P10.07.B ADVANCING DIAGNOSTIC CRITERIA AND CLINICAL INSIGHTS IN PEDIATRIC HIGH-GRADE GLIOMAS, H3 WILDTYPE AND IDH WILDTYPE

Abstract BACKGROUND Diffuse pediatric-type high-grade gliomas, characterised by H3 wildtype and IDH wildtype status, represent a recently recognised and highly aggressive brain tumour entity with distinct molecular and epigenetic signatures. Although incorporated into the WHO CNS5 classification, considerable histopathological and molecular heterogeneity within this group continues to challenge accurate diagnosis and complicates therapeutic decision making. MATERIAL AND METHODS Methylation profiling of 70 samples, including 61 diffuse high grade gliomas (CNS WHO Grade 4, NOS) and 9 controls, was combined with targeted next-generation sequencing (NGS). This integrative approach, alongside clinical, radiological, histopathological, and immunohistochemical data, provided a refined molecular and phenotypic characterization of pediatric high grade gliomas without H3 and IDH mutations, enhancing diagnostic precision and offering key insights into the tumours biology. RESULTS Methylation profiling classified 19.7% (12/61) of cases as Diffuse Pediatric Type High Grade Glioma, H3 and IDH wildtype. Among these, 58.3% (7/12) were classified as pedHGG RTK1, with NGS data revealing PDGFRA alterations and mutations in genes such as NOTCH, MYCN, and TSC1, along with activation of key pathways like RAS/MAPK and PI3K/AKT. The remaining cases included 25% (3/12) as pedHGG RTK2, with alterations in EGFR, FGFR1, PIK3CA, PTCH1, CDKN2C, and components of the RAS/MAPK and PI3K/AKT pathways, and one each as pedHGG MYCN and Diffuse pedHGG subtype B. Notably, loss of H3K27me3 expression was identified in 42% (5/12) of cases, posing a challenge to the WHO CNS5 classification, which currently considers H3K27me3 retention a defining feature. A majority of RTK1 classified tumours demonstrated either mismatch repair deficiency or features of radiation-induced gliomas, suggesting a potential enrichment of these biological alterations within this subgroup. Furthermore, MGMT promoter methylation was detected in only 33% of cases, indicating its limited prognostic or predictive value in this molecular context. CONCLUSION Findings highlight the biological and molecular complexity of pediatric high grade gliomas, H3 wildtype and IDH wildtype, revealing significant gaps in current diagnostic frameworks. They highlight the essential role of molecular diagnostics, particularly methylation profiling, sequencing, and mismatch repair deficiency markers, in improving tumour classification and identifying clinically relevant subgroups. The observed heterogeneity and specific molecular alterations call for a re-evaluation of the WHO classification system to better reflect the biology of this rare and aggressive tumour subtype. A deeper understanding of these molecular features is vital for refining prognostication, guiding targeted therapies, and ultimately improving patient outcomes.