LAUSR

Pediatric supratentorial ependymomas with RELA fusions (RELA-EP) have been identified as a unique novel tumor entity [9, 10]. Fusions between C11orf95 and RELA pathologically activate the NFκB signaling pathway indicated by nuclear accumulation of p65-RelA. Deletions of CDKN2A encoding the negative cell-cycle regulator p16 have been described in a subset of supratentorial ependymomas, associated with worse outcome [2, 5, 7]. We assessed the frequency and prognostic impact of CDKN2A deletions in a cohort of 54 RELA-EP in children treated according to HIT2000-E protocols (for detailed demographic information, see supplementary materials and methods and supplementary table 1). High-resolution, genome-wide copy number profiles were generated by molecular inversion probe (MIP) assay. Chromosomal breaks were identified within the C11orf95 and RELA genes corresponding to fusion transcripts (Fig. 1a, d). All cases showed pathological nuclear accumulation of p65-RelA as a hallmark of RELA-EP (Fig. 1f). Homozygous deletion (complete loss) of CDKN2A was detected in 9 of 54 (16.7%) cases (Fig. 1c); and 8 of these (88.9%) showed a concordant complete loss of p16 protein (Fig. 1g). In one case, few tumor cells expressed p16 protein indicating retained CDKN2A alleles in single cells. Fourteen cases (25.9%) harbored a hemizygous deletion of CDKN2A. In these, p16 protein was retained in 92.9% of cases tested— one case lacked p16 protein expression indicating the inactivation of the second allele by alternative mechanisms. Thirty-one of 54 cases (57.4%) had no deletion of CDKN2A; all showed p16 protein expression (Fig. 1h). Immunohistochemistry for p16, therefore, may serve as a surrogate for complete CDKN2A loss, but cannot differentiate between hemizygous and wild-type status. There was no statistical association between CDKN2A deletions and mitotic activity as previously described in IDH-mutant glioma [1]. The Electronic supplementary material The online version of this article (https ://doi.org/10.1007/s0040 1-020-02169 -z) contains supplementary material.