Abstract
Medulloblastoma (MB) is the most common malignant pediatric brain tumor. While the pathways that are deregulated in MB remain to be fully characterized, amplification and/or overexpression of the MYCN gene, which is has a critical role in cerebellar development as a regulator of neural progenitor cell fate, has been identified in several MB subgroups. Phenotypically, aberrant expression of MYCN is associated with the large-cell/anaplastic MB variant, which accounts for 5-15% of cases and is associated with aggressive disease and poor clinical outcome. To better understand the role of MYCN in MB in vitro and in vivo and to aid the development of MYCN-targeted therapeutics we established tumor-derived neurosphere cell lines from the GTML (Glt1-tTA/TRE-MYCN-Luc) genetically engineered mouse model. A fraction of GTML neurospheres were found to be growth factor independent, expressed CD133 (a marker of neural stem cells), failed to differentiate upon MYCN withdrawal and were highly tumorigenic when orthotopically implanted into the cerebellum. Principal component analyzes using single cell RNA assay data suggested that the clinical candidate aurora-A kinase inhibitor MLN8237 converts GTML neurospheres to resemble non-MYCN expressors. Correlating with this, MLN8237 significantly extended the survival of mice bearing GTML MB allografts. In summary, our results demonstrate that MYCN plays a critical role in expansion and survival of aggressive MB-propagating cells, and establish GTML neurospheres as an important resource for the development of novel therapeutic strategies.
Highlights
In children, medulloblastoma (MB) is the most frequently occurring primitive neuroectodermal tumor (PNET) originating in the brain and has been classified into four major subtypes based on copy number variation and transcription profiles: MBWNT, MBSHH, MBGroup3 and MBGroup4 [1,2]
We found that Nestin, a marker for neural stem/progenitor cells, and the proliferation marker Ki67 were expressed in GTML neurospheres in a MYCN
MYCN, as expected, was barely detectable in M10519 cells treated with dox (Fig. 2B and 2C). These results suggest that MYCN expression results in the expansion of cells with markers typical of neural stem cells and/or progenitors associated with MB tumorigenesis
Summary
Medulloblastoma (MB) is the most frequently occurring primitive neuroectodermal tumor (PNET) originating in the brain and has been classified into four major subtypes based on copy number variation and transcription profiles: MBWNT, MBSHH (sonic hedgehog), MBGroup and MBGroup4 [1,2]. We previously reported a genetically engineered mouse model (GEMM) of MYCN-driven MB (GTML: Glt1-tTA/TREMYCN-Luc) in which MYCN expression is controlled by doxycycline (dox) (Tet-off). Using this system we showed that targeted expression of MYCN induces both classic and large cell anaplastic (LCA) pathology independent of SHH [11]. Neurosphere lines established from GTML mice proliferate robustly, express neuronal markers, and form tumors when orthotopically implanted into the brains of mice [12] These results suggest that GTML neurosphere cultures contain tumorpropagating cells, making this a potentially useful system with which the transformative role of MYCN in MB genesis could be elucidated
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