Abstract Children, adolescent and young adults (CAYA) with relapsed high-grade gliomas (rHGG) share a dismal prognosis. Singular immunotherapeutic interventions like therapeutic vaccines have shown safety and immunogenicity, yet limited clinical efficacy. In the HIT-HGG Rez Immunovac phase I/II study (Eudra-CT 2013-000419-26) we optimzed induction and persistance of immune responses by combining upfront Treg-depletion using metronomic cyclophosphamide followed by therapeutic dendritic cell-based vaccines and subsequent checkpoint inibitor blockade (4x Nivolumab 3 mg/kg and Ipilimumab 1 mg/kg q3wk followed by Nivo mono 6 mg/kg q4wk for up to one year). Primary efficacy endpoint was to reach a 6-month post-relapse overall survival (prOS) of at least 82%. Twenty-five CAYAs with a mean age of 13.8 years [4.3-19.5] were enrolled between 2018-2024. Five patients were excluded due to progression before vaccine start or insufficient tumor material, 20 patients received therapy as per protocol. Fifteen SAEs were noted with no unexpected toxicities. The primary end point was confirmed with a 6-month prOS of 84% and one patient being alive after 31.2 months follow-up. PrOS was significantly improved over a historical control group (HCG, n = 87) from the HIT-HGG database (median OS 9.0 months, range 3.3-31.2 vs. 4.98 months, range 0.26-47.77 in Immunovac vs. HCG patients respectively, p=0.0003). Patients with stable disease at the end of vaccination had a better prOS than those relapsing during the vaccine schedule. Immunomonitoring demonstrated a decline in Tregs and a shift from naïve to memory T-cell subsets under treatment. Vaccine-specific CD4+ and CD8+ T-cell responses were observed in almost all patients, including responses against tumor-specific HLA-class I/II epitopes from the respective immunopeptidomes. Furthermore, spatial omics using cyclic immunofluorescence imaging technology at baseline revealed an immune-cold tumor microenvironment (TME) dominated by myeloid cells (∼50%), predominantly M2-polarized macrophages. T- and NK-cells contributed ∼10% and 15%, respectively. Immune effector cells were spatially dispersed, displayed an exhausted phenotype and were typically embedded within immunosuppressive glial niches or stromal compartments. The tumor compartment exhibited aggressive features, including high Ki67, strong GFAP expression, and frequent p53 alterations, but showed robust HLA-class II expression, revealing potential vulnerability to CD4+ T-cell-mediated responses. In conclusion, our data demonstrate safety and clinical efficacy of an optimized immunotherpeutic regime in CAYA with rHGG. Our immunomonitoring data show substantial T-cell responses and changes in the peripheral immune compartments. Together with insights from the TME these results will shape the design of future immunotherapy trials in HGG.

Objective: Mitotane is an effective treatment for advanced adrenocortical carcinoma (ACC). Given the limited pediatric data available, this study aims to evaluate the associations between mitotane dosage, plasma drug levels, and anthropometric measurements, as well as their potential impact on dosage requirements to optimize therapeutic outcomes in pediatric patients with ACC (pACC). Design and methods: A retrospective, international, multicenter study was conducted on pediatric ACC patients treated with mitotane across 18 centers. Mitotane serum levels were obtained from the Lysosafe Online® database or directly from the centers. Data from the cohort with plasma levels within the target range (≥14 mg/L; n = 319) were analyzed and compared to those with levels outside this range (n = 320). Results: Fifty pediatric patients (60% female) diagnosed between 2004 and 2023 were included, with a median follow-up of 34.5 months and a 10-year overall survival of 33 months. The median age at diagnosis was 8.6 years, with most tumors (84%) hormone-secreting. Among 49 patients undergoing surgery, 31 (62%) achieved R0 resection. The median treatment duration was 18 months, with a median mitotane dose of 87 mg/kg/day in patients within the target plasma level range, showing no significant difference from those outside the range. However, BMI was significantly associated with doses of plasma levels in target range (P = 0.001), as underweight (105.4 mg/kg/day) and healthy weight patients (98.4 mg/kg/day) required higher doses than overweight/obese patients (44.4 mg/kg/day). No significant differences in daily dose levels (mg/kg/day and mg/m2/day) were observed based on body weight. Conclusion: This study supports estimating mitotane dosages in pediatric ACC, emphasizing the need for close monitoring and frequent follow-ups at specialized centers due to individualized dosing and a narrow therapeutic window.

BACKGROUND:The HIT network was established in 2000 to create a population-based structure aiming to improve survival rates and reduce late effects for children with central nervous system (CNS) tumors by conducting comprehensive clinical trials. METHODS:The HIT network currently consists of 10 coordinating trial centers mandated by the German Society for Pediatric Oncology and Hematology (GPOH) to conduct clinical trials and research projects, and to provide counseling to local centers for individual patients. The network is complemented by 11 reference centers (neuropathology, tumor biology, neuroradiology, pediatric neurosurgery, cerebrospinal fluid [CSF], assessments, radiotherapy, genetics), biostatistical support, and currently 72 local treatment sites. RESULTS:Numbers of children and adolescents with newly diagnosed CNS tumors registered to trials and registries increased from approximately 500 to more than 600 per year, corresponding to >95% of affected HIT-eligible children and adolescents in Germany. Clinical counseling and upfront reference assessments ensure homogeneous clinical standards and avoid inadequate treatment of individual patients. Since 2007, the established reference services have been partially re-funded by German health insurances. DISCUSSION:The HIT network provides a unique structure for population-based state-of-the-art diagnostic assessments, treatment recommendations and counseling. It increases the "a priori" accuracy of stratification parameters, and the timely inclusion into clinical trials and tumor-specific registries. Favorable outcomes are achieved within the trials and registry landscape, for example, through consistent reference assessments, reducing the gap to real world data. Resulting data facilitate representative, unbiased high-quality research projects across all CNS tumor entities. Interdisciplinary cooperation and competitive scientific output are enhanced.

Introduction Structured physical exercise programs (PEPs) in pediatric oncology have shown benefits for physical, psychosocial, and cognitive outcomes, yet their integration into routine care varies globally. Objectives This study explores health care professionals' perceptions of PEPs across European, North African, and Arab pediatric oncology centers, comparing attitudes between centers with and without established programs, based on findings from an international survey. Materials and Methods A validated, 14-item, multilanguage questionnaire was distributed to health care professionals involved in pediatric oncology care, including physicians, nurses, physiotherapists, and sport therapists. Responses from 575 professionals (367 European; 208 North African/Arab) were analyzed, comparing perceptions according to region, profession, and PEPs availability at their centers. Results Overall, 97% of respondents with PEPs experience and 91% without considered PEPs beneficial. Perceived benefits extended beyond endurance and strength to mood, self-esteem, and quality of life, particularly among those with prior PEP exposure. European centers without PEP prioritized physical rehabilitation, whereas North African/Arab centers emphasized psychosocial benefits. Significant differences emerged regarding timing, setting, and relationship to physiotherapy. Most favored integrating PEP into standard care, with stronger support among experienced respondents. Conclusion Across regions, health care professionals recognize PEP as a valuable adjunct in pediatric oncology. Cultural norms and resource availability shape program focus, with European centers leaning toward physical rehabilitation and North African/Arab centers prioritizing psychosocial support. Findings highlight the need for balanced, context-sensitive strategies that integrate both physical and psychosocial elements, ensuring equitable access and optimized patient outcomes in diverse health care settings.

Background:Diffuse intrinsic pontine glioma (DIPG) remains uniformly lethal. Stereotactic biopsy confirms the diagnosis and enables molecular profiling. Metastasis along the biopsy track (BTM) has been reported only anecdotally; its prevalence, clinical relevance, and implications for treatment remain unclear. Methods:A multicenter retrospective study in patients with confirmed DIPG and BTM was conducted based on central neuroradiologic review. Radiotherapy schedules were re-assessed to evaluate the feasibility of upfront biopsy track irradiation. Results:Ten children met inclusion criteria (median age 6.8 years). Biopsy route was supratentorial in six and infratentorial in four children, and side-cutting needles were used predominantly. H3F3A mutations were most frequent (n = 8); TP53 alterations were common in tumors with extended molecular profiling available. Median PFS was 8.1 months. Five patients each developed BTM prior to (median 2.7 months) or concurrently with progression of primary tumor. There was no difference in overall survival (median OS 12.0 months) compared with the reference cohort. Estimated BTM prevalence among biopsied DIPG from additional registry data was between 6.9% and 13.0%. Primary biopsy track irradiation proved to be feasible, and comparing the surgical access routes, the infratentorial biopsy track hardly increased radiation exposure of the whole brain. Conclusions:Needle track metastasis is a rare progression pattern in stereotactic biopsied DIPG. Upfront irradiation of the biopsy track may represent a strategy to mitigate the potential risk of BTM. From a dosimetric perspective, an infratentorial approach may therefore be considered, as it was associated with only marginally increased radiation exposure.

BACKGROUND:Due to the novelty and rarity of infant-type hemispheric glioma (IHG), optimal treatment and factors determining clinical outcomes are yet to be established. METHODS:We curated a series of 164 patients with IHG; 155 identified by methodical literature search and nine additional patients contributed by collaborators. RESULTS:All tumors were hemispheric, diagnosed at a median age of 3.4 (0-52) months, and frequently (95%) non-metastatic. One hundred forty-two (86.5%) tumors harbored fusions involving receptor tyrosine kinase (RTK) genes (ALK [67/142, 47%], NTRK1/2/3 [32/142, 22.5%], ROS1 [29/142, 20.4%], MET [13/142, 9.2%], and ABL2 [1/142, 0.7%]). Sixty-four percentage, 20%, and 8% of patients were treated with surgery and adjuvant chemotherapy, surgery-only, and surgery plus targeted therapy, respectively. Five patients received radiation. Three-year event-free survival (EFS) and overall survival (OS) was 49.5% [40.7-60.2] and 79.6% [72.1-87.9], respectively. Twenty-two patients succumbed to disease, of which tumor progression (8/22, 36%) and intra-cranial hemorrhage (5/22, 23%) were the most common causes. Multivariate analysis showed that the factors most associated with an increased risk of death were no treatment except for surgery and presence of residual tumor after definitive surgery. These findings present a challenging dichotomy where surgery is both a serious risk factor for early death and, when successful, a benefit. CONCLUSIONS:Together, these findings show that IHG is a fusion driven tumor of the very young that is survivable even after progression. While optimal primary therapy for patients with IHG has yet to be established, the findings of this meta-analysis suggest treatment should focus on lowering surgical morbidity and improving its success.

Background: The HIT-HGG-2007 trial investigated temozolomide (TMZ) radio-chemotherapy for pediatric patients with high-grade gliomas (pHGG) to demonstrate therapeutic non-inferiority compared to previous intensive radio-chemotherapy regimens (HIT-GBM-C/-D). Methods: Between June 2009 and December 2016, 456 patients were enrolled into this international, prospective, single-arm, multicenter phase II trial in Germany, Austria, and Switzerland of whom 438 patients were evaluable for confirmatory analysis. Patients from the HIT-GBM-C/-D trials served as historic control (n = 439). Tumors of both cohorts with available tissue were re-classified according to the 2021 WHO classification of CNS tumors (n = 140). Results: Regarding event-free-survival (EFS) rate at 6 months, non-inferiority of the HIT-HGG-2007 regimen was confirmed (p = 0.0125). In terms of exploratory analyses, median EFS and overall survival (OS) was 9.5 months (95 % confidence interval [CI95], 8.9-10.4) and 14.7 months (CI95, 13.5-16.3), superior to intensive chemotherapy regimens (p < 0.0001 and p = 0.0328). EFS and OS remained superior after re-classification of tumors according to WHO2021 criteria. TMZ radio-chemotherapy had lower rates of severe hematological, gastrointestinal, and hepatic toxicity compared to HIT-GBM-C/-D. Younger age, WHO grade IV histology, tumor location in the brainstem or basal ganglia, and lower extent of resection were independent adverse risk factors for OS and EFS. MGMT gene promoter methylation status had no impact on EFS and OS. Conclusions: The HIT-HGG-2007 trial demonstrated non-inferiority compared with intensive chemotherapy regimens. MGMT promotor methylation status had no impact on survival. Exploratory analysis supports treatment of newly diagnosed non-pontine pHGG according to the HIT-HGG-2007 regimen due to improved EFS and OS rates together with a favorable toxicity profile.

Background:We assessed clinical features, treatment, and survival of pediatric patients with neurofibromatosis type 1 (NF1) with high-grade glioma (HGG). Methods:Patients from this retrospective cohort study were identified through an international collaborative effort by the SIOPE HGG/DIPG working group. NF1 was diagnosed based on clinical presentation and confirmed by either a pathogenic germline NF1 gene alteration or the exclusion of mismatch repair deficiency. A control cohort without genetic cancer predisposition was matched in a 2:1-ratio from the HIT-HGG database. Results:We identified 29 pediatric patients with NF1-associated HGG. Median age at diagnosis of HGG was 11 years. All but 1 tumor arose outside the optic pathway and included circumscribed and diffuse HGG. Molecular analysis in a subset of tumors identified an enrichment of alterations in CDKN2A, TP53, and ATRX. Event-free and overall survival were as poor as in matched sporadic HGG patients. The prognosis was not superior with upfront radiotherapy compared with delayed radiotherapy. Conclusions:NF1-associated HGGs behave as aggressively as their sporadic counterparts. The relevance of delaying radiotherapy until the time of progression and adjuvant MEK inhibitor treatment needs further investigation.

Aims: Embryonal tumours with PLAGL1 or PLAGL2 amplification (ET, PLAGL) show substantial heterogeneity regarding their clinical characteristics and have been treated inconsistently, resulting in diverse outcomes. In this study, we aimed to evaluate the clinical behaviour of ET, PLAGL and elucidate their response pattern across the different applied treatment regimens. Methods: We conducted an in-depth retrospective analysis of clinical and serial imaging data of 18 patients with ET, PLAGL (nine each of PLAGL1 and PLAGL2 amplified). Results: Patients with PLAGL1-amplified tumours (ET, PLAGL1) had fewer relapses (3/9), while PLAGL2-amplified tumours (ET, PLAGL2) were prone to early relapse or progression (8/9) and to distant, leptomeningeal and intraventricular relapses. Progression-free survival differed significantly between the subtypes (log-rank test, p = 0.0055). Postoperative treatment included chemotherapy (n = 17, various protocols), alone (n = 8) or combined with radiotherapy (n = 9). Responses to chemotherapy were observed in both subtypes, and incomplete resection was not associated with inferior survival. All three survivors with ET, PLAGL2 were treated with induction and high-dose chemotherapy with (n = 1—low-dose CSI and boost) or without (n = 2) radiotherapy, whereas five patients with less intensive chemotherapy relapsed. All six survivors with ET, PLAGL1 were treated with conventional chemotherapy regimens, with (n = 4—local radiotherapy n = 3; CSI and boost n = 1) or without (n = 2) radiotherapy. Two patients with ET, PLAGL1 relapsed after 8 years. Conclusions: Adjuvant therapy should be considered for all ET, PLAGL patients: Patients with ET, PLAGL2 might benefit from intensified chemotherapy regimens. In contrast, patients with ET, PLAGL1 showed superior outcomes without high-dose chemotherapy or craniospinal irradiation.

We prospectively analyzed antifungal use in immunocompromised children through a multicenter 12-week weekly point-prevalence survey in 31 hematology-oncology (HO) and hematopoietic stem cell/solid organ transplant (HSCT/SOT) units of 18 hospitals in 11 European countries. All patients hospitalized and receiving systemic antifungals were included. Ward policies, and weekly ward/patient data were collected. All 21 HO and 10 HSCT/SOT units had prophylaxis policies for high-risk patients (27/31 used azoles, 14/31 echinocandins and 15/31 liposomal amphotericin B [LAMB]). Among 572 courses recorded, prophylaxis was indicated in 439/572 (77%) and treatment in 133/572 (62/133 empirical, 43/133 pre-emptive, 28/133 targeted). Among patients receiving prophylaxis, 56% belonged to the non-high-risk group. Most common reasons for empirical, pre-emptive and targeted treatment were antibiotic-resistant febrile neutropenia (52%), abnormalities on chest-CT with/without positive galactomannan (77%) and candidiasis (82%), respectively. Fluconazole and LAMB were the most frequently prescribed agents both for prophylaxis (31%, 21%) and treatment (32%, 23%). Underdosing of micafungin for treatment in 50% of prescriptions and of fluconazole for treatment and prophylaxis in 70% of cases was noticed. In conclusion, most antifungal prescribing was for prophylaxis, with fluconazole being the main antifungal prescribed. Inadequate doses of antifungal prescribing and prophylaxis of non-high-risk patients could be targets for improvement.

Pediatric high-grade glioma (pedHGG) can occur as first manifestation of cancer predisposition syndromes resulting from pathogenic germline variants in the DNA mismatch repair (MMR) genes MSH2, MSH6, MLH1, and PMS2. The aim of this study was to establish a generalized screening for Lynch syndrome and constitutional MMR deficiency (CMMRD) in pedHGG patients, as the detection of MMR deficiencies (MMRD) may enable the upfront therapeutic use of checkpoint inhibitors and identification of variant carriers in the patients' families. We prospectively enrolled 155 centrally reviewed primary pedHGG patients for MMR-immunohistochemistry (IHC) as part of the HIT-HGG-2013 trial protocol. MMR-IHC results were subsequently compared to independently collected germline sequencing data (whole exome sequencing or pan-cancer DNA panel next-generation sequencing) available in the HIT-HGG-2013, INFORM, and MNP2.0 trials. MMR-IHC could be successfully performed in 127/155 tumor tissues. The screening identified all present cases with Lynch syndrome or CMMRD (5.5%). In addition, MMR-IHC also detected cases with exclusive somatic MMR gene alterations (2.3%), including MSH2 hypermethylation as an alternative epigenetic silencing mechanism. Most of the identified pedHGG MMRD patients had no family history of MMRD, and thus, they represented index patients in their families. Cases with regular protein expression in MMR-IHC never showed evidence for MMRD in DNA sequencing. In conclusion, MMR-IHC presents a cost-effective, relatively widely available, and fast screening method for germline MMRD in pedHGG with high sensitivity (100%) and specificity (96%). Given the relatively high prevalence of previously undetected MMRD cases among pedHGG patients, we strongly recommend incorporating MMR-IHC into routine diagnostics.

Diffuse pediatric-type high-grade gliomas (pedHGG), H3- and IDH-wildtype, encompass three main DNA-methylation-based subtypes: pedHGG-MYCN, pedHGG-RTK1A/B/C, and pedHGG-RTK2A/B. Since their first description in 2017 tumors of pedHGG-RTK2A/B have not been comprehensively characterized and clinical correlates remain elusive. In a recent series of pedHGG with a Gliomatosis cerebri (GC) growth pattern, an increased incidence of pedHGG-RTK2A/B (n = 18) was observed. We added 14 epigenetically defined pedHGG-RTK2A/B tumors to this GC series and provided centrally reviewed radiological, histological, and molecular characterization. The final cohort of 32 pedHGG-RTK2A/B tumors consisted of 25 pedHGG-RTK2A (78

Background:Ependymomas of the spinal cord are rare among children and adolescents, and the individual risk of disease progression is difficult to predict. This study aims to evaluate the prognostic impact of molecular typing on pediatric spinal cord ependymomas. Methods:Eighty-three patients with spinal ependymomas ≤22 years registered in the HIT-MED database (German brain tumor registry for children, adolescents, and adults with medulloblastoma, ependymoma, pineoblastoma, and CNS-primitive neuroectodermal tumors) between 1992 and 2022 were included. Forty-seven tumors were analyzed by DNA methylation array profiling. In 6 cases, HOXB13 and MYCN proteins were detected as surrogate markers for specific methylation classes. Ten patients had NF2-related schwannomatosis. Results:With a median follow-up time of 4.9 years, 5- and 10-year overall survival (OS) were 100% and 86%, while 5- and 10-year progression-free survival (PFS) were 65% and 54%. Myxopapillary ependymoma (SP-MPE, n = 32, 63%) was the most common molecular type followed by spinal ependymoma (SP-EPN, n = 17, 33%) and MYCN-amplified ependymoma (n = 2, 4%). One case could not be molecularly classified, and one was reclassified as anaplastic pilocytic astrocytoma. 5-year PFS did not significantly differ between SP-MPE and SP-EPN (65% vs. 78%, P = .64). MYCN-amplification was associated with early relapses (<2.3 years) in both cases and death in one patient. Patients with SP-MPE subtype B (n = 9) showed a non-significant trend for better 5 years-PFS compared to subtype A (n = 18; 86% vs. 56%, P = .15). The extent of resection and WHO tumor grades significantly influenced PFS in a uni- and multivariate analysis. Conclusions:Molecular typing of pediatric spinal ependymomas aids in identifying very high-risk MYCN-amplified ependymomas. Further insights into the molecular heterogeneity of spinal ependymomas are needed for future clinical decision-making.

Purpose: To explore the perception of physical exercise programs for pediatric oncology patients among childhood cancer care professionals. We also aimed at comparing such perceptions between cultures. Healthcare professionals’ endorsement may be essential for initiating and promoting such programs. Methods: An anonymous survey was designed and administered voluntarily to childhood cancer care professionals (including pediatric oncologists, nurses, and physiotherapists) in European, North-African and Arab pediatric oncology centers. Results: Five-hundred-and twenty-eight professionals from 14 sites answered the survey. Most respondents considered physical exercise programs as a suitable therapeutic approach for pediatric cancer patients with a potential positive contribution to survival (81%), wellbeing (82%), quality of life (80%), and self-esteem (75%). 91% of respondents would also support the future introduction of physical exercise programs into standard pediatric oncological care. There was a comparatively higher appreciation of physical exercise programs among European centers compared to North-African / Arab centers. Conclusion: We registered a broad acceptance of physical exercise programs among all European and North-African / Arab childhood cancer care professionals. The positive perception was independent of any pre-existing experience with such programs and seems therefore representative. This finding may encourage the further promotion of physical exercise programs in pediatric oncology.

Abstract BACKGROUND Malignant transformation (MT) of pediatric low-grade glioma (pLGG) to secondary high-grade glioma (sHGG) is a rare but momentous event. We explored its clinical, epidemiologic, and molecular characteristics and risk factors. METHODS Forty-eight pediatric patients with histology-based or radiological (n=4) diagnosis of primary LGG (median age 10.2 years [range, 0.7–16.9]) and subsequent sHGG were identified within registries of the HIT-network from 1996-2019. For 29 patients, tumor specimens of both pLGG and corresponding sHGG were available for neuropathological review and studied genetically. RESULTS MT occurred after median 4 years (range, 0.5-14) following LGG diagnosis. Its 15-year cumulative incidence was 1.6±0.4% (23/1586 SIOP-LGG-2004 patients), but 11.7±4.1% (9/100) for diffuse pLGG WHO-grade 2 (DG2). Clinical risk factors for MT were spinal location and prior dissemination. MT was accompanied by an increment of nuclear p53 accumulation (20/29 tumors), cytogenetic and molecular alterations. CDKN2A/B homozygous deletions (7/12 tumors), 1p/1q alterations (4/12) and ATRX loss (3/12) were the most common acquired alterations detected in sHGG. 8/16 tumors were unclassifiable by methylation profiling. DG2s (n=19), harboring IDH-1 mutation in 3/9 cases, progressed to diffuse WHO-grade 3 (n=8) or grade 4 HGG (n=9), two developed gliomatosis cerebri. Circumscribed LGG (PA, n=13; GG, n=7; PXA, n=4; DNET, n=1) typically harbored MAPK/ERK pathway alterations (BRAF-/NTRK-/FGFR-fusions, n=5/9; BRAFV600E mutation, n=4/12) and transformed to non-diffuse HGG (n=14), but also to diffuse grade 4 HGG. H3 K27M-/BRAFV600E co-mutation in two of these patients eventually resulted in MT to fatal diffuse midline glioma. Thirty-five sHGG patients received radio-chemotherapy. Overall survival at 5 and 10 years following MT was 49±8% and 31±8%. CONCLUSIONS Our findings underline the importance of careful long-term follow-up of patients with pLGG. Repeated resection/biopsy of recurrent/progressing or “atypical” tumors should be pursued and include molecular typing to identify patients at risk for MT and initiate appropriate treatment.

Abstract BACKGROUND Pediatric central nervous system (CNS) tumors can occur as first manifestations of cancer predisposition syndromes (CPS) resulting from pathogenic variants in DNA mismatch repair (MMR) genes MLH1, MSH2, MSH6, and PMS2. Early detection of MMR deficiencies (MMRD) may warrant the therapeutic use of checkpoint inhibitors and enable genetic consulting for the patient’s families. METHODS We prospectively screened 128 pediatric high-grade gliomas (pedHGG) for MMR protein expression by immunohistochemistry (IHC) as part of the HIT-HGG-2013 clinical trial. We compared IHC results to independently collected patient information on CPS including MMRD to test the method’s efficiency in screening for Lynch syndrome and Constitutional MMRD syndrome CMMRD. RESULTS From 128 successfully tested tumors, ten cases (10/128, 7.8%), showed loss of expression of at least one of the MMR proteins. In seven of the ten affected patients (7/128, 5.5%), genetic testing uncovered heterozygous (6/10) or homozygous (1/10) pathogenic germline variants in MMR genes (MLH1, MSH2 or MSH6) confirming the diagnosis of Lynch syndrome respectively CMMRD. In three cases (3/128 2.3%), MMR alterations were restricted to tumor tissue. The group of diffuse midline gliomas, H3 K27-altered, (67/128, 52.3%) did not show MMR alterations, while either somatic or germline MMR mutations were detected in two of eight patients with diffuse hemispheric glioma, H3 G34-mutant. In 88 patients (88/128, 68.8%) either clinical or molecular-genetic information on CPS could be provided. All ten patients with signs of MMRD were successfully detected by IHC from tumor tissue. CONCLUSIONS IHC for MMR proteins represents a highly sensitive screening method for MMRD in pedHGG. Considering the occurrence of at least 11.5% (7/61) of MMRD in pediatric patients with diffuse high-grade glioma excluding DMG, MMRD IHC should be part of routine diagnostics as cost-effective, broadly available and robust screening method.