Laveniya Satgunaseelan

BeadChip array‐based DNA methylation profiling has been recognized by the World Health Organization (WHO) as a key diagnostic tool for brain tumor classification. While its diagnostic utility has been well established, data on technical reproducibility, interlaboratory comparability, and data interpretation under real‐world diagnostic conditions remain limited. Bridging this gap, we here report the results of an international proficiency test using the Infinium MethylationEPIC v2.0 platform and the corresponding Brain Tumor Classifier version 12.8. Tissue slides of eight FFPE brain tumor samples, covering a representative range of CNS tumor entities, were distributed among 24 laboratories in 10 different countries. Participants were asked to report methylation classes and copy number variation (CNV) profiles. Pre‐array workflows were left to local procedures and results had to be submitted within 15 working days. Technical data reproducibility was high with a median pairwise beta‐value correlation of 0.99 (range 0.93–1.0). In general, participating centers generated high‐quality data, reflected by consistently low detection p ‐values (<0.01). Eighteen of the 24 participating centers (75%) successfully passed the test. Of the six centers that failed the test, two laboratories experienced technical issues that led to misclassification of individual cases and contributed to incorrect CNV reporting. Four additional centers showed substantial discrepancies in the interpretation of diagnostically highly relevant CNVs, whereas methylation classification was not impaired. While accurate DNA quantification proved to be an important pre‐array step, the use of the DNA restoration kit had only minor influence on overall results. Taken together, our interlaboratory performance testing on EPIC v2.0 CNS tumor profiling confirms high reproducibility of tumor classification but reveals the need for harmonized CNV reporting.

Map demonstrating the countries represented by contributing institutions.

Long-term survivors ≥ 5 years (n = 12)

Patients with G34V mutation subtype (n = 9)

Initial chemotherapy combinations

Kaplan-Meier survival plots demonstrating overall survival. (A). Initial chemotherapy (all ages): TMZ-based vs TMZ/CCNU. (B). Initial chemotherapy (under 18 years old): TMZ-based vs TMZ/CCNU. (C). Re-irradiation: yes vs no. (D). ATRX mutation: yes vs no. (E). PDGFRA amplification: yes vs no. (F) TP53 mutation: yes vs no. (G). PDGFRA mutation: yes vs no.

Purpose: Knowledge of prognostic factors and long-term survival in patients with diffuse hemispheric glioma, H3 G34–mutant (DHG, H3 G34), remains limited in this tumor with a poor prognosis.Experimental Design:This retrospective, multi-institutional study investigated prognostic variables for patients with DHG, H3 G34, and their association with progression-free survival (PFS) and overall survival (OS). Uni- and multivariable Cox proportional hazard models were applied with multiple imputed datasets.Results:A total of 153 patients (142 G34R, 9 G34V, 2 via DNA methylation) were included. The median age at diagnosis was 17 years (range, 2–45). Initial gross/near total resection (GTR/NTR) was achieved in 43% of patients. Radiation was given in 91% (85% focal irradiation), and initial chemotherapy was given in 87% [70% temozolomide-based (TMZ), 25% TMZ/lomustine, 5% non-TMZ]. Median OS was 24 months [interquartile range (IQR), 22–28] with a median PFS of 14 months (IQR, 12–19). Twelve patients (8%) were found to be long-term survivors (≥5 years). Exploratory multivariable analysis showed that adjuvant radiotherapy [HR, 0.076; 95% confidence interval (CI), 0.033–0.17] and achieving GTR/NTR compared with < NTR (HR, 0.51; 95% CI, 0.33–0.78) were associated with improved PFS. Multivariable analysis showed improved OS with increasing age at diagnosis (HR, 0.70; 95% CI, 0.57–0.87), initial radiotherapy (HR, 0.38; 95% CI, 0.15–0.96), and initial GTR/NTR compared with < NTR (HR, 0.60; 95% CI, 0.37–0.97).Conclusions:This cohort highlights prognostic factors for patients with DHG, H3 G34, and describes relapse patterns and therapy approaches. Clinical trials and prospective registries are needed to improve outcomes.