PAX5 Alterations in a Consecutive Childhood B-Cell Acute Lymphoblastic Leukemia Cohort Treated Using the ALL IC-BFM 2009 Protocol.

In this study, we aimed to identify patients within our B-ALL cohort with altered PAX5. Our objective was to use a comprehensive analysis approach to characterize the types of genetic changes, determine their origin (somatic/germline), and analyze the clinical outcomes associated with them. A consecutive cohort of 99 patients with B-ALL treated at the Children's Hospital of the UMC Ljubljana according to the ALL IC-BFM 2009 protocol was included in our study. We used RNA sequencing data for gene expression analysis, fusion gene detection and single nucleotide variant identification, multiplex-ligation dependent probe amplification for copy number variation assessment, and Sanger sequencing for germline variant detection. PAX5 was impacted in 33.3% of our patients, with the genetic alterations ranging from CNVs and rearrangements to SNVs. The most common were CNVs, which were found in more than a third of patients, followed by point mutations in 5.2%, and gene rearrangements in 4.1%. We identified eight patients with a PAX5-associated genetic subtype that were previously classified as "B-other", and they showed intermediate outcomes. We showed higher minimal residual disease values at the end of induction and poorer event-free survival in hyperdiploid cases carrying duplications in PAX5 compared to other hyperdiploid cases. We also report an interesting case of a patient with PAX5::FKBP15 and a pathogenic variant in PTPN11 who underwent an early relapse with a monocytic switch. In conclusion, this study provides valuable insights into the presence, frequency, and prognostic significance of diverse PAX5 alterations in B-ALL patients, highlighting the complexity of genetic factors and their impact on patient outcomes.

Paediatric Medical Traumatic Stress in Children with Cancer and their Parents: Difference in Stress Levels Due to Illness and Treatment Factors.

Pediatric medical traumatic stress (PMTS) is a set of children's and their parents' psychological and physiological responses to pain, injuries, serious illnesses, and other experiences with the medical environment. Pediatric cancer patients have the highest prevalence of PMTS as the illness its treatment involve a set of stressors that trigger many negative psychological reactions. The current study examined the difference in levels of traumatic stress in children with cancer and their parents due to medical factors (type of cancer, outcome, duration, and intensity of treatment, time since diagnosis, relapse, and hospitalization in ICU). The study involved 183 parents of 133 children and 62 children and adolescents who were treated between 2009 and 2019 at the Clinical Department of Pediatric Hematology and Oncology of University Children's Hospital in Ljubljana. We collected the data using The Intensity of Treatment Rating Scale 2.0 [IRT-2], PTSD Checklist for Children/Parent [PCL-C/PR], The PTSD Checklist for DSM-5 [PCL-5] and The Child PTSD Symptoms Scale for DSM-5 [CPSS-5]. Traumatic stress symptoms are frequently present in both children and their parents, regardless of the cancer type, treatment duration, and treatment outcome. Children with relapse, children with more intensive treatment, and parents of the latter are at higher risk for PMTS occurrence. Additionally, we found a decreasing trend of traumatic responses after five or more years post-cancer diagnosis. [Table: see text].

Integrative Transcriptomic Profiling of the Wilms Tumor.

Our study aimed to identify relevant transcriptomic biomarkers for the Wilms tumor, the most common pediatric kidney cancer, independent of the histological type and stage. Using next-generation sequencing, we analyzed the miRNA profiles of 74 kidney samples, which were divided into two independent groups: fresh frozen tissue and formalin-fixed paraffin-embedded tissue samples. Subsequent mRNA expression profiling and pathway analysis were performed to establish the interplay and potential involvement of miRNAs and mRNA in the Wilms tumor. Comparative analysis, irrespective of post-dissection tissue processing, revealed 41 differentially expressed miRNAs, with 27 miRNAs having decreased expression and 14 miRNAs having increased expression in the Wilms tumor tissue compared to healthy kidney tissue. Among global mRNA transcriptomic profile differences, cross-sectional analysis suggested a limited list of genes potentially regulated by differentially expressed miRNAs in the Wilms tumor. This study identified the comprehensive miRNA and mRNA profile of the Wilms tumor using next-generation sequencing and bioinformatics approach, providing better insights into the pathogenesis of the Wilms tumor. The identified Wilms tumor miRNAs have potential as biomarkers for the diagnosis and treatment of the Wilms tumor, regardless of histological subtype and disease stage.

Toxicity and Clinical Results after Proton Therapy for Pediatric Medulloblastoma: A Multi-Centric Retrospective Study.

Medulloblastoma is the most common malignant brain tumor in children. Even if current treatment dramatically improves the prognosis, survivors often develop long-term treatment-related sequelae. The current radiotherapy standard for medulloblastoma is craniospinal irradiation with a boost to the primary tumor site and to any metastatic sites. Proton therapy (PT) has similar efficacy compared to traditional photon-based radiotherapy but might achieve lower toxicity rates. We report on our multi-centric experience with 43 children with medulloblastoma (median age at diagnosis 8.7 years, IQR 6.6, M/F 23/20; 26 high-risk, 14 standard-risk, 3 ex-infant), who received active scanning PT between 2015 and 2021, with a focus on PT-related acute-subacute toxicity, as well as some preliminary data on late toxicity. Most acute toxicities were mild and manageable with supportive therapy. Hematological toxicity was limited, even among HR patients who underwent hematopoietic stem-cell transplantation before PT. Preliminary data on late sequelae were also encouraging, although a longer follow-up is needed.

Clinical impacts of copy number variations in B-cell differentiation and cell cycle control genes in pediatric B-cell acute lymphoblastic leukemia: a single centre experience.

BACKGROUND: IKZF1 gene deletions have been identified as a poor prognostic factor in pediatric B-cell acute lymphoblastic leukemia (B-ALL), especially in the presence of co-occurring deletions (IKZF1plus profile). This study aimed to determine the frequency of IKZF1 deletions and deletions in other B-cell differentiation and cell cycle control genes, and their prognostic impact in Slovenian pediatric B-ALL patients. PATIENTS AND METHODS: We studied a cohort of 99 patients diagnosed with B-ALL from January 2012 to December 2020 and treated according to the ALL IC-BFM 2009 protocol. Eighty-eight bone marrow or peripheral blood samples were analysed for copy number variations (CNVs) using the SALSA MLPA P335 ALL-IKZF1 probemix. RESULTS: At least one CNV was detected in more than 65% of analysed samples. The most frequently altered genes were PAX5 and CDKN2A/B (30.7%, 26.1%, and 25.0%, respectively). Deletions in IKZF1 were present in 18.2% of analysed samples and were associated with an inferior 5-year event-free survival (EFS; 54.8% vs. 85.9%, p = 0.016). The IKZF1plus profile was identified in 12.5% of the analysed samples, and these patients had an inferior 5-year EFS than those with deletions in IKZF1 only and those without deletions (50.8% vs. 75.0% vs. 85.9%, respectively, p = 0.049). Overall survival (OS) was also worse in patients with the IKZF1plus profile than those with deletions in IKZF1 only and those without deletions (5-year OS 76.2% vs. 100% vs. 93.0%, respectively). However, the difference between the groups was not statistically significant. CONCLUSIONS: Our results are in concordance with the results obtained in larger cooperative clinical trials. Copy number variations analysis using the SALSA MLPA kit is a reliable tool for initial diagnostic approach in children with B-ALL, even in smaller institutions in low- and middle-income countries.

PATZ1 fusions define a novel molecularly distinct neuroepithelial tumor entity with a broad histological spectrum.

Large-scale molecular profiling studies in recent years have shown that central nervous system (CNS) tumors display a much greater heterogeneity in terms of molecularly distinct entities, cellular origins and genetic drivers than anticipated from histological assessment. DNA methylation profiling has emerged as a useful tool for robust tumor classification, providing new insights into these heterogeneous molecular classes. This is particularly true for rare CNS tumors with a broad morphological spectrum, which are not possible to assign as separate entities based on histological similarity alone. Here, we describe a molecularly distinct subset of predominantly pediatric CNS neoplasms (n = 60) that harbor PATZ1 fusions. The original histological diagnoses of these tumors covered a wide spectrum of tumor types and malignancy grades. While the single most common diagnosis was glioblastoma (GBM), clinical data of the PATZ1-fused tumors showed a better prognosis than typical GBM, despite frequent relapses. RNA sequencing revealed recurrent MN1:PATZ1 or EWSR1:PATZ1 fusions related to (often extensive) copy number variations on chromosome 22, where PATZ1 and the two fusion partners are located. These fusions have individually been reported in a number of glial/glioneuronal tumors, as well as extracranial sarcomas. We show here that they are more common than previously acknowledged, and together define a biologically distinct CNS tumor type with high expression of neural development markers such as PAX2, GATA2 and IGF2. Drug screening performed on the MN1:PATZ1 fusion-bearing KS-1 brain tumor cell line revealed preliminary candidates for further study. In summary, PATZ1 fusions define a molecular class of histologically polyphenotypic neuroepithelial tumors, which show an intermediate prognosis under current treatment regimens.

Diagnostics and treatment of diffuse intrinsic pontine glioma: where do we stand?

INTRODUCTION: Diffuse intrinsic pontine glioma (DIPG) is a rare clinically, neuro-radiologically, and molecularly defined malignancy of the brainstem with a median overall survival of approximately 11 months. Our aim is to evaluate the current tendency for its treatment in Europe in order to develop (inter)national consensus guidelines. METHODS: Healthcare professionals specialized in DIPG were asked to fill in an online survey with questions regarding usual treatment strategies at diagnosis and at disease progression in their countries and/or their centers, respectively. RESULTS: Seventy-four healthcare professionals responded to the survey, of which 87.8% were pediatric oncologists. Only 13.5% of the respondents biopsy all of their patients, 41.9% biopsy their patients infrequently. More than half of the respondents (54.1%) treated their patients with radiotherapy only at diagnosis, whereas 44.6% preferred radiotherapy combined with chemotherapy. When the disease progresses, treatment strategies became even more diverse, and the tendency for no treatment increased from 1.4% at diagnosis to 77.0% after second progression. 36.5% of the healthcare professionals treat children younger than 3 years differently than older children at diagnosis. This percentage decreased, when the disease progresses. Most of the participants (51.4%) included less than 25% of their patients in clinical trials. CONCLUSION: This survey demonstrates a large heterogeneity of treatment regimens, especially at disease progression. We emphasize the need for international consensus guidelines for the treatment of DIPG, possible by more collaborative clinical trials.

Low-dose continuous infusion of factor VIII in patients with haemophilia A.

BACKGROUND: Patients with haemophilia A (HA) or B (HB) can be given prophylactic or on-demand treatment administered by continuous infusion or bolus injections of factor VIII (FVIII) or IX (FIX). In this study we evaluated the efficacy and safety of low-dose continuous infusion of FVIII or FIX. MATERIAL AND METHODS: We studied all eligible patients with HA or HB treated with continuous infusion of factor concentrates over an 18-year period in a single Slovenian Haemophilia Comprehensive Care Centre. Treatment started with a bolus injection of FVIII or FIX, followed by continuous infusion at the initial rate of 2 IU/kg/h of FVIII in HA patients and 4.5 IU/kg/h of FIX in HB patients. The infusion rate was subsequently adjusted according to the indication for therapy. RESULTS: A total of 66 continuous infusions (40 in major surgery, 10 in minor surgery and 16 with bleeding episode) in 46 HA patients and 16 (15 in severe and 1 in mild HA) in eight HB patients were included in the study. During the first week of treatment, the median continuous infusion rates in HA patients undergoing major surgery, minor surgery and a bleeding event were 2.18 (0.75-3.68), 1.48 (1.0-2.54) and 2.24 (1.33-3.93) IU/kg/h, respectively. The median FVIII activities were 0.69 (0.37-1.19), 0.47 (0.39-0.84) and 0.52 (0.36-1.06) IU/mL. After the first week of treatment, even lower doses of FVIII were needed. Red blood cell transfusions had to be administered to three patients (2 with severe and 1 with moderate HA) during the continuous infusion and inhibitors developed in five patients. In HB patients, the median continuous infusion rate was 1.85 (1.07-2.94) IU/kg/h and the median FIX activity was 0.62 (0.30-1.04) IU/mL. Red blood cell transfusions were not required, and thrombophlebitis and inhibitors did not appear. DISCUSSION: Overall, low-dose continuous infusion was shown to be an effective and safe way of treating patients with HA. The protocol used also proved efficient and safe in all HB patients.

Diagnostic accuracy of lipopolysaccharide-binding protein for predicting bacteremia/clinical sepsis in children with febrile neutropenia: comparison with interleukin-6, procalcitonin, and C-reactive protein.

PURPOSE: In febrile neutropenia (FN), no reliable marker has been identified to discriminate between severe infection and other causes of fever early in the clinical course. Since lipopolysaccharide-binding protein (LBP) has proven to be an accurate biomarker of bacteremia/clinical sepsis in critically ill non-immunocompromised infants and children, we performed a prospective study to determine the diagnostic accuracy of LBP in children with FN. METHODS: Concentrations of LBP, procalcitonin (PCT), interleukin-6 (IL-6), and C-reactive protein (CRP) were prospectively measured on two consecutive days in 90 FN episodes experienced by 47 children. Receiver operating characteristic curve analysis was performed for each biomarker to predict bacteremia/clinical sepsis and severe sepsis. RESULTS: Eighteen of the 90 episodes were classified as bacteremia/clinical sepsis. On both days 1 and 2, all biomarkers had a low to intermediate diagnostic accuracy for sepsis, and no significant differences were found between them (area under the curve (AUC) for LBP, 0.648 and 0.714; for PCT, 0.665 and 0.744; for IL-6, 0.775 and 0.775; and for CRP, 0.695 and 0.828). Comparison of their AUCs to the AUC of maximum body temperature on admission (AUC = 0.668) also failed to show any significant differences. In severe sepsis, however, the best diagnostic accuracies were found for IL-6 and PCT (AUC 0.892 and 0.752, respectively), and these were significantly higher than those for LBP (AUC 0.566) on admission. CONCLUSIONS: On admission and 24 h later, the LBP concentration is less accurate for predicting bacteremia/clinical sepsis compared to IL-6, PCT, and CRP.

Treatment of severe human metapneumovirus (hMPV) pneumonia in an immunocompromised child with oral ribavirin and IVIG.

BACKGROUND: The clinical manifestations of human metapneumovirus (hMPV) infection resemble those of respiratory syncytial virus with the most severe disease occurring in infants, the elderly, chronically ill, and immunocompromised hosts. OBSERVATION: We present a case of a 2-year-old girl undergoing intensive chemotherapy for Burkitt lymphoma who developed severe hMPV pneumonia. Rapid and complete recovery was observed after treatment with oral ribavirin and intravenous immunoglobulin. CONCLUSION: As hMPV can cause severe pneumonia in immunocompromised patients and due to the reports of effective treatment with ribavirin, clinical studies to elucidate the role of ribavirin in treatment of hMPV pneumonia may be needed.

Multifocal hepatoblastoma in a 6-month-old girl with trisomy 18: a case report.

INTRODUCTION: Edward's syndrome (trisomy 18) is a rare entity with a reported incidence of 1/3000 to 1/7000 births. Less than 10% of patients survive beyond the first year of life, which may influence the fact that malignant tumors are rarely reported in association with this syndrome. CASE PRESENTATION: The authors report a rare case of a 6-month-old girl with trisomy 18 and multifocal hepatoblastoma. The course of the disease, autopsy results and review of the literature are presented. CONCLUSION: Our case represents the seventh published case of hepatoblastoma in a patient with trisomy 18. All of the seven published cases were women, possibly due to the high preponderance of females among the children with Edward's syndrome and longer survival of females with trisomy 18 compared to males. Since both trisomy 18 and hepatoblastoma are rare conditions, the probability that a child with trisomy 18 will independently develop a hepatoblastoma is very low. Therefore, we believe that the existence of these cases in children with trisomy 18 indicates a significant association. It can be assumed that trisomy 18 potentiates the development of hepatoblastoma. Careful clinical and post-mortem studies are needed to recognize the real frequency of hepatoblastoma in children with trisomy 18, who might die from different causes with unrecognizable hepatoblastoma.

No evidence of association of methylenetetrahydrofolate reductase polymorphism with occurrence of second neoplasms after treatment of childhood leukemia.

Methylenetetrahydrofolate reductase (MTHFR) polymorphisms have been associated not only with the risk for acute lymphoblastic leukemia (ALL) in adults and children, but also with increased methotrexate toxicity. The present study aimed to investigate whether MTHFR polymorphisms modify the risk for development of secondary malignancies in children treated for ALL with protocols that included high-dose methotrexate. MTHFR genotypes were determined in DNA samples isolated from archived bone marrow smears of 15 patients with a second malignancy and a matched control group of 30 patients who did not developed a second malignancy after the treatment for ALL. The frequencies of MTHFR C677T and A1298C genotypes in all patients were: C677T: CC 40%, CT 46.7% and TT 13.3% and A1298C: AA 46.7%, AC 44.4% and CC 8.9%. The relative risk for second malignancy was not significantly increased in ALL patients having at least one polymorphic C667T [odds ratio (OR) 1.51; 95% confidence interval (CI) 0.43-5.31] or one polymorphic A1298C allele (OR 1; 95% CI 0.29?-?3.46). Our study suggests that MTHFR polymorphisms are not associated with increased risk of second cancer in children treated with high-dose methotrexate.

Malignant blue nevus with lymph node metastases in five-year-old girl.

We report an unusual case of a malignant blue nevus in a five-year-old girl, which turned out to be malignant only after the development of lymph node metastases three years after the excision of the primary tumor on the patient's cheek. A functional bilateral neck dissection was performed and the patient is alive with no evidence of disease 8 years after the excision of the primary skin lesion.