High-risk Metastatic Neuroblastoma Research Articles

Patterns of Failure in High-Risk Neuroblastoma with or without Metastatic Site Irradiation

<h3>Purpose/Objective(s)</h3> Neuroblastoma is the most common extracranial malignancy diagnosed in childhood. Despite recent improvements in outcomes and an overall 5-year survival of >80%, the majority of children with high-risk neuroblastoma will relapse despite aggressive treatment. As part of their treatment paradigm, radiation treatment (RT) is recommended to the primary site after surgery, as well as to metastatic sites of disease that continue to be metaiodobenzylguanidine (MIBG)-avid post-induction chemotherapy. While there is an abundance of data supporting the former, data is limited on the role of RT to metastatic sites and thus our practice has generally been to withhold RT to metastatic sites. The aim of this study is to examine patterns of failure in patients with high-risk metastatic neuroblastoma to better determine the value of metastatic site RT. <h3>Materials/Methods</h3> We performed an IRB-approved retrospective review from our institutional database of patients with high-risk neuroblastoma who were treated from 2010-2020. The included patients had MIBG-avid metastatic disease at the time of diagnosis, as well as first relapse. These patients achieved a partial or complete response before first relapse, and did not receive total body radiation or therapy with ¹³¹I-MIBG prior to first relapse. We analyzed the patients who failed and compared the anatomical location of all sites at initial relapse to sites that were MIBG-avid on post-induction chemotherapy scan. <h3>Results</h3> With a median follow up of 38 months, 13 of the 41 patients (32%) treated during this time recurred, and 5 (12%) died. 3 patients who recurred were excluded from the analysis as they did not have sufficient records. Of the 10 patients analyzed with recurrent disease, there were 70 MIBG-avid metastatic sites on post-induction chemotherapy scan, and of these only 11 (16%) received RT. There were 13 total MIBG-avid metastatic sites at time of first recurrence, of which 6 (46%) were in original site present on post-induction chemo MIBG scan, 1 which received radiation treatment and the other 5 (38%) did not. Of the sites that did receive radiation treatment, 10/11 (91%) did not recur. In analysis per patient, 4/10 (40%) recurred initially in metastatic sites that were MIBG-avid on post-induction scan, while the remaining majority recurred in both initial and new sites or new sites only. <h3>Conclusion</h3> In this unique cohort of high-risk neuroblastoma patients who mostly did not receive RT to post-induction chemotherapy MIGB-avid metastatic sites, a little less than half of the patients and sites relapsed in initial areas, while the remaining relapsed at new sites. Those sites that were treated with RT had excellent local control >90%, consistent with prior reports. While this data suggests that RT to metastatic sites on post-induction chemotherapy MIBG scan may improve outcomes, the sample size is limited and caution must be taken to balance potential benefits with risks, especially in situations with several metastatic sites.

Abstract 5218: Oral beta-glucan enhanced anti-ganglioside antibody titer after vaccination against high-risk neuroblastoma: Results of a randomized trial

Abstract Background and Aim: Ganglioside GD2/GD3 vaccine stimulated robust antibody response among patients with high-risk metastatic neuroblastoma (HR-NB) who had prior disease progression (J Clin Oncol 39:215-226, 2020). High anti-GD2-IgG1antibody titer was associated with improved progression-free survival and overall survival in multivariable analyses. A gel formulation of yeast beta-glucan was used as an oral vaccine adjuvant, but its importance remained unproven. Patients and Methods: In a follow-up randomized Phase II trial (Clinicaltrials.gov identifier: NCT00911560), seven vaccine injections at 1, 2, 3, 8, 20, 32 and 52 weeks were administered to patients with HR-NB during their first or subsequent remissions. Each subcutaneous vaccine injection consisted of 30 ug each of GD2 and GD3, which was lactonized and conjugated to keyhole limpet hemocyanin and mixed with the subcutaneous saponin OPT-821 adjuvant at 150 µg/m2. No patients in this analysis received prior ganglioside vaccine. They were randomized to Arm 1 (n=54) receiving no glucan, or Arm 2 (n=53) receiving oral beta-glucan regimen (40 mg/kg/day, 14 days on/14 days off) starting at week 1. From week 6 onwards, all 107 patients received oral beta-glucan regimen through year end or up to disease progression. Serum IgG1 against GD2 and GD3, and IgM against GD2 were measured by ELISA at (right before) each vaccine injection. Wilcoxon rank sum test was used to compare antibody titers between the 2 arms. Results: In both arms, patients had comparable disease status at study entry, with 70% each in first remission. The remaining patients were in second remission after one prior disease progression. Consistently higher antibody response was observed among Arm 2 patients. The primary endpoint was met for anti-GD2-IgG1 titer at vaccine injection #6 at 32 weeks (p=0.08). Per protocol design, statistical significance (p&amp;lt;0.1) of anti-GD2-IgG1titer was observed at vaccine #5 (2.7-fold), #6 (1.4-fold), and #7 (4.1-fold), irrespective of being in first or second remission at study entry. Total anti-GD2-IgG1 antibody exposure (AUC) up to vaccine #7 for Arm 2 was 1.9-fold higher than that of Arm 1 (p=0.05). In contrast, anti-GD2-IgM achieved statistical significance only at vaccine #4 (p=0.001), and for anti-GD3-IgG1, only at vaccine #5 (p=0.07). Oral beta-glucan was well tolerated, and no patient showed pain or neuropathic side effects despite having higher antibody titers. Conclusion: Adding oral yeast beta-glucan as vaccine adjuvant during the first 6 weeks of immunization significantly enhanced the anti-GD2-IgG1 antibody response without added toxicities. Its impact on patient survival will require a longer clinical follow-up. Citation Format: Irene Y. Cheung, Audrey Mauguen, Yi Feng, Govind Ragupathi, Ellen Basu, Stephen S. Roberts, Shakeel Modak, Brian H. Kushner, Nai-Kong V. Cheung. Oral beta-glucan enhanced anti-ganglioside antibody titer after vaccination against high-risk neuroblastoma: Results of a randomized trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5218.

Long-term analysis of children with metastatic neuroblastoma treated in the ENSG5 randomised clinical trial.

The European Neuroblastoma Study Group 5 (ENSG5) trial showed that time-intensive "rapid" induction chemotherapy (COJEC) was superior to "standard" 3-weekly chemotherapy for children with high-risk metastatic neuroblastoma. Long-term outcomes of the ENSG5 trial were analysed. Patients with metastatic neuroblastoma aged ≥12 months were randomly assigned to "standard" or "rapid" induction, receiving the same chemotherapy drugs and doses. Event-free survival (EFS) and overall survival (OS) were analysed and prognostic factors evaluated. Amongst patients surviving >5 years, a population of children with persistent metastatic disease after the end of treatment was identified and described. Ten-year EFS was 18.2% (95% confidence interval: 12.2-25.2) for the "standard" arm and 26.8% (19.5-34.7) for the "rapid" arm (hazard ratio [HR] 0.85, P=0.28). Ten-year OS for the "standard" arm was 19.7% (13.4-26.8) and 28.3% (20.8-36.2) for the "rapid arm" (HR 0.83, P=0.19). There was a trend for worse EFS and OS for patients having MYCN amplification (HR 1.37 and 1.40, respectively) and those with partial and mixed response to induction (HR 1.69 and 1.75 for EFS and 1.66 and 2.00 for OS, respectively). Among 69 patients who survived >5 years, six had persistent metastatic disease after the end of treatment. The benefit of the "rapid" induction regimen seems to be maintained in the long term, although the small number of survivors could justify the lack of statistical significance. MYCN amplification and poor metastatic response to induction could be associated with worse outcomes. A small group of patients with persistent metastatic disease that survived long term has been described.

Inhibition of Microsomal Prostaglandin E Synthase-1 in Cancer-Associated Fibroblasts Suppresses Neuroblastoma Tumor Growth

Despite recent progress in diagnosis and treatment, survival for children with high-risk metastatic neuroblastoma is still poor. Prostaglandin E2 (PGE2)-driven inflammation promotes tumor growth, immune suppression, angiogenesis and resistance to established cancer therapies. In neuroblastoma, cancer-associated fibroblasts (CAFs) residing in the tumor microenvironment are the primary source of PGE2. However, clinical targeting of PGE2 with current non-steroidal anti-inflammatory drugs or cyclooxygenase inhibitors has been limited due to risk of adverse side effects. By specifically targeting microsomal prostaglandin E synthase-1 (mPGES-1) activity with a small molecule inhibitor we could block CAF-derived PGE2 production leading to reduced tumor growth, impaired angiogenesis, inhibited CAF migration and infiltration, reduced tumor cell proliferation and a favorable shift in the M1/M2 macrophage ratio. In this study, we provide proof-of-principle of the benefits of targeting mPGES-1 in neuroblastoma, applicable to a wide variety of tumors. This non-toxic single drug treatment targeting infiltrating stromal cells opens up for combination treatment options with established cancer therapies.

Abstract A02: Neuroblastoma patient-derived orthotopic xenografts: Clinically relevant models for drug testing

Abstract Widespread metastasis is a major problem for the treatment of high-risk neuroblastoma. Relevant neuroblastoma animal models are hence needed to study and target high-risk metastatic neuroblastoma. We developed neuroblastoma patient-derived orthotopic xenografts (PDXs) using viably cryopreserved or fresh patient neuroblastoma fragments which were implanted orthotopically into immunodeficient NSG mice. Immunohistochemistry showed that PDXs retain neuroblastoma markers and a highly infiltrative growth pattern. Importantly, we found distant metastasis to lungs, liver and bone marrow. Single nucleotide polymorphism array analysis confirmed that PDXs maintain patient-specific chromosomal aberrations 1p del, MYCN amp and 17q gain. In vitro cultures established from the PDXs express neuroblastoma markers and retain their tumorigenic and metastatic ability in vivo after orthotopic injection. PDXs were highly vascularized with mouse endothelial cells and one PDX model formed tumor vasculature through co-engrafted human tumor endothelial cells. Tumors also contained pericytes, cancer-associated fibroblasts, macrophages and extracellular components resembling the patient disease. PDXs established in athymic nude mice additionally developed intratumoral lymphatic vessels and contained mouse-derived CD45+ lymphoid cells. Thus, PDXs maintain important tumor microenvironment hallmarks for high stage neuroblastoma. Furthermore, we demonstrate the feasibility of using short-term in vitro cultured PDX-derived cells as a drug-testing model, and show that the HIF2A oncogene is transcriptionally regulated at hypoxia via the PI3K/mTORC2 pathway in these cells. The results establish the PI3K and mTORC2 pathways as potential therapeutic targets of aggressive neuroblastoma. In conclusion, neuroblastoma orthotopic PDXs resemble many clinical aspects of the patient disease, e.g., distant metastases, and PDXs are valuable models for preclinical drug testing. Citation Format: Noémie Braekeveldt, Caroline Wigerup, David Gisselsson, Sofie Mohlin, My Merselius, Siv Beckman, Irene Tadeo, Anna P. Berbegall, Ingrid Öra, Samuel Navarro, Rosa Noguera, Sven Påhlman, Daniel Bexell. Neuroblastoma patient-derived orthotopic xenografts: Clinically relevant models for drug testing. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Pediatric Cancer Research: From Mechanisms and Models to Treatment and Survivorship; 2015 Nov 9-12; Fort Lauderdale, FL. Philadelphia (PA): AACR; Cancer Res 2016;76(5 Suppl):Abstract nr A02.

Serum-circulating miRNAs predict neuroblastoma progression in mouse model of high-risk metastatic disease.

BackgroundCirculating miRNAs have momentous clinical relevance as prognostic biomarkers and in the progression of solid tumors. Recognizing novel candidates of neuroblastoma-specific circulating miRNAs would allow us to identify potential prognostic biomarkers that could predict the switch from favorable to high-risk metastatic neuroblastoma (HR-NB).ResultsUtilizing mouse models of favorable and HR-NB and whole miRnome profiling, we identified high serum levels of 34 and low levels of 46 miRNAs in animals with HR-NB. Preferential sequence homology exclusion of mouse miRNAs identified 25 (11 increased; 14 decreased) human-specific prognostic marker candidates, of which, 21 were unique to HR-NB. miRNA QPCR validated miRnome profile. Target analysis defined the candidate miRNAs' signal transduction flow-through and demonstrated their converged roles in tumor progression. miRNA silencing studies verified the function of select miRNAs on the translation of at least 14 target proteins. Expressions of critical targets that correlate tumor progression in tissue of multifarious organs identify the orchestration of HR-NB. Significant (>10 fold) increase in serum levels of miR-381, miR-548h, and miR-580 identify them as potential prognostic markers for neuroblastoma progression.ConclusionFor the first time, we identified serum-circulating miRNAs that predict the switch from favorable to HR-NB and, further imply that these miRNAs could play a functional role in tumor progression.

MYCN controls an alternative RNA splicing program in high-risk metastatic neuroblastoma

The molecular mechanisms underlying the aggressive behavior of MYCN driven neuroblastoma (NBL) is under intense investigation; however, little is known about the impact of this family of transcription factors on the splicing program. Here we used high-throughput RNA sequencing to systematically study the expression of RNA isoforms in stage 4 MYCN-amplified NBL, an aggressive subtype of metastatic NBL. We show that MYCN-amplified NBL tumors display a distinct gene splicing pattern affecting multiple cancer hallmark functions. Six splicing factors displayed unique differential expression patterns in MYCN-amplified tumors and cell lines, and the binding motifs for some of these splicing factors are significantly enriched in differentially-spliced genes. Direct binding of MYCN to promoter regions of the splicing factors PTBP1 and HNRNPA1 detected by ChIP-seq demonstrates that MYCN controls the splicing pattern by direct regulation of the expression of these key splicing factors. Furthermore, high expression of PTBP1 and HNRNPA1 was significantly associated with poor overall survival of stage4 NBL patients (p ≤ 0.05). Knocking down PTBP1, HNRNPA1 and their downstream target PKM2, an isoform of pro-tumor-growth, result in repressed growth of NBL cells. Therefore, our study reveals a novel role of MYCN in controlling global splicing program through regulation of splicing factors in addition to its well-known role in the transcription program. These findings suggest a therapeutically potential to target the key splicing factors or gene isoforms in high-risk NBL with MYCN-amplification.

Neuroblastoma patient-derived orthotopic xenografts retain metastatic patterns and geno- and phenotypes of patient tumours.

Neuroblastoma is a childhood tumour with heterogeneous characteristics and children with metastatic disease often have a poor outcome. Here we describe the establishment of neuroblastoma patient-derived xenografts (PDXs) by orthotopic implantation of viably cryopreserved or fresh tumour explants of patients with high risk neuroblastoma into immunodeficient mice. In vivo tumour growth was monitored by magnetic resonance imaging and fluorodeoxyglucose–positron emission tomography. Neuroblastoma PDXs retained the undifferentiated histology and proliferative capacity of their corresponding patient tumours. The PDXs expressed neuroblastoma markers neural cell adhesion molecule, chromogranin A, synaptophysin and tyrosine hydroxylase. Whole genome genotyping array analyses demonstrated that PDXs retained patient-specific chromosomal aberrations such as MYCN amplification, deletion of 1p and gain of chromosome 17q. Thus, neuroblastoma PDXs recapitulate the hallmarks of high-risk neuroblastoma in patients. PDX-derived cells were cultured in serum-free medium where they formed free-floating neurospheres, expressed neuroblastoma gene markers MYCN, CHGA, TH, SYP and NPY, and retained tumour-initiating and metastatic capacity in vivo. PDXs showed much higher degree of infiltrative growth and distant metastasis as compared to neuroblastoma SK-N-BE(2)c cell line-derived orthotopic tumours. Importantly, the PDXs presented with bone marrow involvement, a clinical feature of aggressive neuroblastoma. Thus, neuroblastoma PDXs serve as clinically relevant models for studying and targeting high-risk metastatic neuroblastoma.What's new?Neuroblastoma is a childhood tumour with heterogeneous characteristics and children with metastatic disease have a poor outcome. Here, the authors established neuroblastoma patient-derived xenografts (PDXs) by orthotopic implantation of viably cryopreserved or fresh tumour explants of patients with high-risk neuroblastoma into immunodeficient mice. The PDXs retained the genotype and phenotype of patient tumours and exhibited substantial infiltrative growth and metastasis to distant organs including bone marrow. PDX-derived neuroblastoma cells were expanded in vitro and retained tumourigenic and metastatic capacity in vivo. The PDXs may thus represent an important tool for investigating neuroblastoma growth and metastasis as well as drug targeting.

GD2-Targeted Immunotherapy and Radioimmunotherapy

Ganglioside GD2 is a tumor-associated surface antigen found in a broad spectrum of human cancers and stem cells. They include pediatric embryonal tumors (neuroblastoma, retinoblastoma, brain tumors, osteosarcoma, Ewing sarcoma, rhabdomyosarcoma), as well as adult cancers (small cell lung cancer, melanoma, soft tissue sarcomas). Because of its restricted normal tissue distribution, GD2 has been proven safe for antibody targeting. Anti-GD2 antibody is now incorporated into the standard of care for the treatment of high-risk metastatic neuroblastoma. Building on this experience, novel combinations of antibodies, cytokines, cells, and genetically engineered products all directed at GD2 are rapidly moving into the clinic. In this review, past and present immunotherapy trials directed at GD2 will be summarized, highlighting the lessons learned and the future directions.

SP021 Bone marrow minimal residual disease (MRD) was the strongest predictor of survival from high risk metastatic neuroblastoma (NB) following anti-GD2 immunotherapy, when tested in multivariate models that include FcR polymorphism and missing ligand for inhibitory killer-immunoglobulin-like receptor (KIR)

SP021 Bone marrow minimal residual disease (MRD) was the strongest predictor of survival from high risk metastatic neuroblastoma (NB) following anti-GD2 immunotherapy, when tested in multivariate models that include FcR polymorphism and missing ligand for inhibitory killer-immunoglobulin-like receptor (KIR)

Outcomes After Definitive Radiation Treatment of Metastatic Sites in Patients With Stage IV Neuroblastoma

Purpose/Objective(s): In patients (pts) with high-risk metastatic neuroblastoma (NB), the benefit of radiation therapy (RT) to metastatic sites (MS) as part of primary treatment has not been fully investigated. The purpose of this study was to characterize the distribution of MS, pattern of RT for MS, and evaluate patterns of recurrence and outcomes in this patient population. Materials/Methods: The records of all pts diagnosed with stage IV NB (International NB Staging System) between 8/2000 and 1/2010 at Children’s Healthcare of Atlanta were reviewed. Exclusion criteria included: bone-marrow-only MS, total body irradiation, or no imaging follow-up. A total of 37 pts met eligibility. Image-defined MS involvement was documented at the following time points: pre-induction (preI) chemotherapy (CTX), post-induction (postI) CTX, and post-transplant/pre-RT (preRT). MS involvement was further classified into 4 categories based on location: appendicular skeleton, axial skeleton, calvarium, and soft tissue. The median follow-up period for pts without relapse was 61 months (range, 9-113 months). Actuarial rates were determined by the Kaplan-Meier method from start of induction CTX. Time-to-event rates were compared using the log-rank test. Results: The median age at diagnosis was 3.7 years (range, 0.7-20.7). Extent of resection was gross total, subtotal, and biopsy only in 26 pts (70%), 9 pts (24%), and 2 pts (5%), respectively. Five-year (yr) overall survival (OS) for all pts was 67%. All pts received RT to their primary site and regional lymph nodes. The 5-yr local control rate of the primary site was 94%. Thirteen pts (35%) received upfront RT to one MS category: 2 pts (appendicular), 4 pts (axial), 3 pts (calvarium), and 4 pts (soft tissue). The median RT dose to MS was 21.6 Gy (range, 21.0-30.6) at 1.8 Gy/ fraction. Among these pts (n Z 13), infield recurrence occurred in 4 pts (31%), and out-of-field recurrence occurred in 5 pts (38%). There was no difference in 5-yr OS between pts who were treated with or without RT to a MS (73% vs 63%, pZ NS). For pts who had positive MS after induction CTX vs those without, there was no difference in 5-yr OS (64% vs 68%, p Z NS) or 5-yr MS control (56% vs 69%, p Z NS). Of the 78 preI CTX involved MS locations, 71% became negative on postI CTX imaging. Of the 70 negative MS locations on preI CTX imaging, only 3% became positive on postI CTX imaging. Ten pts had preRT imaging, and of the 11 postI CTX positive MS locations, 3 became negative on preRT imaging. Conclusions: Treatment of MS with RT as part of definitive therapy in stage IV NB was not associated with improved OS or MS control. Infield recurrences after RT to MS are substantial, and RT dose may need to be evaluated. The presence of residual MS after induction CTX did not predict poorer survival. Further studies are necessary to clarify the role of RT to MS in the definitive setting. Author Disclosure: S. Kandula: None. R.S. Prabhu: None. H. Katzenstein: None. M. Qayed: None. N. Esiashvili: None.

Two cases of neuroblastoma comprising two distinct clones

We report two cases of high-risk metastatic neuroblastoma, comprising two biologically distinct components in the adrenal primary tumor, which showed clear differences not only histologically but also in MYCN amplification and HA-RAS/TRKA immunoreactivity (Case 1), anaplastic lymphoma kinase (ALK) immunoreactivity (Case 2). These two cases with multiple separated components were similar to cases classified as ganglioneuroblastoma, nodular subtype (GNBn), in terms of composite tumor. Comparable to the GNBn category, the prognosis of the patients described here may depend on the components with unfavorable histology according to International Neuroblastoma Pathology Classification. Further analyses of such composite neuroblastoma cases are important for assessing disease prognosis.

Radiation therapy as part of local control of metastatic neuroblastoma: the St Jude Children's Research Hospital experience

Purpose The purpose of the study was to compare outcomes of pediatric patients with high-risk metastatic neuroblastoma who received radiotherapy (RT) with those of patients who did not. Patients and methods We reviewed the records of 63 patients with newly diagnosed metastatic neuroblastoma treated at our institution (1989-2001) to investigate their characteristics at presentation, dose and field of RT, treatment response, and failure patterns. Results Seventeen patients received RT, and 46 did not. In the RT group, a greater percentage of patients had residual disease before consolidation than did those in the no-RT group (88.2% vs 69.6%, P = .008). Gross total resection was achieved less often in the RT group (65% vs 89%, P = .055), but the 5-year cumulative incidences of local failure were similar (35.3% ± 12.4% vs 32.6% ± 7.1%). Although there was no difference in 5-year event-free survival, overall survival was better in the no-RT group (47.8% ± 7.2% vs 23.5% ± 9.2%, P = .026). Conclusion The addition of RT to the therapy of a group of patients with more residual locoregional disease appeared to improve the local failure rate to approximately that of patients with less residual disease. Radiotherapy may provide even greater benefit to those with less residual disease before consolidation.

Fenretinide-induced caspase-8 activation and apoptosis in an established model of metastatic neuroblastoma

BackgroundResistance of high-risk metastatic neuroblastoma (HR-NB) to high dose chemotherapy (HD-CT) raises a major therapeutic challenge in pediatric oncology. Patients are treated by maintenance CT. For some patients, an adjuvant retinoid therapy is proposed, such as the synthetic retinoid fenretinide (4-HPR), an apoptotic inducer. Recent studies demonstrated that NB metastasis process is enhanced by the loss of caspase-8 involved in the Integrin-Mediated Death (IMD) process. As the role of caspase-8 appears to be critical in preventing metastasis, we aimed at studying the effect of 4-HPR on caspase-8 expression in metastatic neuroblasts.MethodsWe used the human IGR-N-91 MYCN-amplified NB experimental model, able to disseminate in vivo from the primary nude mouse tumor xenograft (PTX) into myocardium (Myoc) and bone marrow (BM) of the animal. NB cell lines, i.e., IGR-N-91 and SH-EP, were treated with various doses of Fenretinide (4-HPR), then cytotoxicity was analyzed by MTS proliferation assay, apoptosis by the propidium staining method, gene or protein expressions by RT-PCR and immunoblotting and caspases activity by colorimetric protease assays.ResultsThe IGR-N-91 parental cells do not express detectable caspase-8. However the PTX cells established from the primary tumor in the mouse, are caspase-8 positive. In contrast, metastatic BM and Myoc cells show a clear down-regulation of the caspase-8 expression. In parallel, the caspases -3, -9, -10, Bcl-2, or Bax expressions were unchanged. Our data show that in BM, compared to PTX cells, 4-HPR up-regulates caspase-8 expression that parallels a higher sensitivity to apoptotic cell death. Stable caspase-8-silenced SH-EP cells appear more resistant to 4-HPR-induced cell death compared to control SH-EP cells. Moreover, 4-HPR synergizes with drugs since apoptosis is restored in VP16- or TRAIL-resistant-BM cells. These results demonstrate that 4-HPR in up-regulating caspase-8 expression, restores and induces apoptotic cell death in metastatic neuroblasts through caspase-8 activation.ConclusionThis study provides basic clues for using fenretinide in clinical treatment of HR-NB patients. Moreover, since 4-HPR induces cell death in caspase-8 negative NB, it also challenges the concept of including 4-HPR in the induction of CT of these patients.