Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive sarcomas and a major cause of mortality in neurofibromatosis type 1 (NF-1). Distinguishing MPNSTs from benign neurofibromas remains challenging. We investigated fibroblast activation protein alpha (FAP) as a malignancy biomarker and theranostic target in peripheral nerve sheath tumors. Therefore, we integrated publicly available bulk transcriptomics, spatial transcriptomics, and single-cell RNA sequencing with immunohistochemistry (IHC) on independent archival samples. We further directly assessed clinical translatability using FAP-targeted PET/CT in an NF-1 patient undergoing work-up for suspected malignant transformation. Across independent bulk datasets, FAP was consistently up-regulated in MPNSTs compared with neurofibromas. In the TCGA sarcoma dataset, FAP varied by histotype but was clearly expressed in MPNSTs. Spatial transcriptomics revealed enrichment of FAP-high regions in MPNSTs and co-localization with tumor cell markers. Single-cell analysis showed FAP expression in MPNST tumor cells and cancer-associated fibroblasts, with the highest levels in neural crest-like tumor subpopulations previously linked to adverse prognosis; pseudotime analysis indicated decreasing FAP expression along trajectories toward Schwann cell precursor-like states linking FAP expression to a more primitive, dedifferentiated tumor cell state. IHC confirmed strong, predominantly tumor cell-intrinsic FAP expression in MPNSTs, with minimal staining in neurofibromas and normal tissues. Plexiform neurofibromas exhibited intermediate FAP expression. In clinical imaging, FAP-PET demonstrated higher tracer uptake in histologically proven MPNSTs than in benign lesions within the same patient, including a neurofibroma that was FDG-avid but FAP-negative, supporting added diagnostic specificity over FDG-PET/CT. In summary, FAP is robustly overexpressed in MPNSTs at transcript and protein levels, potentially concentrates in high-risk tumor cell states, and is detectable by targeted PET imaging. These findings identify FAP as a clinically relevant biomarker for malignancy in NF-1-associated tumors and support implementation of FAP-directed diagnostics and therapeutics in peripheral nerve sheath tumor work-up.

Background:Distal duplication 4q syndrome and distal 10q26 deletion syndrome are rare chromosomal abnormalities associated with complex and overlapping phenotypes. Most cases result from unbalanced inheritance of a parental translocation, yet the specific contributions of each chromosomal segment to the clinical phenotype remain poorly defined. Case presentation:We report on a male preterm infant born at 28 + 6 weeks' gestation who carried a derivative chromosome 10 due to a paternally inherited unbalanced translocation t(4;10)(q31.22;q26.13), leading to a 42.46 Mb duplication of 4q31.22-q35.2 and a 10.77 Mb deletion of 10q26.13-q26.3. The patient presented with severe postnatal linear growth restriction, delayed neurodevelopment, a giant umbilical hernia, bilateral renal hypoplasia, and relative overweight. Thumb anomalies were absent. Methods:Genetic analyses included chromosomal microarray, conventional karyotyping, fluorescence in situ hybridization (FISH), and transcriptome profiling from peripheral blood mononuclear cells. Results:Gene expression analysis confirmed reduced expression of established 10q26-related genes, including FGFR2, EMX2, WDR11, NSMCE4A, and EBF3, and additionally identified reduced expression of DMBT1, CUZD1, CTBP2, CHST15, OAT, and LHPP. These genes are implicated in epithelial-mesenchymal signaling, extracellular matrix organization, developmental regulation, and metabolic pathways. Regional transcriptomic analysis further supported dosage-associated effects, with lower averaged expression across genes represented on the array within the deleted 10q26.13-10q26.3 interval and higher averaged expression across genes within the duplicated 4q31.22-4q35.2 interval compared with the reference dataset. Adjacent flanking regions did not show comparable directional shifts, supporting the interpretation that the observed transcriptional changes were regionally aligned with the structural chromosomal imbalance rather than reflecting a global PBMC expression difference. Conclusion:This case illustrates how extended terminal deletions of 10q can disrupt key structural and metabolic gene networks. To our knowledge, this is the first transcriptomic characterization of a 10.77 Mb deletion in the 10q26.13-q26.3 region together with a large terminal 4q duplication. Integrating functional transcriptomics with clinical and cytogenetic data may enhance our understanding of rare chromosomal disorders and inform individualized management, including reproductive counseling and longitudinal clinical follow-up.

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.

Claudin-6 has emerged as a promising immunotherapeutic target, yet protein-level data in atypical teratoid/rhabdoid tumors (AT/RTs) have been inconsistent. We analyzed 36 well-characterized AT/RT samples and found membranous claudin-6 protein expression in 58% of cases, with striking enrichment in the molecular subgroup AT/RT-TYR (100%) and significantly higher staining scores compared with AT/RT-SHH and AT/RT-MYC. Correspondingly, AT/RT-TYR showed lower CLDN6 promoter methylation. Pilot spatial transcriptomic experiments in AT/RT-TYR confirmed heterogeneous CLDN6 transcription, and co-expression analyses linked CLDN6 to epithelial-mesenchymal transition-related genes. These findings clarify subgroup-specific expression patterns and support claudin-6 as a biologically and therapeutically relevant target in AT/RT-TYR.

Rhabdoid tumours (RT) are malignancies of the central nervous system, kidneys, liver and soft tissues that most commonly affect very young children with survival rates below 30% in high-risk cohorts. Treatment entails surgery, intensive chemotherapy and radiotherapy, associated with substantial short- and long-term toxicities. There is an unmet need to develop targeted therapies for RT to improve patient outcomes and mitigate the toxicities of current therapy. Detailed research followed by a workshop had the objective of enabling the development of targeted therapeutics for RT. Given the inherent commonality of their biology (i.e. biallelic inactivation of SMARCB1 or more rarely SMARCA4) the therapeutic approach should be similar for intra-cranial and extra-cranial tumours. DDB1–CUL4-associated factor 5 is a promising target, and the development of small molecule binders/degraders is a priority. Enhancer of zeste 2 polycomb repressive complex 2 subunit (EZH2) degraders may have greater therapeutic potential than inhibitors. Fibroblast growth factor receptor and platelet-derived growth factor receptor inhibitors may have value in subgroups. Mouse double minute 2 homologue (MDM2) is a priority target for novel therapeutic development and combination trials. Combinations of EZH2, MDM2 inhibitors and selective inhibitors of nuclear export should be evaluated robustly preclinically and drive early clinical studies.

BACKGROUND:Rhabdoid tumors (RT) are among the most aggressive pediatric malignancies, characterized by early onset in life, loss of SWI/SNF complex members (SMARCB1 or SMARCA4), and dismal outcomes despite multimodal therapy. Refractory and relapsing RT remain almost uniformly fatal, and targeted or immune-based approaches have yet to demonstrate clinical benefit. METHODS:To explore novel therapeutic vulnerabilities, we systematically investigated the expression of clinically actionable surface proteins that could serve as targets for antibody-drug conjugates (ADCs), radiopharmaceutical therapy (RPT), or cellular immunotherapies. Based on large-scale transcriptomic analyses, we prioritized FAP (fibroblast activation protein), CXCR4 (chemokine receptor 4), and IL13RA2 (interleukin receptor 13 RA2) and performed comprehensive protein-level validation by immunohistochemistry in an unprecedented cohort of 60 rhabdoid tumors spanning all molecular subgroups (ATRT-TYR, ATRT-SHH, ATRT-MYC, and eMRT). RESULTS:Integrating these data with spatial and single-nucleus transcriptomic profiling, we identified subgroup- and cell-type-specific expression patterns, including heterogeneous FAP distribution between stromal and tumor compartments and a distinct IL13RA2-positive rhabdoid cell population with melanosomal and stem-like features: Overall 51/60 tumors demonstrated positivity, defined as immunohistochemical H-score above or at least equal to 10, for FAP in the stroma and 34/60 tumors for CXCR4, indicating that a substantial fraction of rhabdoid tumors may be amenable to targeted therapies directed against these epitopes. CONCLUSION:These findings define a set of biologically and clinically relevant surface targets in RT and provide a translational blueprint for rational ADC and RPT target development in pediatric cancer.

Pediatric adrenocortical tumors (pACT) are biologically heterogeneous and incompletely classified by histopathology. To elucidate proteome-level tumor organization, we performed mass spectrometry–based proteomic profiling of 83 pACT and 43 normal adrenal samples. Unsupervised clustering identified four distinct molecular subtypes partially overlapping with histological diagnoses. These subtypes reflected discrete biological states: stromal–immune enrichment with reduced steroidogenesis, mitochondrial and steroidogenic activation, IGF/mTOR-driven anabolic reprogramming, and highly proliferative chromatin-remodeled carcinoma states. Proteomic clusters correlated strongly with endocrine phenotype, proliferative index, vascular invasion, and survival, outperforming conventional pathology. A five-protein classifier (DAAM2, CIP2A, TSC2, PALS2, P3H1) reproduced subtype structure with 92.8% cross-validated accuracy. Cluster-specific analysis therapeutic vulnerabilities, including IGF-axis activation and epigenetic and DNA replication targets in aggressive tumors. These data establish a proteome-based classification of pACT integrating metabolic, proliferative, immune features, providing a framework for molecularly guided risk assessment and precision therapy in this rare cancer. Statement of significance Proteome-based classification of pediatric adrenocortical tumors identifies four functional tumor states that integrate metabolism, proliferation, and lineage identity with clinical outcomes. A minimal protein classifier and subtype-specific therapeutic vulnerabilities define a translational framework for molecular stratification and precision therapy in this rare endocrine malignancy. ### Competing Interest Statement Matthias Mann is an indirect investor in Evosep Biosystems. ### Funding Statement The German MET studies were supported by the German Childhood Cancer Foundation (DKS 2021.11, DKS 2024.16, DKS 2025.07, and DKS 2025.16). The MoPACT project was funded by Mitteldeutsche Kinderkrebsforschung. This work was supported by the European Unions EU4Health Programme under the grant agreement number 101248798 and the Max Planck Society for the Advancement of Science. AM is supported by a PhD scholarship from the Onassis Foundation (Scholarship ID: F ZS 031-1/2022-2023) and by an interdisciplinary life science fellowship awarded by the Joachim Herz Stiftung. The funders had no role in the design, data collection, data analysis, and reporting of this study. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The MET studies were conducted in accordance with the Declaration of Helsinki and were approved by the ethics committees of the University of Luebeck (IRB 97125) and Otto-von-Guericke University Magdeburg (IRBs 174/12 and 52/22), Germany. The Molecular basis of Pediatric Adrenocortical Tumors (MoPACT) project (IRB 23-0351) was approved by the Ethics Committee of the Ludwig Maximilians University Munich, Germany. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes All data produced in the present study are available upon reasonable request to the authors

BACKGROUND:Atypical teratoid rhabdoid tumors (ATRTs) are highly aggressive pediatric central nervous system tumors defined by the inactivation of the SMARCB1 gene. Despite the identification of three distinct molecular subtypes, each defined by unique clinical and molecular characteristics, no subtype-specific therapeutic strategies are currently available. This highlights an urgent need to deepen our understanding of the cellular heterogeneity and developmental origins of ATRTs. METHODS:We generated a comprehensive single-nucleus transcriptomic atlas of ATRT samples, integrated it with single-nucleus ATAC-seq and spatial transcriptomics data, and validated our findings experimentally using patient-derived ATRT tumoroid models. RESULTS:Our analyses revealed distinct subtype-specific differentiation trajectories, each resembling different brain progenitor lineages. We identified key transcription factors that appear to drive these developmental pathways. Furthermore, a shared cycling, intermediate precursor cell (IPC)-like cell population, interspersed throughout tumors, was consistently present within all ATRT samples. We demonstrate that these subtype-specific differentiation pathways can be pharmacologically manipulated in patient-derived ATRT tumoroids. By directing tumor cells along their respective subtype-specific trajectories, we were able to induce a shift toward more differentiated, non-proliferative states. CONCLUSIONS:Collectively, our findings show that ATRTs recapitulate fetal brain signaling programs in a subtype-specific manner. This work provides a framework for understanding ATRT heterogeneity and supports the feasibility of maturation-based therapeutic strategies tailored to the molecular subtype of the tumor.

Cutaneous malignant melanoma is a common cancer in adults but extremely rare in young children, affecting fewer than one child per million each year in Europe. Because of its rarity, most treatments for children are adapted from adult therapies, despite possible biological differences. This study aimed to explore the molecular features of a rare and aggressive melanoma in a 16-month-old patient to understand disease progression and treatment resistance. We studied the tumour and metastases of a patient with a melanoma carrying an NRAS mutation, who received chemotherapy and immune checkpoint inhibitor treatment. The patient died 10 months after diagnosis. We used DNA methylation analysis, single-nucleus RNA sequencing, and deep spatial transcriptomic profiling to examine genetic changes, gene activity, and their spatial distribution in both the primary tumour and lymph node metastases. Here, we show that the tumour displayed high genetic and transcriptomic diversity. We identified increases in MITF and BRAF gene copies as likely key drivers of the aggressive disease, which were not detected at diagnosis. We also found activation of biological pathways, including VEGFA and WNT signalling, and abnormal activity of several genes linked to immune therapy response, with marked variation between tumour regions. This case demonstrates that paediatric melanoma can harbour complex and spatially variable molecular changes that contribute to rapid disease progression and treatment failure. Our findings support incorporating detailed spatial transcriptional profiling into clinical assessment to better guide therapy in rare paediatric cancers. Melanoma is a type of skin cancer that is common in adults but extremely rare in very young children. Because it is so uncommon, children are usually treated with approaches designed for adults, which may not work as well. In this case study, we investigated an aggressive case of melanoma in a 16-month-old child to understand why the cancer progressed quickly and did not respond to treatment. We examined samples from the original tumour and its spread to the lymph nodes using advanced techniques to map genetic changes and patterns of gene activity. We found specific changes in key cancer-related genes and signals that likely drove the disease and made treatment ineffective. These results highlight the importance of detailed tumour analysis to guide better treatment strategies for rare childhood cancers. Mucha et al. analyse tumour and metastases samples from a rare and aggressive melanoma in a 16-month-old child using advanced genetic and spatial profiling. They identify key gene changes and signaling pathways that likely drive rapid disease progression and treatment resistance, highlighting the need for detailed tumour mapping in paediatric melanoma.

BACKGROUND:Malignant rhabdoid tumors occasionally develop along cranial nerves, but clinical, histopathological, and molecular features have not been examined in larger series. PROCEDURE:We retrospectively interrogated data from the European Rhabdoid Registry, EU-RHAB, to identify malignant rhabdoid tumors affecting cranial nerves. We retrieved clinical information and reviewed magnetic resonance imaging (MRI) data. Furthermore, histopathological review and molecular profiling were performed. RESULTS:Among 425 patients, we identified a total of 14 harboring malignant rhabdoid tumors with cranial nerve involvement. Median age at diagnosis was 28 months (range: 0-13 years). Various cranial nerves were affected, the trigeminal nerve (n = 4) and the facial and/or vestibulocochlear nerve (n = 5) being most frequently involved. In most cases, the initial clinical and neuroradiological suspicion was schwannoma. Neuroradiology review of magnetic resonance imaging studies confirmed a tumor along the cranial nerve, but signal characteristics with restricted diffusion were rather suggestive of a malignant tumor of high cellularity. Histopathological examination, using among others neurofilament staining confirmed the diagnosis and infiltration of nerve fascicles. DNA methylation profiles demonstrated high similarity with ATRT-MYC as well as extracranial malignant rhabdoid tumors (median calibrated scores: 1.00). CONCLUSIONS:Malignant rhabdoid tumors of the cranial nerves represent a small but clinically distinct group, which initially is often not included in the differential diagnoses of pediatric cranial nerve tumors. Restricted diffusion on MRI may provide an early diagnostic clue. Histopathology and molecular signature are characteristic, but the developmental origin of malignant rhabdoid tumors of the cranial nerves remains to be determined.

DICER1 tumor predisposition syndrome is a genetic condition that increases the risk of developing certain cancer types. While thyroid tumors are the main tumors caused by this condition in adult oncology, children and adolescents with DICER1 germline mutations may suffer from a broader spectrum of tumors, including Sertoli-Leydig cell tumors, pleuropulmonary blastomas, embryonal rhabdomyosarcomas, and pineoblastomas. Although these diseases—many of which are hallmark tumors of DICER1 syndrome and rarely occur sporadically—have been known for several years, the more recent identification of DICER1 mutations in embryonal tumors with multilayered rosettes (ETMR) and DICER1-associated intra- and extracranial sarcomas has expanded the spectrum of tumor types potentially linked to DICER1 syndrome. This review sought to investigate the presence and characteristics of DICER1 mutations in rare CNS tumors and to discuss their potential implications for early recognition of DICER1-related syndromes. To address this, we conducted a comprehensive systematic literature review and analyzed data from our nationwide German database (CNS-InterREST) regarding these entities. When present, DICER1 mutation status, mutation type (somatic vs. germline), and localization within the gene were recorded. Demographic and clinical data—including age at diagnosis and tumor localization—were also evaluated where available. We found that the prevalence of DICER1 mutations in the cohort of ETMR patients included in the CNS-InterREST study was exceedingly low (1/31). The distribution of DICER1 mutations in patients with ETMR or intracranial sarcomas is comparable to that in other previously identified DICER1-mutant tumors. Our literature review demonstrates that within the 248 cases, which include three intracranial DICER1-mutated neoplasias and one reference group, most somatic mutations accumulate in the RNase IIIb domain, while germline mutations are usually evenly distributed throughout the gene. Overall, further research is necessary to unravel the cell-of-origin of the respective tumor types and whether other, hitherto undescribed, genetic factors may contribute to the development of ETMR and DICER1-associated intracranial sarcomas.

Rhabdoid tumors (RT) are among the most aggressive pediatric malignancies, characterized by early onset, loss of SWI/SNF complex members (SMARCB1 or SMARCA4), and dismal outcomes despite multimodal therapy. Refractory and relapsing RT remain almost uniformly fatal, and targeted or immune-based approaches have yet to demonstrate clinical benefit. To explore novel therapeutic vulnerabilities, we systematically investigated the expression of clinically actionable surface proteins that could serve as targets for antibody–drug conjugates (ADCs), radiopharmaceutical therapy (RPT), or cellular immunotherapies. Based on large-scale transcriptomic analyses, we prioritized FAP, CXCR4, and IL13RA2 and performed comprehensive protein-level validation by immunohistochemistry in an unprecedented cohort of 60 rhabdoid tumors spanning all molecular subgroups (ATRT-TYR, ATRT-SHH, ATRT-MYC, and eMRT). Integrating these data with spatial and single-nucleus transcriptomic profiling, we identified subgroup- and cell-type–specific expression patterns, including heterogeneous FAP distribution between stromal and tumor compartments and a distinct IL13RA2-positive rhabdoid cell population with melanosomal and stem-like features. These findings define a set of biologically and clinically relevant surface targets in RT and provide a translational blueprint for rational ADC and RPT target discovery in pediatric cancer.

BACKGROUND:Extracranial malignant rhabdoid tumors (eMRT) are rare, highly aggressive pediatric neoplasms. While the liver is a relatively common anatomic site of presentation, the clinical course of patients with hepatic eMRT (eMRT-L) is not well described. METHODS:We retrospectively analyzed 30 children affected by eMRT-L treated on a consensus regimen provided by the European Rhabdoid Registry (EU-RHAB). Clinical characteristics, radiology features according to the Pre-Treatment Extent of Tumor (PRETEXT) system, treatment details, and outcome were assessed. We employed patients with rhabdoid tumors of the kidney (RTK; n = 30) and other eMRT (n = 60) as controls. RESULTS:Median age at diagnosis was 8 months (range: 0-53 months), 16 of 30 patients (55%) presented with metastatic disease. R0 resection was achieved in seven patients (23%). Most tumors showed PRETEXT Stage ≥3 (66%) and frequently exhibited PRETEXT annotation factors. One-year overall and event-free survivals were both 17% (95% confidence interval: 7.5-37). Compared to RTK and other eMRT, patients with eMRT-L had significantly inferior outcomes (hazard ratios 2.47 and 4.39, respectively). Complete resection and absence of metastases were associated with improved survival. Consolidation therapies (i.e., radiotherapy or high-dose chemotherapy) were only rarely used. CONCLUSIONS:EMRT-L represents a distinct high-risk subgroup within the eMRT spectrum, characterized by inferior survival despite standardized multimodal therapy. Current treatment approaches demonstrate limited efficacy. Our results highlight the urgent need for prospective, collaborative studies to refine risk stratification and to evaluate novel treatment options.

Pediatric adrenocortical tumors (pACTs) are rare neoplasms with significant mortality, presenting diagnostic challenges in distinguishing benign from malignant tumors. Here, we employed a comprehensive multimodal molecular profiling approach to delineate and characterize molecular subgroups and investigated their potential for risk stratification. We profiled 109 pediatric adrenocortical tumors using Illumina methylation arrays and analyzed them together with 105 published samples. For 29 of the 109 tumors, fresh-frozen samples were analyzed with single-nucleus RNA sequencing and seven of these also with single-nucleus ATAC sequencing. Our analysis revealed four molecularly distinct subgroups with differences in clinical presentation, cellular composition and molecular signatures. Importantly, the herein identified high-risk subgroup demonstrated substantially reduced overall survival, with a 5-year survival rate of merely 27% compared to 81-95% in other subgroups. Notably, these tumors showed increased CpG island methylation and aberrant activation of the canonical WNT signaling pathway in gene expression profiling. Additionally, through integration of scRNA-Seq and scATAC-Seq, we uncovered complex transcriptional networks driving tumor biology. Tumor cells resembling the physiological adrenocortical zones exhibited unique gene expression patterns, with the high-risk subgroup characterized by an increased proportion of zona glomerulosa-like cells. These zona glomerulosa-like cells showed dysregulated WNT/CTNNB1 signaling and a proliferative and cell cycle activation signature, which emerge as a critical driver of malignancy. In particular, the glomerulosa-like tumor cells seem to fuel tumor proliferation by a crosstalk with fasciculata/reticularis-like tumor cells, which is unique for the high risk pACT subgroup. Further key molecular alterations include upregulation of cancer associated genes like PVT1 and MDM2, suggesting potential therapeutic vulnerabilities. Moreover, our analysis uncovered transmembrane glycoprotein BAMBI to be specifically upregulated in the high-risk tumors, highlighting its potential as a prognostic indicator. This comprehensive characterization provides a robust framework for risk stratification in pACTs. Specifically, we have unraveled a previously undescribed molecular hub using canonical WNT signaling to drive proliferation in high risk tumors. These changes may open an avenue towards integrating WNT-directed therapies in the clinical setting. Victoria E. Fincke, Maurice Loßner, Marina Kunstreich, Irmengard Sax, Marlena Mucha, Felix Dorn, Maria D. Hernandez Ramirez, Stefan Wudy, Christian Vokuhl, Christoph Slavetinsky, Jörg Fuchs, Michael C. Frühwald, Matthias Schlesner, Antje Redlich, Rainer Claus, Michaela Kuhlen, Pascal D. Johann. Decoding pediatric adrenocortical tumor heterogeneity through multimodal molecular profiling - pathways to improved risk stratification and therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3886.

Pediatric adrenocortical tumors (pACTs) are rare endocrine neoplasms with variable prognosis, commonly associated with germline pathogenic variants (PVs) in the tumor suppressor gene TP53. Here, we report the case of a 3.1-year-old female presenting with virilization and Cushing syndrome due to a left-sided adrenal mass. The tumor was completely resected and confirmed as stage II adrenocortical carcinoma (ACC) based on the Wieneke index. Comprehensive molecular profiling revealed heterozygous germline PVs in BRCA2 [c.9382C>T p.(Arg3128*)] and CHEK2 [c.1232G>A p.(Trp411*)]. These findings suggest a potential role of impaired DNA damage repair in ACC pathogenesis, as both PVs are associated with hereditary breast and ovarian cancer (HBOC) syndromes and genomic instability. This case expands the genetic spectrum of pACT and underscores the importance of advanced molecular analyses in identifying rare germline alterations that may inform personalized treatment strategies and cancer prevention programs. Although no additional treatment was required in this case, BRCA2 status highlights the potential for tailored therapeutic approaches, including poly(ADP-ribose) polymerase (PARP) inhibitors, in selected patients. Further research is warranted to explore the specific contributions of BRCA2 and CHEK2 PVs to ACC tumorigenesis and their implic ations for targeted therapies.

Rhabdoid tumors are aggressive pediatric cancers characterized by significant alterations in gene expression and tumor microenvironment composition. Our study aims to investigate the molecular mechanisms driving tumor aggressiveness and relapse in extracranial malignant rhabdoid tumors (eMRT) and atypical teratoid rhabdoid tumors (ATRT). Using high-resolution single-cell RNA sequencing and spatial transcriptomics, we analyzed gene expression patterns and cellular interactions within the tumor microenvironment, with a particular focus on immune cell composition and activity. Previous research has identified distinct DNA methylation subgroups in eMRT, associated with clinical outcomes such as high- and standard-risk profiles. These subgroups, linked to differences in survival rates, relapse frequencies, and SMARCB1 genetic alterations, provide a foundational framework for understanding tumor heterogeneity. Building on this framework, we explored subgroup-specific gene expression profiles and tumor microenvironmental dynamics, moving beyond methylation-based stratification. Pathway enrichment analysis identified key molecular pathways associated with immune evasion and tumor progression, providing insights into the tumor microenvironment and its role in disease progression. We observed enrichment in pathways such as nuclear-transcribed mRNA catabolic process (nonsense-mediated decay), and cytoplasmic translation, which are crucial for sustaining the high proliferative rates of tumor cells. Unique to the high-risk group in eMRT is the enrichment of pathways such as protein phosphorylation and plasma membrane-bounded cell projection assembly, which are critical for oncogenic signaling and invasive mechanisms. Immune cell characterization revealed a higher macrophage count in the eMRT high-risk group compared to the standard-risk group, aligning with their aggressive phenotype, as macrophages facilitate invasion and immune evasion. The high-risk eMRT group also showed elevated levels of growth factors (FGF9, HGF, PDGFC) and invasion-related proteins (SEMA3D, IL1RAPL1), with reduced ALB levels, which suggest enhanced proliferation and metabolic alterations characteristic of aggressive tumors. These findings offer a deeper understanding of the unique molecular and cellular landscapes of rhabdoid tumors, providing valuable insights to inform potential therapeutic targets. Maria D. Hernandez Ramirez, Marlena Mucha, Victoria E. Fincke, Martin Hasselblatt, Christian Vokuhl, Michael C. Frühwald, Pascal D. Johann. Molecular and immune landscape of extracranial malignant rhabdoid tumors (eMRT) and atypical teratoid rhabdoid tumors (ATRT): Insights into disease mechanisms [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7048.

Purpose: Extracranial malignant rhabdoid tumors (eMRT) are a challenging entity. Despite the use of multimodal treatment approaches, therapy failure occurs in 55% to 67% of these. Molecular markers for identification of patients at increased risk for relapse or refractory (R/R) disease are not available. Clinical characteristics may only insufficiently predict the individual course of disease.Experimental Design: Using the EU-RHAB database, we analyzed a cohort of 121 patients with eMRT clinically. For 81 patients, molecular and clinical data were available, which were further complemented with publicly available DNA molecular data from 92 eMRTs. We aimed to delineate molecular risk factors by dissecting the DNA methylome of these tumors. Moreover, we establish clinical characteristics and treatment details of R/R disease in a subcohort of 80 patients.Results: Using consensus hierarchical clustering, we identified three distinct subgroups, one of which (eMRT standard risk) was associated with significantly improved survival, irrespective of germline status and/or localization. At the transcriptome level, this subgroup was characterized by an overexpression of genes involved in muscle development. A relevant proportion of patients developed distant relapses or progressions; the median time to the event was 4 months, underlining the need for early identification and risk stratification of R/R disease. The overall survival was significantly decreased in patients with progressive disease when compared with relapse cases, and reaching complete remission during salvage therapy provided a survival benefit.Conclusions: Our analysis of eMRT in this comprehensive cohort provides novel insights into the patterns of relapse and integrates molecular and clinical risk factors to guide clinical decision-making.