OBJECTIVE:Drug-resistant epilepsy (DRE) affects approximately one-third of patients with epilepsy. The molecular heterogeneity underlying DRE remains poorly defined, largely due to limited access to resected brain tissue and substantial genetic diversity. Current classifications rely primarily on clinical symptoms and histopathological features rather than molecular mechanisms, constraining mechanistic insight and the development of targeted therapies. This study aimed to develop a transcriptome-based, machine learning-guided framework for molecular classification of DRE. METHODS:We performed comprehensive RNA sequencing on 153 surgically resected samples from 95 patients with DRE. Two transcriptomic subtypes were identified through unsupervised clustering. We also leveraged a weighted correlation network-based framework and systematic transcriptional signature comparison and developed a classification model using machine learning algorithms. RESULTS:Unsupervised clustering revealed two molecular subtypes that diverged from traditional pathological classifications, indicating an alternative transcriptomic basis for epilepsy pathogenesis. A classification model was constructed based on four key differentially regulated pathways: (1) neuroactive ligand-receptor interaction, (2) cAMP signaling, (3) γ-aminobutyric acid (GABA)ergic synapse, and (4) calcium signaling. Among the tested algorithms, the random forest model demonstrated superior performance, achieving 96% classification accuracy with an area under the curve (AUC) of .95. SIGNIFICANCE:These molecular subtypes and their pathways could serve as key molecular hallmarks of epilepsy, offering valuable insights for developing targeted therapies. Moreover, our findings introduce a novel framework for classifying epilepsy based on its molecular nature, potentially connecting the clinical symptoms with the underlying causes more effectively.

OBJECTIVES:Neuroblastoma (NB) is a prevalent pediatric solid malignancy associated with significant morbidity and mortality, largely driven by epigenetic alterations. This review aims to identify novel biomarkers related to long non-coding RNAs (lncRNAs) and DNA methylation in NB to enhance prognostic capabilities. METHODS:We conducted a detailed analysis of the interplay between lncRNAs and DNA methylation in NB, focusing on regulatory variations and their implications for disease progression. Key lncRNAs, including GTL2/MEG3, DALI, NBAT-1, and DLX6-AS1, were examined for their regulation by DNA methylation through cis- and trans-methylation mechanisms. RESULTS:There are clinical and biological implications of lncRNAs in NB and related cancers. Notably, GTL2 and its alias MEG3 are implicated in tumorigenesis through epigenetic modifications, such as hypermethylation, leading to the loss of gene expression and aggressive tumor behavior. Similarly, the interactions of DALI with adjacent genes illustrate the crucial role lncRNAs play in neuronal differentiation and tumor progression, suggesting their potential to impact prognosis through regulatory effects. Furthermore, NBAT-1 emerges as a promising tumor suppressor with strong correlations to NB prognosis, where its methylation-induced silencing is associated with negative outcomes. DLX6-AS1 is also linked to increased NB risk, with expression patterns correlating to disease stage and survival rates; however, more extensive survival data are required to establish its prognostic value. CONCLUSION:This review highlights the potential of lncRNAs as prognostic indicators in NB, emphasizing the need for further research to elucidate their roles and validate them as biomarkers for improved patient outcomes.

Background:Active oxidative phosphorylation is increasingly recognized as a defining metabolic feature of high-risk neuroblastoma (NB). NADH:ubiquinone oxidoreductase subunit S6 (NDUFS6), encoding an essential subunit of mitochondrial respiratory chain complex I, plays a critical role in sustaining oxidative phosphorylation. Nevertheless, its precise contribution to NB pathogenesis and progression remains largely undefined. Methods:To characterize the expression landscape of NDUFS6, publicly available single-cell and bulk RNA sequencing datasets were analyzed, and the GSE49710 dataset was used to evaluate its prognostic significance. Immunohistochemical staining was performed to assess NDUFS6 expression across distinct clinical subgroups. Stable SK-N-BE(2) and SH-SY5Y cell lines with NDUFS6 overexpression or knockdown were generated to conduct functional assays, including Cell Counting Kit-8 (CCK-8) proliferation, colony formation and Transwell assays. Transcriptomic alterations induced by NDUFS6 modulation were profiled by RNA sequencing. In parallel, a structure-based virtual screening of the MedChemExpress (MCE) Bioactive Compound Library Plus was conducted to identify candidate small-molecule inhibitors of NDUFS6, with selected compounds evaluated for cytotoxicity in NB cells. Results:NDUFS6 expression was significantly upregulated in high-risk NB (HR-NB) and was positively associated with disease progression and poor prognosis. Functional assays revealed that NDUFS6 knockdown suppressed proliferation, invasion, and migration of NB cells, whereas its overexpression promoted these malignant behaviors. Transcriptomic analysis revealed that high NDUFS6 expression activated pathways related to energy metabolism and adenosine triphosphate (ATP) synthesis, while concurrently suppressing neuronal differentiation and immune activation. Structure-based virtual screening identified several candidate inhibitors of NDUFS6, including guanosine 5'-triphosphate (disodium salt), 1,4-β-D-xylopentaose, and deferoxamine. Subsequent drug sensitivity assays demonstrated that guanosine-5'-triphosphate (disodium salt) exerted potent inhibitory effects in both MYCN-amplified and non-amplified NB cell lines. These findings underscore the oncogenic role of NDUFS6 and highlight its potential as a therapeutic target for precision treatment in HR-NB. Conclusions:NDUFS6 is significantly upregulated in HR-NB and contributes to tumor aggressiveness by promoting proliferation, migration, and invasion, accompanied by activation of metabolic pathways and suppression of neuronal differentiation and immune responses. Virtual screening identified guanosine 5'-triphosphate (disodium salt) as a potential NDUFS6 inhibitor with efficacy in both MYCN-amplified and non-amplified cells, highlighting NDUFS6 as a promising therapeutic target and providing a rationale for targeted intervention in HR-NB.

BACKGROUND:NTRK-rearranged spindle cell neoplasms (NTRK-RSCNs) are an emerging entity, molecularly characterized by NTRK rearrangements. Advances in molecular testing have revealed that NTRK fusions are prevalent in various tumor types, particularly in soft tissue tumors. CASE REPORT:A 5-year-old boy presented with frequent urination and a low fever. Imaging studies showed a large mass in the right kidney. Laboratory tests revealed hypophosphatemia and hypercalcemia. The patient received chemotherapy and radiation followed by nephrectomy. The tumor exhibited diverse high-grade sarcomatous morphological features distinct from common NTRK-RSCNs and was initially diagnosed as a malignant phosphaturic mesenchymal tumor (PMT) without molecular alteration evidence. Subsequent upgraded next-generation sequencing identified a TPM4::NTRK1 fusion, CDKN2A/B deletion, SDHD and NOTCH3 variants. Based on these molecular findings, the diagnosis was confirmed as an NTRK-rearranged high-grade sarcoma with an unusual histomorphological phenotype. CONCLUSION:This case highlights novel molecular characteristics of NTRK-rearranged high-grade sarcoma and underscores the critical role of comprehensive molecular profiling in diagnosing challenging cases.

OBJECTIVE This study aimed to characterize pediatric idiopathic pulmonary hemosiderosis (IPH) and assess whether adding bronchial artery embolization (BAE) to glucocorticoid (GC) therapy reduces the cumulative risk of disease recurrence. METHODS We conducted a retrospective cohort study of children with IPH between January 2003 and January 2020 from a single center. Demographic, clinicopathologic, therapeutic, and outcomes data were extracted. The primary outcome was cumulative recurrence of hemorrhage, analysed using the Andersen-Gill models. The secondary outcomes included time to first recurrence and change in hemoglobin levels from baseline to six months after treatment. RESULTS A total of 63 patients were included, with 44 (70%) receiving BAE plus GC treatment and 19 (30%) receiving GC treatment alone. The median age at diagnosis was 4.6 years (interquartile range: 2.1–7.6), and 28 (44%) were female. During a median follow-up period of 36 months (range, 0.7–120), 87 relapses occurred in the GC with BAE group versus 24 in the GC without BAE group. The adjusted hazard ratio was 1.66 (95% confidence interval [CI], 0.58–4.74) for the GC with BAE group compared with GC alone. The corresponding hazard ratio for time to first recurrence was 0.98 (95% CI, 0.39–2.44). Changes in hemoglobin levels did not differ significantly between the two groups (–20.5 [95% CI, − 27.4 to − 0.7] vs − 14.0 [95% CI, − 26.7 to 14.4], mean difference − 6.5 [95% CI, − 19.0 to 6.0], P = 0.30). CONCLUSIONS Our data do not support the addition of BAE to conventional GC therapy as routine primary treatment for pediatric IPH.

Gastrointestinal adenocarcinoma is a major cancer type for the digestive system, ranking as the top cause of cancer-related deaths worldwide. While there has been extensive research on mutations in protein-coding regions, the knowledge of the landscape of its non-coding regulatory elements is still insufficient. Combining the analysis of active enhancer profiles and genomic structural variation, we discovered and validated a lineage-specific super-enhancer for MYB in gastrointestinal adenocarcinoma. This super-enhancer is composed of a predominant enhancer e4 and several additional enhancers, whose transcriptional activity is regulated by the direct binding of HNF4A and MYB itself. Suppression of the super-enhancer downregulated the expression of MYB , inhibited downstream Notch signaling and prevented the development of gastrointestinal adenocarcinoma both in vitro and in vivo. Our study uncovers a mechanism driven by non-coding variations that regulate MYB expression in a lineage-specific manner, offering new insights into the carcinogenic mechanism and potential therapeutic strategies for gastrointestinal adenocarcinoma.

Background: Neuroblastoma (NB) is a well-known pediatric malignancy intertwined with neurodevelopment. Previously implicated in neuronal differentiation, Zinc Finger Protein 536 (ZNF536) has emerged as a promising prognostic and immune-related biomarker in our pan-cancer analysis. Methods: Single-cell RNA transcriptome sequencing, bulk transcriptome analysis, and immuno-histochemistry were used to assess ZNF536 expression and its association with prognosis. Cell proliferation, migration, invasion, and differentiation in ZNF536-knockdown NB cell lines were detected to evaluate the effect of ZNF536 on tumor cells. Vascular Endothelial Growth Factor Receptor 2 (VEGFR2), a potential target of ZNF536, and its downstream PI3K/AKT signaling cascade were investigated using transcriptome sequencing, CUT&Tag, quantitative real-time PCR (qRT-PCR), and Western blotting. The role of ZNF536 in tumorigenesis and the potential regulation axis was evaluated in vivo using a BALB/c nude mouse xenograft tumor model. Results: ZNF536 mRNA and protein expression were significantly higher in NB patients with poor prognosis. In vitro, ZNF536 knockdown curtailed proliferation, migration, and invasion of NB cells while fostering differentiation. ZNF536 regulated VEGFR2 expression, thus activating the PI3K-AKT pathway. In vivo, ZNF536 knockdown reduced tumor growth and proliferation via the VEGFR2-PI3K-AKT pathway. Conclusion: ZNF536 resulted as a novel prognostic biomarker in NB, promoting oncogenesis through VEGFR2-PI3K-AKT signaling axis modulation, suggesting its therapeutic potential in managing NB progression.

Background: Infantile fibrosarcoma (IFS) is the most prevalent soft tissue sarcoma in children under 1 year old and is known for its rapid growth. The tumor lacks specific immunohistochemical tumor marker and a general view of tumor microenvironment (TME). Its primary therapeutic intervention places patients at a risk of disability or mutilation. This study aimed to elucidate the universal transcriptional characteristics of IFS and explore novel targets for diagnosis and therapy using single-cell RNA sequencing (scRNA-seq). Methods: Fresh tissue samples of IFS for scRNA-seq were collected from four patients before other treatments were administered. We conducted cell clustering, inferring copy number variation from scRNAseq (InferCNV) analysis, gene differential expression analysis, cell function evaluation, Pearson correlation analysis, and cell-cell and ligand-receptor interaction analysis to investigate the distinct ecosystem of IFS. Results: According to the single-cell resolution data, we depicted the cell atlas of IFS, which comprised 14 cell populations. Through comparison with normal cells, the malignant cells were distinguished, and potential novel markers ( POSTN , IGFBP2 and CTHRC1 ) were identified. We also found four various functional malignant cell subtypes, three of which exhibited cancer stem cells (CSCs) phenotypes, and investigated the interplay between these subtypes and nonmalignant cells in the TME of IFS. Endothelial cells and macrophages were found to dominate the cell-cell communication landscape within the microenvironment, promoting tumorigenesis via multiple receptor-ligand interactions. Conclusions: This study provides a comprehensive characterization of the tumor transcriptome and TME of IFS at the cellular level, offering valuable insights for clinically significant advancements in the immunohistochemical diagnosis and treatment of IFS.

Introduction: Angiosarcoma is an exceedingly rare entity in pediatric population. Herein, we report two pediatric angiosarcoma with novel phenotypic and genotypic profile. Methods: The two patients' information was summarized by clinical data, histopathology, immunohistochemistry, genetic, treatment, and prognosis. Results: Two Chinese children presented with abdominal mass or consumptive hypothyroidism at 2 and 6 years. A patient presented with a unique histopathology of epithelioid AS with smooth muscle hyperplasia, and carried a novel somatic mutation in FAT1 (c.3929C > T/p. Ser1310Leu) along with germ-line variants in CDK8 (c.895A > C/p. Lys299Gln), FANCI (c.3906-07inv/p. Glu1303Lys), and MST1R (c.3581-83delinsACG/p. Arg1194-Ser1195delinsHisGly). The other patient presented with a novel clinical phenotype of consumptive hypothyroidism. They received postoperative treatment and were monitored for 20 and 26 months, showing good recovery. Conclusion: The phenotypic and genotypic spectrum of AS in pediatric population was expanded by these two patients, which requires the accumulating more cases to gain a deeper understanding.

Pediatric-type follicular lymphoma (PTFL) and pediatric nodal marginal zone lymphoma (PNMZL) are two rare indolent B-cell lymphomas with overlapping features. Recently, cases showing hybridizing features of PTFL and PNMZL have been reported. Herein, we retrospectively analyzed the clinicopathologic features of 59 patients, including 39 with PTFL, 5 with PNMZL, and 15 with mixed-type tumors (MTT). And next-generation sequencing analysis was performed on 3 PTFL, 2 PNMZL, and 2 MTT cases. In addition, previously published mutational data of 96 PTFLs, 25 PNMZLs, and 46 MTTs were also analyzed. There were 52 male and 7 female patients, with a median age of 17 years. Most patients (96.6

Background The phenomenon of males exhibiting higher lymphoma incidence and mortality rates than females has been widely recognized, and many survival analyses of certain lymphoma subtypes have incorporated sex as a critical prognostic determinant. However, comprehensive research on these sex-based differences in lymphomas remains limited. Methods This study aimed to conduct an integrated population-based analysis of sex-based disparities across various lymphoma subtypes using the Surveillance, Epidemiology, and End Results (SEER) database. We assessed the age-adjusted incidence rates of cases in the SEER22 and incidence-based mortality rates in the SEER12. The rates were stratified according to subtype, sex, race/ethnicity (incidence and mortality), age at diagnosis, and year of diagnosis (incidence only). Furthermore, we compared the risks of all-cause mortality, cancer-specific mortality, and cardiovascular disease-related mortality based on sex in SEER17. Results Our findings revealed that males had a higher likelihood of developing lymphoma and exhibited higher mortality rates than females for most lymphoma subtypes. These sex-based disparities in incidence and mortality were relatively consistent across five racial/ethnic groups. However, sex-based disparities in cancer incidence varied among age groups, and a trend toward narrowing differences with increasing age was observed in nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL), mantle cell lymphoma, and Burkitt lymphoma. Over the past two decades, trends in sex-based disparities of lymphoma incidence have remained stable. Regarding survival outcomes, males faced an increased risk of death compared to females for most lymphoma subtypes, except for NLPHL. Conclusions The results of our study underscore the complex and multifaceted nature of sex-based disparities in lymphoma and may offer valuable guidance for future research to unravel the potential causes of these differences.

Background: Castleman disease (CD) is a rare lymphoproliferative disease. Idiopathic multicentric CD (iMCD), representing a distinct entity in CD, is partly attributed to autoimmune abnormalities and the hyperplastic process in iMCD involving the immune system. Consequently, iMCD presents a range of overlapping manifestations with connective tissue disorder (CTD), resulting in an inability to tell whether they coexist or imitate each other. Reports of CD combined with CTD are rare, more cases are needed to be summarized and analyzed to improve the efficiency of diagnosis and accelerate the development of novel treatments. Case Description: A male pediatric patient was diagnosed with CTD in October 2019 and had been receiving regular treatment with tocilizumab and glucocorticoid or methotrexate since April 2020. He was further diagnosed with iMCD of the hyaline vascular subtype according to biopsy -proven histopathological features and imaging -proven multiple involvement in August 2021. He received 4 doses of rituximab and then a combination of thalidomide and dexamethasone for about 1 year. His clinical symptoms were well controlled throughout the disease for a long period, but inflammatory markers were repeatedly elevated, which eventually turned normal after switching to siltuximab from July 2023, although a significant elevation of interleukin-6 occurred. Conclusions: We reported a pediatric case diagnosed as CTD and iMCD, whose inflammation finally be well controlled by siltuximab. Hopefully, our work will add insight into such rare situations and it is undoubtedly that the pathophysiological mechanism of CD and CTD coexistence and prediction models of treatment response remains to be explored to facilitate the clinical management and optimal treatment.

AbstractMitochondrial transcription factor A (TFAM) deficiency may cause mtDNA depletion syndrome, which manifests as neonatal liver failure or primary ovarian insufficiency, hearing loss, seizures, and intellectual disability. Treatment focusing on symptomatic management, and the clinical prognosis remains poor. Here, we describe a novel case of TFAM mutation presenting with progressive neonatal cholestasis, hypoglycemia and abnormal amino acid profiling. The patient progressed to liver failure at 6 months of age but did not exhibit neurological involvement. No morphologic abnormalities were observed in muscle biopsy, while mtDNA copy number was reduced in comparison to age- and tissue-matched controls. After liver transplantation, liver biochemistries and blood amino acid profiling normalized three weeks later. Moreover, the boy was doing well post-transplant without any clinical concerns, and his development and neurological examination remain normal 33 months after liver transplantation. Our report suggests that liver transplantation appears to have a favorable profile in such patients.

As a major type of structural variants, tandem duplication plays a critical role in tumorigenesis by increasing oncogene dosage. Recent work has revealed that noncoding enhancers are also affected by duplications leading to the activation of oncogenes that are inside or outside of the duplicated regions. However, the prevalence of enhancer duplication and the identity of their target genes remains largely unknown in the cancer genome. Here, by analyzing whole-genome sequencing data in a non-gene-centric manner, we identify 881 duplication hotspots in 13 major cancer types, most of which do not contain protein-coding genes. We show that the hotspots are enriched with distal enhancer elements and are highly lineage-specific. We develop a HiChIP-based methodology that navigates enhancer-promoter contact maps to prioritize the target genes for the duplication hotspots harboring enhancer elements. The methodology identifies many novel enhancer duplication events activating oncogenes such as ESR1, FOXA1, GATA3, GATA6, TP63, and VEGFA, as well as potentially novel oncogenes such as GRHL2, IRF2BP2, and CREB3L1. In particular, we identify a duplication hotspot on Chromosome 10p15 harboring a cluster of enhancers, which skips over two genes, through a long-range chromatin interaction, to activate an oncogenic isoform of the NET1 gene to promote migration of gastric cancer cells. Focusing on tandem duplications, our study substantially extends the catalog of noncoding driver alterations in multiple cancer types, revealing attractive targets for functional characterization and therapeutic intervention.

Abstract The gastrointestinal adenocarcinoma is a major cancer type for the digestive system, ranking as the top cause of cancer-related deaths worldwide. In contrast to the large body of studies on the protein-coding regions' mutations, the knowledge about the landscape of its non-coding regulatory elements is still insufficient. Combining the analysis of genomic structural variation and active enhancers profile, we discovered and validated a lineage-specific super-enhancer for MYB in gastrointestinal adenocarcinoma. This super-enhancer is constituted by a predominant enhancer e4 and multiple facilitator enhancers, whose transcriptional activity is controlled by the direct binding of HNF4A and MYB itself. Suppression of the super-enhancer downregulated the expression of MYB, inhibited the downstream Notch signaling and prevented the development of gastrointestinal adenocarcinoma in vitro and in vivo. Our study revealed a non-coding variation-based mechanism to affect MYB expression in a lineage specific manner, which provided an inspiring insight into the carcinogenic mechanism and therapeutic strategies for gastrointestinal adenocarcinoma. ### Competing Interest Statement The authors have declared no competing interest.

Background: With the development of modern medicine and the application of multimodal treatment strategies, the survival rate after childhood malignant tumors as well as the incidence of Secondary malignant neoplasm (SMN) have increased. Case report: Patient is a Chinese girl. Initially diagnosed with Wilms tumor affecting her left kidney at the age of 6, she was later found to have TFE3-rearranged renal cell carcinoma (rRCC) affecting her right kidney at the age of 12. Employing NGS technology on specimens obtained from the TFE3-rRCC, we uncovered a significant somatic mutation within PRSS8 gene. This discovery sheds new light on the intricate genetic landscape underlying her condition, paving the way for further exploration and personalized treatment strategies. Conclusion: We consider that the etiology of SMN may be the result of chemoradiotherapy.

OBJECTIVE:To evaluate the efficacy of rituximab (RTX)-containing therapy as first-line as well as rescue treatment for giant cell hepatitis with autoimmune hemolytic anemia (GCH-AHA). METHODS:This retrospective study recruited patients diagnosed with GCH-AHA and treated with conventional immunosuppressor regimens consisting of prednisone or RTX-containing regimes consisting of RTX and prednisone, with or without another immunosuppressor. The primary outcomes were the complete remission (CR) rate and time-period required for CR. The secondary outcomes included relapses and adverse events. RESULTS:Twenty patients (8 females and 12 males; age range 1-26 months), 15 receiving conventional regimens and 5 receiving RTX-containing regimens, were included. The CR rates were 73.3 % (11/15) and 100 % (5/5) in the conventional and RTX-containing groups, respectively. The time-period required for CR was significantly shorter in the RTX-containing group than in the conventional group (6 (3-8) versus 14 (5-25) months, P = 0.015). Relapses occurred in 30.8 % (4/13) of patients in the conventional group; all achieved CR after adding RTX. Relapses occurred in 40.0 % (2/5) of patients in the RTX-containing group; both achieved CR after adding intravenous immune globulins or tacrolimus. Transient low immunoglobulin and infections were recorded in both groups. Treatment withdrawal was achieved in 73.3 % (11/15) and 60.0 % (3/5) of patients receiving conventional and RTX-containing regimens after 36 (2-101) and 22 (4-41) months, respectively. Two patients in conventional group died of disease progression and infection. CONCLUSIONS:RTX-containing first-line therapy achieves CR of GCH-AHA more quickly than the conventional therapy. RTX is efficacious when added to rescue therapy.