High-risk Neuroblastoma Research Articles

Vitamin D Status in Children With High-risk Neuroblastoma.

Vitamin D deficiency has become a matter of concern in pediatric cancer patients. A relationship between neuroblastoma and Vitamin D signaling pathways has been revealed with interest in the antiproliferative and antiinvasive properties of vitamin D. Our aim is to describe the prevalence of Vitamin D deficiency among children with high-risk neuroblastoma (HR-NB) and to explore its association with disease status. In all, 182 patients with HR-NB were managed at our center from 2017 to 2021. Serum 25(OH)D levels were tested at the first blood analysis performed and correlated with clinical data and disease status. One hundred forty-eight (81.4%) had low 25(OH)D levels (48.4% categorized as deficiency (25(OH)D below 20ng/mL) and 33.0% as insufficiency (25(OH)D 20 to 30ng/mL). Median Vitamin D level was 20.2ng/mL. Vitamin D levels were not associated with race or sex. Although malnourished patients had lower median 25(OH)D levels(11.1ng/mL), no statistical association was observed with Vitamin D deficiency. There was no association between Vitamin D levels and disease status. An inverse correlation was found between age and vitamin D levels ( P =0.0040). A concerning high prevalence of low Vitamin D levels affects more than two-thirds of patients with HR-NB in our cohort, regardless of the disease status at the time of evaluation. Older children are at a higher risk for deficient levels of vitamin D.

Critical elements of pediatric neuroblastoma surgery

Neuroblastoma (NB) is the most common solid extracranial malignancy of childhood with an incidence of 1 per 100,000 in the United States compromising approximately 10 % of childhood cancer. Unfortunately, patients with high-risk NG continue to have long-term survival less than 50 %. Both Children's Oncology Group and the International Society of Paediatric Oncology have demonstrated the important role of surgery in the treatment of high-risk NB. Herein, we compose the results of an extensive literature review as well as expert opinion from leaders in pediatric surgical oncology, to present the critical elements of effective surgery for high-risk neuroblastoma.

Repurposing disulfiram, an alcohol-abuse drug, in neuroblastoma causes KAT2A downregulation and in vivo activity with a water/oil emulsion

Neuroblastoma, the most common type of pediatric extracranial solid tumor, causes 10% of childhood cancer deaths. Despite intensive multimodal treatment, the outcomes of high-risk neuroblastoma remain poor. We urgently need to develop new therapies with safe long-term toxicity profiles for rapid testing in clinical trials. Drug repurposing is a promising approach to meet these needs. Here, we investigated disulfiram, a safe and successful chronic alcoholism treatment with known anticancer and epigenetic effects. Disulfiram efficiently induced cell cycle arrest and decreased the viability of six human neuroblastoma cell lines at half-maximal inhibitory concentrations up to 20 times lower than its peak clinical plasma level in patients treated for chronic alcoholism. Disulfiram shifted neuroblastoma transcriptome, decreasing MYCN levels and activating neuronal differentiation. Consistently, disulfiram significantly reduced the protein level of lysine acetyltransferase 2A (KAT2A), drastically reducing acetylation of its target residues on histone H3. To investigate disulfiram’s anticancer effects in an in vivo model of high-risk neuroblastoma, we developed a disulfiram-loaded emulsion to deliver the highly liposoluble drug. Treatment with the emulsion significantly delayed neuroblastoma progression in mice. These results identify KAT2A as a novel target of disulfiram, which directly impacts neuroblastoma epigenetics and is a promising candidate for repurposing to treat pediatric neuroblastoma.

Survival and Local Recurrence Risk in Patients with High-Risk Neuroblastoma Treated with Proton Therapy over a 10 Year Interval

Patients (pts) with high-risk neuroblastoma (HR-NBL) require radiation (RT) to the primary tumor site (PS); approach is standardized within North American paradigms but remains a subject of global study. Long-term experience using proton therapy (PRT) in this population is lacking. We hypothesized that PRT would be associated with low risk of local recurrence (LR) in a large population of pts with HR-NBL spanning > 10 years. Sequential pts with HR-NBL at a single institution received RT to PS and persistent metastatic sites (MS). Dose to PS after subtotal resection (STR) was reduced from 36 Gy to 21.6 Gy in 2019 based on results from the Children's Oncology Group ANBL0532 trial (Liu K, 2019). Analysis using Kaplan Meier method and log rank test was performed with IRB approval. From 9/2010 - 12/2021, 99 pts with HR-NBL received PS RT during first-line therapy; most [78, (79%)] had adrenal primary tumors and 26 (26%) received MS RT. Median age was 48m at RT (R 11m to 17.5y) and 52 (53%) were female. All pts had multi-agent induction chemotherapy (CT) [+ dinutuximab [12 (13%)] and/ or therapeutic 131MIBG [19 (19%)] and resection of primary tumor prior to RT; 34 (34%) patients had STR with residual disease (RD) on post-op imaging, 65 (66%) had gross total resection (GTR). Dose to PS was 21.6 Gy for 78 (79%) pts and 36 Gy for 21 (21%) based on RD and treatment era; PRT was pencil beam [78 (79%)] or double scattered [22 (22%)], combined with IMRT in 2 (2%). With median FU of 4.2 yrs (R 0.5y - 12y), 80 pts (81%) are alive [66 (67%) disease-free, 14 (14%) with disease], 19 (19%) have died. Progressive disease (PD) occurred in 33 (33%), with median time to PD 24m (R 8-116m); two pts (2%) had isolated LR, 25 (25%) distant PD, and 6 (6%) concurrent LR and distant PD. Risk of LR at 10 years was 8%; absolute risk of any LR was 8% (6/78) in 21.6 Gy cohort and 9% (2/21) in 36 Gy cohort (p = NS). After induction CT, 34 (34%) pts had STR with > 1cm3 RD on axial imaging; 18/ 34 (53%) also had MIBG uptake (MIBG+) at PS. Based on treatment era, 21 pts (62%) after STR received 21.6 Gy + boost to RD (36 Gy), and 13 (38%) 21.6 alone. Of those who received 36 Gy (median FU 5.7y), 2/21 (9.5%) had LR with concurrent distant PD; of those who received 21.6 Gy (median FU 3.2y) 4/13 (31%) had LR (2 with concurrent distant PD and 2 LR only) (p = 0.03). In the 21.6 Gy GTR cohort, 2/65 (3%) had LR + distant PD. Of 8 total patients who experienced LR, 5 had MIBG + RD, 1 MIBG- RD, and 2 GTR. We observed excellent outcomes in 99 pts treated with proton radiotherapy for HR-NBL from 2010 through 2021, with 81% of patients alive and 92% free of LR. Our data suggest that LR is rare after GTR and 21.6 Gy, and uncommon among pts with STR treated with 36 Gy. A small number of pts received 21.6 Gy after STR, however, this experience suggests that a subset of pts with RD may require RT dose > 21.6 Gy. Further work is required to further characterize individual management of PS in pts with HR-NBL with regard to extent of RD and biologic disease features.

Association between Radiation Dose and Local Failure in Recurrent High-Risk Neuroblastoma

Historically, standard-of-care (SoC) for high-risk neuroblastoma recurrences is irradiation to 2160 cGy for gross or microscopic disease. Boosting gross disease to 3000-3600 cGy remains controversial. Prior reports have demonstrated worse local control with gross disease irradiated to <3000 cGy. We sought to evaluate whether higher radiation doses delivered to sites of recurrence affected local control. We identified seventy-five high-risk neuroblastoma patients aged 6 months to 26 years that completed definitive treatment between 2007 and 2021 at two large academic centers. Only patients with distant recurrences treated with radiotherapy were included for analysis. Treatments were stratified into 1800-2160 cGy or 3000-3600 cGy dose levels. Local failure after irradiation was defined as progression within the radiation field after salvage RT. The Fine-Gray competing risk model was used to identify cumulative incidence of local failure (CILF) after irradiation with competing risk of death. Thirteen patients experienced recurrence after completion of definitive treatment, representing 30 metastatic lesions. Thirteen lesions received 1800-2160 cGy and seventeen received 3000-3600 cGy. With a median follow-up of 42.0 months (IQR 30.8-60.8), local failure after irradiation occurred in 10% (3/30) of metastatic lesions. Median time to failure was 7 months. All three failures had been treated to 2160 cGy; two in the calvarium and one in the femur. There were no local failures among patients treated to 3000-3600 cGy. The 1-year and 2-year CILF for the low- versus high-dose levels were 15% versus 0% (p<0.01) and 25% versus 0% (p<0.01), respectively. Four of the 13 patients (31%) died at a median time of 12.5 months after first recurrence; all four had distant recurrences. Dose-escalated irradiation ≥3000 cGy was associated with improved local control in recurrent high-risk neuroblastoma; prospective study and validation is warranted.

A Multi-Color Flow Cytometric Assay for Quantifying Dinutuximab Binding to Neuroblastoma Cells in Tumor, Bone Marrow, and Blood.

GD2, a disialoganglioside, is present on the surface of most neuroblastomas, as well as on some other cancers, such as melanoma and osteogenic sarcoma. The anti-GD2 antibody ch14.18 (dinutuximab) has an FDA-registered indication for use as maintenance therapy for high-risk neuroblastoma with cytokines and 13-cis-retinoic acid after myeloablative therapy. Recent studies using immunohistochemistry of tumor or tumor cells in marrow have shown that some neuroblastomas are negative for GD2. Dinutuximab and other anti-GD2 antibodies are increasingly used in combination with cytotoxic chemotherapy for treating relapsed neuroblastoma, so it is important to be able to identify patients with tumor cells with low GD2 expression, as such patients may experience toxicity but not benefit from the antibody therapy. As the most common clinical samples available for relapsed neuroblastoma are bone marrow aspirates, we developed a method to quantify dinutuximab binding density and the frequency of neuroblastoma cells positive for the antibody in bone marrow aspirates. Here, we describe a multi-color flow cytometry assay that employs non-GD2 antibodies to identify neuroblastoma cells in a mixed population (tumor, bone marrow, or blood) and an anti-GD2 antibody to quantify both the frequency and density of GD2 expression on neuroblastoma cells.

Neuroblastoma in the Era of Precision Medicine: A Clinical Review.

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.

Discovery of Novel Oxazepine Derivatives as Akt/ROCK Inhibitors for Growth Arrest and Differentiation Induction in Neuroblastoma Treatment.

Patients with high-risk neuroblastoma face limited treatment choices, typically involving a combination of cytotoxic and differentiation maintenance therapies due to a scarcity of drugs. Evidence suggests that targeted inhibitors may provide opportunities for inducing neuroblastoma differentiation while inhibiting proliferation. Here, we demonstrate the synergistic effect of inhibiting Akt and ROCK in antineuroblastoma and present the design and discovery of a new Akt/ROCK inhibitor, B12. It displays strong antiproliferative effects and excellent differentiation inducing activity against Neuro2a cells. Treatment with B12 results in the arrest of G0/G1 cell cycles, a significant decrease in N-myc protein level, and an increase in differentiation markers. The administration of B12 effectively suppresses xenograft tumor growth and promotes differentiation. Overall, the discovery of B12 based on the Akt/ROCK dual inhibition strategy may provide hope for the development of more effective and targeted therapies for this challenging disease.

Multimodal Therapy with Consolidating Haploidentical Stem Cell Transplantation and Dinutuximab Beta for Patients with High-Risk Neuroblastoma and Central Nervous System Relapse.

Despite highly intensive multimodality treatment regimens, the prognosis of patients with high-risk neuroblastoma (HRNB) and central nervous system (CNS) relapse remains poor. We retrospectively reviewed data from 13 patients with HRNB and CNS relapse who received multimodal therapy with consolidating haploidentical stem cell transplantation (haplo-SCT) followed by dinutuximab beta ± subcutaneous interleukin-2 (scIL-2). Following individual relapse treatment, patients aged 1-21 years underwent haplo-SCT with T/B-cell-depleted grafts followed by dinutuximab beta 20 mg/m2/day × 5 days for 5-6 cycles. If a response was demonstrated after cycle 5 or 6, patients received up to nine treatment cycles. After haplo-SCT, eight patients had a complete response, four had a partial response, and one had a stable disease. All 13 patients received ≥3 cycles of immunotherapy. At the end of the follow-up, 9/13 patients (66.7%) demonstrated complete response. As of July 2023, all nine patients remain disease-free, with a median follow-up time of 5.1 years since relapse. Estimated 5-year event-free and overall survival rates were 55.5% and 65.27%, respectively. Dinutuximab beta ± scIL-2 following haplo-SCT is a promising treatment option with a generally well-tolerated safety profile for patients with HRNB and CNS relapse.

Leptomeningeal Metastasis From Neuroblastoma Revealed by 123I-MIBG SPECT/CT.

A 2-year-old girl with history of high-risk neuroblastoma underwent 123I-MIBG, which showed increased 123I-MIBG in the left side of the head. Contrast-enhanced MRI and cerebral spine fluid cytology confirmed the diagnosis of leptomeningeal metastasis.

Treatment of High-Risk Neuroblastoma with Dinutuximab and Chemotherapy Administered in all Cycles of Induction.

Administration of chemoimmunotherapy using concurrent chemotherapy and an anti-GD2 monoclonal antibody (mAb), dinutuximab (DIN), demonstrated efficacy for the treatment of relapsed and refractory neuroblastoma. Chemoimmunotherapy, using a humanized anti-GD2 mAb, demonstrated a signal of activity in a phase 2 study for the treatment of patients with newly diagnosed high-risk neuroblastoma (HRNBL). In this single-institution retrospective study, patients with HRNBL received an Induction chemotherapy regimen plus DIN in all Induction cycles. Toxicity and response data were abstracted from the electronic medical record. Toxicities were graded by CTCAE v.5.0. The end of Induction (EOI) objective response rate was determined using the Revised International Neuroblastoma Response Criteria. Twenty-seven patients with HRNBL (23 newly diagnosed, 16 females, median age 3.9 years) started Induction chemoimmunotherapy from 27 January 2017 to 28 December 2022. All patients received DIN with all cycles of Induction therapy, and all but one patient completed Induction therapy. The most common non-hematologic grade ≥ 3 toxicities included fever (44%), hypoxemia (20%), and hypoalbuminemia (11%). End of Induction responses included eighteen with a complete response (CR), seven with a partial response (PR), one with progressive disease (PD), and zero with a minor response or stable disease. Twenty-six of twenty-seven patients (96%) completed all Induction cycles and were evaluable for a response. The EOI response of PR or better in the evaluable cohort was 96%. Dinutuximab was well tolerated with all Induction cycles, demonstrated an encouraging EOI response rate, and should be evaluated in a randomized study.

Cost-effectiveness analysis of dinutuximab β for the treatment of high-risk neuroblastoma in China.

Dinutuximab β can be used to treat children with high-risk neuroblastoma (NB). Due to its high price, whether dinutuximab β is cost-effective for the treatment of high-risk NB remains uncertain. Therefore, assessing the cost-effectiveness of dinutuximab β in children with high-risk NB is of high importance. The health utilities and economic outcomes in children with high-risk NB were projected using a partitioned survival model. The individual patient data (IPD) of add-on treatment with dinutuximab β (GD2 group) were derived from the literature, while the IPD of traditional therapy (TT group) were obtained from retrospective data of Shanghai Children's Medical Center. Treatment costs included drugs, adverse event-related expenses, and medical resource use. Utility values were obtained from the literature. Costs and quality-adjusted life-years (QALYs) were measured over a 10-year time horizon. Deterministic sensitivity analyses (DSA) and probabilistic sensitivity analyses (PSA) were also conducted. Compared with the TT group, QALY increased in the GD2 group by 0.72 with an increased cost of $171,269.70, leading to an incremental cost-effectiveness ratio of 236,462.75$/QALY. DSA showed that the price of dinutuximab β was the main factor on the results than other parameters. Compared with the TT group, the GD2 group could not be cost-effective in the PSA at the $37,920/QALY threshold. Results found that dinutuximab β is not a cost-effective treatment option for children with high-risk NB unless its price is significantly reduced.

Target Genes of c-MYC and MYCN with Prognostic Power in Neuroblastoma Exhibit Different Expressions during 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.

Development and validation of a nomogram for predicting survival in intermediate- and high-risk neuroblastoma of the Children's Oncology Group risk stratification.

To develop and validate a nomogram for predicting survival in intermediate- and high-risk neuroblastoma patients and to compare the accuracy of the nomogram in predicting survival with Children's Oncology Group (COG) risk stratification. A total of 885 intermediate- and high-risk neuroblastoma patients were enrolled in this study, including 243 patients from our hospital (the training set) and 642 patients from the TARGET database (the validation set). The factors related to event-free survival (EFS) and overall survival (OS) in neuroblastoma were determined to construct the nomogram by Cox regression analysis. The C-index, calibration curves, and area under the time-dependent receiver operating characteristic curves (AUCs) were used to assess the predictive performance of the nomogram. International Neuroblastoma Staging System stage and Mitosis-karyorrhexis index (MKI) were significant unfavorable factors for EFS, while MKI and MYCN status were significant unfavorable factors for OS. The C-index of the nomogram was 0.621 and 0.586 for predicting EFS, 0.650 and 0.570 for predicting OS in the training and validation sets, respectively. The calibration curves revealed good agreement in the EFS and OS predicted by the nomogram. The AUCs of the nomogram for 1-, 2-, 3-year EFS and OS were 0.633, 0.669, 0.604 and 0.672, 0.670, 0.702 in the training set, respectively. Moreover, the nomogram was able to classify patients into two groups according to risk scores, with the "high-risk" group having a lower survival rate than the "intermediate-risk" group. And the nomogram performed better than the COG risk stratification, which had a C-index of 0.537, 0.502 and 0.565, 0.572 for predicting EFS, OS in the training and validation sets, respectively. We developed and validated a prognostic nomogram for intermediate- and high-risk neuroblastoma patients that clinicians can use to make more informed decisions for individual patients.

Analysis of High-Risk Neuroblastoma Transcriptome Reveals Gene Co-Expression Signatures and Functional Features.

Neuroblastoma represents a neoplastic expansion of neural crest cells in the developing sympathetic nervous system and is childhood's most common extracranial solid tumor. The heterogeneity of gene expression in different types of cancer is well-documented, and genetic features of neuroblastoma have been described by classification, development stage, malignancy, and progression of tumors. Here, we aim to analyze RNA sequencing datasets, publicly available in the GDC data portal, of neuroblastoma tumor samples from various patients and compare them with normal adrenal gland tissue from the GTEx data portal to elucidate the gene expression profile and regulation networks they share. Our results from the differential expression, weighted correlation network, and functional enrichment analyses that we performed with the count data from neuroblastoma and standard normal gland samples indicate that the analysis of transcriptome data from 58 patients diagnosed with high-risk neuroblastoma shares the expression pattern of 104 genes. More importantly, our analyses identify the co-expression relationship and the role of these genes in multiple biological processes and signaling pathways strongly associated with this disease phenotype. Our approach proposes a group of genes and their biological functions to be further investigated as essential molecules and possible therapeutic targets of neuroblastoma regardless of the etiology of individual tumors.

BARD1 germline variants induce haploinsufficiency and DNA repair defects in neuroblastoma.

High-risk neuroblastoma is a complex genetic disease that is lethal in more than 50% of patients despite intense multimodal therapy. Through genome-wide association studies (GWAS) and next-generation sequencing, we have identified common single nucleotide polymorphisms and rare, pathogenic or likely pathogenic germline loss-of-function variants in BARD1 enriched in neuroblastoma patients. The functional implications of these findings remain poorly understood. We correlated BARD1 genotype with expression in normal tissues and neuroblastomas, along with the burden of DNA damage in tumors. To validate the functional consequences of germline pathogenic or likely pathogenic BARD1 variants, we used CRISPR-Cas9 to generate isogenic neuroblastoma (IMR-5) and control (RPE1) cellular models harboring heterozygous BARD1 loss-of-function variants (R112*, R150*, E287fs, and Q564*) and quantified genomic instability in these cells via next-generation sequencing and with functional assays measuring the efficiency of DNA repair. Both common and rare neuroblastoma-associated BARD1 germline variants were associated with lower levels of BARD1 mRNA and an increased burden of DNA damage. Using isogenic heterozygous BARD1 loss-of-function variant cellular models, we functionally validated this association with inefficient DNA repair. BARD1 loss-of-function variant isogenic cells exhibited reduced efficiency in repairing Cas9-induced DNA damage, ineffective RAD51 focus formation at DNA double-strand break sites, and enhanced sensitivity to cisplatin and poly (ADP-ribose) polymerase (PARP) inhibition both in vitro and in vivo. Taken together, we demonstrate that germline BARD1 variants disrupt DNA repair fidelity. This is a fundamental molecular mechanism contributing to neuroblastoma initiation that may have important therapeutic implications.

USP7 Inhibition Suppresses Neuroblastoma Growth via Induction of p53-Mediated Apoptosis and EZH2 and N-Myc Downregulation.

Neuroblastoma (NB) is a pediatric malignancy originating from neural crest cells of the sympathetic nervous system that accounts for 15% of all pediatric cancer deaths. Despite advances in treatment, high-risk NB remains difficult to cure, highlighting the need for novel therapeutic approaches. Ubiquitin-specific protease 7 (USP7) is a deubiquitinase that plays a critical role in tumor suppression and DNA repair, and USP7 overexpression has been associated with tumor aggressiveness in a variety of tumors, including NB. Therefore, USP7 is a potential therapeutic target for NB. The tumor suppressor p53 is a known target of USP7, and therefore reactivation of the p53 pathway may be an effective therapeutic strategy for NB treatment. We hypothesized that inhibition of USP7 would be effective against NB tumor growth. Using a novel USP7 inhibitor, Almac4, we have demonstrated significant antitumor activity, with significant decreases in both cell proliferation and cell viability in TP53 wild-type NB cell lines. USP7 inhibition in NB cells activated the p53 pathway via USP7 and MDM2 degradation, leading to reduced p53 ubiquitination and increased p53 expression in all sensitive NB cells. In addition, USP7 inhibition led to decreased N-myc protein levels in both MYCN-amplified and -nonamplified NB cell lines, but no correlation was observed between MYCN amplification and treatment response. USP7 inhibition induced apoptosis in all TP53 wild-type NB cell lines. USP7 inhibition also induced EZH2 ubiquitination and degradation. Lastly, the combination of USP7 and MDM2 inhibition showed enhanced efficacy. Our data suggests that USP7 inhibition may be a promising therapeutic strategy for children with high-risk and relapsed NB.

Rapid detection of telomerase expression of neuroblastoma in paraffin-embedded tissue: combination of in situ hybridisation and quantitative PCR

Neuroblastoma is a heterogeneous paediatric malignant tumour. Telomere maintenance mechanism (TMM) by telomerase activation or alternative lengthening of telomeres (ALT) is a hallmark of high-risk neuroblastoma. However, the prior assays for telomerase, such as TERT expression by RNA sequencing or microarrays, may not be easy to perform in many histopathology laboratories in hospitals. The aims of this study are to assess the utility of ultrasensitive single-cell RNA in situ hybridisation (RNAscope), immunohistochemistry, and RT-qPCR on formalin-fixed, paraffin-embedded tumour samples as diagnostic tools for detecting TERT expression in neuroblastoma. In this study, we detected MYCN amplification in 22 of 222 cases (10%), TERT rearrangements in 18 of 220 cases (8%), and ALT activation in 39 of 222 cases (18%) using fluorescence in situ hybridisation (FISH). By RNA in situ hybridisation, 36 of 210 (17%) pretreatment neuroblastomas were found to have TERT overexpression, which was significantly associated with the high-risk group (33/78, 42%), TERT rearrangements (16/18, 89%), and MYCN amplification (13/22, 59%). None of the tumours with ALT showed TERT staining. In our study, 19 of the 55 MYCN non-amplified high-risk neuroblastomas displayed TERT mRNA expression, including 13 of the 14 TERT rearrangements, none of the 30 ALT-positive cases, and a significant proportion (6/11, 55%) that did not have the aforementioned genomic anomalies. RT-qPCR results correlated well with RNAscope levels (Spearman's rho=0.621, p<0.001, n=94). In conclusion, TERT RNA in situ hybridisation and RT-qPCR are suitable methods to evaluate TERT expression in neuroblastoma. The combination of detection of the genomic alterations and TERT mRNA expression is a powerful strategy for TMM activation detection, which can categorise neuroblastomas into multiple clinical subgroups for risk stratification in routine histopathology practice.

Homoharringtonine as a PHGDH inhibitor: Unraveling metabolic dependencies and developing a potent therapeutic strategy for high-risk neuroblastoma

Neuroblastoma, a childhood cancer affecting the sympathetic nervous system, continues to challenge the development of potent treatments due to the limited availability of druggable targets for this aggressive illness. Recent investigations have uncovered that phosphoglycerate dehydrogenase (PHGDH), an essential enzyme for de novo serine synthesis, serves as a non-oncogene dependency in high-risk neuroblastoma. In this study, we show that homoharringtonine (HHT) acts as a PHGDH inhibitor, inducing intricate alterations in cellular metabolism, and thus providing an efficient treatment for neuroblastoma. We have experimentally verified the reliance of neuroblastoma on PHGDH and employed molecular docking, thermodynamic evaluations, and X-ray crystallography techniques to determine the bond interactions between HHT and PHGDH. Administering HHT to treat neuroblastoma resulted in effective cell elimination in vitro and tumor reduction in vivo. Metabolite and functional assessments additionally disclosed that HHT treatment suppressed de novo serine synthesis, initiating intricate metabolic reconfiguration and oxidative stress in neuroblastoma. Collectively, these discoveries highlight the potential of targeting PHGDH using HHT as a potent approach for managing high-risk neuroblastoma.

Identification of an Ultra-High-Risk Subgroup of Neuroblastoma Patients within the High-Risk Cohort Using a Computed Tomography-Based Radiomics Approach

To identify ultra-high-risk (UHR) neuroblastoma patients who experienced disease-related mortality within 18 months of diagnosis within the high-risk cohort using computed tomography (CT)-based radiomics analysis. A retrospective analysis was conducted on 105 high-risk neuroblastoma patients, divided into a training set (n=74) and a test set (n=31). Radiomics features were extracted and selected from arterial phase CT images, and an optimal radiomics signature was established using the support vector machine algorithm. Evaluation metrics, including area under the curve (AUC) and 95% confidence interval (CI), were calculated. Furthermore, the fit and clinical benefit of the signature, along with its correlation with overall survival (OS), were analyzed. The optimal radiomics signature comprised 11 features. In the training set, AUC and accuracy were 0.911 (95% CI: 0.840-0.982) and 0.892, respectively. In the test set, AUC and accuracy were 0.828 (95% CI: 0.669-0.987) and 0.839, respectively. There was no significant difference between predicted probability and actual probability, and the signature demonstrated net benefit. The concordance index of this signature for predicting OS was 0.743 (95% CI: 0.672-0.814) in the training set and 0.688 (95% CI: 0.566-0.810) in the test set. Moreover, the signature achieved AUC values of 0.832, 0.863, and 0.721 for 1-year, 2-year, and 3-year OS in the training set, and 0.870, 0.836, and 0.638 in the test set for the respective time periods. The utilization of CT-based radiomics signature to identify an UHR subgroup of neuroblastoma patients within the high-risk cohort can help aid in predicting early disease progression.