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Large datasets in paediatric oncology are inherently rare. Therefore, it is paramount to fully exploit all available data, which are distributed over several resources, including biomaterials, images, clinical trials, and registries. With privacy-preserving record linkage (PPRL), personalised or pseudonymised datasets can be merged, without disclosing the patients' identities. Although PPRL is implemented in various settings, use case descriptions are currently fragmented and incomplete. The present paper provides a comprehensive overview of current and future use cases for PPRL in paediatric oncology. We analysed the literature, projects, and trial protocols, identified use cases along a hypothetical patient journey, and discussed use cases with paediatric oncology experts. To structure PPRL use cases, we defined six key dimensions: distributed personalised records, pseudonymisation, distributed pseudonymised records, record linkage, linked data, and data analysis. Selected use cases were described (a) per dimension and (b) on a multi-dimensional level. While focusing on paediatric oncology, most aspects are also applicable to other (particularly rare) diseases. We conclude that PPRL is a key concept in paediatric oncology. Therefore, PPRL strategies should already be considered when starting research projects, to avoid distributed data silos, to maximise the knowledge derived from collected data, and, ultimately, to improve outcomes for children with cancer.

Background: Liquid biopsies are revolutionary tools used to detect tumor-specific genetic alterations in body fluids, including the use of cell-free DNA (cfDNA) for molecular diagnosis in cancer patients. In brain tumors, cerebrospinal fluid (CSF) cfDNA might be more informative than plasma cfDNA. Here, we assess the use of CSF cfDNA in pediatric embryonal brain tumors (EBT) for molecular diagnosis., Methods: The CSF cfDNA of pediatric patients with medulloblastoma ( n = 18), ATRT ( n = 3), ETMR ( n = 1), CNS NB FOXR2 ( n = 2) and pediatric EBT NOS ( n = 1) (mean cfDNA concentration 48 ng/mL; range 4-442 ng/mL) and matched tumor genomic DNA were sequenced by WES and/or a targeted sequencing approach to determine single-nucleotide variations (SNVs) and copy number alterations (CNA). A specific capture covering transcription start sites (TSS) of genes of interest was also used for nucleosome footprinting in CSF cfDNA., Results: 15/25 CSF cfDNA samples yielded informative results, with informative CNA and SNVs in 11 and 15 cases, respectively. For cases with paired tumor and CSF cfDNA WES ( n = 15), a mean of 83 (range 1-160) shared SNVs were observed, including SNVs in classical medulloblastoma genes such as SMO and KMT2D. Interestingly, tumor-specific SNVs (mean 18; range 1-62) or CSF-specific SNVs (mean 5; range 0-25) were also observed, suggesting clonal heterogeneity. The TSS panel resulted in differential coverage profiles across all 112 studied genes in 7 cases, indicating distinct promoter accessibility., Conclusion: CSF cfDNA sequencing yielded informative results in 60% (15/25) of all cases, with informative results in 83% (15/18) of all cases analyzed by WES. These results pave the way for the implementation of these novel approaches for molecular diagnosis and minimal residual disease monitoring.

Background: To assess whether expectant observation of infants ≤ 90 days old with small suprarenal masses (sSRMs) could avoid unnecessary surgery without impacting outcome. Methods: Infants ≤ 90 days with a ≤ 5 cm mass, without midline extension or lymph node or distant spread were registered (ClinicalTrials.org:NCT01728155). Once staging was completed, they were followed with ultrasound, MRI and urinary catecholamines. Surgical resection was only planned if there was a ≥40% mass volume increase or for a mass persisting after 48 weeks of the planned observation. Results: Over a 5-year period, 128 infants were registered. No infant had detectable MYCN amplification in the peripheral blood. Surgery was performed in 39 (30.5%) patients, in 18 during and in 21 after the planned 48-week observation, and 74% were confirmed to be neuroblastomas. Non-life-threatening surgical complications occurred in two cases. The 3-year overall survival and event-free survival were 100% and 87.1%, respectively. The 16 events observed were volume increase (N = 11) and progression to neuroblastoma stage MS (N = 5). Patients with solid masses or MIBG-positive masses had lower EFS. Conclusions : Expectant observation for infants with sSRMs with clinical follow-up and timely imaging (including MRI scan) is safe and effective, allowing surgery to be avoided in the majority of them.

Ornithine decarboxylase (ODC1), a critical regulatory enzyme in polyamine biosynthesis, is a direct transcriptional target of MYCN, amplification of which is a powerful marker of aggressive neuroblastoma. A single nucleotide polymorphism (SNP), G316A, within the first intron of ODC1 , results in genotypes wildtype GG, and variants AG/AA. CRISPR-cas9 technology was used to investigate the effects of AG clones from wildtype MYCN -amplified SK-N-BE(2)-C cells and the effect of the SNP on MYCN binding, and promoter activity was investigated using EMSA and luciferase assays. AG clones exhibited decreased ODC1 expression, growth rates, and histone acetylation and increased sensitivity to ODC1 inhibition. MYCN was a stronger transcriptional regulator of the ODC1 promoter containing the G allele, and preferentially bound the G allele over the A. Two neuroblastoma cohorts were used to investigate the clinical impact of the SNP. In the study cohort, the minor AA genotype was associated with improved survival, while poor prognosis was associated with the GG genotype and AG/GG genotypes in MYCN -amplified and non-amplified patients, respectively. These effects were lost in the GWAS cohort. We have demonstrated that the ODC1 G316A polymorphism has functional significance in neuroblastoma and is subject to allele-specific regulation by the MYCN oncoprotein.

Despite intensive treatment, 50% of children with high-risk neuroblastoma (HR-NB) succumb to their disease. Progression through current trials evaluating the efficacy of new treatments for children with HR disease usually depends on an inadequate response to induction chemotherapy, assessed using imaging modalities. In this study, we sought to identify circulating biomarkers that might be detected in a simple blood sample to predict patient response to induction chemotherapy. Since exosomes released by tumor cells can drive tumor growth and chemoresistance, we tested the hypothesis that exosomal microRNA (exo-miRNAs) in blood might predict response to induction chemotherapy. The exo-miRNAs expression profile in plasma samples collected from children treated in HR-NBL-1/SIOPEN before and after induction chemotherapy was compared to identify a three exo-miRs signature that could discriminate between poor and good responders. Exo-miRNAs expression also provided a chemoresistance index predicting the good or poor prognosis of HR-NB patients., Competing Interests: The authors declare no conflict of interest.

Simple Summary: Neuroblastoma is the third most common type of childhood cancer but is responsible for over 15% of all pediatric cancer deaths. Children with advanced disease have less than a 50% survival rate and are in desperate need of new treatment options. Protein phosphatase 2A (PP2A) is a protein that inhibits tumors, but it is turned off in neuroblastoma. We have previously shown that we can activate PP2A in neuroblastoma with certain drugs and decrease tumor growth, but these drugs also affect the function of the immune system. We now have new small molecules that activate PP2A without upsetting the immune system. We observed a decrease in tumor growth, proliferation, and motility with our novel PP2A activators. We feel that a better understanding of how activating PP2A inhibits neuroblastoma will lead to better treatments for these children. Background: Neuroblastoma (NB) remains one of the deadliest pediatric solid tumors. Recent advancements aimed at improving outcomes have been insufficient, and patients with high-risk NB continue to have a poor prognosis. Protein phosphatase 2A (PP2A) is a tumor suppressor protein downregulated in many cancers, including NB. PP2A activation has been shown to affect the malignant phenotype in other solid tumors. The present studies aim to investigate the effects of two novel PP2A activators as a NB therapeutic. Methods: Four established NB cell lines and a patient-derived xenoline were utilized to study the effect on cell viability, proliferation, motility, and in vivo tumor growth using two novel tricyclic sulfonamide PP2A activators, ATUX-3364 and ATUX-8385. Results: ATUX-3364 and ATUX-8385 increased PP2A activity. These PP2A activators led to decreased viability, proliferation, and motility of NB cells. Treatment of animals bearing NB tumors with ATUX-3364 or ATUX-8385 resulted in decreased tumor growth in MYCN-amplified SK-N-BE(2) tumors. At the molecular level, PP2A-based reactivation led to dephosphorylation of MYCN-S62 and decreased MYCN protein expression. Conclusions: PP2A activators decreased NB cell viability, proliferation, and motility. In vivo experiments show that PP2A activators have more significant effects on tumorigenesis in MYCN-amplified tumors. Finally, phosphorylation of MYCN protein was decreased following treatment with novel sulfonamide PP2A activators. These data and mechanistic insights may be useful for developing new PP2A-based therapies that target MYCN for the treatment of NB. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma (NBL) is the most common extracranial cancer in children. High-risk NBL is characterized by bone marrow metastasis and MYCN oncogene amplification and is associated with a dismal five-year overall survival of approximately 50%, despite an intensive multimodal treatment regimen including chemotherapy, surgery, high-dose chemotherapy with autologous stem cell rescue, and radiation. Herein, we systemically reviewed contemporary therapy schemes and their challenges as well as recent advancement in effective treatments for NBL. Due to its relative rarity, we hope this review can serve as a useful guidebook and potentially provide new insights to researchers and clinicians interested in this field. Neuroblastoma (NBL) cells highly express disialoganglioside GD2, which is restricted and weakly expressed in selected healthy cells, making it a desirable target of immunotherapy. Over the past two decades, application of dinutuximab, an anti-GD2 monoclonal antibody (mAb), has been one of the few new therapies to substantially improve outcomes to current levels. Given the persistent challenge of relapse and therapeutic resistance, there is an urgent need for new effective and tolerable treatment options for high-risk NBL. Recent breakthroughs in immune checkpoint inhibitor (ICI) therapeutics have not translated into high-risk NBL, like many other major pediatric solid tumors. Given the suppressed tumor microenvironment (TME), single ICIs like anti-CTLA4 and anti-PD1 have not demonstrated significant antitumor response rates. Meanwhile, emerging studies are reporting novel advancements in GD2-based therapies, targeted therapies, nanomedicines, and other immunotherapies such as adoptive transfer of natural killer (NK) cells and chimeric antigen receptors (CARs), and these hold interesting promise for the future of high-risk NBL patient care. Herein, we summarize the current state of the art in NBL therapeutic options and highlight the unique challenges posed by NBL that have limited the successful adoption of immune-modifying therapies. Through this review, we aim to direct the field's attention to opportunities that may benefit from a combination immunotherapy strategy. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma, a cancer of the peripheral nervous system, is the most common and deadly tumor that occurs in childhood. In order to cure more patients with neuroblastoma, alternative treatment approaches based on biological understanding of the disease are fundamental. In this review, we discuss the recent developments in neuroblastoma research, with emphasis on new molecular technique approaches for understanding the molecular and cellular interactions within the tumor microenvironment and the development of new treatment modalities holding promise for future treatment regimens. Neuroblastoma is a peripheral nervous system tumor that almost exclusively occurs in young children. Although intensified treatment modalities have led to increased patient survival, the prognosis for patients with high-risk disease is still around 50%, signifying neuroblastoma as a leading cause of cancer-related deaths in children. Neuroblastoma is an embryonal tumor and is shaped by its origin from cells within the neural crest. Hence, neuroblastoma usually presents with a low mutational burden and is, in the majority of cases, driven by epigenetically deregulated transcription networks. The recent development of Omic techniques has given us detailed knowledge of neuroblastoma evolution, heterogeneity, and plasticity, as well as intra- and intercellular molecular communication networks within the neuroblastoma microenvironment. Here, we discuss the potential of these recent discoveries with emphasis on new treatment modalities, including immunotherapies which hold promise for better future treatment regimens. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma is the most common cancer in infants and the most common solid tumor outside the brain in children, and responds poorly to current therapies. The sphingolipid-modifying drug opaganib, which has anticancer and anti-inflammatory activity in many preclinical models, was tested for inhibition of neuroblastoma cell proliferation in cell culture and in tumors growing in mice. Opaganib inhibited cell proliferation regardless of the MYCN gene status of the neuroblastoma cells. Treatment of tumor-bearing mice with opaganib in combination with the established drugs irinotecan and temozolomide reduced tumor growth and increased survival compared with irinotecan plus temozolomide alone. Because opaganib has already demonstrated safety in patients with cancer, this new drug may provide improved therapy for neuroblastoma patients. Neuroblastoma (NB), the most common cancer in infants and the most common solid tumor outside the brain in children, grows aggressively and responds poorly to current therapies. We have identified a new drug (opaganib, also known as ABC294640) that modulates sphingolipid metabolism by inhibiting the synthesis of sphingosine 1-phosphate (S1P) by sphingosine kinase-2 and elevating dihydroceramides by inhibition of dihydroceramide desaturase. The present studies sought to determine the potential therapeutic activity of opaganib in cell culture and xenograft models of NB. Cytotoxicity assays demonstrated that NB cells, including cells with amplified MYCN, are effectively killed by opaganib concentrations well below those that accumulate in tumors in vivo. Opaganib was shown to cause dose-dependent decreases in S1P and hexosylceramide levels in Neuro-2a cells, while concurrently elevating levels of dihydroceramides. As with other tumor cells, opaganib reduced c-Myc and Mcl-1 protein levels in Neuro-2a cells, and also reduced the expression of the N-Myc protein. The in vivo growth of xenografts of human SK-N-(BE)2 cells with amplified MYCN was suppressed by oral administration of opaganib at doses that are well tolerated in mice. Combining opaganib with temozolomide plus irinotecan, considered the backbone for therapy of relapsed or refractory NB, resulted in increased antitumor activity in vivo compared with temozolomide plus irinotecan or opaganib alone. Mice did not lose additional weight when opaganib was combined with temozolomide plus irinotecan, indicating that the combination is well tolerated. Opaganib has additive antitumor activity toward Neuro-2a tumors when combined with the checkpoint inhibitor anti-CTLA-4 antibody; however, the combination of opaganib with anti-PD-1 or anti-PD-L1 antibodies did not provide increased antitumor activity over that seen with opaganib alone. Overall, the data demonstrate that opaganib modulates sphingolipid metabolism and intracellular signaling in NB cells and inhibits NB tumor growth alone and in combination with other anticancer drugs. Amplified MYCN does not confer resistance to opaganib, and, in fact, the drug attenuates the expression of both c-Myc and N-Myc. The safety of opaganib has been established in clinical trials with adults with advanced cancer or severe COVID-19, and so opaganib has excellent potential for treating patients with NB, particularly in combination with temozolomide and irinotecan or anti-CTLA-4 antibody. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma (NB) presents with two patterns of disease: with or without metastasis. Both types of disease presentation include tumors with high-risk (HR) features. The management of HR-NB includes chemotherapy, surgery, radiotherapy, and anti-GD2 immunotherapy. Anti-GD2 monoclonal antibodies (mAbs) have significantly improved the outcome of HR-NB patients but they are mostly effective against disease affecting the bone and/or bone marrow (known as the osteomedullary compartment), and less so against soft tissue disease. The question arises as to whether anti-GD2 immunotherapy might benefit HR-NB patients with disease compounded by only soft tissue. In this retrospective review, we found that achieving first complete remission with chemotherapy, surgery, and radiotherapy does not prevent the risk of relapse. However, adding anti-GD2 mAbs once the patient has achieved complete remission significantly decreases the chances of relapse by 80%. Our study provides further support to indicate anti-GD2 mAbs in all cases with HR-NB. Neuroblastoma presents with two patterns of disease: locoregional or systemic. The poor prognostic risk factors of locoregional neuroblastoma (LR-NB) include age, MYCN or MDM2-CDK4 amplification, 11q, histology, diploidy with ALK or TERT mutations, and ATRX aberrations. Anti-GD2 immunotherapy has significantly improved the outcome of high-risk (HR) NB and is mostly effective against osteomedullary minimal residual disease (MRD), but less so against soft tissue disease. The question is whether adding anti-GD2 monoclonal antibodies (mAbs) benefits patients with HR-NB compounded by only soft tissue. We reviewed 31 patients treated at SJD for HR-NB with no osteomedullary involvement at diagnosis. All tumors had molecular genetic features of HR-NB. The outcome after first-line treatment showed 25 (80.6%) patients achieving CR. Thirteen patients remain in continued CR, median follow-up 3.9 years. We analyzed whether adding anti-GD2 immunotherapy to first-line treatment had any prognostic significance. The EFS analysis using Cox models showed a HR of 0.20, p = 0.0054, and an 80% decrease in the risk of relapse in patients treated with anti-GD2 immunotherapy in the first line. Neither EFS nor OS were significantly different by CR status after first-line treatment. In conclusion, adding treatment with anti-GD2 mAbs at the stage of MRD helps prevent relapse that unequivocally portends poor survival. [ABSTRACT FROM AUTHOR]

Simple Summary: Approximately half of all infants and children newly diagnosed with neuroblastoma are at high risk for relapse despite aggressive multi-modal therapy. Polychemotherapy usually provokes a good initial response; however, minimal residual disease with the dissemination of a few resistant tumor cells frequently causes relapse. The number of long-term survivors of high-risk disease has remained unsatisfactorily poor with survival rates as low as 50% after first-line therapy and below 20% in the relapse situation. Applying extensive state-of-the-art methods and workflows for metabolomics and proteomics, this study characterized the modes of action of the metabolic inhibitor L-glyceraldehyde in a panel of neuroblastoma cell lines as well as fibroblasts used as a control. Our analyses revealed that L-glyceraldehyde arrests proliferation and induces neuroblastoma cell death by a multi-modal mechanism that involves oxidative stress and the inhibition of nucleotide biosynthesis. Our data provide the molecular basis for the rationale design of multi-modal therapies involving metabolic targets. Glyceraldehyde (GA) is a three-carbon monosaccharide that can be present in cells as a by-product of fructose metabolism. Bruno Mendel and Otto Warburg showed that the application of GA to cancer cells inhibits glycolysis and their growth. However, the molecular mechanism by which this occurred was not clarified. We describe a novel multi-modal mechanism by which the L-isomer of GA (L-GA) inhibits neuroblastoma cell growth. L-GA induces significant changes in the metabolic profile, promotes oxidative stress and hinders nucleotide biosynthesis. GC-MS and 13C-labeling was employed to measure the flow of carbon through glycolytic intermediates under L-GA treatment. It was found that L-GA is a potent inhibitor of glycolysis due to its proposed targeting of NAD(H)-dependent reactions. This results in growth inhibition, apoptosis and a redox crisis in neuroblastoma cells. It was confirmed that the redox mechanisms were modulated via L-GA by proteomic analysis. Analysis of nucleotide pools in L-GA-treated cells depicted a previously unreported observation, in which nucleotide biosynthesis is significantly inhibited. The inhibitory action of L-GA was partially relieved with the co-application of the antioxidant N-acetyl-cysteine. We present novel evidence for a simple sugar that inhibits cancer cell proliferation via dysregulating its fragile homeostatic environment. [ABSTRACT FROM AUTHOR]

Simple Summary: Cellular use of important fuel sources like glucose is disrupted in most human cancers. The MYC family of pro-cancer proteins is known to contribute to these shifts in metabolism. Relatively little is known about any such shifts in fatty acid use, which is unfortunate because fatty acids, particularly unsaturated fatty acids, are rich fuel sources and can also serve important roles in cellular survival and immune system function. Here, we aimed to better understand the role of unsaturated fatty acid synthesis in pediatric neuroblastoma, a devastating disease in children that is driven by the MYC family member MYCN, where less than half of high-risk patients survive. We discovered that the levels of omega-3, omega-6, and omega-9 unsaturated fatty acids and the genes that synthesize them are highly disturbed in aggressive neuroblastoma. We further connected these disrupted patterns to MYCN and microRNAs, a special class of small RNAs important in neuroblastoma and other cancers. This work provides new connections between fatty acid synthesis and poor outcomes in neuroblastoma, providing a better understanding of how this disease may use fatty acids as it develops and progresses and some potential new considerations for treatment. MYCN amplification (MNA) and disruption of tumor suppressor microRNA (TSmiR) function are key drivers of poor outcomes in neuroblastoma (NB). While MYCN and TSmiRs regulate glucose metabolism, their role in de novo fatty acid synthesis (FAS) and unsaturated FAS (UFAS) remains poorly understood. Here, we show that FAS and UFAS (U/FAS) genes FASN, ELOVL6, SCD, FADS2, and FADS1 are upregulated in high-risk (HR) NB and that their expression is associated with lower overall survival. RNA-Seq analysis of human NB cell lines revealed parallel U/FAS gene expression patterns. Consistent with this, we found that NB-related TSmiRs were predicted to target these genes extensively. We further observed that both MYC and MYCN upregulated U/FAS pathway genes while suppressing TSmiR host gene expression, suggesting a possible U/FAS regulatory network between MYCN and TSmiRs in NB. NB cells are high in de novo synthesized omega 9 (ω9) unsaturated fatty acids and low in both ω6 and ω3, suggesting a means for NB to limit cell-autonomous immune stimulation and reactive oxygen species (ROS)-driven apoptosis from ω6 and ω3 unsaturated fatty acid derivatives, respectively. We propose a model in which MYCN and TSmiRs regulate U/FAS and play an important role in NB pathology, with implications for other MYC family-driven cancers. [ABSTRACT FROM AUTHOR]

Simple Summary: Identifying the genetic cause of a retinoblastoma tumour can help doctors determine any future cancer risk for the patient or their family. Somatic testing has historically used DNA from the tumour, after the eye has been removed. We and others have previously shown cell-free DNA (cfDNA) from the tumour is present in the eye fluid of retinoblastoma patients. In this study we tested eye fluid from 68 patients, collected at different points in their treatment. Measurable levels of cfDNA were found in all 11 samples of eye fluid taken from patients who had received less than three cycles of chemotherapy, and 95% (21/22) of causal genetic variants were identified. Eye fluid collected later in the treatment had 150 times less cfDNA and only 46% of genetic variants (25/54) could be detected. Eye fluid sampling early in treatment is therefore likely to be required for successful somatic testing in retinoblastoma patients undergoing eye conservation treatment. The identification of somatic RB1 variation is crucial to confirm the heritability of retinoblastoma. We and others have previously shown that, when tumour DNA is unavailable, cell-free DNA (cfDNA) derived from aqueous humour (AH) can be used to identify somatic RB1 pathogenic variation. Here we report RB1 pathogenic variant detection, as well as cfDNA concentration in an extended cohort of 75 AH samples from 68 patients. We show cfDNA concentration is highly variable and significantly correlated with the collection point of the AH. Cell-free DNA concentrations above 5 pg/µL enabled the detection of 93% of known or expected RB1 pathogenic variants. In AH samples collected during intravitreal chemotherapy treatment (Tx), the yield of cfDNA above 5 pg/µL and subsequent variant detection was low (≤46%). However, AH collected by an anterior chamber tap after one to three cycles of primary chemotherapy (Dx1+) enabled the detection of 75% of expected pathogenic variants. Further limiting our analysis to Dx1+ samples taken after ≤2 cycles (Dx ≤ 2) provided measurable levels of cfDNA in all cases, and a subsequent variant detection rate of 95%. Early AH sampling is therefore likely to be important in maximising cfDNA concentration and the subsequent detection of somatic RB1 pathogenic variants in retinoblastoma patients undergoing conservative treatment. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma is a tumor arising from the sympathetic nervous system, and epidermal growth factor (EGF) influences its growth and metastatic behavior. We demonstrated that cathepsin D (CD) counteracts EGF-induced neuroblastoma cell growth in 2D by downregulating EGFR/MAPK signaling. Aggressive NB is highly metastatic, and whether CD is involved in the survival of metastatic NB clones is not known. Here, we addressed how CD differentially affects cell growth in suspension versus the adherent condition. To reproduce tumor heterogeneity, we co-cultured transgenic clones silenced for or overexpressing CD. We found that the Over CD clone had an advantage for growth in suspension, while the CD knocked-down clone was favored for the adherent growth in 2D. This dual role of CD suggests that clonal evolution may produce subclones with different CD levels, conferring survival and growth advantages depending on the metastatic step. We propose that epigenetic regulation of CD expression could be an additional strategy to prevent NB metastases. Neuroblastoma (NB) is an embryonal tumor arising from the sympathetic central nervous system. The epidermal growth factor (EGF) plays a role in NB growth and metastatic behavior. Recently, we have demonstrated that cathepsin D (CD) contrasts EGF-induced NB cell growth in 2D by downregulating EGFR/MAPK signaling. Aggressive NB is highly metastatic to the bone and the brain. In the metastatic process, adherent cells detach to form clusters of suspended cells that adhere once they reach the metastatic site and form secondary colonies. Whether CD is involved in the survival of metastatic NB clones is not known. Therefore, in this study, we addressed how CD differentially affects cell growth in suspension versus the adherent condition. To mimic tumor heterogeneity, we co-cultured transgenic clones silenced for or overexpressing CD. We compared the growth kinetics of such mixed clones in 2D and 3D models in response to EGF, and we found that the Over CD clone had an advantage for growth in suspension, while the CD knocked-down clone was favored for the adherent growth in 2D. Interestingly, on switching from 3D to 2D culture conditions, the expression of E-cadherin and of N-cadherin increased in the KD-CD and Over CD clones, respectively. The fact that CD plays a dual role in cancer cell growth in 2D and 3D conditions indicates that during clonal evolution, subclones expressing different level of CD may arise, which confers survival and growth advantages depending on the metastatic step. By searching the TCGA database, we found up to 38 miRNAs capable of downregulating CD. Interestingly, these miRNAs are associated with biological processes controlling cell adhesion and cell migration. The present findings support the view that during NB growth on a substrate or when spreading as floating neurospheres, CD expression is epigenetically modulated to confer survival advantage. Thus, epigenetic targeting of CD could represent an additional strategy to prevent NB metastases. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma (NB), a prevalent childhood cancer, presents challenges in treatment due to its cellular diversity and the presence of tumor-derived endothelial cells (TECs) associated with chemoresistance. We lack specific biomarkers for TECs, hindering effective therapies. We developed a stiffness-based in vitro platform simulating arterial and venous conditions to address this gap. Notably, adrenergic NB cells transdifferentiated into TECs where there was an arterial-like stiffness, while mesenchymal cells did not. This platform facilitated the identification of Globotriaosylceramide (GB3) as a novel TEC biomarker. Moreover, we harnessed Shiga toxin-functionalized nanoparticles for the specific targeting of GB3-positive cells, showing promise for future therapeutic strategies. Our study provides insights into NB heterogeneity, offers a predictive tool for assessing aggressiveness, and introduces potential targets for precision therapies. Neuroblastoma (NB) is a childhood cancer in sympathetic nervous system cells. NB exhibits cellular heterogeneity, with adrenergic and mesenchymal states displaying distinct tumorigenic potentials. NB is highly vascularized, and blood vessels can form through various mechanisms, including endothelial transdifferentiation, leading to the development of tumor-derived endothelial cells (TECs) associated with chemoresistance. We lack specific biomarkers for TECs. Therefore, identifying new TEC biomarkers is vital for effective NB therapies. A stiffness-based platform simulating human arterial and venous stiffness was developed to study NB TECs in vitro. Adrenergic cells cultured on arterial-like stiffness transdifferentiated into TECs, while mesenchymal state cells did not. The TECs derived from adrenergic cells served as a model to explore new biomarkers, with a particular focus on GB3, a glycosphingolipid receptor implicated in angiogenesis, metastasis, and drug resistance. Notably, the TECs unequivocally expressed GB3, validating its novelty as a marker. To explore targeted therapeutic interventions, nanoparticles functionalized with the non-toxic subunit B of the Shiga toxin were generated, because they demonstrated a robust affinity for GB3-positive cells. Our results demonstrate the value of the stiffness-based platform as a predictive tool for assessing NB aggressiveness, the discovery of new biomarkers, and the evaluation of the effectiveness of targeted therapeutic strategies. [ABSTRACT FROM AUTHOR]

Simple Summary: The aim of this study was to analyze the safety and effectiveness of ICIs used in 42 pediatric patients with various types of highly advanced malignancies. Good outcomes were achieved in patients with malignant cutaneous melanoma or Hodgkin lymphoma. Age > 14 years and good performance status were favorable prognostic factors. Background/aim: The role of immune checkpoint inhibitors (ICIs; anti-PD1) in the treatment of childhood cancers is still evolving. The aim of this nationwide retrospective study was to assess the safety and effectiveness of ICIs used in a group of 42 patients, with a median age of 13.6 years, with various types of advanced malignancies treated in pediatric oncology centers in Poland between 2015 and 2023. Results: The indications for treatment with anti-PD1 were as follows: Hodgkin lymphoma (11); malignant skin melanoma (9); neuroblastoma (8); and other malignancies (14). At the end of follow-up, complete remission (CR) was observed in 37.7% (15/42) of children and disease stabilization in 9.5% (4/42), with a mean survival 3.6 (95% CI = 2.6–4.6) years. The best survival (OS = 1.0) was observed in the group of patients with Hodgkin lymphoma. For malignant melanoma of the skin, neuroblastoma, and other rare malignancies, the estimated 3-year OS values were, respectively, 0.78, 0.33, and 0.25 (p = 0.002). The best progression-free survival value (0.78) was observed in the group with malignant melanoma. Significantly better effects of immunotherapy were confirmed in patients ≥ 14 years of age and good overall performance ECOG status. Severe adverse events were observed in 30.9% (13/42) patients. [ABSTRACT FROM AUTHOR]

Simple Summary: In our glioblastoma patients treated with standard therapy, the TERTp C250T mutation occurred less frequently than the C228T mutation. Patients with the C250T mutation had better prognosis than those with either TERTp-wt or TERTp C228T mutations, even when adjusted for key glioblastoma prognostic factors. This may be due to the lesser involvement of the C250T mutation in telomere- and chromosome-related pathways, as evidenced by the results of a gene enrichment analysis adjusted for MGMTp methylation status: TERTp C250T was differentially enriched compared to TERTp-wt and C228T. There were no differences according to TERTp mutation status in the mutations or copy number variants of other genes commonly present in glioblastoma. The biological pathways by which TERTp and MGMTp exert their effects are independent. The aim of this study was to determine how TERTp mutations impact glioblastoma prognosis. Materials and Methods: TERTp mutations were assessed in a retrospective cohort of 258 uniformly treated glioblastoma patients. RNA-sequencing and whole exome sequencing results were available in a subset of patients. Results: Overall, there were no differences in outcomes between patients with mutated TERTp-wt or TERTp. However, we found significant differences according to the type of TERTp mutation. Progression-free survival (mPFS) was 9.1 months for those with the C250T mutation and 7 months for those with either the C228T mutation or TERTp-wt (p = 0.016). Overall survival (mOS) was 21.9 and 15 months, respectively (p = 0.026). This differential effect was more pronounced in patients with MGMTp methylation (mPFS: p = 0.008; mOS: p = 0.021). Multivariate analysis identified the C250T mutation as an independent prognostic factor for longer mOS (HR 0.69; p = 0.044). We found no differences according to TERTp mutation status in molecular alterations common in glioblastoma, nor in copy number variants in genes related to alternative lengthening of telomeres. Nevertheless, in the gene enrichment analysis adjusted for MGMTp methylation status, some Reactome gene sets were differentially enriched, suggesting that the C250T mutation may exert a lesser effect on telomeres or chromosomes. Conclusions: In our series, patients exhibiting the C250T mutation had a more favorable prognosis compared to those with either TERPp-wt or TERTp C228T mutations. Additionally, our findings suggest a reduced involvement of the C250T mutation in the underlying biological mechanisms related to telomeres. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastomas are the most frequent neoplasms of infancy worldwide, accounting for 8% to 10% of all childhood cancers. Long-term survival with high-risk-type tumours is poor. Given their heterogeneous biological and clinical nature, tumours display mixed cell populations with some showing spontaneous regression and others metastatic activity. There remains a lack of specific biomarkers for early diagnosis, prognosis and personalised patient-targeted therapies. Therefore, the identification of neuroblastoma biomarkers, in the form of gene signatures that distinguish high-risk from low-risk patients is essential. We have demonstrated that activation of NTRK1 by TP53-dependent repression of PTPN6 expression, is significantly associated with favourable relapse-free survival in neuroblastoma. In this investigation, we have identified and validated a seventeen-gene signature for neuroblastoma prognosis using differentially expressed genes, upon the activation of NTRK1-PTPN6-TP53. A random survival forest model was used to construct a gene signature which was then assessed across validation datasets. Results showed that the gene signature was statistically significantly associated with favourable event-free survival (EFS). Patients with higher prognostic scores had significantly lower EFS than those with lower prognostic scores. Importantly, each individual gene was significantly associated with EFS in an independent manner. Moreover, the gene-set was significantly associated with lack of MYCN amplification, diagnosis before 18 months of age, differentiated tumour histology and presence of NTRK1-PTPN6-TP53 activation. These results strongly suggest that the gene signature is associated with good-prognosis neuroblastomas and may have value for patient risk stratification and personalised targeted therapies. Neuroblastoma is the most common extracranial solid tumour in children, comprising close to 10% of childhood cancer-related deaths. We have demonstrated that activation of NTRK1 by TP53 repression of PTPN6 expression is significantly associated with favourable survival in neuroblastoma. The molecular mechanisms by which this activation elicits cell molecular changes need to be determined. This is critical to identify dependable biomarkers for the early detection and prognosis of tumours, and for the development of personalised treatment. In this investigation we have identified and validated a gene signature for the prognosis of neuroblastoma using genes differentially expressed upon activation of the NTRK1-PTPN6-TP53 module. A random survival forest model was used to construct a gene signature, which was then assessed across validation datasets using Kaplan–Meier analysis and ROC curves. The analysis demonstrated that high BASP1, CD9, DLG2, FNBP1, FRMD3, IL11RA, ISGF10, IQCE, KCNQ3, and TOX2, and low BSG/CD147, CCDC125, GABRB3, GNB2L1/RACK1 HAPLN4, HEBP2, and HSD17B12 expression was significantly associated with favourable patient event-free survival (EFS). The gene signature was associated with favourable tumour histology and NTRK1-PTPN6-TP53 module activation. Importantly, all genes were significantly associated with favourable EFS in an independent manner. Six of the signature genes, BSG/CD147, GNB2L1/RACK1, TXNDC5, FNPB1, B3GAT1, and IGSF10, play a role in cell differentiation. Our findings strongly suggest that the identified gene signature is a potential prognostic biomarker and therapeutic target for neuroblastoma patients and that it is associated with neuroblastoma cell differentiation through the activation of the NTRK1-PTPN6-TP53 module. [ABSTRACT FROM AUTHOR]

Simple Summary: Tumor differentiation therapy using retinoids has a long history of clinical development and an established role as a part of the standard treatment for high-risk neuroblastoma. However, basic and clinical science related to this treatment is still immature in multiple respects. In the present review, we systematically expand on the basic and clinical science of retinoids for neuroblastoma treatment, off-label issues related to the treatment and possible countermeasures, and finally, prospects for this therapy. Retinoids are vitamin A derivatives and include trans-retinoic acid, isotretinoin, tamibarotene, and bexarotene, all of which are currently available for clinical use. The clinical development of retinoid therapy for neuroblastoma has a history spanning more than four decades. The most promising agent is isotretinoin, which can contribute to improving event-free survival in patients with high-risk neuroblastoma by approximately 10% when administered over six months as maintenance therapy. Although isotretinoin is regarded as an essential component in the standard clinical management of high-risk neuroblastoma, its use for this purpose in the US and EU is off-label. To promote isotretinoin use in Japan as a treatment for neuroblastoma, our clinical research team is planning to launch an investigator-initiated, registration-directed clinical trial. The present review article discusses the basic science behind retinoid therapy, pre-clinical/clinical evidence on neuroblastoma, the concept of the proposed clinical trial, and prospects for this therapy. [ABSTRACT FROM AUTHOR]

Simple Summary: Solid thoracic tumors in children are rare, with neuroblastoma being the most commonly diagnosed tumor entity. A safe and complete tumor resection is often not possible with a unilateral cervical approach, sternotomy, thoracotomy, or bilateral thoracotomy (single or staged) in certain localizations of the thorax, such as the cervicothoracic junction, the posterior mediastinum, or bilateral dorsal thoracic tumors. In adults, trap-door thoracotomy has been established and evaluated as a safe surgical approach for tumors of the cervicothoracic junction and clamshell thoracotomy as a safe surgical approach for posterior mediastinal or bilateral dorsal thoracic tumors. The aim of this study was to evaluate the surgical and oncologic implications of trap-door thoracotomy and clamshell thoracotomy in children with solid tumors in these specific localizations. These findings may have a positive impact on the quality of care and patient safety for children with solid thoracic tumors. Solid tumors of the cervicothoracic junction, the posterior mediastinum, or bilateral dorsal thoracic tumors represent a challenge in pediatric surgical oncology. The aim of this study was to evaluate trap-door thoracotomy and clamshell thoracotomy as surgical approaches. A single-center retrospective study of children with solid tumors in these specific localizations was performed. From 2015 to 2023, 26 children (17 girls; 9 boys) were treated at a median age of 54 months (range 8–229). Tumor resection was performed for neuroblastoma (n = 11); metastatic disease (n = 7); malignant rhabdoid tumor (n = 4); Ewing sarcoma (n = 1); inflammatory myofibroblastic tumor (n = 1); rhabdomyosarcoma (n = 1); and neurofibroma (n = 1). The surgical goal of macroscopic complete excision was achieved in all of the 14 children who underwent trap-door thoracotomy and in 11 of the 12 children who underwent clamshell thoracotomy. There were no major complications. At a median follow-up of 8 months (range 0–60), the disease was under local control or in complete remission in 66.7% of the children. In conclusion, surgical resection of solid tumors of the cervicothoracic junction in children can be performed safely and successfully with trap-door thoracotomy and with clamshell thoracotomy for posterior mediastinal or bilateral dorsal thoracic tumors. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma (NB) is the most frequent solid extracranial tumor in children and the most frequently diagnosed cancer during infancy. A genetic modification, a gain of 17q chromosome arm, is the most common modification in neuroblastoma. Substantial research has been performed on 17q's role in neuroblastoma development and its clinical significance. This paper aims to make a comprehensive review of this evidence. The main findings of the review are: (1) current knowledge supports that 17q gain is involved in the development of neuroblastoma and (2) 17q gain is an important clinical marker independently and in association with other genetic modifications. Neuroblastoma (NB) is the most frequent extracranial solid childhood tumor. Despite advances in the understanding and treatment of this disease, the prognosis in cases of high-risk NB is still poor. 17q gain has been shown to be the most frequent genomic alteration in NB. However, the significance of this remains unclear because of its high frequency and association with other genetic modifications, particularly segmental chromosomal aberrations, 1p and 11q deletions, and MYCN amplification, all of which are also associated with a poor clinical prognosis. This work reviewed the evidence on the clinical and biological significance of 17q gain. It strongly supports the significance of 17q gain in the development of NB and its importance as a clinically relevant marker. However, it is crucial to distinguish between whole and partial chromosome 17q gains. The most important breakpoints appear to be at 17q12 and 17q21. The former distinguishes between whole and partial chromosome 17q gain; the latter is a site of IGF2BP1 and NME1 genes that appear to be the main oncogenes responsible for the functional effects of 17q gain. [ABSTRACT FROM AUTHOR]

Simple Summary: Elevated expression of BIRC5/survivin, a protein crucial for cell division, is linked to unfavorable outcomes in neuroblastoma (NB), a prevalent pediatric tumor. Novel therapeutic approaches are consequently warranted. Therefore, we examined the effects of S12 and LQZ-7I, small molecules that disrupt survivin's interaction with itself and other proteins, on NB cells in vitro and in vivo. We demonstrate that these drugs effectively suppress the growth of NB cells, partly by hindering cell division. In contrast, incomplete inhibition of BIRC5 transcription did not diminish NB growth. We conclude that inhibitors of survivin protein interaction hold promise as a novel therapeutic class against NB and should be further investigated. Increased expression of BIRC5/survivin, a crucial regulator of the mitotic spindle checkpoint, is associated with poor prognosis in neuroblastoma (NB), the most common extracranial tumor of childhood. Transcriptional inhibitors of survivin have been tested in adult cancers and inhibitors of survivin homodimerization are emerging. We compared genetic inhibition of survivin transcription with the inhibition of survivin homodimerization by S12 and LQZ-7I, chosen from a larger panel of survivin dimerization inhibitors with activity against NB cells. Mice hemizygous for Birc5 were crossed with NB-prone TH-MYCN mice to generate Birc5+/-/MYCNtg/+ mice. The marked decrease of survivin transcription in these mice did not suffice to attenuate the aggressiveness of NB, even when tumors were transplanted into wild-type mice to assure that immune cell function was not compromised by the lack of survivin. In contrast, viability, clonogenicity and anchorage-independent growth of NB cells were markedly decreased by S12. S12 administered systemically to mice with subcutaneous NB xenotransplants decreased intratumoral hemorrhage, albeit not tumor growth. LQZ-7I, which directly targets the survivin dimerization interface, was efficacious in controlling NB cell growth in vitro at markedly lower concentrations compared to S12. LQZ-7I abrogated viability, clonogenicity and anchorage-independent growth, associated with massively distorted mitotic spindle formation. In vivo, LQZ-7I effectively reduced tumor size and cell proliferation of NB cells in CAM assays without apparent toxicity to the developing chick embryo. Collectively, these findings show that inhibiting survivin homodimerization with LQZ-7I holds promise for the treatment of NB and merits further investigation. [ABSTRACT FROM AUTHOR]

Simple Summary: Surgery remains the main treatment for thoracic neurogenic tumors. However, with the development of the minimally invasive approach, few studies so far have defined the indications for thoracoscopy in these tumors. We conducted a large retrospective multicenter study to define the role of thoracoscopy for neurogenic tumors. We identified 119 patients from 15 centers over 20 years. The median operative time was 2 h, and the length of stay was 4 days. The conversion rate was 11%, the long-term complication rate (Horner's and back issues) was 11%, the residue rate was 11.8%, the relapse rate was 7.5%, and 4% of patients died of their disease. The risk for conversion was associated with the need for chemotherapy. Complications were related to the IDRFs, the residue rates associated with persisting bronchial or vascular IDRF, and size but not on multivariate analysis. Recurrences were associated with neuroblastoma histology and unfavorable tumor biology. Objectives: To assess the efficacy of thoracoscopy and the outcome for children with thoracic neurogenic tumors. Methods: We performed a retrospective review of 15 European centers between 2000 and 2020 with patients who underwent thoracoscopy for a neurogenic mediastinal tumor. We assessed preoperative data, complications, and outcomes. Results were expressed with the median and range values. Results: We identified 119 patients with a median age of 4 years old (3 months–17 years). The diameter was 5.7 cm (1.1–15). INRG stage was L1 n = 46, L2 n = 56, MS n = 5, M n = 12. Of 69 patients with image-defined risk factors (IDRF), 29 had only (T9–T12) locations. Twenty-three out of 34 patients with preoperative chemotherapy had an 18 mm (7–24) decrease in diameter. Seven out of 31 patients lost their IDRF after chemotherapy. Fourteen had a conversion to thoracotomy. The length of the hospital stay was 4 days (0–46). The main complications included chylothorax (n = 7) and pneumothorax (n = 5). Long-term complications included Horner's syndrome (n = 5), back pain, and scoliosis (n = 5). Pathology was 53 neuroblastomas, 36 ganglioneuromas, and 30 ganglioneuroblastomas. Fourteen had a postoperative residue. With a median follow-up of 21 months (4–195), 9 patients had a recurrence, and 5 died of disease. Relapses were associated with tumor biology, histology, and the need for chemotherapy (p = 0.034, <0.001, and 0.015, respectively). Residues were associated with preoperative IDRF (excluding T9–T12 only) and the need for preoperative chemotherapy (p = 0.04 and 0.020). Conclusion: Our results show that thoracoscopy is safe, with good outcomes for thoracic neurogenic tumors in selected cases. Surgical outcomes are related to the IDRFs, whereas oncologic outcomes are related to tumor histology and biology. [ABSTRACT FROM AUTHOR]

Simple Summary: Copy number abnormalities (CNAs) are frequent in neuroblastoma and used to determine treatments. Little is known about the role of structural variants (SVs) in disease progression. Optical genome mapping (OGM) uses label patterns to scan the genome which can be used to identify SVs in cancer. Currently, the detection of SVs still relies on standard cytogenetic techniques. We investigated the utility of OGM to detect SVs in neuroblastoma cell lines and tumours and compared it with standard cytogenetic techniques. OGM confirmed known CNAs and identified novel SVs of potential clinical significance. Background: Neuroblastoma is the most common extracranial solid tumour in children, accounting for 15% of paediatric cancer deaths. Multiple genetic abnormalities have been identified as prognostically significant in neuroblastoma patients. Optical genome mapping (OGM) is a novel cytogenetic technique used to detect structural variants, which has not previously been tested in neuroblastoma. We used OGM to identify copy number and structural variants (SVs) in neuroblastoma which may have been missed by standard cytogenetic techniques. Methods: Five neuroblastoma cell lines (SH-SY5Y, NBLW, GI-ME-N, NB1691 and SK-N-BE2(C)) and two neuroblastoma tumours were analysed using OGM with the Bionano Saphyr® instrument. The results were analysed using Bionano Access software and compared to previous genetic analyses including G-band karyotyping, FISH (fluorescent in situ hybridisation), single-nucleotide polymorphism (SNP) array and RNA fusion panels for cell lines, and SNP arrays and whole genome sequencing (WGS) for tumours. Results: OGM detected copy number abnormalities found using previous methods and provided estimates for absolute copy numbers of amplified genes. OGM identified novel SVs, including fusion genes in two cell lines of potential clinical significance. Conclusions: OGM can reliably detect clinically significant structural and copy number variations in a single test. OGM may prove to be more time- and cost-effective than current standard cytogenetic techniques for neuroblastoma. [ABSTRACT FROM AUTHOR]

Simple Summary: Wilms tumor (WT), as the most common renal tumor in childhood, is treated very successfully within prospective trials and studies. The association with various genetic alterations has been studied, but it is still unclear why a WT develops in a specific child and why some children suffer from relapse. Genetic risk and external, environmental factors need further attention. Parental age at birth plays a role in various childhood diseases, so it is reasonable to also investigate whether older parental age is a risk factor for the development of childhood renal tumors. We could demonstrate that parental age has no correlation with the incidence of renal tumors in childhood. Despite excellent outcomes, many open questions remain about Wilms tumor (WT). Influences and risk factors for tumorigenesis, as well as tumor aggressiveness and recurrence, are not fully understood. Parental age plays a role in various childhood diseases and is also discussed as a risk factor for childhood cancer. We analyzed both maternal and paternal age at birth as risk factors for the occurrence of Wilms and non-Wilms tumors in children and investigated whether older maternal or paternal age is associated with a higher tumor incidence. During 1990 and 2019 we collected data from 3991 patients from the multicenter studies SIOP9/GPO, SIOP 93-01/GPOH, and SIOP 2001/GPOH, of whom maternal and paternal age was available in 2277 cases. Data from the Federal Statistical Office containing live births in Germany from 1990–2019 served as a comparative database. For maternal age at birth, the control data yielded 22,451,412 cases and for paternal age yielded 19,046,314 cases. Comparing maternal and paternal ages of the study patients with those of the control data, we confirmed that higher parental age is not correlated with the incidence of renal tumors in childhood. Mean ages of fathers and mothers in patients and the control cohort increased between 1991 and 2019 (fathers: 30.28 vs. 34.04; mothers: 27.68 vs. 29.79 in the patient group and 31.29 vs. 34.23 and 28.88 vs. 32.67 in the control group, respectively) without higher numbers of patients with kidney cancer over time. No influence was found for the subtype of cancer nor for syndromes. In addition, overall survival of patients is independent of the year of diagnosis and the age of the parents but depends on histology type and stage in WT. [ABSTRACT FROM AUTHOR]

Simple Summary: Transcription factors (TFs) can reprogram cellular states by modulating the expression of their target genes and establishing gene expression programs under homeostasis and diseases manifestation. In neuroblastoma, the TF MYCN has been recorded as dysregulated, presenting both aberrant expression and genomic abnormalities across its coding locus. Herein, we computationally investigated the gene expression characteristics that distinguish neuroblastoma-MYCN-amplified from neuroblastoma non-MYCN-amplified cancer cells, and we addressed the upregulation of several metabolism-related TF-encoding genes. Moreover, cistromic computational assessments of MYCN revealed its direct binding across regulatory sequences that reside in cis proximity to several of those genes. These results illuminate substantial mechanistic interrelationships between the key driver of neuroblastoma and a wealth of transcriptional regulators in cancer cells. Cancer is a disease caused by (epi)genomic and gene expression abnormalities and characterized by metabolic phenotypes that are substantially different from the normal phenotypes of the tissues of origin. Metabolic reprogramming is one of the key features of tumors, including those established in the human nervous system. In this work, we emphasize a well-known cancerous genomic alteration: the amplification of MYCN and its downstream effects in neuroblastoma phenotype evolution. Herein, we extend our previous computational biology investigations by conducting an integrative workflow applied to published genomics datasets and comprehensively assess the impact of MYCN amplification in the upregulation of metabolism-related transcription factor (TF)-encoding genes in neuroblastoma cells. The results obtained first emphasized overexpressed TFs, and subsequently those committed in metabolic cellular processes, as validated by gene ontology analyses (GOs) and literature curation. Several genes encoding for those TFs were investigated at the mechanistic and regulatory levels by conducting further omics-based computational biology assessments applied on published ChIP-seq datasets retrieved from MYCN-amplified- and MYCN-enforced-overexpression within in vivo systems of study. Hence, we approached the mechanistic interrelationship between amplified MYCN and overexpression of metabolism-related TFs in neuroblastoma and showed that many are direct targets of MYCN in an amplification-inducible fashion. These results illuminate how MYCN executes its regulatory underpinnings on metabolic processes in neuroblastoma. [ABSTRACT FROM AUTHOR]

Simple Summary: Patients with high-risk neuroblastoma, especially those whose disease either does not respond or recurs, have a poor chance of survival. Additionally, the majority of survivors of modern day high-risk neuroblastoma therapy develop significant late-effects. In an effort to improve survival, new agents with less toxicity are being explored. As technologies advance, the ability to design therapies for specific molecular targets on neuroblastoma are crucial. The most notable existing targeted therapies for high-risk neuroblastoma thus far are ALK inhibitors and anti-GD2 therapies, both of which have become mainstays of treatment both in the upfront and relapsed settings. Development of additional molecular targeted therapy has proven challenging, with limited successes. Adding to this difficulty is the heterogeneity of high-risk neuroblastoma, especially in the relapsed setting. This heterogeneity makes the numbers of patients eligible for specific targeted therapy small, complicating optimal clinical trial design. Here, we review the known molecular targets of neuroblastoma and their clinical implications. The latest advances in treatment for patients with neuroblastoma are constantly being incorporated into clinical trials and clinical practice standards, resulting in incremental improvements in the survival of patients over time. Survivors of high-risk neuroblastoma (HRNBL), however, continue to develop treatment-related late effects. Additionally, for the majority of the nearly 50% of patients with HRNBL who experience relapse, no curative therapy currently exists. As technologies in diagnostic and molecular profiling techniques rapidly advance, so does the discovery of potential treatment targets. Here, we discuss the current clinical landscape of therapies for neuroblastoma in the era of precision medicine. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma (NB) is the most common extracranial solid pediatric cancer, with highly observed metastasis at diagnosis. Primary NBs generally have favorable outcomes with current therapies; however, high-risk, metastatic NBs are resistant and result in relapsed disease. Patients with metastatic NBs, thus, have a lower overall survival. Thus, to develop effective therapies for metastatic NBs, it is essential to better understand the biology of metastasis in NB. This requires the utilization of pre-clinical animal models, importantly, immunocompetent models that have an intact immune system and better reflect the disease physiology. However, currently, there are few good animal models available for studying HR-NB metastasis. This study aimed to generate efficient and appropriate mouse models to study HR-NB metastasis. Our newly developed and validated mouse cell lines are relevant and promising tools to study (high-risk) HR-NB metastasis and identify novel therapeutic targets for metastatic NBs; thus, they could benefit the NB research community. NB, being a highly metastatic cancer, is one of the leading causes of cancer-related deaths in children. Increased disease recurrence and clinical resistance in patients with metastatic high-risk NBs (HR-NBs) result in poor outcomes and lower overall survival. However, the paucity of appropriate in vivo models for HR-NB metastasis has limited investigations into the underlying biology of HR-NB metastasis. This study was designed to address this limitation and develop suitable immunocompetent models for HR-NB metastasis. Here, we developed several highly metastatic immunocompetent murine HR-NB cell lines. Our newly developed cell lines show 100% efficiency in modeling experimental metastasis in C57BL6 mice and feature metastasis to the sites frequently observed in humans with HR-NB (liver and bone). In vivo validation demonstrated their specifically gained metastatic phenotype. The in vitro characterization of the cell lines showed increased cell invasion, acquired anchorage-independent growth ability, and resistance to MHC-I induction upon IFN-γ treatment. Furthermore, RNA-seq analysis of the newly developed cells identified a differentially regulated gene signature and an enrichment of processes consistent with their acquired metastatic phenotype, including extracellular matrix remodeling, angiogenesis, cell migration, and chemotaxis. The presented newly developed cell lines are, thus, suitable and promising tools for HR-NB metastasis and microenvironment studies in an immunocompetent system. [ABSTRACT FROM AUTHOR]

Simple Summary: More than 40% of high-risk neuroblastoma (NB) cases are characterized by aberrations in the transcription factors MYCN and c-MYC. Nevertheless, the developmental stage and mechanisms through which MYCN and c-MYC contribute to the onset and progression of the malignancy are not fully understood. In this study, we selected different c-MYC and MYCN targets with clinical or biological relevance and prognostic value to model multigene transcriptional risk scores. We found that the modeled scores accurately classify patients into groups with different outcomes, with high-risk-scoring patients exhibiting worse clinical features. Furthermore, target genes associated with poor prognosis showed a higher expression in sympathoblasts than in chromaffin cells during the sympathoadrenal development. Deregulation of the MYC family of transcription factors c-MYC (encoded by MYC), MYCN, and MYCL is prevalent in most human cancers, with an impact on tumor initiation and progression, as well as response to therapy. In neuroblastoma (NB), amplification of the MYCN oncogene and over-expression of MYC characterize approximately 40% and 10% of all high-risk NB cases, respectively. However, the mechanism and stage of neural crest development in which MYCN and c-MYC contribute to the onset and/or progression of NB are not yet fully understood. Here, we hypothesized that subtle differences in the expression of MYCN and/or c-MYC targets could more accurately stratify NB patients in different risk groups rather than using the expression of either MYC gene alone. We employed an integrative approach using the transcriptome of 498 NB patients from the SEQC cohort and previously defined c-MYC and MYCN target genes to model a multigene transcriptional risk score. Our findings demonstrate that defined sets of c-MYC and MYCN targets with significant prognostic value, effectively stratify NB patients into different groups with varying overall survival probabilities. In particular, patients exhibiting a high-risk signature score present unfavorable clinical parameters, including increased clinical risk, higher INSS stage, MYCN amplification, and disease progression. Notably, target genes with prognostic value differ between c-MYC and MYCN, exhibiting distinct expression patterns in the developing sympathoadrenal system. Genes associated with poor outcomes are mainly found in sympathoblasts rather than in chromaffin cells during the sympathoadrenal development. [ABSTRACT FROM AUTHOR]

Simple Summary: This study found that high levels of STMN1 expression in neuroblastoma indicate malignant potential, proliferation potency, and poor prognosis. STMN1 knockdown inhibited neuroblastoma cell growth regardless of MYCN overexpression. The study suggests that assessing STMN1 expression could be a powerful indicator of prognosis and a promising therapeutic candidate against refractory neuroblastoma. Background. Stathmin 1 (STMN1), a marker for immature neurons and tumors, controls microtubule dynamics by destabilizing tubulin. It plays an essential role in cancer progression and indicates poor prognosis in several cancers. This potential protein has not been clarified in clinical patients with neuroblastoma. Therefore, this study aimed to assess the clinical significance and STMN1 function in neuroblastoma with and without MYCN amplification. Methods. Using immunohistochemical staining, STMN1 expression was examined in 81 neuroblastoma samples. Functional analysis revealed the association among STMN1 suppression, cellular viability, and endogenous or exogenous MYCN expression in neuroblastoma cell lines. Result. High levels of STMN1 expression were associated with malignant potential, proliferation potency, and poor prognosis in neuroblastoma. STMN1 expression was an independent prognostic factor in patients with neuroblastoma. Furthermore, STMN1 knockdown inhibited neuroblastoma cell growth regardless of endogenous and exogenous MYCN overexpression. Conclusion. Our data suggest that assessing STMN1 expression in neuroblastoma could be a powerful indicator of prognosis and that STMN1 might be a promising therapeutic candidate against refractory neuroblastoma with and without MYCN amplification. [ABSTRACT FROM AUTHOR]

Simple Summary: The insulin-like growth factor 1 receptor (IGF1R) is a receptor tyrosine kinase (RTK) widely expressed in many cancers, including the pediatric cancer neuroblastoma. Aberrant activation of the anaplastic lymphoma kinase (ALK) RTK by activating point mutations or amplification is identified in 5–12% of neuroblastomas. Here, we investigated IGF1R in ALK-driven neuroblastoma, with the aim of understanding its contribution and exploring its potential for targeted therapy. Using ALK-driven neuroblastoma cell lines, we show that ALK-mutated cells are more sensitive to IGF1R inhibition than ALK-amplified cells, and a synergistic effect is obtained when combining ALK and IGF1R inhibitors. Mechanistically, in ALK-mutated neuroblastoma cells, both ALK and IGF1R contribute significantly to the activation of the downstream PI3K-AKT and RAS-MAPK signaling pathways. Our results suggest that differential activation of signaling downstream of the ALK and IGF1R pathways is in part due to preferential recruitment of adaptor proteins. Aberrant activation of anaplastic lymphoma kinase (ALK) by activating point mutation or amplification drives 5–12% of neuroblastoma (NB). Previous work has identified the involvement of the insulin-like growth factor 1 receptor (IGF1R) receptor tyrosine kinase (RTK) in a wide range of cancers. We show here that many NB cell lines exhibit IGF1R activity, and that IGF1R inhibition led to decreased cell proliferation to varying degrees in ALK-driven NB cells. Furthermore, combined inhibition of ALK and IGF1R resulted in synergistic anti-proliferation effects, in particular in ALK-mutated NB cells. Mechanistically, both ALK and IGF1R contribute significantly to the activation of downstream PI3K-AKT and RAS-MAPK signaling pathways in ALK-mutated NB cells. However, these two RTKs employ a differential repertoire of adaptor proteins to mediate downstream signaling effects. We show here that ALK signaling led to activation of the RAS-MAPK pathway by preferentially phosphorylating the adaptor proteins GAB1, GAB2, and FRS2, while IGF1R signaling preferentially phosphorylated IRS2, promoting activation of the PI3K-AKT pathway. Together, these findings reveal a potentially important role of the IGF1R RTK in ALK-mutated NB and that co-targeting of ALK and IGF1R may be advantageous in clinical treatment of ALK-mutated NB patients. [ABSTRACT FROM AUTHOR]

Simple Summary: Cryopreservation of ovarian tissue and transplantation are the only available fertility techniques to preserve fertility and endocrine function for prepubertal girls with cancer who require immediate cancer treatment. The primary objective of this systematic review was to identify and critically appraise the existing evidence regarding targeted minimal infiltrative disease detection in harvested ovarian tissues and identify markers that may be of value for assessment before autotransplantation, thereby facilitating fertility restoration in childhood cancer survivors. While the majority of malignancies were found to be at low risk of containing malignant cells in ovarian tissue, more studies are needed to ensure safe implementation of future fertility restoration in clinical practice. Background: Ovarian tissue cryopreservation and transplantation are the only available fertility techniques for prepubertal girls with cancer. Though autotransplantation carries a risk of reintroducing malignant cells, it can be avoided by identifying minimal infiltrative disease (MID) within ovarian tissue. Methods: A broad search for peer-reviewed articles in the PubMed database was conducted in accordance with PRISMA guidelines up to March 2023. Search terms included 'minimal residual disease', 'cryopreservation', 'ovarian', 'cancer' and synonyms. Results: Out of 542 identified records, 17 were included. Ovarian tissues of at least 115 girls were evaluated and categorized as: hematological malignancies (n = 56; 48.7%), solid tumors (n = 42; 36.5%) and tumors of the central nervous system (n = 17; 14.8%). In ovarian tissue of 25 patients (21.7%), MID was detected using RT-qPCR, FISH or multicolor flow cytometry: 16 of them (64%) being ALL (IgH rearrangements with/without TRG, BCL-ABL1, EA2-PBX1, TEL-AML1 fusion transcripts), 3 (12%) Ewing sarcoma (EWS-FLI1 fusion transcript, EWSR1 rearrangements), 3 (12%) CML (BCR-ABL1 fusion transcript, FLT3) and 3 (12%) AML (leukemia-associated immunophenotypes, BCR-ABL1 fusion transcript) patients. Conclusion: While the majority of malignancies were found to have a low risk of containing malignant cells in ovarian tissue, further studies are needed to ensure safe implementation of future fertility restoration in clinical practice. [ABSTRACT FROM AUTHOR]

Simple Summary: Next-generation sequencing (NGS) is a cutting-edge technology that enables rapid, high-throughput sequencing of DNA and RNA. Researchers and clinicians can identify genetic mutations, gene fusions, and other alterations that may drive cancer growth. This is particularly important in precision oncology as it is applied in the context of Molecular Tumor Boards (MTBs). The latter are multidisciplinary teams of experts who use NGS and bioinformatics tools to analyze patients' genetic profiles and develop personalized treatment recommendations for cancer patients. Thus, a crucial process for MTB decision-making is the analysis, compilation, and presentation of high-dimensional sequencing data, which are used for both preparation of and case presentation to all stakeholders. MIRACUM-Pipe precisely addresses these requirements and offers an easy-to-use, one-prompt standardized solution to analyze NGS data, including quality control, variant calling, copy number estimation, annotation, visualization, and report generation. (1) Background: Next-generation sequencing (NGS) of patients with advanced tumors is becoming an established method in Molecular Tumor Boards. However, somatic variant detection, interpretation, and report generation, require in-depth knowledge of both bioinformatics and oncology. (2) Methods: MIRACUM-Pipe combines many individual tools into a seamless workflow for comprehensive analyses and annotation of NGS data including quality control, alignment, variant calling, copy number variation estimation, evaluation of complex biomarkers, and RNA fusion detection. (3) Results: MIRACUM-Pipe offers an easy-to-use, one-prompt standardized solution to analyze NGS data, including quality control, variant calling, copy number estimation, annotation, visualization, and report generation. (4) Conclusions: MIRACUM-Pipe, a versatile pipeline for NGS, can be customized according to bioinformatics and clinical needs and to support clinical decision-making with visual processing and interactive reporting. [ABSTRACT FROM AUTHOR]

Simple Summary: Dinutuximab beta is an antibody approved for the maintenance treatment of patients with high-risk neuroblastoma. It is being investigated in combination with different established chemotherapy regimens in various clinical settings. We reviewed the clinical charts of 25 patients with relapsed/refractory neuroblastoma who had failed one or more second-line treatments and were given compassionate use treatment with dinutuximab beta long-term infusion combined with two induction chemotherapy regimens (N5 and N6), recommended by German guidelines. We found no unexpected severe toxicities. Grade 3/4 pain was reported in treatment cycle 1 by four patients, which was reduced to no patients by cycles 3 and 4. Almost half (48%) of patients had a complete, partial or minor response to treatment, despite previous treatment failures. Therefore, combining long-term infusion of dinutuximab beta with these chemotherapy regimens during earlier treatment phases may be beneficial for patients with relapsed/refractory neuroblastoma and should be further evaluated in clinical trials. The anti-disialoganglioside (GD2) monoclonal antibody dinutuximab beta is approved for the maintenance treatment of high-risk neuroblastoma. Dinutuximab beta combined with different chemotherapy regimens is being investigated in various clinical settings. We conducted a retrospective clinical chart review of 25 patients with relapsed/refractory neuroblastoma who had failed ≥1 second-line therapy and received compassionate use treatment with dinutuximab beta long-term infusion combined with the induction chemotherapy regimens N5 (cisplatin, etoposide, vindesine) and N6 (vincristine, dacarbazine, ifosfamide, doxorubicin) recommended by the German Pediatric Oncology and Hematology Group [GPOH] guidelines. The treatment did not result in any unexpected severe toxicities or in any major treatment delays. Grade 3/4 pain was reported by 4/25 patients in cycle 1, decreasing to 0/9 patients in cycles 3 and 4. The median follow-up was 0.6 years. The best response in this group was 48% (12/25 patients), which included three patients with minor responses. At 1 year, the estimated event-free survival was 27% (95% confidence interval [CI] 8–47) and overall survival was 44% (95% CI 24–65). Combining long-term infusion of dinutuximab beta with N5 and N6 chemotherapy demonstrated an acceptable safety profile and encouraging objective response rates in heavily pretreated patients with high-risk neuroblastoma, warranting further evaluation in clinical trials. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma is a pediatric tumor originating from the precursors of sympathetic nerves. The disease is known for its high heterogeneity. Hence, developing adequate preclinical models reflecting the complex biology of neuroblastoma is particularly challenging. This paper describes the current status of the available neuroblastoma models with their strengths and limitations, and demonstrates the future perspectives for preclinical neuroblastoma research. Preclinical in vitro and in vivo models remain indispensable tools in cancer research. These classic models, including two- and three-dimensional cell culture techniques and animal models, are crucial for basic and translational studies. However, each model has its own limitations and typically does not fully recapitulate the course of the human disease. Therefore, there is an urgent need for the development of novel, advanced systems that can allow for efficient evaluation of the mechanisms underlying cancer development and progression, more accurately reflect the disease pathophysiology and complexity, and effectively inform therapeutic decisions for patients. Preclinical models are especially important for rare cancers, such as neuroblastoma, where the availability of patient-derived specimens that could be used for potential therapy evaluation and screening is limited. Neuroblastoma modeling is further complicated by the disease heterogeneity. In this review, we present the current status of preclinical models for neuroblastoma research, discuss their development and characteristics emphasizing strengths and limitations, and describe the necessity of the development of novel, more advanced and clinically relevant approaches. [ABSTRACT FROM AUTHOR]

Simple Summary: Cell lines serve as tools for understanding molecular signatures of predictive biomarkers. However, the use of highly passaged commercial cell lines has to be questioned, as these do not closely resemble the originating tumor. Here, we established patient-derived cell lines from MPM tumors and performed whole exome and mRNA sequencing to understand genomic and transcriptomic differences between MPM tumors, patient-derived cell lines, and commercial cell lines. Our results show that the genome and transcriptome of tumors correlate to a higher degree with patient-derived cell lines than with commercial cell lines. These findings are of major relevance for the scientific community in regard to using cell line models to study predictive biomarkers and to interpret preclinical results which cannot be translated in clinical practice. Cell lines are extensively used to study cancer biology. However, the use of highly passaged commercial cell lines has to be questioned, as they do not closely resemble the originating tumor. To understand the reliability of preclinical models for Malignant pleural mesothelioma (MPM) studies, we have performed whole transcriptome and whole exome analyses of fresh frozen MPM tumors and compared them to cell lines generated from these tumors, as well as commercial cell lines and a preclinical MPM mouse model. Patient-derived cell lines were generated from digested fresh tumors and whole exome sequencing was performed on DNA isolated from formalin-fixed, paraffin-embedded (FFPE) tumor samples, corresponding patient-derived cell lines, and normal tissue. RNA sequencing libraries were prepared from 10 fresh frozen tumor samples, the 10 corresponding patient-derived cell lines, and 7 commercial cell lines. Our results identified alterations in tumor suppressor genes such as FBXW7, CDKN2A, CDKN2B, and MTAP, all known to drive MPM tumorigenesis. Patient-derived cell lines correlate to a high degree with their originating tumor. Gene expressions involved in multiple pathways such as EMT, apoptosis, myogenesis, and angiogenesis are upregulated in tumor samples when compared to patient-derived cell lines; however, they are downregulated in commercial cell lines compared to patient-derived cell lines, indicating significant differences between the two model systems. Our results show that the genome and transcriptome of tumors correlate to a higher degree with patient-derived cell lines rather than commercial cell lines. These results are of major relevance for the scientific community in regard to using cell lines as an appropriate model, resembling the pathway of interest to avoid misleading results for clinical applications. [ABSTRACT FROM AUTHOR]

Simple Summary: Penile squamous cell carcinomas harbouring mutations of TP53 have an increased risk of lymph node metastases and an impaired prognosis, but the mutational analysis of the TP53 gene is not available in many pathology laboratories. Although p53 immunohistochemistry (IHC) has been proposed as an alternative to the molecular analysis, the current method of evaluation of p53 IHC has many inaccuracies. The aim of our study was to determine, in a series of 40 penile tumours, if a recently described pattern-based framework of p53 IHC evaluation correlates better than the classical method with the TP53 mutational status. Our results show that the new method has a very good correlation with TP53 mutations (95% sensitivity; 92% specificity), higher than that of the classical method, and can be considered as a reliable surrogate of the TP53 mutational status. This new framework can help clinicians to better define risk groups and refine treatment strategies. p53 immunohistochemistry (IHC) has been proposed as a surrogate for TP53 mutations in penile squamous cell carcinomas (PSCC). We aimed to evaluate the performance of a pattern-based evaluation of p53 IHC in PSCC. Human papilloma virus (HPV) DNA testing, p16 and p53 IHC, and whole exome sequencing were performed in a series of 40 PSCC. p53 IHC was evaluated following a pattern-based framework and conventional p53 IHC evaluation. Out of 40 PSCC, 12 (30.0%) were HPV-associated, and 28 (70.0%) were HPV-independent. The agreement between the p53 IHC pattern-based evaluation and TP53 mutational status was almost perfect (k = 0.85). The sensitivity and accuracy of the pattern-based framework for identifying TP53 mutations were 95.5% and 92.5%, respectively, which were higher than the values of conventional p53 IHC interpretation (54.5% and 70.0%, respectively), whereas the specificity was the same (88.9%). In conclusions, the pattern-based framework improves the accuracy of detecting TP53 mutations in PSCC compared to the classical p53 IHC evaluation. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma mostly affects young children and despite intensive treatment, many children die of progressive disease. It remains challenging to identify those patients at risk. Analyzing blood, as liquid biopsies, is not invasive and can help to identify these patients. We studied whether RNA molecules can be detected in these liquid biopsies. In blood plasma, RNA can be free-floating or packed in small particles, 'extracellular vesicles'. We present a workflow to analyze this cell-free RNA from small volumes of blood plasma of children with neuroblastoma. We have used neuroblastoma-specific markers and markers involved in cell proliferation. These latter genes can be upregulated in many different tumor types. We demonstrate that both types of markers have a higher expression in patients with metastatic disease, compared to healthy controls and patients with localized disease. These findings are essential for future studies on cell-free RNA, hopefully leading to improved survival for these patients. Neuroblastoma affects mostly young children, bearing a high morbidity and mortality. Liquid biopsies, e.g., molecular analysis of circulating tumor-derived nucleic acids in blood, offer a minimally invasive diagnostic modality. Cell-free RNA (cfRNA) is released by all cells, especially cancer. It circulates in blood packed in extracellular vesicles (EV) or attached to proteins. We studied the feasibility of analyzing cfRNA and EV, isolated by size exclusion chromatography (SEC), from platelet-poor plasma from healthy controls (n = 40) and neuroblastoma patients with localized (n = 10) and metastatic disease (n = 30). The mRNA content was determined using several multiplex droplet digital PCR (ddPCR) assays for a neuroblastoma-specific gene panel (PHOX2B, TH, CHRNA3) and a cell cycle regulation panel (E2F1, CDC6, ATAD2, H2AFZ, MCM2, DHFR). We applied corrections for the presence of platelets. We demonstrated that neuroblastoma-specific markers were present in plasma from 14/30 patients with metastatic disease and not in healthy controls and patients with localized disease. Most cell cycle markers had a higher expression in patients. The mRNA markers were mostly present in the EV-enriched SEC fractions. In conclusion, cfRNA can be isolated from plasma and EV and analyzed using multiplex ddPCR. cfRNA is an interesting novel liquid biopsy-based target to explore further. [ABSTRACT FROM AUTHOR]

Simple Summary: Specific recurrent segmental aberrations (SCA) have been associated with poor survival in neuroblastoma (NB). The effect of many other not recurrent SCA has not been clarified yet. Here we examined the role of the loss of chromosome 10q in a cohort of 260 NB primary tumors, observing a lower survival rate for the patients expressing this rare SCA. We identified a cluster of 75 genes in the long arm of chromosome 10, whose deletion was associated with poorer survival. Six genes, and in particular CCSER2, appeared to be good candidates to play a role as tumor suppressor genes in NB and possible targets for future therapeutic interventions. Neuroblastoma (NB) is a tumor affecting the peripheral sympathetic nervous system that substantially contributes to childhood cancer mortality. Despite recent advances in understanding the complexity of NB, the mechanisms determining its progression are still largely unknown. Some recurrent segmental chromosome aberrations (SCA) have been associated with poor survival. However, the prognostic role of most SCA has not yet been investigated. We examined a cohort of 260 NB primary tumors at disease onset for the loss of chromosome 10q, by array-comparative genomic hybridization (a-CGH) and Single Nucleotide Polymorphism (SNP) array and we found that 26 showed 10q loss, while the others 234 displayed different SCA. We observed a lower event-free survival for NB patients displaying 10q loss compared to patients with tumors carrying other SCA. Furthermore, analyzing the region of 10q loss, we identified a cluster of 75 deleted genes associated with poorer outcome. Low expression of six of these genes, above all CCSER2, was significantly correlated to worse survival using in silico data from 786 NB patients. These potential tumor suppressor genes can be partly responsible for the poor prognosis of NB patients with 10q loss. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma is a rare type of cancer that usually affects children. The high-risk patients' expected survival rate is less than 50%. One reason is the lack of precision in the standard treatment protocol: a one-size-fits-all multi-modal therapy. The study presented in this paper was designed to address this deficit by optimising the use of two chemotherapeutic agents—vincristine and cyclophosphamide—during induction chemotherapy—the part of the protocol that shrinks the primary tumour before surgical removal. We combined a mathematical model and an optimisation algorithm to identify the best chemotherapy schedules for a cohort of virtual patients with different initial tumour compositions. Our results reveal novel strategies to exploit a pair of drugs with different levels of efficacy, provide a platform on which to individualise induction chemotherapy, and lay the foundation for a personalised therapy that leverages targeted therapies, multi-region sequencing, liquid biopsies, and modern computational methods to improve today's multi-modal therapy. Neuroblastoma is the most common extra-cranial solid tumour in children. Despite multi-modal therapy, over half of the high-risk patients will succumb. One contributing factor is the one-size-fits-all nature of multi-modal therapy. For example, during the first step (induction chemotherapy), the standard regimen (rapid COJEC) administers fixed doses of chemotherapeutic agents in eight two-week cycles. Perhaps because of differences in resistance, this standard regimen results in highly heterogeneous outcomes in different tumours. In this study, we formulated a mathematical model comprising ordinary differential equations. The equations describe the clonal evolution within a neuroblastoma tumour being treated with vincristine and cyclophosphamide, which are used in the rapid COJEC regimen, including genetically conferred and phenotypic drug resistance. The equations also describe the agents' pharmacokinetics. We devised an optimisation algorithm to find the best chemotherapy schedules for tumours with different pre-treatment clonal compositions. The optimised chemotherapy schedules exploit the cytotoxic difference between the two drugs and intra-tumoural clonal competition to shrink the tumours as much as possible during induction chemotherapy and before surgical removal. They indicate that induction chemotherapy can be improved by finding and using personalised schedules. More broadly, we propose that the overall multi-modal therapy can be enhanced by employing targeted therapies against the mutations and oncogenic pathways enriched and activated by the chemotherapeutic agents. To translate the proposed personalised multi-modal therapy into clinical use, patient-specific model calibration and treatment optimisation are necessary. This entails a decision support system informed by emerging medical technologies such as multi-region sequencing and liquid biopsies. The results and tools presented in this paper could be the foundation of this decision support system. [ABSTRACT FROM AUTHOR]

Simple Summary: Medulloblastoma is one of the most common types of brain tumors in children. During and after treatment with surgery, chemotherapy, and/or radiotherapy, children with this disease are monitored with imaging and cerebrospinal fluid analysis for the detection of tumor cells. These methods are not always sensitive or specific enough to confirm or rule out residual disease. Here, we develop a laboratory test based on the genetic makeup of medulloblastomas in 12 children. We analyze liquid biopsies (cerebrospinal fluid and blood plasma) for specific genetic fragments leaking from the individual tumors and find molecular traces of disease in 75% (9/12) of children overall. None of the children had malignant cells in the cerebrospinal fluid. We propose that this test could open up new technical possibilities to track measurable residual disease in children with medulloblastoma in order to further risk-adapt treatment, but first, larger studies of the approach at standardized time points are warranted. Medulloblastoma is a malignant embryonal tumor of the central nervous system (CNS) that mainly affects infants and children. Prognosis is highly variable, and molecular biomarkers for measurable residual disease (MRD) detection are lacking. Analysis of cell-free DNA (cfDNA) in cerebrospinal fluid (CSF) using broad genomic approaches, such as low-coverage whole-genome sequencing, has shown promising prognostic value. However, more sensitive methods are needed for MRD analysis. Here, we show the technical feasibility of capturing medulloblastoma-associated structural variants and point mutations simultaneously in cfDNA using multiplexed droplet digital PCR (ddPCR). Assay sensitivity was assessed with a dilution series of tumor in normal genomic DNA, and the limit of detection was below 100 pg of input DNA for all assays. False positive rates were zero for structural variant assays. Liquid biopsies (CSF and plasma, n = 47) were analyzed from 12 children with medulloblastoma, all with negative CSF cytology. MRD was detected in 75% (9/12) of patients overall. In CSF samples taken before or within 21 days of surgery, MRD was detected in 88% (7/8) of patients with localized disease and in one patient with the metastasized disease. Our results suggest that this approach could expand the utility of ddPCR and complement broader analyses of cfDNA for MRD detection. [ABSTRACT FROM AUTHOR]

Simple Summary: Tumor segmentation is a key step in oncologic imaging processing. We have recently developed a model to detect and segment neuroblastic tumors on MR images based on deep learning architecture nnU-Net. In this work, we performed an independent validation of the automatic segmentation tool with a large heterogeneous dataset. We reviewed the automatic segmentations and manually edited them when necessary. We were able to show that the automatic network was able to locate and segment the primary tumor on the T2 weighted images in the majority of cases, with an extremely high agreement between the automatic tool and the manually edited masks. The time needed for manual adjustment was very low. Objectives. To externally validate and assess the accuracy of a previously trained fully automatic nnU-Net CNN algorithm to identify and segment primary neuroblastoma tumors in MR images in a large children cohort. Methods. An international multicenter, multivendor imaging repository of patients with neuroblastic tumors was used to validate the performance of a trained Machine Learning (ML) tool to identify and delineate primary neuroblastoma tumors. The dataset was heterogeneous and completely independent from the one used to train and tune the model, consisting of 300 children with neuroblastic tumors having 535 MR T2-weighted sequences (486 sequences at diagnosis and 49 after finalization of the first phase of chemotherapy). The automatic segmentation algorithm was based on a nnU-Net architecture developed within the PRIMAGE project. For comparison, the segmentation masks were manually edited by an expert radiologist, and the time for the manual editing was recorded. Different overlaps and spatial metrics were calculated to compare both masks. Results. The median Dice Similarity Coefficient (DSC) was high 0.997; 0.944–1.000 (median; Q1–Q3). In 18 MR sequences (6%), the net was not able neither to identify nor segment the tumor. No differences were found regarding the MR magnetic field, type of T2 sequence, or tumor location. No significant differences in the performance of the net were found in patients with an MR performed after chemotherapy. The time for visual inspection of the generated masks was 7.9 ± 7.5 (mean ± Standard Deviation (SD)) seconds. Those cases where manual editing was needed (136 masks) required 124 ± 120 s. Conclusions. The automatic CNN was able to locate and segment the primary tumor on the T2-weighted images in 94% of cases. There was an extremely high agreement between the automatic tool and the manually edited masks. This is the first study to validate an automatic segmentation model for neuroblastic tumor identification and segmentation with body MR images. The semi-automatic approach with minor manual editing of the deep learning segmentation increases the radiologist's confidence in the solution with a minor workload for the radiologist. [ABSTRACT FROM AUTHOR]

Simple Summary: Beckwith–Wiedemann Syndrome (BWS) is one of the five most frequent syndromes predisposing to Wilms tumor (WT). BWS can be underdiagnosed if the phenotype is mild, and WT can be the presenting symptom. The purpose of this review is to summarize the available data in the current literature on the diagnosis, surveillance, treatment strategies, and outcome of WT in the presence of BWS. The awareness of clinical and pathological features of WT suggestive of BWS-associated WT can lead to prompt genetic counselling and, consequently, to the most appropriate treatment. Beckwith–Wiedemann Syndrome (BWS) is a pediatric overgrowth disorder involving a predisposition to embryonal tumors. Most of the tumors associated with BWS occur in the first 8–10 years of life, and the most common is Wilms tumor (WT). BWS clinical heterogeneity includes subtle overgrowth features or even silent phenotypes, and WT may be the presenting symptom of BWS. WT in BWS individuals exhibit distinct characteristics from those of sporadic WT, and the management of these patients needs a peculiar approach. The most important feature is a higher risk of developing bilateral disease at some time in the course of the illness (synchronous bilateral disease at diagnosis or metachronous recurrence after initial presentation with unilateral disease). Accordingly, neoadjuvant chemotherapy is the recommended approach also for BWS patients with unilateral WT to facilitate nephron-sparing surgical approaches. This review emphasizes the importance of early BWS recognition, particularly if a WT has already occurred, as this will result in an urgent consideration of first-line cancer therapy. [ABSTRACT FROM AUTHOR]

Simple Summary: Approximately 50% of high-risk neuroblastomas (NB) relapse within two years after the end of treatment. The prognosis for relapsed or refractory patients is poor, and additional therapeutic options are needed. The identification of ALK somatic mutations or amplification plays an important role in the treatment of relapsed/refractory patients. The aim of our study was to evaluate the genomic status of patients with relapsed/refractory NB and to employ ALK Tyrosine Kinase Inhibitors (TKIs) in patients with targetable ALK mutations. In the era of precision medicine, ALK inhibitors may play an important role in the treatment of high-risk, ALK-mutated, NB patients. Neuroblastoma (NB) is the most common extracranial solid tumor in childhood. Patients with relapsed/refractory disease have a poor prognosis, and additional therapeutic options are needed. Mutations and amplifications in the ALK (Anaplastic Lymphoma Kinase) gene constitute a key target for treatment. Our goal, within the Italian project of PeRsonalizEdMEdicine (PREME), was to evaluate the genomic status of patients with relapsed/refractory NB and to implement targeted therapies in those with targetable mutations. From November 2018 to November 2021, we performed Whole Exome Sequencing or Targeted Gene Panel Sequencing in relapsed/refractory NB patients in order to identify druggable variants. Activating mutations of ALK were identified in 8(28.57%) of 28 relapsed/refractory NB patients. The mutation p.F1174L was found in six patients, whereas p.R1275Q was found in one and the unknown mutation p.S104R in another. Three patients died before treatment could be started, while five patients received crizotinib: two in monotherapy (one with p.F1174L and the other with p.S104R) and three (with p.F1174L variant) in combination with chemotherapy. All treated patients showed a clinical improvement, and one had complete remission after two cycles of combined treatment. The most common treatment-related toxicities were hematological. ALK inhibitors may play an important role in the treatment of ALK-mutated NB patients. [ABSTRACT FROM AUTHOR]

Simple Summary: Systems biology analysis of gene expression across solid tumors identifies a 27 signature that centers on EZH2, the key component of polycomb repressive complex 2. Our study demonstrates that targeting the 27 gene network by the combination of BIRC5 and EZH2 inhibitors achieves remarkable anticancer synergy that is independent of the histone methyltransferase activity of EZH2.Globlal gene expression analysis in combination with pathway analysis shows that combination of YM155, the BIRC5 inhibitor, with EZH2 inhibitors induces unfolded protein response. Targeting multiple molecules in the same biological network may maximize therapeutic efficacy. In this study, we identified a 27-gene module that is highly expressed in solid tumors, encoding actionable targets including EZH2 and BIRC5. The combination of EZH2 inhibitors and a BIRC5 inhibitor, YM155, results in a remarkable synergistic effect. The action of EZH2 inhibitors in this process is independent of the histone methyltransferase activity of polycomb repressive complex 2. Our study reveals a potential therapeutic approach for treating solid tumors by simultaneously targeting EZH2 and BIRC5. [ABSTRACT FROM AUTHOR]

Simple Summary: Central nervous system (CNS) metastatic spread in neuroblastoma (NB) is rare and occurs more often at relapse/progression. In this retrospective study, we reviewed the CNS imaging of all the patients treated at the Bambino Gesù Children Hospital over a 25-year period. CNS metastasis in NB is confirmed to be rare, occurring in 4.7% of patients, and associated with advanced disease and bone skull involvement. In the last decade, the involvement of CNS at relapse has been observed more frequently, supporting the rising concern of the impact of immunotherapy in the pattern of relapse in high risk (HR) NB. Further studies are needed to confirm a higher CNS relapse risk in the immunotherapy era as well as the need for including CNS imaging in follow-up. Central nervous system (CNS) metastatic spread in neuroblastoma (NB) is rare and occurs more often at relapse/progression. We report on CNS involvement in high risk (HR) NB over 25 years. For this retrospective study, we reviewed the CNS imaging of all the patients treated at Bambino Gesù Children Hospital from 1 July 1996 to 30 June 2022. A total of 128 patients with HR NB were diagnosed over 26 years. Out of 128 patients, CNS metastatic spread occurred in 6 patients: 3 patients presented a metastatic spread at diagnosis, while in 3 patients, CNS was involved at relapse. Overall, the rate of occurrence of CNS spread is 4.7% with the same distribution at diagnosis and at relapse, namely 2.3%. Interestingly, CNS spread at diagnosis was observed only before 2012, whereas CNS was observed at relapse only after 2012, in the immunotherapy era. CNS metastases presented similar imaging features at diagnosis and at relapse, with a peculiar hemorrhagic aspect and mainly hemispheric localization in patients with bone skull involvement at the time of diagnosis. The outcome is dismal, and 3 out of 6 patients died for progressive disease. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroblastoma is the most common extracranial pediatric tumor. Although children with low- and intermediate-risk neuroblastoma, which correspond to approximately half of all newly diagnosed cases, have a good event-free and overall survival, high-risk neuroblastoma can be extremely aggressive and hard-to-treat tumors. In neuroblastoma, p53 and TAp73 act as safeguards against malignant transformation, but they are commonly inhibited by negative regulators, such as MDMs, Itch, and Aurora kinase A. This review focuses on the relevant tumor suppressor role of p53 and TAp73 in neuroblastoma, further addressing their connection with crucial biomarkers of poor prognosis, such as N-MYC, and their great potential as therapeutic targets. Neuroblastoma (NB) is an embryonic cancer that develops from neural crest stem cells, being one of the most common malignancies in children. The clinical manifestation of this disease is highly variable, ranging from spontaneous regression to increased aggressiveness, which makes it a major therapeutic challenge in pediatric oncology. The p53 family proteins p53 and TAp73 play a key role in protecting cells against genomic instability and malignant transformation. However, in NB, their activities are commonly inhibited by interacting proteins such as murine double minute (MDM)2 and MDMX, mutant p53, ΔNp73, Itch, and Aurora kinase A. The interplay between the p53/TAp73 pathway and N-MYC, a known biomarker of poor prognosis and drug resistance in NB, also proves to be decisive in the pathogenesis of this tumor. More recently, a strong crosstalk between microRNAs (miRNAs) and p53/TAp73 has been established, which has been the focused of great attention because of its potential for developing new therapeutic strategies. Collectively, this review provides an updated overview about the critical role of the p53/TAp73 pathway in the pathogenesis of NB, highlighting encouraging clues for the advance of alternative NB targeted therapies. [ABSTRACT FROM AUTHOR]

Simple Summary: Until recently, the prevalence of hereditary cancer in children was estimated to be very low. However, recent studies suggest that at least 10% of pediatric cancer patients have a germline mutation in a cancer predisposition gene. It has been shown that most of these mutations affect splicing, a process by which different transcripts of the same gene are produced. The splicing process is very important, as it regulates many aspects of cellular proliferation, survival, and differentiation. Hereditary cancer genes are highly prone to splicing alterations, and among them there are several genes that may contribute to the development of pediatric solid tumors when mutated in the germline. In this review, we analyze the importance of the splicing-disrupting mutations in pediatric solid cancer and inherited predisposition syndromes. The therapies developed to correct aberrant splicing in cancer are also discussed. The prevalence of hereditary cancer in children was estimated to be very low until recent studies suggested that at least 10% of pediatric cancer patients carry a germline mutation in a cancer predisposition gene. A significant proportion of pathogenic variants associated with an increased risk of hereditary cancer are variants affecting splicing. RNA splicing is an essential process involved in different cellular processes such as proliferation, survival, and differentiation, and alterations in this pathway have been implicated in many human cancers. Hereditary cancer genes are highly susceptible to splicing mutations, and among them there are several genes that may contribute to pediatric solid tumors when mutated in the germline. In this review, we have focused on the analysis of germline splicing-disrupting mutations found in pediatric solid tumors, as the discovery of pathogenic splice variants in pediatric cancer is a growing field for the development of personalized therapies. Therapies developed to correct aberrant splicing in cancer are also discussed as well as the options to improve the diagnostic yield based on the increase in the knowledge in splicing. [ABSTRACT FROM AUTHOR]

Simple Summary: The treatment of high-risk neuroblastoma patients with anti-GD2 antibodies has improved survival, and it is an established treatment strategy; however, many patients still experience a late relapse. One disadvantage of passive immunotherapy is the absence of a memory response. Therefore, developing an active immunotherapy leading to a sustained immune response may provide a solution and prevent the occurrence of late relapses following anti-GD2 antibody therapy. Here, we describe the first-in-man compassionate use of the ganglidiomab vaccine following passive immunotherapy with an anti-GD2 antibody (dinutuximab beta) in seven neuroblastoma patients. The vaccine was well-tolerated, and all patients not pre-treated by haploidentical transplantation developed vaccine-specific immune responses. (1) Background: High-risk neuroblastoma (HR-NB) is associated with a poor prognosis despite a multimodal high-intensity treatment regimen, including immunotherapy with anti-GD2 monoclonal antibodies (mAb). Here, we investigated the effects of an anti-idiotypic vaccine based on the mAb ganglidiomab that structurally mimics GD2. (2) Methods: Patients with HR-NB treated with anti-GD2 mAb dinutuximab beta and who achieved complete remission after frontline or salvage therapy were offered the vaccine (0.5 mg ganglidiomab adsorbed to Alhydrogel®). Side effects (CTCAE v4.03) and immune responses were determined on each visit. We also evaluated the time to relapse or progression until the last follow-up. (3) Results: Seven HR-NB patients (five frontlines, two relapsed) received 6–22 subcutaneous injections every two weeks. Six of the seven patients showed an immune response. The non-responding patient had a haploidentical stem cell transplantation as part of the previous treatment. No fever, pain, neuropathy, or toxicities ≥ grade 3 occurred during or post-treatment. All immunized patients did not experience relapses or progressions of their neuroblastoma. (4) Conclusions: This is the first-in-man use of the ganglidiomab vaccine, which was well-tolerated, and all patients not pre-treated by haploidentical transplantation developed vaccine-specific immune responses. These findings provide an important basis for the design of prospective clinical trials. [ABSTRACT FROM AUTHOR]

Simple Summary: Merkel cell polyomavirus is a widespread pathogen, strongly associated with the highly aggressive Merkel cell carcinoma. In this review paper we aim to examine a possible association between Merkel cell polyomavirus and the emergence of different types of cancer. Evidently, Merkel cell polyomavirus is the major oncogenic factor for Merkel cell carcinomas, but no establishing results can be derived about the virus' prevalence and role in the pathogenesis of other malignant diseases. Most certainly, it is imperative that the virus' biology, infection cycle, as well as its interactions with the host, are further investigated. Identifying a link between a virus and tumorigenesis is crucial and could lead to new virus-targeted therapeutic approaches with one final target: cancer elimination. Merkel cell polyomavirus (MCPyV), the sole member of Polyomavirus associated with oncogenesis in humans, is the major causative factor of Merkel cell carcinoma (MCC), a rare, neuroendocrine neoplasia of the skin. Many aspects of MCPyV biology and oncogenic mechanisms remain poorly understood. However, it has been established that oncogenic transformation is the outcome of the integration of the viral genome into the host DNA. The high prevalence of MCPyV in the population, along with the detection of the virus in various human tissue samples and the strong association of MCPyV with the emergence of MCC, have prompted researchers to further investigate the role of MCPyV in malignancies other than MCC. MCPyV DNA has been detected in several different non-MCC tumour tissues but with significantly lower prevalence, viral load and protein expression. Moreover, the two hallmarks of MCPyV MCC have rarely been investigated and the studies have produced generally inconsistent results. Therefore, the outcomes of the studies are inadequate and unable to clearly demonstrate a direct correlation between cellular transformation and MCPyV. This review aims to present a comprehensive recapitulation of the available literature regarding the association of MCPyV with oncogenesis (MCC and non-MCC tumours). [ABSTRACT FROM AUTHOR]