Genomic and Transcriptomic Analyses Reveals ZNF124 as a Critical Regulator in Highly Aggressive Medulloblastomas

Medulloblastoma (MB) is the most common malignant pediatric brain tumor, however, the mechanisms underlying tumorigenesis in different MB subgroups remain incompletely understood. Although previous studies of MB predisposition have been conducted in tertiary referral centers primarily in Caucasian cohorts, it is not unclear clear whether there exist population-specific genetic alterations in MBs. In this study, we investigated the contribution of genomic and transcriptomic alterations to the risk of malignant MB in the Chinese population (designated as the Asian cohort). We analyze the genomic and transcriptomic alterations of the Asian MB cohort by using a combination of whole-exome sequencing (WES) and RNA-deep-sequencing. In addition, we integrate publicly available data with the Asian MB cohort and identify a subset of potential MB-driving genes specifically enriched in each of the MB subgroups. We further characterize a newly identified group-3-enriched transcriptional regulator, ZNF124, and demonstrate that ZNF124 is critical for the growth of the most aggressive group-3 MB cells. Together, our analyses indicate conserved yet distinct genetic alterations and gene expression patterns of MBs between different ethnic groups. Our studies further provide an important resource for identifying potential tumor-driving factors in MBs, enhancing our understanding of the disease process for developing ethnically targeted therapies in patients with MB.

Download Full-text

TBIO-15. MODELING DEVELOPMENTAL GENE EXPRESSION DYNAMICS AT CELLULAR RESOLUTION TO INTERPRET PEDIATRIC BRAIN TUMOR TRANSCRIPTIONAL PROGRAMS

Neuro-Oncology ◽

10.1093/neuonc/noaa222.842 ◽

2020 ◽

Vol 22 (Supplement_3) ◽

pp. iii469-iii469

Author(s):

Selin Jessa ◽

Nisha Kabir ◽

Maria Vladoiu ◽

Steven Hébert ◽

Michael D Taylor ◽

...

Keyword(s):

Brain Tumor ◽

Single Cell ◽

Time Course ◽

Expression Patterns ◽

Pediatric Brain Tumor ◽

Cell Types ◽

Pediatric Brain Tumors ◽

Developmental Time ◽

Genetic Alterations ◽

Pediatric Brain

Abstract A central challenge in understanding the biology of pediatric brain tumors is defining the cellular and molecular context where oncogenesis occurs. We hypothesize that spatiotemporally restricted cell types are uniquely susceptible to specific genetic alterations, which alter normal neurodevelopmental programs and ultimately lead to oncogenesis. The resulting tumors retain some transcriptomic features of their lineage of origin. To delineate these origins, we assembled a densely sampled developmental time course of the mouse forebrain and pons, doubling our recently published single-cell atlas. This dataset comprises >100,000 cells at 9 timepoints from E10-P6. However, while single cell transcriptomics reveal rich gene dynamics during cell differentiation, interpretation of individual genes can be challenging due to data sparsity. Leveraging this time-series, we present strategies to model and visualize the expression of a given gene across differentiation of distinct lineages. We demonstrate an interactive web app to interrogate the expression of genes or gene sets during brain development, extract temporally correlated genes, and search active transcription factor regulatory modules. Finally, we profile the expression of core transcriptional programs of several pediatric brain tumor entities during development. Our analyses reveal genes with restricted expression patterns that elucidate tumor etiology. More broadly, these resources harness single cell data to enable exploration of neurodevelopmental gene programs with great relevance to pediatric brain tumor oncogenesis.

Download Full-text

EMBR-07. MYC BUT NOT MYCN GENERATES AGGRESSIVE GROUP 3 MEDULLOBLASTOMA BY ARF PATHWAY SUPPRESSION

Neuro-Oncology ◽

10.1093/neuonc/noab090.025 ◽

2021 ◽

Vol 23 (Supplement_1) ◽

pp. i7-i7

Author(s):

Oliver Mainwaring ◽

Holger Weishaupt ◽

Sonja Hutter ◽

Miao Zhao ◽

Gabriela Rosén ◽

...

Keyword(s):

Tumor Development ◽

Glutamate Transporter ◽

Pediatric Brain Tumor ◽

P53 Gene ◽

Hsp90 Inhibitor ◽

Hsp90 Inhibition ◽

Group 3 ◽

Pediatric Brain ◽

Hsp90 Protein

Abstract Medulloblastoma (MB), the most common malignant pediatric brain tumor, often harbor MYC amplifications and arise in the presence of a functional p53 suppressor protein. To elucidate the mechanism behind this inexplicable tumor development we generated an inducible, immunocompetent transgenic mouse model of MYC-driven MB. Tumors driven from the glutamate transporter promoter molecularly resembled aggressive Group 3 MB driven by an enriched photoreceptor program. They developed embryonically in a monoclonal fashion in the presence of a functional unmutated p53 gene. Compared to MYCN-expressing MB driven from the same promoter, we discovered pronounced silencing of the ARF suppressor upstream of p53. We similarly found significant methylation of the ARF promoter in MYC-amplified as compared to MYCN-amplified human MB samples. While MYCN-driven tumor malignancy was more sensitive to ARF depletion, it dramatically increased metastatic spread of MYC-driven tumors. DNMT inhibition could restore ARF levels in MYC-expressing tumors but did not show any therapeutic advantage in tumors in vivo. Computational modeling suggested the HSP90 protein to act as a more specific target and ARF could indeed be restored by the HSP90 inhibitor onalespib that promoted increased survival in our inducible animal model suggesting that HSP90 inhibition could be potentially used in patients affected by MYC-driven ARF-silenced cancer.

Download Full-text

IMMU-48. IMMUNOMODULATION WITH RESIQUIMOD REPOLARIZES THE IMMUNE MICROENVIRONMENT TO INHIBIT MEDULLOBLASTOMA PROGRESSION

Neuro-Oncology ◽

10.1093/neuonc/noab196.407 ◽

2021 ◽

Vol 23 (Supplement_6) ◽

pp. vi103-vi103

Author(s):

Christopher Park ◽

Coral Del Mar ◽

Morrent Thang ◽

Taylor Dismuke ◽

Duhyeong Hwang ◽

...

Keyword(s):

Tumor Growth ◽

Tumor Progression ◽

Median Survival ◽

Sonic Hedgehog ◽

Expression Patterns ◽

Myeloid Cells ◽

Pediatric Brain Tumor ◽

Toll Like Receptor ◽

Paracrine Signaling ◽

Pediatric Brain

Abstract BACKGROUND New, non-cytotoxic treatments may improve outcomes for medulloblastoma, the most common malignant pediatric brain tumor. Sonic hedgehog (SHH) subgroup medulloblastoma, which includes subtypes with poor prognosis, can be modeled in immunocompetent, transgenic mice. These models are ideal for preclinical testing of immunotherapies. Here, we show that immune stimulation using resiquimod, a small molecule agonist of Toll-like receptor 7 and 8 (TLR 7/8), alters myeloid populations in medulloblastoma and significantly reduces tumor growth. METHOD We generated mice with medulloblastoma by interbreeding the hGFAP-Cre and SmoM2 mouse lines. The resultant hGFAP-Cre/SmoM2 (G-Smo) mice develop SHH medulloblastoma with 100% frequency. We analyzed myeloid populations and demonstrated TLR7/8 expression patterns in G-Smo tumors. We then compared survival of untreated G-Smo mice versus G-Smo mice treated with three doses of resiquimod at postnatal days 10, 12, and 14. We also assessed pharmacodynamic effects at progressive intervals after a single dose. RESULTS Approximately 10% of cells in G-Smo medulloblastomas were myeloid cells, and these cells were the only cells that expressed TLR7/8. Resiquimod slowed tumor growth and increased the survival of mice with medulloblastoma. Untreated median survival was 14.5 days (n=12), compared to resiquimod-treated median survival of 37 days (n=10; p=0.0003). All treated mice eventually demonstrated tumor progression. Immunohistochemistry for IBA1, a pan-macrophage marker, demonstrated significant increase in myeloid cells within the tumor by 24 hours after treatment (p=0.0178), however the IGF1+ fraction of myeloid cells decreased (p=0.0275). CONCLUSION Resiquimod prolongs survival in mice with SHH-driven medulloblastoma, demonstrating the potential for therapies that target myeloid cells to produce significant anti-tumor effects. Myeloid-derived IGF-1 has been shown to support tumor progression and resiquimod may act by disrupting this paracrine signaling.

Download Full-text

TMOD-30. IDENTIFYING NEW DRIVERS OF GROUP 3 MEDULLOBLASTOMA

Neuro-Oncology ◽

10.1093/neuonc/noaa215.980 ◽

2020 ◽

Vol 22 (Supplement_2) ◽

pp. ii234-ii234

Author(s):

Meher Beigi Masihi ◽

Catherine Lee ◽

Alexandra Garancher ◽

Grace Furnari ◽

Robert Wechsler-Reya

Keyword(s):

Brain Tumor ◽

Molecular Mechanisms ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Pediatric Brain Tumor ◽

Genetic Alterations ◽

Sleeping Beauty ◽

Tumor Formation ◽

Group 3

Abstract Medulloblastoma (MB) is the most common malignant childhood brain tumor. MB can be divided into four major subgroups – WNT, Sonic hedgehog (SHH), Group 3 (G3-MB), and Group 4 (G4-MB) – that exhibit distinct genetic alterations, gene expression profiles, and clinical outcomes. Patients with G3-MB have the worst prognosis, and a deeper understanding of this form of the disease is critical for development of new therapies. Most G3-MBs express high levels of the MYC oncogene, suggesting that MYC plays an important role in tumorigenesis. However, MYC overexpression is not sufficient to drive tumor formation. To identify genes that cooperate with MYC to promote development of G3-MB, we performed an in vivo mutagenesis screen using mice expressing the Sleeping Beauty (SB) transposon. Cerebellar stem cells isolated from transposon/transposase-expressing transgenic mice were infected with viruses encoding Myc, and these cells were transplanted into the cerebellum of adult hosts. Tumors that arose were subjected to DNA and RNA sequencing to identify candidate genes, and these genes were subjected to functional analysis to determine whether they could cooperate with Myc to drive G3-MB. These studies identified the transcription factor Ras-responsive element binding protein 1 (Rreb1) as a potent Myc-cooperating gene. Tumors driven by Myc and Rreb1 (MR tumors) resemble G3-MB at a histological and molecular level. Moreover, RREB1 is overexpressed in human G3-MB, and knockdown of RREB1 expression impairs growth of G3-MB cell lines and patient-derived xenografts. Ongoing studies are aimed at identifying the molecular mechanisms by which Rreb1 contributes to tumor growth. Our studies demonstrate an important role for RREB1 in G3-MB, and provide a new model that can be used to identify therapeutic targets and develop more effective and less toxic therapies for this devastating pediatric brain tumor.

Download Full-text

TMOD-28. MYC GENERATES AGGRESSIVE MEDULLOBLASTOMA BY HSP90 PATHWAY ACTIVATION AND ARF SILENCING

Neuro-Oncology ◽

10.1093/neuonc/noab196.889 ◽

2021 ◽

Vol 23 (Supplement_6) ◽

pp. vi221-vi221

Author(s):

Oliver Mainwaring ◽

Holger Weishaupt ◽

Sonja Hutter ◽

Miao Zhao ◽

Anna Borgenvik ◽

...

Keyword(s):

Mouse Model ◽

Tumor Development ◽

Pediatric Brain Tumor ◽

P53 Gene ◽

Hsp90 Inhibitor ◽

Pathway Activation ◽

Group 3 ◽

P53 Tumor Suppressor Protein ◽

Pediatric Brain

Abstract Medulloblastoma, the most common malignant pediatric brain tumor, often shows amplification or overexpression of the MYC transcription factor and arises in the presence of a functional p53 tumor suppressor protein. To elucidate the mechanism behind this inexplicable tumor development we generated an inducible, immunocompetent transgenic mouse model of MYC-expressing medulloblastoma. Aggressive tumors developed clonally in the presence of an unaltered p53 gene that molecularly resembled Group 3 medulloblastoma. Compared to MYCN-expressing medulloblastoma driven from the same promoter, we instead discovered pronounced and MIZ1-independent silencing of the ARF suppressor, which was also suppressed in MYC-amplified as compared to MYCN-amplified human medulloblastoma. While MYCN-driven tumor malignancy was more sensitive to ARF depletion, it dramatically increased metastatic spread of MYC-driven tumors. DNMT inhibition could restore ARF levels in MYC-expressing tumors but did not show any therapeutic advantage in tumors in vivo. Bioinformatics analysis further showed a strong correlation of the HSP90 pathway with MYC in human Group 3 MB and in the MYC-driven mouse model. The HSP90 inhibitor Onalespib showed significant selectivity for targeting MYC-driven as compared to MYCN-driven tumors. The drug promoted ARF restoration and increased the survival in our animal model which suggests that it could be potentially used in the treatment of MYC-driven ARF-silenced brain cancer patients.

Download Full-text

In Vivo and Ex Vivo Pediatric Brain Tumor Models: An Overview

Frontiers in Oncology ◽

10.3389/fonc.2021.620831 ◽

2021 ◽

Vol 11 ◽

Author(s):

Zhiqin Li ◽

Sigrid A. Langhans

Keyword(s):

Brain Tumor ◽

Brain Tumors ◽

Drug Development ◽

Pediatric Brain Tumor ◽

Genetic Alterations ◽

Benign Tumors ◽

Tumor Type ◽

Tumor Models ◽

Pediatric Brain

After leukemia, tumors of the brain and spine are the second most common form of cancer in children. Despite advances in treatment, brain tumors remain a leading cause of death in pediatric cancer patients and survivors often suffer from life-long consequences of side effects of therapy. The 5-year survival rates, however, vary widely by tumor type, ranging from over 90% in more benign tumors to as low as 20% in the most aggressive forms such as glioblastoma. Even within historically defined tumor types such as medulloblastoma, molecular analysis identified biologically heterogeneous subgroups each with different genetic alterations, age of onset and prognosis. Besides molecularly driven patient stratification to tailor disease risk to therapy intensity, such a diversity demonstrates the need for more precise and disease-relevant pediatric brain cancer models for research and drug development. Here we give an overview of currently available in vitro and in vivo pediatric brain tumor models and discuss the opportunities that new technologies such as 3D cultures and organoids that can bridge limitations posed by the simplicity of monolayer cultures and the complexity of in vivo models, bring to accommodate better precision in drug development for pediatric brain tumors.

Download Full-text