Medulloblastoma Subtypes Research Articles

Medulloblastoma in the age of molecular subgroups: a review.

Medulloblastoma is the most common pediatric malignant brain tumor. Advances in molecular profiling have uncovered significant heterogeneity among medulloblastomas and led to the identification of four distinct subgroups (wingless [WNT], sonic hedgehog [SHH], group 3, and group 4) that represent distinct disease entities in both underlying biology and clinical characteristics. The rapidly expanding repertoire of tools to study developmental and cancer biology is providing a wealth of knowledge about these embryonal tumors and is continuously refining the understanding of this complex cancer. In this review, the history of discovery in medulloblastoma is discussed, setting a foundation to outline the current state of understanding of the molecular underpinnings of this disease, with a focus on genomic events that define the aforementioned subgroups and evolving areas of focus, such as the cell of origin of medulloblastoma and medulloblastoma subtypes. With these recent discoveries in mind, the current state of medulloblastoma treatment and clinical trials is reviewed, including a novel risk stratification system that accounts for the molecular biomarkers of patients with a high risk for refractory disease. Lastly, critical areas of focus for future basic science and clinical research on this disease are discussed, such as the complexities of medulloblastoma metastases and recurrence as well as the priorities and strategies to implement in future clinical trials.

P01.01 The role of exosome miRNA between tumor cells and microglias during the progression of medulloblastoma

Abstract BACKGROUND Medulloblastoma (MB) is the most common malignant paediatric brain tumor. Recent studies show that M2 cells were relative more abundant in Shh subtype of MBs compared with other three subtypes. It’s known that M2 cells have close relationship with many tumors’ progression. But if they play any role in the progression of Shh subtype of MB is not yet clear. Many studies demonstrate that exosomes carring miRNAs have close relationship with tumor invasion. The aim of present study is to clarify the role of exosome miRNA between tumor cells and microglias during the progression of Shh subtype of medulloblastoma. MATERIAL AND METHODS Immunofluerescence staining using iNOS and Arg1, which is M1 and M2 specific marker, respectively, was performed in four subtypes of MBs. After coculture of exosomes extracted from Shh subtype of MB cell (DAOY) with microglia cell (BV2), Q-PCR and ELISA assay were done to evaluate the polarization status of the microglia. Transwell and scratch assay were then performed to detect the migration ability of DAOY cell after treatment of exosomes from polirized M2 cells. MiRNA sequencing by Ion Proton technology was then done to analyze the miRNAs expression level between Shh subtype and other subtype of MBs. Transformation assay was used to overexpress and inhibit the expression of these miRNAs respectively to further clarify the role of exosome miRNA in the polarization of BV2 cells. RESULTS M2 cells were observed more abundant than other three subtypes of tumors, supporting that M2 cells play some role in this subtype of MBs. Exosomes of DAOY cells can induce the polarization of M2 cells. The polarized M2 cells can improved the migration and invasion ability of DAOY cell. Dozens of miRNAs were identified with different expression level between Shh subtype of MBs and other subtype of MB cells. Among them, 4 miRNAs were reported to be related with polariztion of M2 in many other lesions. Three of the 4 miRNAs can induce the polarization of M2 in present study. CONCLUSION Our study demonstrated exosome miRNA play a critical role between tumor cells and microglias during the progression of Shh subtype of medulloblastoma.

Simplified Molecular Subtyping of Medulloblastoma for Reduced Cost and Improved Turnaround Time.

Molecular subtyping of medulloblastoma (MB) has become increasingly important for prognosis and management. Typically this involves detailed molecular genetic testing which may not be available in all centers. The purpose of the present study was to find a simplified approach to assign molecular subtypes of MB for routine use in centers with more limited resources. The molecular subtypes of MBs from 32 Thai patients, aged 0.5 to 35 years, were first determined by NanoString. These results were then compared with those obtained using a combination of limited immunohistochemistry (IHC) (β-catenin, GAB-1, YAP-1, p75-NGFR, OTX2) and CTNNTB exon 3 mutation analysis. By NanoString assay, there were 6 MBs (19%) in the wingless (WNT) group, 8 (25%) in the sonic hedgehog (SHH) group, 7 (22%) in group 3, and 11 (34%) in group 4. Although β-catenin immunostaining missed 4/6 WNT MBs, CTNNTB mutation analysis confirmed all WNT MB cases with amplifiable DNA. The IHC panel correctly assigned all the other molecular subtypes, except for 1 MB in group 4. Thus, our protocol was able to correctly categorized 31/32 cases or 97% of cases. Our study is the first to report molecular subtypes of MB in Southeast Asia. We found that molecular subgroups of MBs can be reliably assigned using a limited IHC panel of β-catenin, GAB-1, YAP-1, p75-NGFR, OTX2, together with CTNNTB exon 3 mutation analysis. This simplified approach incurs lower cost and faster turnaround time compared with more elaborate molecular methodologies and should be beneficial to centers with reduced laboratory resources.

Abstract 2865: AsiDNATM, a novel DNA repair inhibitor to sensitize aggressive medulloblastoma subtypes

Abstract Purpose: Medulloblastoma is paediatric tumor of the cerebellum. It represents the most frequent malignant brain tumor in childhood. It can be classified into four disparate molecular subgroups but current therapies are not yet tailored to their specificities. This is particularly important for subgroups with worse prognosis as Group3. Patients who survive often present severe treatment-related morbidity, which can be largely attributed to brain radiation in ages of development. It is therefore important to improve treatment efficacy in more aggressive subgroups as well as reduce treatment-related morbidity across all subgroups. We investigated whether AsiDNATM, a radiosensitizer in clinical trials which works through DNA repair inhibition, could be used to improve radiotherapy efficacy with no additional toxicity allowing radiation dose de-escalation in groups with better prognosis and improved radiotherapy efficacy in those with high-risk disease. Experimental design: The cytotoxic effect of AsiDNATM was determined in a panel of 4 human medulloblastoma cell lines from different subgroups and different P53 status. We analyzed the ability of AsiDNATM to reach brain tissue and toxicity of treatments with or without irradiation in pups (10 days old mice). Distribution of AsiDNA was estimated by quantification of Cy-5-AsiDNA in brain tissues. Radiation toxicity was estimated by survival to high doses, monitoring of circulating growth hormone (GH), weight loss, bones size at adult age and neuro histology analysis. We assessed the AsiDNATM radio-enhancing efficacy in Group3 xenografted preclinical models using X-RAD 320 (PXI Inc) X-ray irradiator. Local brain irradiation was performed with an image-guided X-ray system (SARPP, Xstrahl Ldt), in which dosimetry and animal setting were controlled at each treatment session. Results: AsiDNATM treatments exhibit an additive effect to radiation in all cell lines, leading to an approximate 2-fold reduction of the radiation dose to reach 50% survival. Cy5-AsiDNA distributed to pup brain but not to adult brain devoid of tumors. However, in adults AsiDNATM distributed in tumors in brain through permeability of tumor vessels. Irradiation toxicity in pups was confirmed by a significant decrease in GH secretion and associated reduction of bone size. The lethal dose after irradiation was 20 Gy. No increase of irradiation toxicity was observed with AsiDNATM. In vivo, AsiDNATM alone significantly enhances survival rates (p=0.005) and increases radiotherapy efficacy. When combined with radiotherapy, AsiDNATM led to delay in tumor growth and survival improvement as compared to radiotherapy alone. Conclusion: Overall, our current results show AsiDNATM as an attractive candidate to improve radiation therapy in medulloblastoma in both standard- and high-risk patients with no indication of additional toxicity in healthy developing brain tissues. Citation Format: Sofia Ferreira, Chloe Foray, Sophie Heinrich, Celio Pouponnot, Marie Dutreix. AsiDNATM, a novel DNA repair inhibitor to sensitize aggressive medulloblastoma subtypes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2865.

Differential Expression of Genes for Ubiquitin Ligases in Medulloblastoma Subtypes.

Using publically available datasets on gene expression in medulloblastoma (MB) subtypes, we selected genes for ubiquitin ligases and identified statistically those that best predicted each of the four major MB subgroups as separate disease entities. We identify a gene coding for an ubiquitin ligase, ZNRF3, whose overexpression alone can predict the WNT subgroup for 100% in the Pfister dataset. For the SHH subgroup, we identify a gene for a regulatory subunit of the protein phosphatase 2A (PP2A), PPP2R2C, as the major predictor among the E3 ligases genes. The ubiquitin and ubiquitin-like conjugation database (UUCD) lists PPP2R2C as coding for a Cullin Ring ubiquitin ligase adaptor. For group 3 MBs, the best ubiquitin ligase predictor was PPP2R2B, a gene which codes for another regulatory subunit of the PP2A holoenzyme. For group 4, the best E3 gene predictors were MID2, ZBTB18, and PPP2R2A, which codes for a third PP2A regulatory subunit. Heatmap analysis of the E3 gene data shows that expression of ten genes for ubiquitin ligases can be used to classify MBs into the four major consensus subgroups. This was illustrated by analysis of gene expression of ubiquitin ligases of the Pfister dataset and confirmed in the dataset of Cavalli. We conclude that genes for ubiquitin ligases can be used as genetic markers for MB subtypes and that the proteins coded for by these genes should be investigated as subtype specific therapeutic targets for MB.

Abstract A45: Tumor suppressor miR-584-5p regulates MYC and sensitizes MYC-amplified medulloblastoma to vincristine and ionizing radiation

Abstract Despite recent improvements in the overall survival of medulloblastoma (MB), only a modest percentage of patients survive high-risk disease. Furthermore, the toxicity associated with current therapies inflicts debilitating lifelong side effects on surviving MB patients. Using an unbiased microRNA screen, we identified miR-584-5p as a potent tumor suppressor that uniquely sensitizes MB to both ionizing radiation (IR) and vincristine (VCR), which are commonly used in combination to treat MB. Our studies revealed that miR-584-5p inhibited MB stem cell proliferation and consequently growth in vivo. Notably, we discovered that miR-584-5p imparts its tumor-suppressor and therapy-sensitizing effects by inhibiting histone deacetylase 1 (HDAC1), eukaryotic translation initiation factor 4e family member 3 (eIF4E3), and C-MYC. We demonstrated that miR-584-5p overexpression or the knockdown of its target genes including eIF4E3 and HDAC1 resulted in G2-M arrest, DNA damage, spindle defects, and apoptosis in MB cells. Meta-analysis of normal brain and MB tumor samples showed that while miR-584-5p is highly expressed in normal brain and cerebellum, it is highly downregulated across all MB subtypes. miR-584-5p targets, on the other hand, are highly overexpressed in MB tumors compared to normal cerebellum. Taken together, this study highlights a previously unidentified role for miR-584-5p, eIF4E3, and HDAC1 in regulating microtubule dynamics and DNA repair pathways. In addition, our study identifies novel targets that can affect the therapeutic efficacy of VCR and IR in MB and sets the stage for a new paradigm of treating medulloblastoma using microRNA-based therapeutics. Citation Format: Nourhan Abdelfattah, Subapriya Rajamanickam, Santosh Timilsina, Panneerdoss Subbarayalu, Benjamin Onyeagucha, Manjeet Rao. Tumor suppressor miR-584-5p regulates MYC and sensitizes MYC-amplified medulloblastoma to vincristine and ionizing radiation [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr A45.

GFI1's role in DNA repair suggests implications for tumour cell response to treatment.

Despite recent advances in cancer treatment through personalized and precision medicine and new avenues such as immunotherapy and chimeric antibodies, the induction of DNA damage either through irradiation or specific compounds remains the primary approach to kill tumour cells. Improvements in our understanding of how tumour cells respond to DNA damage, and especially how this response differs from that of normal cells, are crucial to the development of better and more efficient therapies. We have recently shown that the activity of the oncogenic transcription factor GFI1, which is required for the development and maintenance of T and B cell leukemia, increases the ability of tumour cells to repair their DNA following damage (Vadnais et al. Nat Commun 9(1):1418). GFI1 accomplishes this by regulating the post-translational modifications (PTM) of key DNA repair proteins, including MRE11 and 53BP1, by the methyltransferase PRMT1. Here, GFI1 acts as an accessory protein required for the interaction between the enzyme and its substrates. This has implications for the treatment response of tumour cells overexpressing GFI1, which includes T cell leukemia, neuroendocrine lung carcinomas and aggressive subtypes of medulloblastoma, and suggests that targeting GFI1's activity and with this its capacity to aid DNA repair may open avenues for new therapeutic approaches.

Reproducibility of academic preclinical translational research: lessons from the development of Hedgehog pathway inhibitors to treat cancer.

Academic translational research is growing at a great pace at a time in which questions have been raised about the reproducibility of preclinical findings. The development of Hedgehog (HH) pathway inhibitors for the treatment of cancer over the past two decades offers a case study for understanding the root causes of failure to predict clinical outcomes arising from academic preclinical translational research. Although such inhibitors were once hoped to be efficacious in up to 25% of human cancer, clinical studies showed responses only in basal cell carcinoma and the HH subtype of medulloblastoma. Close examination of the published studies reveals limitations in the models used, lack of quantitative standards, utilization of high drug concentrations associated with non-specific toxicities and improper use of cell line and mouse models. In part, these issues arise from scientific complexity, for example, the failure of tumour cell lines to maintain HH pathway activity in vitro, but a greater contributing factor appears to be the influence of unconscious bias. There was a strong expectation that HH pathway inhibitors would make a profound impact on human cancer and experiments were designed with this assumption in mind.

Abstract 4138: CK1δ-BRD4 pathway as novel therapeutic target for SHH subtype of medulloblastoma

Abstract Background: Medulloblastoma is the most common malignant pediatric brain tumor with variable prognosis due to its clinical and genomic heterogeneity. Despite decades of treatment advances, nearly 40% of children experience tumour recurrence, and 30% will die from this disease. Thus, new drugs and drug combinations need to be developed that effectively treat medulloblastoma. Casein kinase 1δ (CK1δ) is a serine/threonine kinase that controls cell cycle progression, signal transduction and neurogenesis, and we previously reported high levels of CK1δ in mouse models of medulloblastoma. BRD4 is an epigenetic reader protein that controls expression of several oncogenes. We wanted to determine the mechanism through which CK1δ regulates BRD4 activity in medulloblastoma and assess whether a combination of CK1δ and BRD4 inhibitors could be more effective in decreasing medulloblastoma progression in vivo. Methods: Granule cell progenitors (GCPs), medulloblastoma Ptch-/-P53-/- mouse cells and SUFU-/- mouse embryonic fibroblasts were incubated with the CK1δ inhibitor SR-1277 and the phosphorylation levels of BRD4 were determined by WB and its binding to chromatin by ChIP. In vitro phosphorylation studies with human BRD4 purified from E. coli were performed and whether genetic disruption of CK1δ reduces BRD4 phosphorylation was tested. We combined BRD4 and CK1δ inhibitors (JQ1 and SR-1277, respectively) and analysed Gli1 mRNA expression and EdU incorporation. We deleted CK1δ in GCPs in Ptch-/-;P53-/- mouse models of medulloblastoma by breeding these mice with a Tg(Atoh1-Cre);CK1δfl/fl strain. Furthermore, we intracranially transplanted human SHH medulloblastoma PDX cells expressing luciferase in mice, to validate the combination therapy. Results: CK1δ inhibitor treatment and CK1δ knockout reduced BRD4 phosphorylation, suggesting that CK1δ phosphorylates BRD4 and is required for BRD4 recruitment to chromatin. In vitro phosphorylation studies with purified human BRD4 and CK1δ confirmed CK1δ-mediated BRD4 phosphorylation and CK1δ is required for BRD4 phosphorylation at serine 492 and 494 in vivo. Using loss of function studies in GCPs, both CK1δ and BET inhibition reduced BRD4 association with Gli1 promoter, thereby reducing Gli1 mRNA levels. In addition, SR-1277 suppressed SHH signaling downstream of SUFU of medulloblastoma cells, in vitro and in vivo. We also found synergy by combining these compounds in vitro and in reducing tumor burden in the in vivo model with intracranial allografts. Conclusion: Together, our studies validate the CK1δ-BRD4 pathway as a novel target in medulloblastoma. The significance of our work is underscored by the possibility that simultaneous CK1δ-BRD4 inhibition could overcome the resistance observed with BRD4 inhibitors and enhance therapeutic benefit to patients. We are also the first group to demonstrate that CK1δ acts downstream of SHH signalling in SHH-driven medulloblastoma. Citation Format: Sze Kiat Tan, Clara Penas, Cheng-Ming Chiang, David Robbins, Nagi Ayad. CK1δ-BRD4 pathway as novel therapeutic target for SHH subtype of medulloblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4138.

Poliovirus Receptor (CD155) Expression in Pediatric Brain Tumors Mediates Oncolysis of Medulloblastoma and Pleomorphic Xanthoastrocytoma.

Poliovirus oncolytic immunotherapy is a putatively novel approach to treat pediatric brain tumors. This work sought to determine expression of the poliovirus receptor (PVR), CD155, in low-grade and malignant pediatric brain tumors and its ability to infect, propagate, and inhibit cell proliferation. CD155 expression in pleomorphic xanthoastrocytoma (PXA), medulloblastoma, atypical teratoid rhabdoid tumor, primitive neuroectodermal tumor, and anaplastic ependymoma specimens was assessed. The ability of the polio: rhinovirus recombinant, PVSRIPO, to infect PXA (645 [BRAF V600E mutation], 2363) and medulloblastoma (D283, D341) cells were determined by viral propagation measurement and cell proliferation. PVR mRNA expression was evaluated in 763 medulloblastoma and 1231 normal brain samples. CD155 was expressed in all 12 patient specimens and in PXA and medulloblastoma cell lines. One-step growth curves at a multiplicity of infection of 10 demonstrated productive infection and peak plaque formation units at 5-10 hours. PVSRIPO infection significantly decreased cellular proliferation in 2363, 645, and D341 cell lines at 48 hours (p < 0.05) and resulted in cell death. PVR expression was highest in medulloblastoma subtypes Group 3γ, WNTα, and WNTβ (p < 0.001). This proof-of-concept in vitro study demonstrates that PVSRIPO is capable of infecting, propagating, prohibiting cell proliferation, and killing PXA and Group 3 medulloblastoma.

Incorporating Advances in Molecular Pathology Into Brain Tumor Diagnostics.

Recent advances in molecular pathology have reshaped the practice of brain tumor diagnostics. The classification of gliomas has been restructured with the discovery of isocitrate dehydrogenase (IDH) 1/2 mutations in the vast majority of lower grade infiltrating gliomas and secondary glioblastomas (GBM), with IDH-mutant astrocytomas further characterized by TP53 and ATRX mutations. Whole-arm 1p/19q codeletion in conjunction with IDH mutations now define oligodendrogliomas, which are also enriched for CIC, FUBP1, PI3K, NOTCH1, and TERT-p mutations. IDH-wild-type (wt) infiltrating astrocytomas are mostly primary GBMs and are characterized by EGFR, PTEN, TP53, NF1, RB1, PDGFRA, and CDKN2A/B alterations, TERT-p mutations, and characteristic copy number alterations including gains of chromosome 7 and losses of 10. Other clinically and genetically distinct infiltrating astrocytomas include the aggressive H3K27M-mutant midline gliomas, and smaller subsets that occur in the setting of NF1 or have BRAF V600E mutations. Low-grade pediatric gliomas are both genetically and biologically distinct from their adult counterparts and often harbor a single driver event often involving BRAF, FGFR1, or MYB/MYBL1 genes. Large scale genomic and epigenomic analyses have identified distinct subgroups of ependymomas tightly linked to tumor location and clinical behavior. The diagnosis of embryonal neoplasms also integrates molecular testing: (I) 4 molecularly defined, biologically distinct subtypes of medulloblastomas are now recognized; (II) 3 histologic entities have now been reclassified under a diagnosis of "embryonal tumor with multilayered rosettes (ETMR), C19MC-altered"; and (III) atypical teratoid/rhabdoid tumors (AT/RT) now require SMARCB1 (INI1) or SMARCA4 (BRG1) alterations for their diagnosis. We discuss the practical use of contemporary biomarkers for an integrative diagnosis of central nervous system neoplasia.

Single cell coagulomes as constituents of the oncogene-driven coagulant phenotype in brain tumours

Molecular profiling of human cancers revealed a startling diversity in disease-causing mechanisms superseding histological and anatomical commonalities. The emerging molecular subtypes and disease entities are often driven by distinct oncogenic pathways and their effectors, including those acting extracellularly on the vascular and coagulation systems. Indeed, several oncogenic mutations such as those affecting protein-coding genes (RAS, EGFR, PTEN, TP53) and non-coding RNA (microRNA) regulate multiple effectors of the coagulation system (coagulome), including tissue factor, protease activated receptors, clotting factors, mediators of platelet function and fibrinolysis. This is exemplified by differential coagulome profiles in the molecular subtypes of glioblastoma, medulloblastoma and other human tumours. There is mounting clinical evidence that the mutational status of cancer driver genes such as KRAS or IDH1 may influence the risk of venous thromboembolism in patients with colorectal, lung or brain cancers. Notably, single cell sequencing in glioblastoma revealed a remarkable intra-tumoural heterogeneity of cancer cell populations with regard to their individual coagulomes, suggesting a combinatorial and dynamic nature of the global pro-thrombotic phenotype. We suggest that the cellular complexity of specific cancers may define their mechanisms of interactions with the coagulation system, and the risks of thrombosis. Thus, more biologically- based, disease-specific and personalized approaches may be needed to diagnose and manage cancer-related thrombosis.

The FDA approved PI3K inhibitor GDC-0941 enhances invitro the anti-neoplastic efficacy of Axitinib against c-myc-amplified high-risk medulloblastoma.

Aberrant receptor kinase signalling and tumour neovascularization are hallmarks of medulloblastoma development and are both considered valuable therapeutic targets. In addition to VEGFR1/2, expression of PDGFR α/β in particular has been documented as characteristic of metastatic disease correlating with poor prognosis. Therefore, we have been suggested that the clinically approved multi‐kinase angiogenesis inhibitor Axitinib, which specifically targets these kinases, might constitute a promising option for medulloblastoma treatment. Indeed, our results delineate anti‐neoplastic activity of Axitinib in medulloblastoma cell lines modelling the most aggressive c‐myc‐amplified Non‐WNT/Non‐SHH and SHH‐TP53‐mutated tumours. Exposure of medulloblastoma cell lines to Axitinib results in marked inhibition of proliferation and profound induction of cell death. The differential efficacy of Axitinib is in line with target expression of medulloblastoma cells identifying VEGFR 1/2, PDGFR α/β and c‐kit as potential markers for drug application. The high specificity of Axitinib and the consequential low impact on the haematopoietic and immune system render this drug ideal multi‐modal treatment approaches. In this context, we demonstrate that the clinically available PI3K inhibitor GDC‐0941 enhances the anti‐neoplastic efficacy of Axitinib against c‐myc‐amplified medulloblastoma. Our findings provide a rational to further evaluate Axitinib alone and in combination with other therapeutic agents for the treatment of most aggressive medulloblastoma subtypes.

HDL nanoparticles targeting sonic hedgehog subtype medulloblastoma

Medulloblastoma is the most common paediatric malignant brain cancer and there is a need for new targeted therapeutic approaches to more effectively treat these malignant tumours, which can be divided into four molecular subtypes. Here, we focus on targeting sonic hedgehog (SHH) subtype medulloblastoma, which accounts for approximately 25% of all cases. The SHH subtype relies upon cholesterol signalling for tumour growth and maintenance of tumour-initiating cancer stem cells (CSCs). To target cholesterol signalling, we employed biomimetic high-density lipoprotein nanoparticles (HDL NPs) which bind to the HDL receptor, scavenger receptor type B-1 (SCARB1), depriving cells of natural HDL and their cholesterol cargo. We demonstrate uptake of HDL NPs in SCARB1 expressing medulloblastoma cells and depletion of cholesterol levels in cancer cells. HDL NPs potently blocked proliferation of medulloblastoma cells, as well as hedgehog-driven Ewing sarcoma cells. Furthermore, HDL NPs disrupted colony formation in medulloblastoma and depleted CSC populations in medulloblastoma and Ewing sarcoma. Altogether, our findings provide proof of principle for the development of a novel targeted approach for the treatment of medulloblastoma using HDL NPs. These findings present HDL-mimetic nanoparticles as a promising therapy for sonic hedgehog (SHH) subtype medulloblastoma and possibly other hedgehog-driven cancers.

MRI features as a helpful tool to predict the molecular subgroups of medulloblastoma: state of the art.

Medulloblastoma is the most common malignant pediatric brain tumor. Medulloblastoma should not be viewed as a single disease, but as a heterogeneous mixture of various subgroups with distinct characteristics. Based on genomic profiles, four distinct molecular subgroups are identified: Wingless (WNT), Sonic Hedgehog (SHH), Group 3 and Group 4. Each of these subgroups are associated with specific genetic aberrations, typical age of onset as well as survival prognosis. Magnetic resonance imaging (MRI) is performed for all patients with brain tumors, and has a key role in the diagnosis, surgical guidance and follow up of patients with medulloblastoma. Several studies indicate MRI as a promising tool for early detection of medulloblastoma subgroups. The early identification of the subgroup can influence the extent of surgical resection, radiotherapy and chemotherapy targeted treatments. In this article, we review the state of the art in MRI-facilitated medulloblastoma subgrouping, with a summary of the main MRI features in medulloblastoma and a brief discussion on molecular characterization of medulloblastoma subgroups. The main focus of the article is MRI features that correlate with medulloblastoma subtypes, as well as features suggestive of molecular subgroups. Finally, we briefly discuss the latest trends in MRI studies and latest developments in molecular characterization.

Basal Suppression of the Sonic Hedgehog Pathway by the G-Protein-Coupled Receptor Gpr161 Restricts Medulloblastoma Pathogenesis

Sonic hedgehog (Shh) determines cerebellar granule cell (GC) progenitor proliferation and medulloblastoma pathogenesis. However, the pathways regulating GC progenitors during embryogenesis before Shh production by Purkinje neurons and their roles in tumorigenesis remain unclear. The cilium-localized G-protein-coupled receptor Gpr161 suppresses Shh-mediated signaling in the neural tube. Here, bydeleting Gpr161 in mouse neural stem cells orGC progenitors, we establish Gpr161 as a tumor suppressor in Shh subtype medulloblastoma. Irrespective of Shh production in the cerebellum, Gpr161 deletion increased downstream activity of the Shh pathway by restricting Gli3-mediated repression, causing more extensive generation and proliferation of GC progenitors. Moreover, earlier deletion of Gpr161 during embryogenesis increased tumor incidence and severity. GC progenitor overproduction during embryogenesis from Gpr161 deletion was cilium dependent, unlike normal development. Low GPR161 expression correlated with poor survival ofSHH subtype medulloblastoma patients. Gpr161 restricts GC progenitor production by preventing premature and Shh-dependent pathway activity, highlighting the importance of basal pathway suppression in tumorigenesis.

Inhibition of polo-like kinase 4 (PLK4): a new therapeutic option for rhabdoid tumors and pediatric medulloblastoma.

Rhabdoid tumors (RT) are highly aggressive and vastly unresponsive embryonal tumors. They are the most common malignant CNS tumors in infants below 6 months of age. Medulloblastomas (MB) are embryonal tumors that arise in the cerebellum and are the most frequent pediatric malignant brain tumors. Despite the advances in recent years, especially for the most favorable molecular subtypes of MB, the prognosis of patients with embryonal tumors remains modest with treatment related toxicity dreadfully high. Therefore, new targeted therapies are needed.The polo-like kinase 4 (PLK4) is a critical regulator of centriole duplication and consequently, mitotic progression. We previously established that PLK4 is overexpressed in RT and MB. We also demonstrated that inhibiting PLK4 with a small molecule inhibitor resulted in impairment of proliferation, survival, migration and invasion of RT cells.Here, we showed in MB the same effects that we previously described for RT. We also demonstrated that PLK4 inhibition induced apoptosis, senescence and polyploidy in RT and MB cells, thereby increasing the susceptibility of cancer cells to DNA-damaging agents. In order to test the hypothesis that PLK4 is a CNS druggable target, we demonstrated efficacy with oral administration to an orthotropic xenograft model.Based on these results, we postulate that targeting PLK4 with small-molecule inhibitors could be a novel strategy for the treatment of RT and MB and that PLK4 inhibitors (PLK4i) might be promising agents to be used solo or in combination with cytotoxic agents.

TMOD-12. AN ONCOLYTIC MEASLES VIRUS SENSITIVE GROUP 3 MEDULLOBLASTOMA MODEL IN IMMUNE-COMPETENT MICE

Modified measles virus (MV) has demonstrated oncolytic efficacy in xenograft models of multiple tumor types using human cells in immune-deficient mice. However, MV does not grow in murine cells, thus, no murine tumor cell line that is oncogenic, grows in immune-competent mice, and is infectable by MV exists. The lack of such a model prevents an examination of the effect of anti-MV immunity and other possible effects of the immune system on MV oncotherapy. We have developed a murine Group 3 medulloblastoma cell line that allows replication of, and is killed by, MV in-vivo. Cerebellar stem cells isolated from P6 human CD46-transgenic immune-competent mice were transduced to express Sendai virus C-protein, c-Myc-Td tomato and GFI1B-GFP proteins. The cell line stably expresses higher levels of C-myc and GFI1B proteins but identical levels of N-myc and p53 when compared to unmodified cerebellar stem cells. Measles virus infects and replicates in the cell line, and cells are killed in a dose and time dependent manner with 55%, 85% and 92% cell death at MOI of 0.1, 1 and 10 respectively after 96h of MV infection. The cells express stem cell makers and NPR3, an established marker for the group 3 subtype of medulloblastoma. This cell line is highly tumorigenic when orthotopically implanted into the brain of syngeneic immune-competent mice. A single intratumoral injection of MV results in significant prolongation of median survival compared to mice injected with heat-killed virus (P=0.0027). Sixty percent of MV-treated mice did not die during the period of study and showed no evidence of tumor at autopsy. Autopsy of brains from inactivated virus treated animals revealed tumors resembling group 3 medulloblastoma in all mice. This is the first cell line and tumor model of any type available for testing the effect of an intact immune system on MV therapy.

MiR-449a is a potential epigenetic biomarker for WNT subtype of medulloblastoma

Objective: To identify the candidate epigenetic biomarkers of Wnt subtype of medulloblastoma(MB). Methods: MicroRNAs(miRNAs) expression array was used to detect the expression of miRNAs in MB cell lines with or without treatment by demethylation reagent. Nanostring gene expression array was used to detect the expression level of mRNA in 45 samples of primary MB. Molecular subtyping was performed based on the NanoString data. The status of methylation was confirmed by methylation specific PCR. The expression of candidate miRNA was confirmed by real-time PCR. Results: All 45 MBs except one were classified into the four molecular groups: 4 in WNT group, 8 in SHH group, 16 were in Group3 and 16 in Group4. Methylation specific PCR (MSP) assay confirmed miR-449a was silenced due to aberrant DNA methylation in MB cell lines.WNT subtype of MBs showed relatively higher expression of miR-449a comparing with other subgroups. Conclusion: MiR-449a, a candidate tumor suppressor gene regulated by hypermethylation, is a novel potential epigenetic marker for WNT subtype of MBs.

Integrative Genomics Characterize Medulloblastoma Subtypes

Abstract Integrative genomic analysis characterized the genomic landscapes of medulloblastoma subtypes.