MYC-driven MB. We previously reported that eukaryotic Elongation Factor-2 Kinase (eEF2K) is a pivotal regulator of MYC-driven tumor adaptation to nutrient deprivation (ND). Our data indicate that the eEF2K 3’ untranslated region (UTR) harbors a potential binding site for let-7 . In addition, eEF2K mRNA and let-7 miRNA expression negatively correlates in MB, suggesting regulation of the former by the latter. We therefore hypothesized that let-7 down-regulation induces eEF2K expression in MB, thereby supporting MYC-driven MB adaptation to ND and tumor progression.

Keywords: Medicine;

ISSN: 1522-8517

E-ISSN: 1523-5866

DOI: 10.1093/neuonc/noy148.124

In: Neuro-Oncology, 2014, Vol. 16(suppl3), pp.iii28-iii28

Description: BACKGROUND: Tumor recurrence is the main cause of death for children with medulloblastoma, the most common malignant childhood brain tumor. The MYCN oncogene is a poor prognosis marker and is amplified in the molecularly defined SHH and Group 4 subgroups but rarely in WNT and Group 3 subgroups of human medulloblastoma. Recent findings on childhood brain tumor relapse mechanisms suggest spatiotemporal differences within these four subgroups. SOX9 is a transcription factor that is important for glial fate determination in the brain but has also been found to promote tumor metastasization. We previously showed how expression of SOX9 correlates well with human SHH tumors but only few scattered SOX9-positive cells are found in SHH-independent Group 3 and Group 4 human medulloblastoma. METHODS: In order to study recurrence processes experimentally, we used a previously described transgenic Tet-OFF (Glt1-tTA) inducible model of MYCN-driven SHH-independent medulloblastoma (GTML mouse). To recreate metastatic recurrence we further used a Tet-ON (SOX9-rtTA) model that drives MYCN expression from the SOX9 promoter upon doxycycline treatment. RESULTS: By crossing the GTML Tet-OFF model with a Tet-ON transgene we managed to study rare SOX9-positive tumor cells after SOX9-negative tumor cells were first depleted using doxycycline. SOX9-positive GTML cells were tumorigenic and reinitiated distant recurrences over time. The SOX9-positive cells further showed an increased resistance to MYCN-targeted therapies. Relapses showed similar histopathology but presented generally higher levels SOX9 as compared to primary GTML tumors. A similar correlation was found in Group 3 and Group 4 medulloblastoma patients where isolated metastases had consistently higher SOX9 levels as compared to corresponding primary tumors. Finally, we overexpressed SOX9 in normal cerebellar stem cells transduced with mutationally stabilized MYCN-T58A and injected them back into the cerebellum of adult mice. Surprisingly, SOX9-positive MYCN-T58A brain tumors migrated and developed in the forebrain in contrast to the cerebellar stem cells transduced with MYCN-T58A only. CONCLUSIONS: Our findings suggest that increased levels of SOX9 drives migration in MYCN-driven medulloblastoma. Rare SOX9-positive tumor cells show an increased therapy resistance and are alone capable of reinitiating childhood brain tumors. Further characterization of SOX9-positive cells in Group 3 and Group 4 tumors could help us understand what drives metastatic medulloblastoma relapse and could lead to new therapies directed against these particularly serious cell types. SECONDARY CATEGORY: Tumor Biology.

Keywords: Medicine;

ISSN: 1522-8517

E-ISSN: 1523-5866

DOI: 10.1093/neuonc/nou208.20

In: Neuro-Oncology, 2018, Vol. 20(suppl2), pp.i140-i140

Description: Abstract BACKGROUND The MYC oncogenes contribute to more than 50% of all human cancers, but their therapeutic targeting has proven challenging. MYC/MYCN amplification in childhood medulloblastoma (MB) and neuroblastoma (NB) determine aggressive disease and high mortality, underlying the need for novel and effective therapies. MYC-driven transformation is energy demanding and impairs cell survival under nutrient deprivation (ND), a characteristic stress condition within the tumor microenvironment. We recently identified eukaryotic Elongation Factor 2 Kinase (eEF2K) as a pivotal mediator of the adaptive response of tumor cells to ND. We therefore hypothesized that eEF2K facilitates the adaptation of MYC/MYCN amplified MB/NB to ND, and that inhibiting this pathway can impair tumor progression. RESULTS Analyzing publicly available genomic databases and tissue microarrays, we found that high eEF2K expression and activity are strongly predictive of poor outcome in MB and NB (p〈0.001), and correlate with MYC/MYCN amplification (p〈0.001). Inhibition of eEF2K significantly decreases survival of MYC/MYCN amplified MB/NB cell lines in vitro under ND. Combination of eEF2K knockdown and caloric restriction determines a twofold growth decrease of MYCN amplified NB mouse xenografts. Finally, eEF2K inactivation significantly attenuated the ability of tumor cells to engage fatty acid oxidation under ND, suggesting a link between eEF2K, MYC transformation and lipid metabolism. CONCLUSIONS eEF2K represents a critical mediator for the adaptive response of MYC/MYCN amplified tumors to acute metabolic stress, and is therefore a promising therapeutic target. Future studies will combine eEF2K pharmacological inhibition with caloric restriction mimetics, as eEF2K activity appears to be critical under ND.

Keywords: Medicine;

ISSN: 1522-8517

E-ISSN: 1523-5866

DOI: 10.1093/neuonc/noy059.501

In: Neuro-Oncology, 2015, Vol. 17(suppl3), pp.iii20-iii20

ISSN: 1522-8517

E-ISSN: 1523-5866

DOI: 10.1093/neuonc/nov061.79

Source: Oxford University Press