Atypical teratoid/rhabdoid tumors (AT/RT) are the most common malignant brain tumors during infancy and associated with a dismal prognosis. The majority of patients suffer from tumor progression or recurrence, but underlying mechanisms remain unknown. To better understand such mechanisms, we performed single-nucleus RNA sequencing (snRNAseq) of eight paired primary tumors and recurrences. Tumor cells and cells of the tumor microenvironment (TME) were analyzed separately. Potentially therapy-resistant tumor cells were identified through the comparison of global gene expression profiles between primary and recurrent tumor cell populations using CIBERSORT. Histopathology, in vitro experiments, bulk RNA sequencing, and survival analysis were performed for validation. Paired primary and recurrent AT/RT showed significant differences in their gene expression profiles. Potentially therapy-resistant AT/RT-MYC tumor cells revealed changes in the extracellular matrix (ECM) as well as altered developmental processes and immune signaling pathways. Respective gene signatures were correlated with inferior survival in AT/RT-MYC patients. Tumor cells of relapsed AT/RT-MYC underwent partial epithelial-mesenchymal transition (pEMT), a feature that was confirmed by immunohistochemistry (IHC) and by analyzing AT/RT cells after standard therapy in vitro. Together, we identified potential mechanisms of tumor relapse and therapy resistance in AT/RT, which could be employed to improve therapy in future.

Abstract Purpose Ataxia is the most prevalent motor impairment in children following surgical resection of posterior fossa tumour. The aim of this study was to examine the prevalence, severity, trajectory and risk factors for ataxia in children following surgical resection of posterior fossa tumours. Methods Prospective observational multicentre cohort study, including children (aged < 18 years) undergoing surgical resection of a posterior fossa tumour in 36 European centres across 15 countries. Children were assessed using two ataxia scales; the Brief ataxia rating scale (BARS) and the Scale for the assessment and rating of ataxia (SARA). Assessments were undertaken pre-operatively, post-operative follow up 1 (2-weeks), post-operative follow up 2 (2-months) and post-operative follow up 3 (1-year) after surgery. Results 756 participants who underwent initial surgical management participated in the study, and 122 completed BARS assessment at all timepoints. In this group of children, ataxia was present in 40 (32.8%) participants pre-operatively, and 49 (40.2%) at post-operative follow up 1. Over 90% of participants demonstrated independent walking by 1 year post-operatively (defined by BARS/SARA gait item ≤3). Children with medulloblastomas and post-operative mutism demonstrated more severe ataxia compared to the remaining children. Conclusion Pre-operative ataxia, medulloblastoma and postoperative mutism were identified as the strongest predictors of persistent ataxia. Pre-operative ataxia has been identified for the first time as an indicator of long-term ataxia. This knowledge could be used to identify potential rehabilitation needs and highlights the benefits of an MDT pre-operative assessment.

INTRODUCTION:Standard Ewing sarcoma treatment includes chemotherapy and local therapy. Previous studies found no average treatment effect of combined local therapy (radiotherapy and surgery) versus surgery alone, leaving it unclear which patients benefit from combined therapy. METHODS:We analysed patients with localized Ewing sarcoma from the EURO-E.W.I.N.G. 99 and Ewing 2008 trials. Using causal survival forests validated through repeated k-fold cross-validation, we predicted individualized treatment effects (ITEs) for 5-year event-free survival (5y EFS) comparing combined therapy to surgery alone. We developed a personalized treatment rule and evaluated ITE predictors using Shapley additive explanations analysis. RESULTS:Of 1501 patients, 719 received combination therapy and 782 surgery alone. Predicted ITEs showed substantial heterogeneity (range: -0.7-1.0 years). We found that 19.4% of patients derived clinically meaningful benefit (5y EFS increase ≥6 months) from combined therapy. In the quartile predicted to benefit least, hazards of any event were increased by 53% (HR 1.53, 95% CI 1.06-2.20, p = 0.02). Conversely, in the quartile predicted to benefit most, hazards were reduced by 40% (HR 0.60, 95% CI 0.42-0.85, p = 0.004). In patients undergoing combined treatment, surgical margin status was not associated with treatment benefit. A simulation analysis showed that applying our treatment strategy could improve EFS. DISCUSSION:Our predicted ITEs can help identify patients benefiting from combined therapy. The finding that patients with non-wide surgical margins did not benefit from combined therapy challenges current decision-making practices based on traditional subgroup analyses.

Postoperative speech impairment (POSI) and cranial nerve deficits (CND) are common complications of pediatric posterior fossa (PF) tumor surgery. Intraoperative MRI (ioMRI) has proven a useful tool in achieving gross total resection. The risk of POSI and CND with ioMRI remains unclear, making it the primary scope of this study. Additionally, we assessed whether POSI was associated with CND. We prospectively included pediatric patients undergoing PF tumor surgery in 36 centers across 15 European countries. Neurological status and speech were assessed preoperatively and 1–4 weeks postoperatively. Surgical details, including tumor location and use of ioMRI, were recorded within 72 h of surgery. Postoperative CND were categorized as 0, 1, 2, or ≥ 3 nerves affected; POSI as habitual, reduced speech, or mutism. Proportional odds models estimated odds ratios (OR) for 1) POSI with stepwise adjustment for tumor location and age, and 2) CND with adjustment for preoperative CND and tumor location. Subgroup analyses assessed systematic differences, missing data, center-level effects, and histology adjustment. Of 790 primary PF tumor surgeries, 141 (18

Postoperative speech impairment (POSI) is a core symptom of cerebellar mutism syndrome (CMS) and is a common complication after the resection of paediatric posterior fossa (PF) tumours. Preoperative glucocorticoids (pGC) are considered standard treatment to reduce tumour oedema; in addition, glucocorticoids are often administered intraoperatively (iGC) to reduce both postoperative nausea and vomiting. The study aims to investigate whether the occurrence of POSI may be associated with pGC and iGC. In a prospective observational multicentre study, we included children with a PF tumour requiring either resection or open biopsy. The use of pGC and iGC, including drug type and dose, was registered. Postoperative speech status was classified as mutism, reduced speech, or habitual speech, where mutism and reduced speech were considered POSI of higher and lower severity, respectively. Proportional odds logistic regression with adjustment for tumour type, tumour location, and age was used to analyse the occurrence of POSI associated with glucocorticoids (GC). From August 2014 to November 2024, we recruited 810 children, of whom 605 were included in the primary analysis. We found no association between the use of GC (pGC nor iGC) and the occurrence of POSI. The result did not change when adjusting for tumour type, tumour location, and age. The analysis included both a comparison between using and not using pGC (OR 1.06 [95

Cerebellar Mutism Syndrome (CMS) is a neurological complication of posterior fossa (PF) tumour surgery in children, and postoperative speech impairment (POSI) is the cardinal symptom of CMS. The role of tumour volume on the risk of POSI remains unexplored. This study investigates the association between tumour volume and the risk of POSI. We included 360 patients from the European CMS study with available preoperative T1-weighted contrast-enhanced brain MRI. Speech status was assessed within two weeks postoperatively and categorised into three levels: habitual speech, severely reduced speech, and mutism. Tumour volumes were calculated using the BrainLab Elements SmartBrush™, a semi-automated segmentation tool. We used proportional odds models to estimate the odds ratio (OR) with adjustments for tumour location, pathology, and age. Based on the primary analysis, a risk stratification model for medulloblastoma patients was constructed, and the optimal volume cut-off was determined with Youden's Index. We found no effect of the overall tumour volume on the risk of POSI. This result did not change when adjusted for tumour location, pathology, and age. We found an association between tumour volume of medulloblastoma and the risk of POSI (unadjusted OR of 1.04 per increase in cm3 (95% CI 1.01;1.07, p = 0.01)), which did not change when adjusting for tumour location and age. The risk stratification cut-off for the tumour volume of medulloblastoma was calculated to be 16,5 cm3. Patients with medulloblastoma and preoperative tumour volumes below 16,5 cm3 had an absolute risk of 13% for POSI (low-risk group), whereas patients with preoperative tumour volumes above 16,5 cm3 had an absolute risk of 50% for POSI (high-risk group). Our data showed an association between preoperative tumour volume and the risk of POSI in children with medulloblastoma, while no association was found for the volume of other tumour types. We suggest a straightforward cut-off risk model for assessing the risk of POSI in children with medulloblastoma based on preoperative tumour volume. This approach can aid clinicians in informing patients and parents about the complications related to CMS following PF tumour surgery in children. ID NCT02300766 (October 2014).

Background/Objectives: The anti-GD2 monoclonal antibody dinutuximab beta has become standard of care maintenance therapy for high-risk neuroblastoma (HR-NB) in the first-line setting and is also approved in the relapsed/refractory setting. We present a retrospective review of 37 children with HR-NB included in the Hungarian Childhood Cancer Registry who received dinutuximab beta (first-line maintenance therapy, n = 31; relapsed/refractory, n = 6). Methods: All patients received dinutuximab beta continuously over the first 10 days of each 35-day cycle, with dosing based on body surface area/weight. Five cycles were planned, with further cycles administered at the treating physician's discretion. Results: At data cutoff, the overall disease control rate was 54.1% (20/37) (complete response, 51.4% (19/37); partial response, 0.0% (0/37), stable disease, 2.7% [1/37]); two patients (5.4%) had progressive disease, and 15 patients (40.5%) had died. The 5-year overall survival (OS) and event-free survival (EFS) rates in the overall population were 63.3% (95% confidence interval, 49.1-81.7) and 56.2% (95% confidence interval, 42.1-75.0), respectively. Grade 3 or 4 adverse events (including blood and lymphatic system disorders, hypoxia, hypotension, and capillary leak syndrome) were generally consistent with dinutuximab beta's known safety profile. Conclusions: Dinutuximab beta was an effective immunotherapy for patients with HR-NB in routine clinical practice, with a generally manageable side effect profile.

Risk-adapted treatment protocols conferred remarkable improvement in the survival rates of pediatric acute lymphoblastic leukemia/lymphoma (ALL/LBL). Nevertheless, clinical management is still challenging in certain molecular subgroups and in the presence of alterations associated with an increased rate of relapse. In this study, disease-relevant genomic and transcriptomic profiles were established in a prospective, multicenter, real-world cohort involving 192 children diagnosed with ALL/LBL. Gene fusions were detected in 34.9% of B-ALL and 46.4% of T-ALL patients, with novel chimeric genes involving JAK2, KMT2A, PAX5, RUNX1, and NOTCH1, and with KMT2A-rearranged patients displaying the worst 3-year event-free survival (P 1/4 .019). Nonsynonymous mutations were uncovered in 74.9% of the analyzed patients, and pairwise scrutiny of genetic lesions revealed recurrent clonal selection mechanisms commonly converging on the same pathway (eg, Ras, JAK/ STAT, and Notch) in individual patients. Investigation of matched diagnostic and relapse samples unraveled complex subclonal variegation, and mutations affecting the NT5C2, TP53, CDKN2A, and PIK3R1 genes, emerging at the time of relapse. TP53 and CREBBP mutations, even as subclonal aberrations, were associated with shorter 3-year event-free survival among all patients with B-ALL (TP53 mutant vs wild-type: P = .008, CREBBP mutant vs wild-type: P = .010), and notably, B-ALL patients showing no measurable residual disease on day 33 could be further stratified based on TP53 mutational status (P < .001). Our in-depth molecular characterization performed across all risk groups identified novel opportunities for molecularly targeted therapy in 55.9% of high-risk and 31.6% of standard/intermediate-risk patients. (c) 2025 THE AUTHORS. Published by Elsevier Inc. on behalf of the United States & Canadian Academy of Pathology. This is an open access article under the CC BY-NC-ND license (http://creativecommons. org/licenses/by-nc-nd/4.0/).

Over the last decade, organoid research has emerged as a transformative field in biomedical science, offering unparalleled opportunities in disease modeling, pharmacological testing, and tissue engineering [...]

Posterior fossa group A (PFA) ependymoma is a lethal brain cancer diagnosed in infants and young children. The lack of driver events in the PFA linear genome led us to search its 3D genome for characteristic features. Here, we reconstructed 3D genomes from diverse childhood tumor types and uncovered a global topology in PFA that is highly reminiscent of stem and progenitor cells in a variety of human tissues. A remarkable feature exclusively present in PFA are type B ultra long-range interactions in PFAs (TULIPs), regions separated by great distances along the linear genome that interact with each other in the 3D nuclear space with surprising strength. TULIPs occur in all PFA samples and recur at predictable genomic coordinates, and their formation is induced by expression of EZHIP. The universality of TULIPs across PFA samples suggests a conservation of molecular principles that could be exploited therapeutically.

Abstract Atypical teratoid/rhabdoid tumors (AT/RT) are the most common malignant brain tumors during infancy. They split into four molecular types. The major three (AT/RT-SHH, AT/RT-TYR, and AT/RT-MYC) all carry mutations in SMARCB1, the fourth smaller type is characterized by SMARCA4 mutations (AT/RT-SMARCA4). AT/RT are associated with a dismal outcome, and 50% of the patients suffer a progress or recurrence of the tumor. Molecular mechanisms of disease recurrence are currently unclear. Here, we identified molecular characteristics of AT/RT primaries and recurrences, which could help to improve the therapy for relapsed patients. Previously, we investigated tumor tissue from 26 patients revealing signatures of recurrences in comparison to matched primary tumor samples using histology, DNA methylation analysis, and bulk RNA sequencing. To further identify mechanisms regarding tumor progression and recurrence, we performed single-cell RNA (scRNA) sequencing for five of these primary-recurrence pairs so far using formalin-fixed paraffin-embedded (FFPE) tissue. Using Uniform Manifold Approximation and Projection (UMAP) for dimensionality reduction, we observed a distinct separation in the gene expression profiles of primary and recurrent tumor cells. In addition, we aimed to identify primary tumor cells which potentially persisted after therapy and developed to recurrences. Therefore, we identified primary tumor cells showing highest similarity to recurrent tumor cells using CIBERSORT and trajectory analysis. Differentially expressed genes of these specific primary tumor cells were investigated. They are involved in cell migration, -adhesion, -proliferation, and -stability, among others. Overall, molecular characteristics that occur in the course of the disease were identified, which may help to define new therapeutic targets for AT/RT recurrences.

Abstract BACKGROUND Rhabdoid-Tumor-Predisposition-Syndrome is due to germline mutations in SMARCB1 (rarely SMARCA4). We pursued a comprehensive clinical and (epi-)genetic characterization of RTPS families. METHODS 90 affected children from 16 countries were compared to 252 with sporadic rhabdoid tumors (01/2004 - 01/2021). Tumors and blood were examined for structural or single nucleotide variants (SV, SNV) in SMARCB1/SMARCA4. In 83% (70/84) of RTPS1-patients complete genetic data were available. Trio-exome (n=25) and 850k-methylation analysis of tumors (52/84) and blood (n=51), parents (n=44) and unaffected siblings (n=4) was performed. RESULTS RTPS1 and 2 were diagnosed in 84 (48 females, 36 males) and 6 patients respectively. Median age at diagnosis (RTPS1 only) was 5 (0 – 203) months (18 months, 0-211 in controls, p<0,001). ATRT were present in 55% (46/84), extracranial, extrarenal malignant RT (eMRT) and RT of the kidney (RTK) in 8% (7/84). 26% (22/84) presented with synchronous tumors (SYN). Homozygous SVs were less frequent in the RTPS-tumors [20% (14/84) vs 47% (82/174), p<0,001]. ATRT-MYC was significantly more common among RTPS1 while ATRT-SHH was less common. Trio blood DNA-methylation analysis separated samples of parents but not siblings from RTPS patients. Likely (DNA-methylation) age was a major confounder. The analysis of trio-exomes is ongoing. 5-year-OS and -EFS in RTPS1-patients were 18,4±4.5% and 14±3,9%, (controls: 48,2 ±3,5%, 40,3±3,2%). VOD (venoocclusive disease) was significant in patients < 2 months [18,5% (12/65) vs. 3,3% (3/92), p=0.005]. RTPS1-patients rarely achieved a CR [32% (27/84) vs. 52% (129/249), p=0,002]. In an adjusted multivariate model prognostic factors were male sex [HR: 1.7 (1.0 – 2.9)], age ≤12 months [HR: 2.1 (1.0 – 4.3)], and diagnosis of ATRT [HR: 0.6 (0.3 – 0.9)]. CONCLUSION We describe significant positive outcome predictors such as age >12 months, absence of SYN, diagnosis of ATRT, localized disease, PGV located at C-termini and female sex in RTPS1-patients.

A major obstacle to the implantation of ex vivo engineered tissues is the incorporation of functional vascular supply to support the growth of new tissue and to minimize ischemic injury. Existing prevascularization systems, such as arteriovenous (AV) loop-based systems, require microsurgery, limiting their use to larger animals. We aimed to develop an implantable device that can be prevascularized to enable vascularization of tissues in small rodents, and test its application on the vascularization of embryonic kidneys. Implanting the chamber between the abdominal aorta and the inferior vena cava, we detected endothelial cells and vascular networks after 48 h of implantation. Loading the chamber with collagen I (C), Matrigel (M), or Matrigel + vascular endothelial growth factor) (MV) had a strong influence on vascularization speed: Chambers loaded with C took 7 days to vascularize, 4 days for chambers with M, and 2 days for chambers with MV. Implantation of E12.5 mouse embryonic kidneys into prevascularized chambers (C, MV) was followed with significant growth and ureteric branching over 22 days. In contrast, the growth of kidneys in non-prevascularized chambers was stunted. We concluded that our prevascularized chamber is a valuable tool for vascularizing implanted tissues and tissue-engineered constructs. Further optimization will be necessary to control the directional growth of vascular endothelial cells within the chamber and the vascularization grade.

Background. Ewing sarcoma (EwS) is a rare and highly malignant bone tumor primarily affecting children, adolescents, and young adults. The pelvis, trunk, and lower extremities are the most common sites, while EwS of the sacrum as a primary site is very rare, and only few studies focusing on this location are published. Due to the anatomical condition, local treatment is challenging in sacral malignancies. We analyzed factors that might influence the outcome of patients suffering from sacral EwS. Methods. We retrospectively analyzed data of the GPOH EURO-E.W.I.N.G 99 trial and the EWING 2008 trial, with a cohort of 124 patients with localized or metastatic sacral EwS. The study endpoints were overall survival (OS) and event-free survival (EFS). OS and EFS were calculated using the Kaplan–Meier method, and univariate comparisons were estimated using the log-rank test. Hazard ratios (HRs) with respective 95% confidence intervals (CIs) were estimated in a multivariable Cox regression model. Results. The presence of metastases (3y-EFS: 0.33 vs. 0.68; P<0.001; HR = 3.4, 95% CI 1.7 to 6.6; 3y-OS: 0.48 vs. 0.85; P<0.001; HR = 4.23, 95% CI 1.8 to 9.7), large tumor volume (≥200 ml) (3y-EFS: 0.36 vs. 0.69; P=0.02; HR = 2.1, 95% CI 1.1 to 4.0; 3y-OS: 0.42 vs. 0.73; P=0.04; HR = 2.1, 95% CI 1.03 to 4.5), and age ≥18 years (3y-EFS: 0.41 vs. 0.60; P=0.02; HR = 2.6, 95% CI 1.3 to 5.2; 3y-OS: 0.294 vs. 0.59; P=0.01; HR = 2.92, 95% CI 1.29 to 6.6) were revealed as adverse prognostic factors. Conclusion. Young age seems to positively influence patients` survival, especially in patients with primary metastatic disease. In this context, our results support other studies, stating that older age has a negative impact on survival. Tumor volume, metastases, and the type of local therapy modality have an impact on the outcome of sacral EwS. Level of evidence: Level 2. This trial is registered with NCT00020566 and NCT00987636.

Background: Our knowledge about the attitudes of healthcare staff to palliative care in pediatric oncology is scarce. We aimed to assess their perceptions of palliative care in Hungary and find answers to the question of how to provide good palliative care for children. Method: Physicians (n = 30) and nurses (n = 43) working in the field of pediatric oncology (12 of them specialized in hospice care) were interviewed. Palliative care practice (communication, integration of palliative care, professionals’ feelings and attitudes, and opportunities for improvement) was assessed by semi-structured interviews evaluated in a mixed quantitative and qualitative way by narrative categorical content analysis and thematic analysis. Results: All providers displayed high negative emotions, positive evaluations, and used many active verbs. Nurses showed higher levels of denial, more self-references, and were more likely to highlight loss. Physicians emphasized the importance of communication regarding adequate or inadequate palliative care. Hospice specialists showed a higher passive verb rate, a lower self-reference, a lower need for psychological support, and a greater emphasis on teamwork and professional aspects. Conclusion: Our results show that nurses are more emotionally stressed than doctors in palliative care in pediatric oncology. To our knowledge, a study comparing doctors and nurses in this field has yet to be carried out. Our results suggest that pediatric oncological staff can positively evaluate a child’s palliative care despite the emotional strain. Regarding hospices, professional practice in palliative care may be a protective factor in reducing emotional distress and achieving professional well-being.

BACKGROUND:The term gliomatosis cerebri (GC), a radiology-defined highly infiltrating diffuse glioma, has been abandoned since molecular GC-associated features could not be established. METHODS:We conducted a multinational retrospective study of 104 children and adolescents with GC providing comprehensive clinical and (epi-)genetic characterization. RESULTS:Median overall survival (OS) was 15.5 months (interquartile range, 10.9-27.7) with a 2-year survival rate of 28%. Histopathological grading correlated significantly with median OS: CNS WHO grade II: 47.8 months (25.2-55.7); grade III: 15.9 months (11.4-26.3); grade IV: 10.4 months (8.8-14.4). By DNA methylation profiling (n = 49), most tumors were classified as pediatric-type diffuse high-grade glioma (pedHGG), H3-/IDH-wild-type (n = 31/49, 63.3%) with enriched subclasses pedHGG_RTK2 (n = 19), pedHGG_A/B (n = 6), and pedHGG_MYCN (n = 5), but only one pedHGG_RTK1 case. Within the pedHGG, H3-/IDH-wild-type subgroup, recurrent alterations in EGFR (n = 10) and BCOR (n = 9) were identified. Additionally, we observed structural aberrations in chromosome 6 in 16/49 tumors (32.7%) across tumor types. In the pedHGG, H3-/IDH-wild-type subgroup TP53 alterations had a significant negative effect on OS. CONCLUSIONS:Contrary to previous studies, our representative pediatric GC study provides evidence that GC has a strong predilection to arise on the background of specific molecular features (especially pedHGG_RTK2, pedHGG_A/B, EGFR and BCOR mutations, chromosome 6 rearrangements).