Purpose High-grade astrocytoma with piloid features (HGAP) is a recently described brain tumor entity defined by a specific DNA methylation profile. HGAP has been proposed to be integrated in the upcoming World Health Organization classification of central nervous system tumors expected in 2021. In this series, we present the first single-center experience with this new entity. Methods During 2017 and 2020, six HGAP were identified. Clinical course, surgical procedure, histopathology, genome-wide DNA methylation analysis, imaging, and adjuvant therapy were collected. Results Tumors were localized in the brain stem (n = 1), cerebellar peduncle (n = 1), diencephalon (n = 1), mesencephalon (n = 1), cerebrum (n = 1) and the thoracic spinal cord (n = 2). The lesions typically presented as T1w hypo- to isointense and T2w hyperintense with inhomogeneous contrast enhancement on MRI. All patients underwent initial surgical intervention. Three patients received adjuvant radiochemotherapy, and one patient adjuvant radiotherapy alone. Four patients died of disease, with an overall survival of 1.8, 9.1, 14.8 and 18.1 months. One patient was alive at the time of last follow-up, 14.6 months after surgery, and one patient was lost to follow-up. Apart from one tumor, the lesions did not present with high grade histology, however patients showed poor clinical outcomes. Conclusions Here, we provide detailed clinical, neuroradiological, histological, and molecular pathological information which might aid in clinical decision making until larger case series are published. With the exception of one case, the tumors did not present with high-grade histology but patients still showed short intervals between diagnosis and tumor progression or death even after extensive multimodal therapy.

2020) Neuropathology and Applied Neurobiology 46, 28-47 DNA methylation-based classification of paediatric brain tumours DNA methylation-based machine learning algorithms represent powerful diagnostic tools that are currently emerging for several fields of tumour classification. For various reasons, paediatric brain tumours have been the main driving forces behind this rapid development and brain tumour classification tools are likely further advanced than in any other field of cancer diagnostics. In this review, we will discuss the main characteristics that were important for this rapid advance, namely the high clinical need for improvement of paediatric brain tumour diagnostics, the robustness of methylated DNA and the consequential possibility to generate high-quality molecular data from archival formalin-fixed paraffin-embedded pathology specimens, the implementation of a single array platform by most laboratories allowing data exchange and data pooling to an unprecedented extent, as well as the high suitability of the data format for machine learning. We will further discuss the four most central output qualities of DNA methylation profiling in a diagnostic setting (tumour classification, tumour sub-classification, copy number analysis and guidance for additional molecular testing) individually for the most frequent types of paediatric brain tumours. Lastly, we will discuss DNA methylation profiling as a tool for the detection of new paediatric brain tumour classes and will give an overview of the rapidly growing family of new tumours identified with the aid of this technique.

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Pituicytoma (PITUI), granular cell tumor (GCT), and spindle cell oncocytoma (SCO) are rare tumors of the posterior pituitary. Histologically, they may be challenging to distinguish and have been proposed to represent a histological spectrum of a single entity. We performed targeted next-generation sequencing, DNA methylation profiling, and copy number analysis on 47 tumors (14 PITUI; 12 GCT; 21 SCO) to investigate molecular features and explore possibilities of clinically meaningful tumor subclassification. We detected two main epigenomic subgroups by unsupervised clustering of DNA methylation data, though the overall methylation differences were subtle. The largest group (n = 23) contained most PITUIs and a subset of SCOs and was enriched for pathogenic mutations within genes in the MAPK/PI3K pathways (12/17 [71%] of sequenced tumors: FGFR1 (3), HRAS (3), BRAF (2), NF1 (2), CBL (1), MAP2K2 (1), PTEN (1)) and two with accompanying TERT promoter mutation. The second group (n = 16) contained most GCTs and a subset of SCOs, all of which mostly lacked identifiable genetic drivers. Outcome analysis demonstrated that the presence of chromosomal imbalances was significantly associated with reduced progression-free survival especially within the combined PITUI and SCO group (p = 0.031). In summary, we observed only subtle DNA methylation differences between posterior pituitary tumors, indicating that these tumors may be best classified as subtypes of a single entity. Nevertheless, our data indicate differences in mutation patterns and clinical outcome. For a clinically meaningful subclassification, we propose a combined histo-molecular approach into three subtypes: one subtype is defined by granular cell histology, scarcity of identifiable oncogenic mutations, and favorable outcome. The other two subtypes have either SCO or PITUI histology but are segregated by chromosomal copy number profile into a favorable group (no copy number changes) and a less favorable group (copy number imbalances present). Both of the latter groups have recurrent MAPK/PI3K genetic alterations that represent potential therapeutic targets.

Diffuse paediatric-type high-grade glioma, H3-wildtype and IDH-wildtype (pHGG) is a rare and aggressive brain tumor characterized by a specific DNA methylation profile. It was recently introduced in the 5th World Health Organization classification of central nervous system tumors of 2021. Clinical data on this tumor is scarce. This is a case series, which presents the first clinical experience with this entity. We compiled a retrospective case series on pHGG patients treated between 2015 and 2022 at our institution. Data collected include patients’ clinical course, surgical procedure, histopathology, genome-wide DNA methylation analysis, imaging and adjuvant therapy. Eight pHGG were identified, ranging in age from 8 to 71 years. On MRI tumors presented with an unspecific intensity profile, T1w hypo- to isointense and T2w hyperintense, with inhomogeneous contrast enhancement, often with rim enhancement. Three patients died of the disease, with overall survival of 19, 28 and 30 months. Four patients were alive at the time of the last follow-up, 4, 5, 6 and 79 months after the initial surgery. One patient was lost to follow-up. Findings indicate that pHGG prevalence might be underestimated in the elderly population.

Background DNA methylation profiling has emerges as a powerful approach to CNS tumor classification and the discovery of novel, molecularly distinct entities. With the release of the 12.5 version of the Heidelberg Brain Tumor Classifier, some unclassifiable cases can be assigned to novel methylation classes. We retrospectively reviewed our databases and identified 16 previously unclassifiable cases, all of which belong to the provisional methylation class “adult-type diffuse high-grade glioma, IDH-wildtype, subtype F (HGG_F)”. Material and Methods We clinically, radiologically and morphologically characterized 16 HGG_F cases and compared them to 347 glioblastomas. We additionally analyzed copy-number alterations and performed DNA exome sequencing. Results Median age at diagnosis of the 12 males and 4 females was 65 years. Upon initial diagnostic workup, specimens were classified as CNS tissue with reactive changes (n=3) or suspicious for the infiltration zone of a diffuse glioma (n = 13). None of the cases demonstrated endothelial proliferation or necrosis and 10/16 tumors had flat copy number profiles. Radiological characteristics were reminiscent of gliomatosis cerebri in eight cases and 9/9 cases had normal FET-PET scans. Whole-exome sequencing revealed genetic alterations frequently found in IDH-wildtype glioblastomas, including TERT promoter mutations in 11/14 (78.6%) and PIK3 mutations (10/14, 71.4%). Outcome was significantly better compared to TCGA IDH-wildtype glioblastomas with a median progression-free survival of 58 months and overall survival of 73 months (both p<0.001). Conclusion We provide evidence that TERT promoter mutations in diffusely infiltrating gliomas without further morphological or molecular signs of high-grade glioma should be interpreted in the context of the clinico-radiological presentation as well as epigenetic prolife and may not be suitable as standalone diagnostic marker for glioblastoma, IDH wildtype.

Background Sarcomas are a heterogeneous group of rare malignant tumors with more than 100 subtypes. Accurate diagnosis remains challenging due to a lack of characteristic molecular or histomorphological hallmarks. A DNA methylation-based tumor profiling classifier for sarcomas (known as sarcoma classifier) from the German Cancer Research Center (Deutsches Krebsforschungszentrum) is now employed in selected cases to guide tumor classification and treatment decisions at our institution. Data on the usage of the classifier in daily clinical routine are lacking. Methods In this single-center experience, we describe the clinical course of five sarcoma cases undergoing thorough pathological and reference pathological examination as well as DNA methylation-based profiling and their impact on subsequent treatment decisions. We collected data on the clinical course, DNA methylation analysis, histopathology, radiological imaging, and next-generation sequencing. Results Five clinical cases involving DNA methylation-based profiling in 2021 at our institution were included. All patients’ DNA methylation profiles were successfully matched to a methylation profile cluster of the sarcoma classifier’s dataset. In three patients, the classifier reassured diagnosis or aided in finding the correct diagnosis in light of contradictory data and differential diagnoses. In two patients with intracranial tumors, the classifier changed the diagnosis to a novel diagnostic tumor group. Conclusions The sarcoma classifier is a valuable diagnostic tool that should be used after comprehensive clinical and histopathological evaluation. It may help to reassure the histopathological diagnosis or indicate the need for thorough reassessment in cases where it contradicts previous findings. However, certain limitations (non-classifiable cases, misclassifications, unclear degree of sample purity for analysis and others) currently preclude wide clinical application. The current sarcoma classifier is therefore not yet ready for a broad clinical routine. With further refinements, this promising tool may be implemented in daily clinical practice in selected cases.