Abstract
Of nine ependymoma molecular groups detected by DNA methylation profiling, the posterior fossa type A (PFA) is most prevalent. We used DNA methylation profiling to look for further molecular heterogeneity among 675 PFA ependymomas. Two major subgroups, PFA-1 and PFA-2, and nine minor subtypes were discovered. Transcriptome profiling suggested a distinct histogenesis for PFA-1 and PFA-2, but their clinical parameters were similar. In contrast, PFA subtypes differed with respect to age at diagnosis, gender ratio, outcome, and frequencies of genetic alterations. One subtype, PFA-1c, was enriched for 1q gain and had a relatively poor outcome, while patients with PFA-2c ependymomas showed an overall survival at 5years of > 90%. Unlike other ependymomas, PFA-2c tumors express high levels of OTX2, a potential biomarker for this ependymoma subtype with a good prognosis. We also discovered recurrent mutations among PFA ependymomas. H3 K27M mutations were present in 4.2%, occurring only in PFA-1 tumors, and missense mutations in an uncharacterized gene, CXorf67, were found in 9.4% of PFA ependymomas, but not in other groups. We detected high levels of wildtype or mutant CXorf67 expression in all PFA subtypes except PFA-1f, which is enriched for H3 K27M mutations. PFA ependymomas are characterized by lack of H3 K27 trimethylation (H3 K27-me3), and we tested the hypothesis that CXorf67 binds to PRC2 and can modulate levels of H3 K27-me3. Immunoprecipitation/mass spectrometry detected EZH2, SUZ12, and EED, core components of the PRC2 complex, bound to CXorf67 in the Daoy cell line, which shows high levels of CXorf67 and no expression of H3 K27-me3. Enforced reduction of CXorf67 in Daoy cells restored H3 K27-me3 levels, while enforced expression of CXorf67 in HEK293T and neural stem cells reduced H3 K27-me3 levels. Our data suggest that heterogeneity among PFA ependymomas could have clinicopathologic utility and that CXorf67 may have a functional role in these tumors.
Highlights
Ependymomas are neuroepithelial tumors of the central nervous system (CNS), presenting in both adults and children, but accounting for almost 10% of all pediatric CNS tumors and up to 30% of CNS tumors in children under three years [4,28,40]
Further heterogeneity among posterior fossa type A (PFA)-1 and PFA-2 ependymomas was observed with both types of clustering analysis; in total, we discovered nine minor subtypes, six within PFA-1 and three within PFA-2 (Fig. 1)
The present study aimed to discover molecular heterogeneity of potential clinical and biological relevance among PFA ependymomas, and DNA methylation profiling yielded two subgroups and nine subtypes
Summary
Ependymomas are neuroepithelial tumors of the central nervous system (CNS), presenting in both adults and children, but accounting for almost 10% of all pediatric CNS tumors and up to 30% of CNS tumors in children under three years [4,28,40]. Among PF ependymomas, two of three molecular groups, PFA (PF-EPN-A) and PFB (PF-EPN-B), account for most tumors; PF-SE tumors are rare, mostly showing the morphology of a subependymoma [32]. PFA tumors are found mainly in infants and young children (median age ≈ 3yrs) and have a relatively poor outcome, while PFB tumors are generally found in young adults (median age ≈ 30yrs) and are associated with a better prognosis [32,52]. Two observations informed the hypotheses behind our experiments: (i) two robust subgroups of PFA ependymomas emerged in a pilot study when a series of infant ependymomas from the St. Jude RT1 trial were analyzed by DNA methylation profiling and gene expression profiling [37], and (ii) a review of published sequencing datasets for PF ependymomas revealed recurrent single nucleotide variants (SNVs) in an uncharacterized gene, CXorf, which is expressed at high levels in PFA ependymomas [34]. Through a collaboration involving the DNA methylation profiling of 675 PFA ependymomas, we tested two broad hypotheses: (i) PFA ependymomas are heterogeneous and composed of subgroups with clinical utility and (ii) CXorf is implicated in the pathogenesis of PFA ependymoma
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