BCOR::CREBBP fusion in malignant neuroepithelial tumor of CNS expands the spectrum of methylation class CNS tumor with BCOR/BCOR(L1)-fusion.

Methylation class "CNS tumor with BCOR/BCOR(L1)-fusion" was recently defined based on methylation profiling and tSNE analysis of a series of 21 neuroepithelial tumors with predominant presence of a BCOR fusion and/or characteristic CNV breakpoints at chromosome 22q12.31 and chromosome Xp11.4. Clear diagnostic criteria are still missing for this tumor type, specially that BCOR/BCOR(L1)-fusion is not a consistent finding in these tumors despite being frequent and that none of the Heidelberger classifier versions is able to clearly identify these cases, in particular tumors with alternative fusions other than those involving BCOR, BCORL1, EP300 and CREBBP. In this study, we introduce a BCOR::CREBBP fusion in an adult patient with a right temporomediobasal tumor, for the first time in association with methylation class "CNS tumor with BCOR/BCOR(L1)-fusion" in addition to 35 cases of CNS neuroepithelial tumors with molecular and histopathological characteristics compatible with "CNS tumor with BCOR/BCOR(L1)-fusion" based on a comprehensive literature review and data mining in the repository of 23 published studies on neuroepithelial brain Tumors including 7207 samples of 6761 patients. Based on our index case and the 35 cases found in the literature, we suggest the archetypical histological and molecular features of "CNS tumor with BCOR/BCOR(L1)-fusion". We also present four adult diffuse glioma cases including GBM, IDH-Wildtype and Astrocytoma, IDH-Mutant with CREBBP fusions and describe the necessity of complementary molecular analysis in "CNS tumor with BCOR/BCOR(L1)-alterations for securing a final diagnosis.

Treatment response as surrogate to predict risk for disease progression in pediatric medulloblastoma with persistent MRI lesions after first-line treatment.

BACKGROUND: This study aims at clarifying the impact of persistent residual lesions following first-line treatment for pediatric medulloblastoma. METHODS: Data on 84 pediatric patients with medulloblastoma and persistent residual lesions on centrally reviewed MRI at the end of first-line therapy were analyzed. RESULTS: Twenty patients (23.8%) had residual lesions in the tumor bed (R+/M0), 51 (60.7%) had distant lesions (R0/M+) and 13 (15.5%) had both (R+/M+). Overall response to first-line therapy was minor or partial (≥25% reduction, MR/PR) for 64 (76.2%) and stable disease (SD) for 20 patients (23.8%). Five-year post-primary-treatment progression-free (pptPFS) and overall survival (pptOS) were superior after MR/PR (pptPFS: 62.5±7.0%[MR/PR] vs. 35.9±12.8%[SD], p=0.03; pptOS: 79.7±5.9[MR/PR] vs. 55.5±13.9[SD], p=0.04). Further, R+/M+ was associated with a higher risk for progression (5-year pptPFS: 22.9±17.9%[R+,M+] vs. 72.4±12.0%[R+,M0]; p=0.03). Watch-and-wait was pursued in 58 patients, while n=26 received additional treatments (chemotherapy only, n=19; surgery only, n=2; combined, n=3; valproic acid, n=2), and their outcomes were not superior to watch-and-wait (5-year pptPFS: 58.5±7.7% vs. 51.6±10.7% p=0.71; 5-year pptOS: 76.3±6.9% vs. 69.8±9.7%, p=0.74). For the whole cohort, five-year pptPFS by molecular subgroup (58 cases) were WNT: 100%, SHH: 50.0±35.4%, Group-4, 52.5±10.5, Group-3 54.2±13.8%; (p=0.08). CONCLUSION: Overall response and extent of lesions can function as surrogate parameters to predict outcomes in pediatric MB patients with persistent lesions after first-line therapy. Especially in case of solitary persistent medulloblastoma MRI lesion, additional therapy was not beneficial. Therefore, treatment response, extent/kind of residual lesions and further diagnostic information needs consideration for indication of additional treatments for persisting lesions.

Modulated critical currents of spin-transfer torque-induced resistance changes in NiCu/Cu multilayered nanowires.

We present a novel method combining anodic aluminum oxide template synthesis and nanolithography to selectively deposit vertically patterned magnetic nanowires on a Si substrate. With this approach we fabricated three-dimensional nanowire-based spin valve devices without the need of complex etching processes or additional spacer coating. Through this method, we successfully obtained NiCu/Cu multilayered nanowire arrays with a controlled sequence along the long axis of the nanowires. Both magnetic switching and excitation phenomena driven by spin-polarized currents were clearly demonstrated in our NiCu/Cu multilayered nanowires. Moreover, the critical currents for switching and excitation were observed to be modulated in an oscillatory manner by the magnetic field in the nanowire-based devices. We present a toy model to qualitatively explain these observations.

Ependymoma from Benign to Highly Aggressive Diseases: A Review.

Ependymomas comprise biologically distinct tumor types with respect to age distribution, (epi)genetics, localization, and prognosis. Multimodal risk-stratification, including histopathological and molecular features, is essential in these biologically defined tumor types. Gross total resection (GTR), achieved with intraoperative monitoring and neuronavigation, and if necessary, second-look surgery, is the most effective treatment. Adjuvant radiation therapy is mandatory in high-risk tumors and in case of residual tumor. There is yet growing evidence that some ependymal tumors may be cured by surgery alone. To date, the role of chemotherapy is unclear and subject of current studies.Even though standard therapy can achieve reasonable survival rates for the majority of ependymoma patients, long-term follow-up still reveals a high probability of relapse in certain biological entities.With increasing knowledge of biologically distinct tumor types, risk-adapted adjuvant therapy gains importance. Beyond initial tumor control, and avoidance of therapy-induced morbidity for low-risk patients, intensified treatment for high-risk patients comprises another challenge. With identification of specific risk features regarding molecular alterations, targeted therapy may represent an option for individualized treatment modalities in the future.

Pineal anlage tumor: clinical and diagnostic features, and rationales for treatment.

PURPOSE: To provide a treatment-focused review and develop basic treatment guidelines for patients diagnosed with pineal anlage tumor (PAT). METHODS: Prospectively collected data of three patients with pineal anlage tumor from Germany was combined with clinical details and treatment information from 17 published cases. RESULTS: Overall, 20 cases of PAT were identified (3 not previously reported German cases, 17 cases from published reports). Age at diagnosis ranged from 0.3 to 35.0 (median: 3.2 ± 7.8) years. All but three cases were diagnosed before the age of three years. For three cases, metastatic disease at initial staging was described. All patients underwent tumor surgery (gross-total resection: 9, subtotal resection/biopsy: 9, extent of resection unknown: 2). 15/20 patients were alive at last follow-up. Median follow-up for 10/15 surviving patients with available follow-up and treatment data was 2.4 years (0.3-6.5). Relapse was reported for 3 patients within 0.8 years after diagnosis. Five patients died, 3 after relapse and 2 from early postoperative complications. Two-year-progression-free- and -overall survival were 65.2 ± 12.7% and 49.2 ± 18.2%, respectively. All 4 patients who received intensive chemotherapy including high-dose chemotherapy combined with radiotherapy (2 focal, 2 craniospinal [CSI]) had no recurrence. Focal radiotherapy- and CSI-free survival rates in 13 evaluable patients were 46.2% (6/13) and 61.5% (8/13), respectively. CONCLUSION: PAT is an aggressive disease mostly affecting young children. Therefore, adjuvant therapy using intensive chemotherapy and considering radiotherapy appears to comprise an appropriate treatment strategy. Reporting further cases is crucial to evaluate distinct treatment strategies.

Radiation Therapy Plays an Important Role in the Treatment of Atypical Teratoid/Rhabdoid Tumors: Analysis of the EU-RHAB Cohorts and Their Precursors.

PURPOSE: Atypical teratoid/rhabdoid tumor (AT/RT) is a rare malignancy of the central nervous system in young children with a dismal prognosis. Prognostic markers have been extensively investigated but have not been validated. The role of radiation therapy (RT) remains controversial. We evaluated the impact of RT as part of multimodality treatment by analyzing data of a European AT/RT cohort. METHODS AND MATERIALS: We retrospectively analyzed data of the European Registry for Rhabdoid Tumors and its precursors. Primary endpoints were progression-free survival (PFS) and overall survival (OS). Potential impact of prognostic factors was analyzed using univariable and multivariable Cox regression analyses with RT as a time-dependent factor. RESULTS: Data of 186 children (118 male, 68 female) treated from 1990 to 2016 were evaluable. The median age at diagnosis was 1.57 years (range, 0.01-26.70 years); 47% (87/186) of the patients were under the age of 18 months. Sixty-nine percent (128/186) received RT (focal RT, n = 93; craniospinal treatment with local boost, n = 34; spinal irradiation, n = 1). The median follow-up duration of the entire cohort was 1.73 years (range, 0.06-20.11 years). The estimated PFS and OS rates were 48% (95% CI, 41%-55%) and 72% (95% CI, 65%-78%) at 1 year and 33% (95% CI, 26%-40%) and 49% (95% CI, 41%-56%) at 2 years, respectively. On multivariable analysis, RT was an independent significant prognostic factor for PFS (hazard ratio, 0.45; 95% CI, 0.27-0.75; P = .002) and OS (hazard ratio, 0.54; 95% CI, 0.32-0.93; P = .025). CONCLUSIONS: This analysis confirms the relevance of local therapies. RT was an independent prognostic factor for outcomes in children experiencing AT/RT. However, long-term sequelae have to be carefully evaluated and considered given the young age at time of RT.

Improved prognostic stratification of patients with isocitrate dehydrogenase-mutant astrocytoma.

Prognostic factors and standards of care for astrocytoma, isocitrate dehydrogenase (IDH)-mutant, CNS WHO grade 4, remain poorly defined. Here we sought to explore disease characteristics, prognostic markers, and outcome in patients with this newly defined tumor type. We determined molecular biomarkers and assembled clinical and outcome data in patients with IDH-mutant astrocytomas confirmed by central pathology review. Patients were identified in the German Glioma Network cohort study; additional cohorts of patients with CNS WHO grade 4 tumors were identified retrospectively at two sites. In total, 258 patients with IDH-mutant astrocytomas (114 CNS WHO grade 2, 73 CNS WHO grade 3, 71 CNS WHO grade 4) were studied. The median age at diagnosis was similar for all grades. Karnofsky performance status at diagnosis inversely correlated with CNS WHO grade (p < 0.001). Despite more intensive treatment upfront with higher grade, CNS WHO grade was strongly prognostic: median overall survival was not reached for grade 2 (median follow-up 10.4 years), 8.1 years (95% CI 5.4-10.8) for grade 3, and 4.7 years (95% CI 3.4-6.0) for grade 4. Among patients with CNS WHO grade 4 astrocytoma, median overall survival was 5.5 years (95% CI 4.3-6.7) without (n = 58) versus 1.8 years (95% CI 0-4.1) with (n = 12) homozygous CDKN2A deletion. Lower levels of global DNA methylation as detected by LINE-1 methylation analysis were strongly associated with CNS WHO grade 4 (p < 0.001) and poor outcome. MGMT promoter methylation status was not prognostic for overall survival. Histomolecular stratification based on CNS WHO grade, LINE-1 methylation level, and CDKN2A status revealed four subgroups of patients with significantly different outcomes. In conclusion, CNS WHO grade, global DNA methylation status, and CDKN2A homozygous deletion are prognostic in patients with IDH-mutant astrocytoma. Combination of these parameters allows for improved prediction of outcome. These data aid in designing upcoming trials using IDH inhibitors.

Clinically relevant molecular hallmarks of PFA ependymomas display intratumoral heterogeneity and correlate with tumor morphology.

Posterior fossa type A (PF-EPN-A, PFA) ependymoma are aggressive tumors that mainly affect children and have a poor prognosis. Histopathology shows significant intratumoral heterogeneity, ranging from loose tissue to often sharply demarcated, extremely cell-dense tumor areas. To determine molecular differences in morphologically different areas and to understand their clinical significance, we analyzed 113 PF-EPN-A samples, including 40 corresponding relapse samples. Cell-dense areas ranged from 0 to 100% of the tumor area and displayed a higher proportion of proliferating tumor cells (p < 0.01). Clinically, cell density was associated with poor progression-free and overall survival (pPFS = 0.0026, pOS < 0.01). Molecularly, tumor areas with low and high cell density showed diverging DNA methylation profiles regarding their similarity to distinct previously discovered PF-EPN-A subtypes in 9/21 cases. Prognostically relevant chromosomal changes at 1q and 6q showed spatial heterogeneity within single tumors and were significantly enriched in cell-dense tumor areas as shown by single-cell RNA (scRNA)-sequencing as well as copy number profiling and fluorescence in situ hybridization (FISH) analyses of different tumor areas. Finally, spatial transcriptomics revealed cell-dense areas of different tumors to be more similar than various different areas of the same tumor. High-density areas distinctly overexpressed genes encoding histone proteins, WNT5A, TGFB1, or IGF2. Relapsing tumors displayed a higher proportion of cell-dense areas (p = 0.036), a change in PF-EPN-A methylation subtypes (13/32 patients), and novel chromosome 1q gains and 6q losses (12/32 cases) compared to corresponding primary tumors. Our data suggest that PF-EPN-A ependymomas habor a previously unrecognized intratumoral heterogeneity with clinical implications, which has to be accounted for when selecting diagnostic material, inter alia, by histological evaluation of the proportion of cell-dense areas.

p16 Immunohistochemistry as a Screening Tool for Homozygous CDKN2A Deletions in CNS Tumors.

The 2021 World Health Organization classification of tumors of the central nervous system emphasizes the significance of molecular parameters for an integrated diagnosis. Homozygous deletion of cyclin-dependent kinase inhibitor 2a (CDKN2A) has been associated with an adverse prognosis in IDH -mutant gliomas, supratentorial ependymomas, meningiomas, and MPNST. In this study, we examined the value of p16 protein immunohistochemistry as a rapid and cost-effective screening tool for a homozygous CDKN2A deletion. Genetic analyses for CDKN2A in 30 pleomorphic xanthoastrocytomas, 32 IDH -wild-type high-grade gliomas, 40 supratentorial ependymomas with ZFTA-RELA gene fusion, 21 IDH-mutant astrocytomas, and 24 meningiomas were performed mainly by a molecular inversion probe assay, a high-resolution, quantitative technology for the assessment of chromosomal copy number alterations. Immunohistochemistry for p16 proved to have a high positive predictive value (range 90% to 100%) and an overall low negative predictive value (range 22% to 93%) for a homozygous CDKN2A deletion. In a setting where molecular testing is limited for cost and time reasons, p16 immunohistochemistry serves as a useful and rapid screening tool for identifying cases that should be subjected to further molecular testing for CDKN2A deletions.

Perinatally diagnosed congenital craniopharyngiomas in the KRANIOPHARYNGEOM trials.

Background: Craniopharyngiomas (CPs) are rare embryonic tumors. Clinical presentation and outcome of patients perinatally diagnosed with congenital CP (cCP) are not clear and refer mainly to a few case reports in the literature. The aim of this study was to analyze clinical presentation and outcome in patients with cCP. Study design: Three hundred and sixty-one patients diagnosed with adamantinomatous CP were recruited 2007-2022 in KRANIOPHARYNGEOM 2007/Registry 2019 and prospectively observed. In two cases, cCP was diagnosed prenatally and in one case on the second day of life. Pre- and perinatal diagnostic findings, postnatal evaluation, and therapeutic interventions and outcome in these three cases of cCP were analyzed. Results: All patients survived. One patient developed psychomotor retardation and a mild hemiparesis. Prenatal routine ultrasound examination led to the diagnosis of cCP. Tumor resection was performed during the early postnatal period (range: 11-51 days of age). Functional capacity, measured by Fertigkeitenskala-Münster-Heidelberg (FMH) was reduced in three and behavioral parameters, measured by the Strength and Difficulties Questionnaire (SDQ) were abnormal in two cases. Conclusion: cCP is a rare diagnosis with a prevalence of 0.83% in our study group. Compared to cases reported in the literature, the presented cases were treated immediately and had a better prognosis. Based on improvements of diagnostic and therapeutic techniques, prenatal diagnosis of cCP should lead to transfer prior to delivery of cCP patients to a specialized center for delivery and postnatal treatment of newborns with sellar masses by a multidisciplinary team to secure the improved prognosis of these patients. Significance statement: We previously reported that lower event-free survival rates after craniopharyngioma are associated with younger age at diagnosis. Perinatally diagnosed congenital craniopharyngiomas are very rare. This article presents three unique cases with congenital craniopharyngioma, comparing their diagnostics, therapy, and development. All three cases had surgery during the early postnatal period with sparing of the posterior hypothalamus. In each case, endocrinopathy was present at follow-up. Low functional capacity was reported in all cases and an abnormal total difficulties score in two cases. Compared to the literature, the presented cases had better prognosis in morbidity and mortality. This report and the review of the literature confirm the importance of a multidisciplinary approach in the diagnostic and treatment of the very rare condition of congenital craniopharyngioma.

[Pediatric brain tumors]. / Kindliche Hirntumoren.

Pediatric central nervous system (CNS) tumors differ in their relative frequency, location, histology, biological behavior and prognosis from tumors in adults. Accurate neuropathological classification of CNS tumors is essential for therapeutic decisions and inclusion in therapy optimization studies. Tissue samples are analyzed by standardized conventional histological, immunohistological and molecular pathological methods and diagnosed according to the current World Health Organization (WHO) classification for CNS tumors (2021). By identifying characteristic genetic alterations and specific epigenetic signatures, the precision in the classification of pediatric brain tumors has significantly improved in recent years. The WHO classification allows a worldwide uniform, standardized classification of brain tumors and forms the basis for comparability of international epidemiological and clinical data. In some tumor types, such as childhood gliomas and embryonal tumors, key molecules and signaling pathways have been identified in recent years that represent starting points for new mechanism-based therapeutic modalities in the treatment of these patients.

Clinical outcome of pediatric medulloblastoma patients with Li-Fraumeni syndrome.

BACKGROUND: The prognosis for Li-Fraumeni syndrome (LFS) patients with medulloblastoma (MB) is poor. Comprehensive clinical data for this patient group is lacking, challenging the development of novel therapeutic strategies. Here, we present clinical and molecular data on a retrospective cohort of pediatric LFS MB patients. METHODS: In this multinational, multicenter retrospective cohort study, LFS patients under 21 years with MB and class 5 or class 4 constitutional TP53 variants were included. TP53 mutation status, methylation subgroup, treatment, progression free- (PFS) and overall survival (OS), recurrence patterns, and incidence of subsequent neoplasms were evaluated. RESULTS: The study evaluated 47 LFS individuals diagnosed with MB, mainly classified as DNA methylation subgroup "SHH_3" (86%). The majority (74%) of constitutional TP53 variants represented missense variants. The 2- and 5-year (y-) PFS were 36% and 20%, and 2- and 5y-OS were 53% and 23%, respectively. Patients who received postoperative radiotherapy (RT) (2y-PFS: 44%, 2y-OS: 60%) or chemotherapy before RT (2y-PFS: 32%, 2y-OS: 48%) had significantly better clinical outcome then patients who were not treated with RT (2y-PFS: 0%, 2y-OS: 25%). Patients treated according to protocols including high-intensity chemotherapy and patients who received only maintenance-type chemotherapy showed similar outcomes (2y-PFS: 42% and 35%, 2y-OS: 68% and 53%, respectively). CONCLUSIONS: LFS MB patients have a dismal prognosis. In the presented cohort use of RT significantly increased survival rates, whereas chemotherapy intensity did not influence their clinical outcome. Prospective collection of clinical data and development of novel treatments are required to improve the outcome of LFS MB patients.

Neurosurgical morbidity in pediatric supratentorial midline low-grade glioma: Results from the German LGG studies.

Surgical resection is a mainstay of treatment for pediatric low-grade glioma (LGG) within all current therapy algorithms, yet associated morbidity is scarcely reported. As supratentorial midline (SML) interventions are particularly challenging, we investigated the frequency of neurosurgical complications/new impairments aiming to identify their risk factors. Records were retrospectively analyzed from 318 patients with SML-LGG from successive German multicenter LGG studies, undergoing surgery between May 1998 and June 2020. Exactly 537 operations (230 resections, 167 biopsies, 140 nontumor procedures) were performed in 318 patients (54% male, median age: 7.6 years at diagnosis, 9.5 years at operation, 11% NF1, 42.5% optic pathway glioma). Surgical mortality rate was 0.93%. Applying the Drake classification, postoperative surgical morbidity was observed following 254/537 (47.3%) and medical morbidity following 97/537 (18.1%) patients with a 40.1% 30-day persistence rate for newly developed neurological deficits (65/162). Neuroendocrine impairment affected 53/318 patients (16.7%), visual deterioration 34/318 (10.7%). Postsurgical morbidity was associated with patient age <3 years at operation, tumor volume ≥80 cm3 , presence of hydrocephalus, complete resection, surgery in centers with less than median reported tumor-related procedures and during the earlier study period between 1998 and 2006, while the neurosurgical approach, tumor location, NF1 status or previous nonsurgical treatment were not. Neurosurgery-associated morbidity was frequent in pediatric patients with SML-LGG undergoing surgery in the German LGG-studies. We identified patient- and institution-associated factors that may increase the risk for complications. We advocate that local multidisciplinary teams consider the planned extent of resection and surgical skills.

A multi-institutional retrospective pooled outcome analysis of molecularly annotated pediatric supratentorial ZFTA-fused ependymoma.

Background: ZFTA-RELA (formerly known as c11orf-RELA) fused supratentorial ependymoma (ZFTAfus ST-EPN) has been recognized as a novel entity in the 2016 WHO classification of CNS tumors and further defined in the recent 2021 edition. ZFTAfus ST-EPN was reported to portend poorer prognosis when compared to its counterpart, YAP1 ST-EPN in some previously published series. The aim of this study was to determine the treatment outcome of molecularly confirmed and conventionally treated ZFTAfus ST-EPN patients treated in multiple institutions. Methods: We conducted a retrospective analysis of all pediatric patients with molecularly confirmed ZFTAfus ST-EPN patients treated in multiple institutions in 5 different countries (Australia, Canada, Germany, Switzerland, and Czechia). Survival outcomes were analyzed and correlated with clinical characteristics and treatment approaches. Results: A total of 108 patients were collated from multiple institutions in 5 different countries across three continents. We found across the entire cohort that the 5- and 10-year PFS were 65% and 63%, respectively. The 5- and 10-year OS of this cohort of patients were 87% and 73%. The rates of gross total resection (GTR) were high with 84 out of 108 (77.8%) patients achieving GTR. The vast majority of patients also received post-operative radiotherapy, 98 out of 108 (90.7%). Chemotherapy did not appear to provide any survival benefit in our patient cohort. Conclusion: This is the largest study to date of contemporaneously treated molecularly confirmed ZFTAfus ST-EPN patients which identified markedly improved survival outcomes compared to previously published series. This study also re-emphasizes the importance of maximal surgical resection in achieving optimal outcomes in pediatric patients with supratentorial ependymoma.

Molecular characterisation defines clinically-actionable heterogeneity within Group 4 medulloblastoma and improves disease risk-stratification.

Group 4 tumours (MBGrp4) represent the majority of non-WNT/non-SHH medulloblastomas. Their clinical course is poorly predicted by current risk-factors. MBGrp4 molecular substructures have been identified (e.g. subgroups/cytogenetics/mutations), however their inter-relationships and potential to improve clinical sub-classification and risk-stratification remain undefined. We comprehensively characterised the paediatric MBGrp4 molecular landscape and determined its utility to improve clinical management. A clinically-annotated discovery cohort (n = 362 MBGrp4) was assembled from UK-CCLG institutions and SIOP-UKCCSG-PNET3, HIT-SIOP-PNET4 and PNET HR + 5 clinical trials. Molecular profiling was undertaken, integrating driver mutations, second-generation non-WNT/non-SHH subgroups (1-8) and whole-chromosome aberrations (WCAs). Survival models were derived for patients ≥ 3 years of age who received contemporary multi-modal therapies (n = 323). We first independently derived and validated a favourable-risk WCA group (WCA-FR) characterised by ≥ 2 features from chromosome 7 gain, 8 loss, and 11 loss. Remaining patients were high-risk (WCA-HR). Subgroups 6 and 7 were enriched for WCA-FR (p < 0·0001) and aneuploidy. Subgroup 8 was defined by predominantly balanced genomes with isolated isochromosome 17q (p < 0·0001). While no mutations were associated with outcome and overall mutational burden was low, WCA-HR harboured recurrent chromatin remodelling mutations (p = 0·007). Integration of methylation and WCA groups improved risk-stratification models and outperformed established prognostication schemes. Our MBGrp4 risk-stratification scheme defines: favourable-risk (non-metastatic disease and (i) subgroup 7 or (ii) WCA-FR (21% of patients, 5-year PFS 97%)), very-high-risk (metastatic disease with WCA-HR (36%, 5-year PFS 49%)) and high-risk (remaining patients; 43%, 5-year PFS 67%). These findings validated in an independent MBGrp4 cohort (n = 668). Importantly, our findings demonstrate that previously established disease-wide risk-features (i.e. LCA histology and MYC(N) amplification) have little prognostic relevance in MBGrp4 disease. Novel validated survival models, integrating clinical features, methylation and WCA groups, improve outcome prediction and re-define risk-status for ~ 80% of MBGrp4. Our MBGrp4 favourable-risk group has MBWNT-like excellent outcomes, thereby doubling the proportion of medulloblastoma patients who could benefit from therapy de-escalation approaches, aimed at reducing treatment induced late-effects while sustaining survival outcomes. Novel approaches are urgently required for the very-high-risk patients.

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

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.

Supratentorial CNS-PNETs in children; a Swedish population-based study with molecular re-evaluation and long-term follow-up.

BACKGROUND: Molecular analyses have shown that tumours diagnosed as supratentorial primitive neuro-ectodermal tumours of the central nervous system (CNS-PNETs) in the past represent a heterogenous group of rare childhood tumours including high-grade gliomas (HGG), ependymomas, atypical teratoid/rhabdoid tumours (AT/RT), CNS neuroblastoma with forkhead box R2 (FOXR2) activation and embryonal tumour with multi-layered rosettes (ETMR). All these tumour types are rare and long-term clinical follow-up data are sparse. We retrospectively re-evaluated all children (0-18 years old) diagnosed with a CNS-PNET in Sweden during 1984-2015 and collected clinical data. METHODS: In total, 88 supratentorial CNS-PNETs were identified in the Swedish Childhood Cancer Registry and from these formalin-fixed paraffin-embedded tumour material was available for 71 patients. These tumours were histopathologically re-evaluated and, in addition, analysed using genome-wide DNA methylation profiling and classified by the MNP brain tumour classifier. RESULTS: The most frequent tumour types, after histopathological re-evaluation, were HGG (35%) followed by AT/RT (11%), CNS NB-FOXR2 (10%) and ETMR (8%). DNA methylation profiling could further divide the tumours into specific subtypes and with a high accuracy classify these rare embryonal tumours. The 5 and 10-year overall survival (OS) for the whole CNS-PNET cohort was 45% ± 12% and 42% ± 12%, respectively. However, the different groups of tumour types identified after re-evaluation displayed very variable survival patterns, with a poor outcome for HGG and ETMR patients with 5-year OS 20% ± 16% and 33% ± 35%, respectively. On the contrary, high PFS and OS was observed for patients with CNS NB-FOXR2 (5-year 100% for both). Survival rates remained stable even after 15-years of follow-up. CONCLUSIONS: Our findings demonstrate, in a national based setting, the molecular heterogeneity of these tumours and show that DNA methylation profiling of these tumours provides an indispensable tool in distinguishing these rare tumours. Long-term follow-up data confirms previous findings with a favourable outcome for CNS NB-FOXR2 tumours and poor chances of survival for ETMR and HGG.

Risk prediction in early childhood sonic hedgehog medulloblastoma treated with radiation-avoiding chemotherapy: Evidence for more than 2 subgroups.

BACKGROUND: The prognostic impact of clinical risk factors and DNA methylation patterns in sonic hedgehog (SHH)-activated early childhood desmoplastic/nodular medulloblastoma (DMB) or medulloblastoma with extensive nodularity (MBEN) were evaluated to better identify patients at risk for relapse. METHODS: One hundred and forty-four patients with DMB (n = 99) or MBEN (n = 45) aged <5 years and treated with radiation-sparing approaches, including intraventricular methotrexate in 132 patients were evaluated. RESULTS: Patients with DMB had less favorable 5-year progression-free survival than MBEN (5y-PFS, 71% [DMB] vs. 93% [MBEN]). Patients aged >3 years were associated with more unfavorable 5y-PFS (47% [>3 years] vs. 85% [<1 year] vs. 84% [1-3 years]). DNA methylation profiles available (n = 78) were reclassified according to the 2021 WHO classification into SHH-1 (n = 39), SHH-2 (n = 38), and SHH-3 (n = 1). Hierarchical clustering delineated 2 subgroups among SHH-2: SHH-2a (n = 19) and SHH-2b (n = 19). Patients with SHH-2b medulloblastoma were older, predominantly displayed DMB histology, and were more often located in the cerebellar hemispheres. Chromosome 9q losses were more frequent in SHH-2b, while few chromosomal alterations were observed in SHH-2a. SHH-2b medulloblastoma carried a significantly increased relapse risk (5y-PFS: 58% [SHH-2b] vs. 83% [SHH-1] vs. 95% [SHH-2a]). Subclassification of SHH-2 with key clinical and cytogenetic characteristics was confirmed using 2 independent cohorts (total n = 188). Gene mutation analysis revealed a correlation of SHH-2a with SMO mutations. CONCLUSIONS: These data suggest further heterogeneity within early childhood SHH-DMB/MBEN: SHH-2 splits into a very low-risk group SHH-2a enriched for MBEN histology and SMO mutations, and SHH-2b comprising older DMB patients with a higher risk of relapse.

A Sequential Targeting Strategy Interrupts AKT-Driven Subclone-Mediated Progression in Glioblastoma.

PURPOSE: Therapy resistance and fatal disease progression in glioblastoma are thought to result from the dynamics of intra-tumor heterogeneity. This study aimed at identifying and molecularly targeting tumor cells that can survive, adapt, and subclonally expand under primary therapy. EXPERIMENTAL DESIGN: To identify candidate markers and to experimentally access dynamics of subclonal progression in glioblastoma, we established a discovery cohort of paired vital cell samples obtained before and after primary therapy. We further used two independent validation cohorts of paired clinical tissues to test our findings. Follow-up preclinical treatment strategies were evaluated in patient-derived xenografts. RESULTS: We describe, in clinical samples, an archetype of rare ALDH1A1+ tumor cells that enrich and acquire AKT-mediated drug resistance in response to standard-of-care temozolomide (TMZ). Importantly, we observe that drug resistance of ALDH1A1+ cells is not intrinsic, but rather an adaptive mechanism emerging exclusively after TMZ treatment. In patient cells and xenograft models of disease, we recapitulate the enrichment of ALDH1A1+ cells under the influence of TMZ. We demonstrate that their subclonal progression is AKT-driven and can be interfered with by well-timed sequential rather than simultaneous antitumor combination strategy. CONCLUSIONS: Drug-resistant ALDH1A1+/pAKT+ subclones accumulate in patient tissues upon adaptation to TMZ therapy. These subclones may therefore represent a dynamic target in glioblastoma. Our study proposes the combination of TMZ and AKT inhibitors in a sequential treatment schedule as a rationale for future clinical investigation.