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. 2023 Oct 3;25(10):1871-1882.
doi: 10.1093/neuonc/noad055.

Optimizing biomarkers for accurate ependymoma diagnosis, prognostication, and stratification within International Clinical Trials: A BIOMECA study

Rebecca J Chapman  1 , David R Ghasemi  2   3   4 , Felipe Andreiuolo  5   6   7 , Valentina Zschernack  5 , Arnault Tauziede Espariat  8   9 , Francesca R Buttarelli  10 , Felice Giangaspero  10   11 , Jacques Grill  9 , Christine Haberler  12 , Simon M L Paine  13 , Ian Scott  13 , Thomas S Jacques  14   15 , Martin Sill  2 , Stefan Pfister  2   3   4 , John-Paul Kilday  16   17 , Pierre Leblond  18 , Maura Massimino  19 , Hendrik Witt  2   3   4 , Piergiorgio Modena  20 , Pascale Varlet  8   9 , Torsten Pietsch  5 , Richard G Grundy  1 , Kristian W Pajtler  1   2   3   4 , Timothy A Ritzmann  1
Affiliations

Optimizing biomarkers for accurate ependymoma diagnosis, prognostication, and stratification within International Clinical Trials: A BIOMECA study

Rebecca J Chapman et al. Neuro Oncol. .

Abstract

Background: Accurate identification of brain tumor molecular subgroups is increasingly important. We aimed to establish the most accurate and reproducible ependymoma subgroup biomarker detection techniques, across 147 cases from International Society of Pediatric Oncology (SIOP) Ependymoma II trial participants, enrolled in the pan-European "Biomarkers of Ependymoma in Children and Adolescents (BIOMECA)" study.

Methods: Across 6 European BIOMECA laboratories, we evaluated epigenetic profiling (DNA methylation array); immunohistochemistry (IHC) for nuclear p65-RELA, H3K27me3, and Tenascin-C; copy number analysis via fluorescent in situ hybridization (FISH) and MLPA (1q, CDKN2A), and MIP and DNA methylation array (genome-wide copy number evaluation); analysis of ZFTA- and YAP1-fusions by RT-PCR and sequencing, Nanostring and break-apart FISH.

Results: DNA Methylation profiling classified 65.3% (n = 96/147) of cases as EPN-PFA and 15% (n = 22/147) as ST-ZFTA fusion-positive. Immunohistochemical loss of H3K27me3 was a reproducible and accurate surrogate marker for EPN-PFA (sensitivity 99%-100% across 3 centers). IHC for p65-RELA, FISH, and RNA-based analyses effectively identified ZFTA- and YAP-fused supratentorial ependymomas. Detection of 1q gain using FISH exhibited only 57% inter-center concordance and low sensitivity and specificity while MIP, MLPA, and DNA methylation-based approaches demonstrated greater accuracy.

Conclusions: We confirm, in a prospective trial cohort, that H3K27me3 immunohistochemistry is a robust EPN-PFA biomarker. Tenascin-C should be abandoned as a PFA marker. DNA methylation and MIP arrays are effective tools for copy number analysis of 1q gain, 6q, and CDKN2A loss while FISH is inadequate. Fusion detection was successful, but rare novel fusions need more extensive technologies. Finally, we propose test sets to guide future diagnostic approaches.

Keywords: Ependymoma; biomarkers; brain tumors; neuro-oncology; paediatric.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1.
Figure 1.
DNA-methylation profiling results. t-SNE plot visualizing DNA-methylation based clustering of the BIOMECA cohort. Central t-SNE: Molecular groups, satellite t-SNEs: molecular subgroup for PFA, ST-ZFTA and PFB. Samples are colorized according to the best available prediction.
Figure 2.
Figure 2.
Evaluation of methods to assess copy number alterations.
Figure 3.
Figure 3.
Evaluation of IHC for H3K27me3 and TNC as potential surrogate PFA markers. Posterior fossa tumors classed as PFA (A) and PFB (B) via DNA-methylation array (score ≥0.9) and the IHC result as assessed per center. (C) H3K27me3 and TNC results in cases not classified by DNA-methylation array. (D) Representative H3K27me3 staining in a PFA (i) and PFB (ii) case. (E) Representative TNC pericellular (i) and perivascular (ii) expression. (D-E) Magnification X40, scale bars 50μm. Representative negative controls (Diii, H3K27me3; Eiii, TNC).
Figure 4.
Figure 4.
Comparison of methods used to assess ZFTA- (A) and YAP1- (B) fusions in supratentorial tumors.
See this image and copyright information in PMC

References

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Supplementary concepts