Carotenoids from Different Pumpkin Varieties Exert a Cytotoxic Effect on Human Neuroblastoma SH-SY5Y Cells.

Plants, including pumpkins (Cucurbita spp.), are an interesting source of nutrients and bioactives with various health benefits. In this research, carotenoid extracts obtained from the pulp of eight pumpkin varieties, belonging to the C. moschata and C. maxima species, were tested for cytotoxicity on SH-SY5Y neuroblastoma cells. The results showed that pumpkin bioactives exert a cytotoxic action against the tested cells, in particular Butternut extract at a 100 µM (53.69 µg/mL) concentration after 24 h of treatment and Mantovana extract at 50 µM (26.84 µg/mL) after 48 h. Moreover, the carotenoid extracts also showed interesting in vitro antioxidant activity, evaluated by ABTS and ORAC assays. To fully characterize the qualitative and quantitative profile of carotenoids in the tested extracts, a high-performance chromatographic technique was performed, revealing that pumpkin pulp carotenoids were mainly represented by ß-carotene, mono- and di-esterified hydroxy- and epoxy-carotenoids. Moreover, the carotenoid dataset was also useful for discriminating samples from two different species. In conclusion, the results of the present study highlight the potential anti-cancer activity of pumpkin carotenoid extracts and the possibility of using them as chemotherapeutic adjuvants.

Phase I study of safety and efficacy of allogeneic natural killer cell therapy in relapsed/refractory neuroblastomas post autologous hematopoietic stem cell transplantation.

Despite low incidence, neuroblastoma, an immunologically cold tumor, is the most common extracranial solid neoplasm in pediatrics. In relapsed/refractory cases, the benefits of autologous hematopoietic stem cell transplantation (auto-HSCT) and other therapies are limited. Natural killer (NK) cells apply cytotoxicity against tumor cells independently of antigen-presenting cells and the adaptive immune system. The primary endpoint of this trial was to assess the safety of the injection of allogenic, ex vivo-expanded and primed NK cells in relapsed/refractory neuroblastoma patients after auto-HSCT. The secondary endpoint included the efficacy of this intervention in controlling tumors. NK cells were isolated and primed ex vivo (by adding interleukin [IL]-2, IL-15, and IL-21) in a GMP-compliant CliniMACS system and administered to four patients with relapsed/refractory MYCN-positive neuroblastoma. NK cell injections (1 and 5 × 107 cells/kg in the first and second injections, respectively) were safe, and no acute or sub-acute adverse events were observed. During the follow-up period, one complete response (CR) and one partial response (PR) were observed, while two cases exhibited progressive disease (PD). In follow-up evaluations, two died due to disease progression, including the case with a PR. The patient with CR had regular growth at the 31-month follow-up, and another patient with PD is still alive and receiving chemotherapies 20 months after therapy. This therapy is an appealing and feasible approach for managing refractory neuroblastomas post-HSCT. Further studies are needed to explore its efficacy with higher doses and more frequent administrations for high-risk neuroblastomas and other immunologically cold tumors.Trial registration number: irct.behdasht.gov.ir (Iranian Registry of Clinical Trials, No. IRCT20201202049568N1).

Salvage chemotherapy regimens with arsenic trioxide for relapsed or refractory neuroblastoma: a promising approach.

In patients with relapsed or refractory neuroblastoma (NB), the limited efficacy of conventional chemotherapies necessitates the exploration of new treatment options. Previous studies have highlighted the anti-tumor properties of arsenic trioxide (ATO) in high-risk NB (HR-NB). This study aims to assess the effectiveness and safety of ATO combined with salvage chemotherapy regimens, featuring cyclophosphamide and topotecan, as a foundational treatment for children with relapsed or refractory NB. Eleven patients (four relapsed, seven refractory NB) were retrospectively analyzed for efficacy and treatment relevance. Salvage treatments, incorporating ATO (0.18 mg/kg daily for 8 h intravenously on days 1 to 10), were administered upon disease progression or relapse, with assessments conducted every two cycles. Treatments had 63.6% efficacy, with six cases of partial response, one case of stable disease, and four cases of disease progression. The overall response rate was 54.5%, and the disease control rate was 63.6%. Importantly, the systemic toxicity experienced by patients following salvage chemotherapy with ATO was mild. Salvage chemotherapy regimens featuring ATO demonstrated potential for prolonging disease stabilization for relapsed or refractory HR-NB patients, exhibiting both favorable efficacy and safety profiles. This suggests further clinical exploration and promotion of this therapeutic approach in the treatment of NB.

Point 1. The inadequate effectiveness of traditional chemotherapy in individuals with recurrent or resistant neuroblastoma (NB) necessitates the investigation of novel therapeutic approaches. Point 2. Arsenic trioxide (ATO)-based salvage treatments are both effective and less toxic in relapsed or refractory NB. Point 3. Salvage chemotherapy regimens incorporating ATO have shown promise in extending disease stabilization in relapsed or refractory high-risk NB patients, with favorable efficacy and safety profiles, which suggests further clinical exploration and promotion of this therapeutic approach in the treatment of NB.

Dose-Dependent Alterations of Lysosomal Activity and Alpha-Synuclein in Peripheral Blood Monocyte-Derived Macrophages and SH-SY5Y Neuroblastoma Cell Line by upon Inhibition of MTOR Protein Kinase - Assessment of the Prospects of Parkinson's Disease Therapy.

To date, the molecular mechanisms of the common neurodegenerative disorder Parkinson's disease (PD) are unknown and, as a result, there is no neuroprotective therapy that may stop or slow down the process of neuronal cell death. The aim of the current study was to evaluate the prospects of using the mTOR molecule as a potential target for PD therapy due to the dose-dependent effect of mTOR kinase activity inhibition on cellular parameters associated with, PD pathogenesis. The study used peripheral blood monocyte-derived macrophages and SH-SY5Y neuroblastoma cell line. As a result, we have for the first time showed that inhibition of mTOR by Torin1 only at a concentration of 100 nM affects the level of the lysosomal enzyme glucocerebrosidase (GCase), encoded by the GBA1 gene. Mutations in GBA1 are considered a high-risk factor for PD development. This concentration led a decrease in pathological phosphorylated alpha-synuclein (Ser129), an increase in its stable tetrameric form with no changes in the lysosomal enzyme activities and concentrations of lysosphingolipids. Our findings suggest that inhibition of the mTOR protein kinase could be a promising approach for developing therapies for PD, particularly for GBA1-associated PD.

Effectiveness of Nutrition Support Team Intervention in Pediatric Patients with Cancer.

Malnutrition in children with cancer is associated with poor prognosis. This study aimed to determine whether nutritional support team (NST) interventions prevent adverse events and improve the nutritional status in pediatric patients admitted for cancer treatment. This was a historical cohort study of pediatric patients with acute lymphocytic leukemia, acute myeloid leukemia, neuroblastoma, or brain tumor who received chemotherapy or underwent hematopoietic stem cell transplantation. Patients admitted between June 2013 and October 2014 were classified into the intervention group. Those admitted between January 2011 and December 2012 were classified into the control group. We created a homogeneous probability model using the inverse probability of treatment weighting method, and compared outcomes. A total of 75 patients were included in the study (38 and 37 in the intervention and control groups, respectively). The intervention group had significantly fewer incidents of nothing by mouth (nil per os [NPO]) (p=0.037) and days of NPO (p=0.046) than the control group. There was no significant difference between the intervention and control groups regarding the change in body mass index z-score between admission and discharge (p=0.376). NST interventions for children with cancer were associated with a reduction in the number of NPO occurrences and NPO days. These findings suggest that NST interventions contribute to continued oral intake.

Decoding pediatric spinal tumors: a single-center retrospective case series on etiology, presentation, therapeutic strategies, and outcomes.

INTRODUCTION: Spinal tumors (ST) often result in dire prognosis, carrying risks such as permanent paralysis, sensory loss, and sphincter dysfunction. Data on their incidence and etiology in pediatric populations are markedly scant. Our study investigates the etiology, clinical manifestation, treatment, and outcomes of pediatric ST. METHODS: We conducted a retrospective review of our institutional pediatric oncology and neurosurgery database, examining 14 patients under 18 years admitted with ST due to oncological diseases since 2005. We analyzed the clinical presentations, evaluations, molecular diagnostics and treatments for these patients. RESULTS: The study spanned 15 years and included 14 pediatric patients, each diagnosed with distinct spinal tumor entity. The mean patient age was approximately 19.6 ± 10.1 months. Severe axial pain along the vertebral column was observed in 13 patients, while acute neurological deterioration manifested in 7 patients. As a first-line intervention, 13 patients underwent decompressive surgery through laminectomy and tumor resection, and only one patient received chemotherapy solely. Before surgery, seven patients were unable to walk; post-surgery, six of them regained their ability to ambulate. The diagnosis encompassed a range of neoplasms: two instances of Ewing sarcoma, 3 instances of teratoma, one case presenting an atypical teratoid Rhabdoid tumor, two instances each of low-grade astrocytoma and neuroblastoma, and single instances of ependymoma, meningioma, rhabdomyosarcoma, and embryonal tumors with multilayered rosettes (ETMRs). Three patients succumbed two years after initiating therapy. CONCLUSION: Despite their rarity, intraspinal tumors in pediatric patients pose substantial therapeutic challenges. The intertwined complexities of the disease entity and the patient's neurological status demand swift initiation of an individualized therapeutic strategy. This crucial step helps optimize outcomes for this patient cohort, who frequently grapple with debilitating health conditions. Inclusion of these patients within a registry is mandatory to optimize treatment outcomes due to their rarity in pediatric population.

Etched-suppressed gold nanorods providing highly distinctive plasmonic patterns: Towards multiplex analysis of neuroblastoma biomarkers.

BACKGROUND: On-site monitoring of vanillylmandelic acid (VMA), homovanillic acid (HVA), and dopamine (DA) as key diagnostic biomarkers for a wide range of neurological disorders holds utmost significance in clinical settings. Numerous colorimetric sensors with mechanistic approaches based on aggregation or silver metallization have been introduced for this purpose. However, these mechanisms have drawbacks, such as sensitivity to environmental factors and probe toxicity. Therefore, there is a great demand for a robust yet non-toxic colorimetric sensor that employs a novel route to monitor these biomarkers effectively. RESULTS: Here, we present a single-component multi-colorimetric probe based on the controllable etching suppression of gold nanorods (AuNRs) upon exposure to the mild etchant N-bromosuccinimide (NBS), designed to accurately detect and discriminate VMA, HVA, DA, and their corresponding mixtures, i.e. , VMA: HVA, VMA:DA, HVA:DA, and VMA:HVA:DA. To enhance the sensitivity and automation capabilities of the designed multi-colorimetric sensor, two machine learning techniques were employed: linear discriminant analysis (LDA) for the qualitative classification and partial least-squares regression (PLSR) for the quantitative analysis of pure biomarkers and their mixtures. The outcomes revealed a high correlation between measured and predicted values, covering a linear range of 0.8-25, 1.2-25, and 2.7-100 µmol L-1, with remarkably low detection limits of 0.260, 0.397, and 0.913 µmol L-1 for VMA, HVA, and DA, respectively. Lastly, the performance of the probe was validated by successfully detecting the neuroblastoma biomarker VMA:HVA in human urine. SIGNIFICANCE: Our designed multi-colorimetric probe introduces a rapid, cost-effective, user-friendly, non-toxic, and non-invasive approach to detecting and discriminating not only the pure biomarkers but also their corresponding binary and ternary mixtures. The distinctive response profiles produced by the probe in the presence of different mixture ratios can indicate various disease states in patients, which is highly crucial in clinical diagnostics.

Oncolytic virotherapy augments self-maintaining natural killer cell line cytotoxicity against neuroblastoma.

BACKGROUND: Neuroblastoma is the most common extracranial solid tumor in children and accounts for 15% of pediatric cancer related deaths. Targeting neuroblastoma with immunotherapies has proven challenging due to a paucity of immune cells in the tumor microenvironment and the release of immunosuppressive cytokines by neuroblastoma tumor cells. We hypothesized that combining an oncolytic Herpes Simplex Virus (oHSV) with natural killer (NK) cells might overcome these barriers and incite tumor cell death. METHODS: We utilized MYCN amplified and non-amplified neuroblastoma cell lines, the IL-12 expressing oHSV, M002, and the human NK cell line, NK-92 MI. We assessed the cytotoxicity of NK cells against neuroblastoma with and without M002 infection, the effects of M002 on NK cell priming, and the impact of M002 and priming on the migratory capacity and CD107a expression of NK cells. To test clinical applicability, we then investigated the effects of M002 and NK cells on neuroblastoma in vivo. RESULTS: NK cells were more attracted to neuroblastoma cells that were infected with M002. There was an increase in neuroblastoma cell death with the combination treatment of M002 and NK cells both in vitro and in vivo. Priming the NK cells enhanced their cytotoxicity, migratory capacity and CD107a expression. CONCLUSIONS: To the best of our knowledge, these investigations are the first to demonstrate the effects of an oncolytic virus combined with self-maintaining NK cells in neuroblastoma and the priming effect of neuroblastoma on NK cells. The current studies provide a deeper understanding of the relation between NK cells and neuroblastoma and these data suggest that oHSV increases NK cell cytotoxicity towards neuroblastoma.

Exploring the effects of three-finger toxins from Naja ashei venom on neuronal and immunological cancer cell membranes.

Three-finger proteins are the most abundant toxins in the venom of Naja ashei, a snake species from the Elapidae family. This research aimed to describe the effects of varying charges of these proteins, isolated from Naja ashei venom using SEC and IEX chromatography. The study examined how differently charged three-finger toxin fractions interact with and affect neuroblastoma (SK-N-SH) and promyeloblast (HL-60) cells, as well as model Langmuir membranes and liposomes designed to mimic cellular lipid composition. Findings revealed that protein surface charges significantly impact cell survival (MTT assay), membrane damage (lactate dehydrogenase release, malondialdehyde formation), and the structural and electrochemical properties of model membranes (Langmuir membranes and zeta potential for liposomes and cancer cell lines). Results indicated that SK-N-SH cells, characterized by a higher negative charge on their cell membranes, interacted more effectively with positively charged toxins than HL-60 cells. However, the mechanism of these electrostatic interactions is complex. The research demonstrated that electrostatic and mechanical membrane modifications induced by venom proteins can significantly affect cell metabolism. Additionally, the total charge of the membrane, influenced by polar lipid components and phospholipid saturation, plays a decisive role in toxin interaction.

Prenyl Pterocarpans from Algerian Bituminaria bituminosa and Their Effects on Neuroblastoma.

The pterocarpan fraction from aerial parts of Bituminaria bituminosa was investigated for both chemical characterization and biological evaluation. Chemical studies were in accordance with the literature data on Bituminaria genus resulting in the identification of typical 4,8-prenyl pterocarpans. Three new members, bituminarins A-C (1-3), were isolated along with main bitucarpin A (4), erybraedin C (5) and erybraedin D (6) already reported from this plant. Further, biological studies evidenced antiproliferative properties of the most abundant pterocarpans 4 and 5 on neuroblastoma SH-SY5Y cell line, in agreement with previously described antiproliferative activity of these compounds against cancer cell lines other than neuroblastoma. The structure and the stereochemistry of the new molecules was determined by extensive spectroscopic analysis and chemical derivatization methods. The biological investigation was carried out by using an assay platform based on a live-cell imaging system revealing an apoptotic cell death induction.

Current and Emerging Biomarkers: Impact on Risk Stratification for Neuroblastoma.

Neuroblastoma has heterogenous clinical presentations that are reflected by several well-defined clinical factors and biomarkers. Combinations of these clinical and biologic prognostic factors have been used for decades to generate classifiers to stratify patients into risk groups (low, intermediate, and high), which in turn are used to inform and tailor treatment as reported in the new NCCN Clinical Practice Guidelines in Oncology for Neuroblastoma. Risk classification uses clinical features, such as age and tumor stage, along with the most significant prognostic tumor biomarkers, including histologic features (differentiation and mitosis-karyorrhexis index), MYCN amplification status, chromosomal copy number alterations (segmental or numerical), and ploidy (DNA content). Recent next-generation sequencing approaches have identified additional tumor-specific genetic factors that have potential roles as prognostic and predictive biomarkers. These emerging biomarkers include telomerase maintenance mechanisms, such as telomerase reverse transcription (TERT) expression and alternative lengthening of telomeres (ALT) status. Somatic alterations of genes, including mutations in the anaplastic lymphoma kinase gene ALK, detected in >10% of patients with newly diagnosed disease, have both prognostic and predictive roles in determining eligibility for targeted therapies (eg, ALK tyrosine kinase inhibitors). In addition to diagnostic tumor-derived biomarkers, significant effort is being directed toward identification of markers to predict response to chemotherapy and immunotherapies. With the increasing use of GD2-containing immunotherapy regimens, efforts are aimed at identifying host or tumor microenvironment immune correlatives that can serve as predictive biomarkers. Understanding the potential role of liquid biopsies as biomarkers during and following treatment, including sequential circulating tumor DNA or tumor-specific mRNA transcripts, is expected to enhance the ability to predict recurrences and also inform understanding of tumor evolution and therapy resistance. These and other emerging biomarkers will lead to refinement and optimization of future neuroblastoma risk classification systems.

Autophagic degradation of CDK4 is responsible for G0/G1 cell cycle arrest in NVP-BEZ235-treated neuroblastoma.

BACKGROUND: CDK4 is highly expressed and associated with poor prognosis and decreased survival in advanced neuroblastoma (NB). Targeting CDK4 degradation presents a potentially promising therapeutic strategy compared to conventional CDK4 inhibitors. However, the autophagic degradation of the CDK4 protein and its anti-proliferation effect in NB cells has not been mentioned. RESULTS: We identified autophagy as a new pathway for the degradation of CDK4. Firstly, autophagic degradation of CDK4 is critical for NVP-BEZ235-induced G0/G1 arrest, as demonstrated by the overexpression of CDK4, autophagy inhibition, and blockade of autophagy-related genes. Secondly, we present the first evidence that p62 binds to CDK4 and then enters the autophagy-lysosome to degrade CDK4 in a CTSB-dependent manner in NVP-BEZ235 treated NB cells. Similar results regarding the interaction between p62 and CDK4 were observed in the NVP-BEZ235 treated NB xenograft mouse model. CONCLUSIONS: Autophagic degradation of CDK4 plays a pivotal role in G0/G1 cell cycle arrest in NB cells treated with NVP-BEZ235.

GSTO2 ameliorates human neuroblastoma cell apoptosis, inflammation, ferroptosis, and oxidative stress by upregulating GPX4 expression in intracerebral hemorrhage.

Intracerebral hemorrhage (ICH) is a severe hemorrhagic stroke and induces severe secondary neurological injury. However, its pathogenesis remains to be explored. The present work investigates the role of glutathione S-transferase omega 2 (GSTO2) in ICH and the underlying mechanism. Human neuroblastoma cells (SK-N-SH) were stimulated using hemin to mimic ICH-like injury. Protein expression levels of GSTO2 and glutathione peroxidase 4 (GPX4) were detected by western blot analysis assay. Cell viability was assessed by cell counting kit-8 assay. Cell proliferation was investigated by 5-ethynyl-2'-deoxyuridine assay. Cell apoptosis was analyzed by flow cytometry. Interleukin-6 and tumor necrosis factor-α levels were quantified by enzyme-linked immunosorbent assays. Fe2+ colorimetric assay kit was used to detect Fe2+ level. A cellular reactive oxygen species (ROS) assay kit was used to detect ROS levels. Malondialdehyde (MDA) level was assessed using the MDA content assay kit. GSH level was quantified using the GSH assay kit. Co-immunoprecipitation assay was performed to identify the association between GSTO2 and GPX4. Hemin stimulation suppressed SK-N-SH cell proliferation and promoted cell apoptosis, cell inflammation, ferroptosis, and oxidative stress. GSTO2 expression was downregulated in hemin-treated SK-N-SH cells in comparison with the control group. In addition, ectopic GSTO2 expression counteracted hemin-induced inhibitory effect on cell proliferation and promoting effects on cell apoptosis, inflammation, ferroptosis, and oxidative stress. Moreover, GSTO2 was associated with GPX4 in SK-N-SH cells. GPX4 silencing attenuated GSTO2 overexpression-induced effects on hemin-stimulated SK-N-SH cell injury. GSTO2 ameliorated SK-N-SH cell apoptosis, inflammation, ferroptosis, and oxidative stress by upregulating GPX4 expression in ICH, providing a therapeutic strategy for ICH.

Altered methylation of imprinted genes in neuroblastoma: implications for prognostic refinement.

BACKGROUND: Neuroblastoma (NB) is a complex disease, and the current understanding of NB biology is limited. Deregulation in genomic imprinting is a common event in malignancy. Since imprinted genes play crucial roles in early fetal growth and development, their role in NB pathogenesis could be suggested. METHODS: We examined alterations in DNA methylation patterns of 369 NB tumours at 49 imprinted differentially methylated regions (DMRs) and assessed its association with overall survival probabilities and selected clinical and genomic features of the tumours. In addition, an integrated analysis of DNA methylation and allele-specific copy number alterations (CNAs) was performed, to understand the correlation between the two molecular events. RESULTS: Several imprinted regions with aberrant methylation patterns in NB were identified. Regions that underwent loss of methylation in > 30% of NB samples were DMRs annotated to the genes NDN, SNRPN, IGF2, MAGEL2 and HTR5A and regions with gain of methylation were NNAT, RB1 and GPR1. Methylation alterations at six of the 49 imprinted DMRs were statistically significantly associated with reduced overall survival: MIR886, RB1, NNAT/BLCAP, MAGEL2, MKRN3 and INPP5F. RB1, NNAT/BLCAP and MKRN3 were further able to stratify low-risk NB tumours i.e. tumours that lacked MYCN amplification and 11q deletion into risk groups. Methylation alterations at NNAT/BLCAP, MAGEL2 and MIR886 predicted risk independently of MYCN amplification or 11q deletion and age at diagnosis. Investigation of the allele-specific CNAs demonstrated that the imprinted regions that displayed most alterations in NB tumours harbor true epigenetic changes and are not result of the underlying CNAs. CONCLUSIONS: Aberrant methylation in imprinted regions is frequently occurring in NB tumours and several of these regions have independent prognostic value. Thus, these could serve as potentially important clinical epigenetic markers to identify individuals with adverse prognosis. Incorporation of methylation status of these regions together with the established risk predictors may further refine the prognostication of NB patients.

Joint single-cell genetic and transcriptomic analysis reveal pre-malignant SCP-like subclones in human neuroblastoma.

BACKGROUND: Neuroblastoma (NB) is a heterogeneous embryonal malignancy and the deadliest tumor of infancy. It is a complex disease that can result in diverse clinical outcomes. In some children, tumors regress spontaneously. Others respond well to existing treatments. But for the high-risk group, which constitutes approximately 40% of all patients, the prognosis remains dire despite collaborative efforts in basic and clinical research. While its exact cellular origin is still under debate, NB is assumed to arise from the neural crest cell lineage including multipotent Schwann cell precursors (SCPs), which differentiate into sympatho-adrenal cell states eventually producing chromaffin cells and sympathoblasts. METHODS: To investigate clonal development of neuroblastoma cell states, we performed haplotype-specific analysis of human tumor samples using single-cell multi-omics, including joint DNA/RNA sequencing of sorted single cells (DNTR-seq). Samples were also assessed using immunofluorescence stainings and fluorescence in-situ hybridization (FISH). RESULTS: Beyond adrenergic tumor cells, we identify subpopulations of aneuploid SCP-like cells, characterized by clonal expansion, whole-chromosome 17 gains, as well as expression programs of proliferation, apoptosis, and a non-immunomodulatory phenotype. CONCLUSION: Aneuploid pre-malignant SCP-like cells represent a novel feature of NB. Genetic evidence and tumor phylogeny suggest that these clones and malignant adrenergic populations originate from aneuploidy-prone cells of migrating neural crest or SCP origin, before lineage commitment to sympatho-adrenal cell states. Our findings expand the phenotypic spectrum of NB cell states. Considering the multipotency of SCPs in development, we suggest that the transformation of fetal SCPs may represent one possible mechanism of tumor initiation in NB with chromosome 17 aberrations as a characteristic element.

Vitronectin Levels in the Plasma of Neuroblastoma Patients and Culture Media of 3D Models: A Prognostic Circulating Biomarker?

Vitronectin is a glycoprotein present in plasma and the extracellular matrix that is implicated in cell migration. The high amount of vitronectin found in neuroblastoma biopsies has been associated with poor prognosis. Moreover, increased vitronectin levels have been described in the plasma of patients with different cancers. Our aim was to assess vitronectin as a potential circulating biomarker of neuroblastoma prognosis. Vitronectin concentration was quantified using ELISA in culture media of four neuroblastoma cell lines grown in a monolayer and in 3D models, and in the plasma of 114 neuroblastoma patients. Three of the neuroblastoma cell lines secreted vitronectin to culture media when cultured in a monolayer and 3D models. Vitronectin release was higher by neuroblastoma cells cultured in 3D models than in the monolayer and was still elevated when cells were grown in 3D scaffolds with cross-linked vitronectin. Vitronectin secretion occurred independently of cell numbers in cultures. Its concentration in the plasma of neuroblastoma patients ranged between 52.4 and 870 µg/mL (median, 218 µg/mL). A ROC curve was used to establish a cutoff of 361 µg/mL, above which patients over 18 months old had worse prognosis (p = 0.0018). Vitronectin could be considered a new plasma prognostic biomarker in neuroblastoma and warrants confirmation in collaborative studies. Drugs inhibiting vitronectin interactions with cells and/or the extracellular matrix could represent a significant improvement in survival for neuroblastoma patients.

Preclinical spheroid models identify BMX as a therapeutic target for metastatic MYCN nonamplified neuroblastoma.

The development of targeted therapies offers new hope for patients affected by incurable cancer. However, multiple challenges persist, notably in controlling tumor cell plasticity in patients with refractory and metastatic illness. Neuroblastoma (NB) is an aggressive pediatric malignancy originating from defective differentiation of neural crest-derived progenitors with oncogenic activity due to genetic and epigenetic alterations and remains a clinical challenge for high-risk patients. To identify critical genes driving NB aggressiveness, we performed combined chromatin and transcriptome analyses on matched patient-derived xenografts (PDXs), spheroids, and differentiated adherent cultures derived from metastatic MYCN nonamplified tumors. Bone marrow kinase on chromosome X (BMX) was identified among the most differentially regulated genes in PDXs and spheroids versus adherent models. BMX expression correlated with high tumor stage and poor patient survival and was crucial to the maintenance of the self-renewal and tumorigenic potential of NB spheroids. Moreover, BMX expression positively correlated with the mesenchymal NB cell phenotype, previously associated with increased chemoresistance. Finally, BMX inhibitors readily reversed this cellular state, increased the sensitivity of NB spheroids toward chemotherapy, and partially reduced tumor growth in a preclinical NB model. Altogether, our study identifies BMX as a promising innovative therapeutic target for patients with high-risk MYCN nonamplified NB.

Clinical significance of sarcopenia in children with neuroblastic tumors.

PURPOSE: To elucidate the clinical significance of sarcopenia in children with neuroblastic tumors (NTs). METHODS: We conducted a retrospective observational study and analyzed the z-scores for height, body weight, body mass index, and skeletal muscle index (HT-z, BW-z, BMI-z, and SMI-z) along with the clinical characteristics of 36 children with NTs. SMI-z was calculated from 138 computed tomography scans at diagnosis, during treatment, and at follow-up. The International Neuroblastoma Risk Group classification was used to identify high-risk groups. We analyzed the data at diagnosis for prognostic analysis and changes over time after diagnosis in the HT-z, BW-z, BMI-z, and SMI-z groups. RESULTS: Among the four parameters at diagnosis, only SMI-z predicted overall survival (hazard ratio, 0.58; 95% confidence interval, 0.34-0.99). SMI-z, HT-z, and BW-z significantly decreased over time after diagnosis (P < 0.05), while BMI-z did not (P = 0.11). In surviving high-risk NT cases without disease, SMI-z, HT-z, and BW-z significantly decreased over time (P < 0.05), while BMI-z did not (P = 0.43). CONCLUSION: In children with NT, the SMI-z at diagnosis was a significant prognostic factor and decreased during treatment and follow-up along with HT-z and BW-z. Monitoring muscle mass is important because sarcopenia may be associated with growth impairment.

Characterizing Relationships between T-cell Inflammation and Outcomes in Patients with High-Risk Neuroblastoma According to Mesenchymal and Adrenergic Signatures.

Recent insights have identified adrenergic (ADRN) and mesenchymal (MES) cell lineages as distinct biologic cell types and T-cell inflammation as a prognostic marker in neuroblastoma. We hypothesized that elucidating unique and overlapping aspects of these biologic features could serve as novel biomarkers for informing ongoing efforts to improve therapeutic approaches for children with high-risk neuroblastoma. We identified lineage-specific, single-stranded super-enhancers to define ADRN and MES specific genes. Publicly available RNA-seq of diagnostic tumor biopsies was used in Discovery and Validation cohorts. Each tumor was assigned a relative MES score and T-cell inflammation (TCI) score. Survival was assessed using the Kaplan-Meier method, and differences were assessed by the log-rank test. Inflammation scores were correlated with MES scores and anticorrelated with MYCN-amplification in both cohorts. Among patients with high-risk, ADRN tumors, those with TCI tumors had superior overall survival to those with non-inflamed tumors. A similar, but nonsignificant, trend was observed in the Validation cohort. Conversely, there was no difference according to TCI status in the MES cohort in either the Discover or Validation cohorts. High-inflammation scores were correlated with improved survival in some patients with high-risk, ADRN but not MES neuroblastoma. Our findings bolster support for further developing T-cell-based and immunotherapy-based approaches for children with high-risk neuroblastoma of varying MES and ADRN expression. SIGNIFICANCE: Adrenergic (ADRN) and mesenchymal (MES) lineages are distinct biologic cell types in neuroblastoma. We defined ADRN and MES specific genes and found that high-risk, ADRN tumors harboring elevated T-cell inflammation signatures had superior overall survival. Our findings bolster support for further developing immunotherapy-based approaches for children with high-risk neuroblastoma.