Why does neuroblastoma attract us so much?
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Animals↗
Biomedical subjects
B De Bernardi
Publications and source records attributed to B De Bernardi.
Quality assessment of genetic markers used for therapy stratification.
PURPOSE: Therapy stratification based on genetic markers is becoming increasingly important, which makes commitment to the highest possible reliability of the involved markers mandatory. In neuroblastic tumors, amplification of the MYCN gene is an unequivocal marker that indicates aggressive tumor behavior and is consequently used for therapy stratification. To guarantee reliable and standardized quality of genetic features, a quality-assessment study was initiated by the European Neuroblastoma Quality Assessment (ENQUA; connected to International Society of Pediatric Oncology) Group. MATERIALS AND METHODS: One hundred thirty-seven coded specimens from 17 tumors were analyzed in 11 European national/regional reference laboratories using molecular techniques, in situ hybridization, and flow and image cytometry. Tumor samples with divergent results were re-evaluated. RESULTS: Three hundred fifty-two investigations were performed, which resulted in 23 divergent findings, 17 of which were judged as errors after re-evaluation. MYCN analyses determined by Southern blot and in situ hybridization led to 3.7% and 4% of errors, respectively. Tumor cell content was not indicated in 32% of the samples, and 11% of seemingly correct MYCN results were based on the investigation of normal cells (eg, Schwann cells). Thirty-eight investigations were considered nonassessable. CONCLUSION: This study demonstrated the importance of revealing the difficulties and limitations for each technique and problems in interpreting results, which are crucial for therapeutic decisions. Moreover, it led to the formulation of guidelines that are applicable to all kinds of tumors and that contain the standardization of techniques, including the exact determination of the tumor cell content. Finally, the group has developed a common terminology for molecular-genetic results.
Biomarkers, Tumor↗
Intratumoural heterogeneity of 1p deletions and MYCN amplification in neuroblastomas.
BACKGROUND: At least three genetic hallmarks identify aggressive tumour behaviour in neuroblastomas; amplification of the oncogene MYCN; deletion (loss of heterozygosity [LOH]) at the short arm of chromosome 1 (del1p36), seen in approximately 28% of the cases; and di-tetraploidy. The MYCN oncogene is amplified in approximately 23% of all neuroblastomas and becomes important for the stratification of therapy in localised and 4s tumours. Up to now, it has been believed that the genetic constellation of neuroblastic tumours is stable and does not alter during tumour evolution or during tumour progression. PROCEDURE: Using fluorescence in situ hybridisation techniques (FISH) to investigate different tumour areas on touch preparations and histological sections, we show that genetic heterogeneity can be detected in neuroblastomas, especially in tumours detected by urinary mass screening. CONCLUSION: The identification of such cell clones is important, because the MYCN amplification and/or the deletion at 1p36 appear to be responsible for aggressive local growth and development of metastases.
Chromosomes, Human, Pair 1↗
Molecular alterations in a case of bilateral adrenal neuroblastoma.
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Adrenal Gland Neoplasms↗
A 7-year-old girl with hip pain and leg weakness.
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Bone Neoplasms↗
Megatherapy combining I(131) metaiodobenzylguanidine and high-dose chemotherapy with haematopoietic progenitor cell rescue for neuroblastoma.
Despite the use of aggressive chemotherapy, stage 4 high risk neuroblastoma still has very poor prognosis which is estimated at 25%. Metabolic radiotherapy with I(131) MIBG appears a feasible option to enhance the effects of chemotherapy. Seventeen patients having MIBG-positive residual disease received 4.1-11.1 mCi/kg of I(131) MIBG 7-10 days before initiating the high-dose chemotherapy cycle consisting of busulphan 16 mg/kg and melphalan 140 mg/m(2) followed by PBSC infusion. We compared the toxicity in these patients to that seen in 15 control subjects with neuroblastoma who underwent a PBSC transplant without MIBG therapy. We observed greater toxic involvement of the gastrointestinal system in children treated with I(131) MIBG: grade 2 or 3 mucositis developed in 13/17 patients treated with I(131) MIBG and in 9/15 treated without it. Grade 1-2 gastrointestinal toxicity occurred in 12/17 children given MIBG and in 5/15 of the controls. One child receiving I(131) MIBG developed transient interstitial pneumonia. Another child who also received I(131) MIBG after PBSC rescue developed fatal pneumonia after the third course of metabolic radiotherapy. Our experience indicates that MIBG can be included in the high-dose chemotherapy regimens followed by PBSC rescue for children with residual neuroblastoma taking up MIBG. Attention should be paid to avoiding lung complications. Prospective studies are needed to demonstrate the real efficacy of this treatment.
3-Iodobenzylguanidine↗
Exclusion of candidate genes and chromosomal regions in familial neuroblastoma.
Two families with recurrence of neuroblastoma one Italian and one British with three and two affected children respectively were genotyped using polymorphic markers on chromosome 1 spanning the p32-p36 region frequently deleted in neuroblastoma tumor cells. Linkage to this region was excluded by haplotype inspection and negative lod scores. Furthermore, the exclusion of genes involved in neurocristopathies sometimes associated with neuroblastoma was carried out by typing the Italian family with polymorphic markers located in or near the corresponding genes. Finally, linkage analysis in the two families showed negative lod scores for markers spanning the 16p12-13 chromosomal region where a locus for familial neuroblastoma has been recently mapped. Our findings indicate that different genes are involved in the pathogenesis of familial neuroblastoma.
Chromosome Mapping↗
In vivo cytoreduction studies and cell sorting--enhanced tumor-cell detection in high-risk neuroblastoma patients: implications for leukapheresis strategies.
PURPOSE: To improve autologous leukapheresis strategies in high-risk neuroblastoma (NB) patients with extensive bone marrow involvement at diagnosis. PATIENTS AND METHODS: Anti-G(D2) immunocytochemistry (sensitivity, 1 in 10(5) to 10(6) leukocytes) was used to evaluate blood and bone marrow disease at diagnosis and during the recovery phase of the first six chemotherapy cycles in 57 patients with stage 4 NB and bone marrow disease at diagnosis. A total of 42 leukapheresis samples from the same patients were evaluated with immunocytology, and in 24 of these patients, an anti-G(D2) immunomagnetic enrichment step was used to enhance tumor-cell detection. RESULTS: Tumor cytoreduction was much faster in blood compared with bone marrow (3.2 logs after the first cycle and 2.1 logs after the first two cycles, respectively). Bone marrow disease was often detectable throughout induction, with a trend to plateau after the fourth cycle. By direct anti-G(D2) immunocytology, a positive leukapheresis sample was obtained in 7% of patients after either the fifth or sixth cycle; when NB cell immunomagnetic enrichment was applied, 25% of patients had a positive leukapheresis sample (sensitivity, 1 in 10(7) to 10(8) leukocytes). CONCLUSION: Standard chemotherapy seems to deliver most of its in vivo purging effect within the first four cycles. In patients with overt marrow disease at diagnosis, postponing hematopoietic stem-cell collection beyond this point may not be justified. Tumor-cell clearance in blood seems to be quite rapid, and earlier collections via peripheral-blood leukapheresis might be feasible. Immunomagnetically enhanced NB cell detection can be highly sensitive and can indicate whether ex vivo purging should be considered.
Adolescent↗
False-negative metaiodobenzylguanidine scintigraphy at diagnosis of neuroblastoma.
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3-Iodobenzylguanidine↗
Features and outcome of neuroblastoma detected before birth.
BACKGROUND/PURPOSE: The growing use of routine ultrasonography during pregnancy is leading to an increasing number of prenatally diagnosed neuroblastomas. Optimal strategy has not yet been defined for these patients, because knowledge on this particular neuroblastoma (NB) population is still limited. However, definite guidelines are needed to avoid inadequate treatment. The authors analyzed the cases of antenatally detected NB (ADNB) reported in the Italian Neuroblastoma Registry during the past 6 years to elucidate the features of this subset of NB. METHODS: The Italian Neuroblastoma Registry was reviewed for the period January 1993 to December 1998 to collect clinical, radiographic, surgical, and histopathological data on ADNB cases. NB stage was evaluated according to INSS criteria. All patients had undergone imaging (computed tomography or magnetic resonance imaging) of the primary tumor and bone marrow biopsy before surgical resection. RESULTS: Seventeen patients were identified. Primary tumour site was adrenal glands in 16 cases and retroperitoneal ganglia in 1. Stage distribution was stage I, 13 cases; stage II-A, 1 case; stage II-B, 1 case; stage IV-S, 2 cases. All cases underwent primary tumour resection. Mean age at surgery was 4 weeks. Resection of primary tumor was radical in 16 cases, partial in 1. All tumors were characterised by favourable histology according to Shimada classification. N-myc gene amplification was studied in 14 patients. N-myc amplification was detected only in a newborn with stage II-A NB, who died of massive bleeding 2 days after tumor resection. DNA index and 1p deletion were studied in 11 and 8 patients, respectively. Both diploidy and deletion of 1p were observed in a newborn who subsequently died of disease progression despite surgery, chemotherapy, and radiation therapy. Fourteen of 17 patients currently are alive and free of disease, and one with IV-S NB and short follow-up is alive with disease. CONCLUSIONS: Our data give evidence that in most cases infants with ADNB represent a subset of patients with excellent outcome. Aggressive treatment may not always be necessary. Infants with ADNB with unfavorable features should undergo early surgical excision, whereas patients with favourable features could be observed awaiting spontaneous regression of the mass, reserving delayed surgery for tumors that increase in size or do not regress.
Adrenal Gland Neoplasms↗
Alveolar soft part sarcoma in children and adolescents: A report from the Soft-Tissue Sarcoma Italian Cooperative Group.
BACKGROUND: Alveolar soft part sarcoma (ASPS) is a rare malignant tumor and little is known about its clinical features and management. We report on a series of 19 pediatric patients managed over 20 years. PATIENTS AND METHODS: Primary conservative surgery was performed in all patients and was radical in nine, non-radical in three; seven patients underwent biopsy alone (3 unresectable tumors, 4 metastatic disease). In two cases radical surgery was performed after primary chemotherapy. Radiotherapy was delivered to 8 patients, chemotherapy to 15. RESULTS: After a median follow-up of 74 months, the five-year survival was 80% for the whole series, 91% for patients with localized disease, 100% for patients with tumor < or = 5 cm, and 31% for those > 5 cm; 16 of 19 patients were alive (12 of 12 with grossly-resected tumor in first continuous remission). Chemotherapy achieved two partial remission among seven evaluable patients. CONCLUSIONS: Pediatric ASPS has a more favorable prognosis than its adult counterpart. In this series, tumor size correlates with metastatic disease at onset and is the major factor influencing survival. Surgery is the mainstay of therapy. The effectiveness of adjuvant therapy remains to be established, though radiotherapy may be advisable in cases of inadequate surgery.
Adolescent↗
Quantitative determination of sst2 gene expression in neuroblastoma tumor predicts patient outcome.
Neuroblastoma (NB) is the most common pediatric neuroendocrine tumor, and it is characterized by a quite variable clinical course. We previously found a great variability in the expression of somatostatin receptor type 2 (sst2) in several human NB cell lines and primary tumors. In this report we investigated whether expression of sst2 is somehow related to clinical outcome. We performed a retrospective study on 54 patients with a maximum follow-up of 100 months. The concentration of specific messenger ribonucleic acid (mRNA) for sst2 was measured by competitive RT-PCR and validated, in a small subset of samples, by quantitative imaging of gene (in situ hybridization) and protein (immunohistochemistry) expression. We found that sst2 mRNA was variably expressed in all NB tumors (range, 2.5 x 10(5) to 8 x 10(9) molecules/microg RNA) with a relevant reduction in the more advanced stage (P < 0.01). Analysis of Kaplan-Meier curves indicated that sst2 expression is positively related to the overall (P < 0.0001) and event-free (P < 0.0001) survival. Expression of sst2 was negatively related to tumor stage (P < 0.02) and MYCN amplification (P < 0.001), a poor prognostic factor. However, the prognostic information derived from sst2 is apparently independent from MYCN amplification, as assessed by stratifying sst2 values according to MYCN. In addition, the expression of sst2 was the only significant prognostic factor (P < 0.02) when it was included in a multivariate model containing other well known prognostic factors such as age, stage, and MYCN amplification. Hence, we propose that sst2 expression represents a new prognostic marker for NB. The main clinical value of a quantitative measure of sst2 lies in its ability to detect patients at low risk, independently from other prognostic factor, including MYCN amplification.
Brain Neoplasms↗
[Intractable diarrhea and neuroblastoma: report of a clinical case].
One of the typical presentations of neuroblastoma is intractable diarrhea or wdha (watery diarrhea, hypokalemia, achloridria). The case admitted to our Pediatric Surgery Department presented watery diarrhea due to VIP hyperincretion caused by a stage 1 neuroblastoma, whose ablation allowed a complete resolution of the clinical conditions. This case report can be useful in the discussion of the differential diagnosis of the most common clinical pictures.
Adrenal Gland Neoplasms↗
Stage-independent expression and genetic analysis of tp73 in neuroblastoma.
The tp73 gene, a tp53 homologue, has been sub-regionally mapped at 1p36.3, a chromosomal region frequently deleted in neuroblastoma. Due to its chromosomal localization and to the mono-allelic expression observed in some neuroblastoma cell lines, it was proposed that tp73 might be involved in the pathogenesis of neuroblastoma. Functional assays have demonstrated that tp73 can inhibit cell proliferation and induce apoptosis. The role of this gene in tumorigenesis, however, is still unclear. We analyzed tp73 expression in 95 sporadic neuroblastoma samples by RT-PCR and we detected the tp73 transcript in 46 cases (48.4%), without significant correlation with age, clinical stage or 3-year overall survival. A genetic polymorphism in the 2nd exon of tp73 was utilized to identify the transcribed allele in tumor-cell samples. Expression from only one of the tp73 alleles was found in 13 out of 16 heterozygous tumors, while in 3 samples both alleles were present. Genotype analysis of 73 patients and 150 controls showed a significant deviation (p = 0.0308) from the Hardy-Weinberg equilibrium for a tp73 allele only among neuroblastoma patients. The absence of correlation between tp73 expression and clinical stage, age and survival suggests that this gene does not play an essential function in the clinical course of the disease. However, the distribution of genomic tp73 alleles in patients indicates that a role of this gene in the development of neuroblastoma cannot be completely ruled out. Int. J. Cancer (Pred. Oncol.) 84:365-369, 1999.
Alleles↗
Expression of B-myb in neuroblastoma tumors is a poor prognostic factor independent from MYCN amplification.
The transcription factors of the Myb family are expressed in several tissues and play an important role in cell proliferation, differentiation, and survival In this study, the expression of A-myb, B-myb, and c-myb was investigated in a group of 64 neuroblastomas at different dinical stages by a sensitive reverse transcription-PCR tchnique and correlated with patients' survival. All of the myb genes were frequently expressed in neuroblastoma tumors. Interestingly, the expression of B-myb, which was detected in 33 cases, was associated with an increased risk of death (P = 0.027 in a univariate analysis), whereas there was no correlation with A-myb and c-myb expression. In addition, in a multivariate Cox regression analysis that included myb gene expression, MYCN status, age at diagnosis, and tumor staging, MYCN amplification and B-myb expression were independently associated to an increased risk (P < 0.01 and P = 0.015, respectively). In overall survival curves obtained by stratifying the neuroblastoma cases on the basis of MYCN status and B-myb expression, the group of patients without MYCN amplification and positive for B-myb expression had worse survival probability than that without MYCN amplification and nonexpressing B-myb (P < 0.01). In summary, these findings provide the first demonstration that B-myb expression can be a useful prognostic marker in human neuroblastoma. Moreover, B-myb expression has a prognostic value complementary to MYCN amplification and can identify a group of high-risk patients that would not be predicted on the basis of the MYCN status only.
Cell Cycle Proteins↗
Neuroblastoma in two siblings supports the role of 1p36 deletion in tumor development.
Familial neuroblastoma occurs rarely. We studied a family with three children; one of them has a disseminated (stage 4) and another has a localized (stage 2) neuroblastoma. We observed subtelomeric locus D1Z2 (1p36) deletion in both tumors by using double-color fluorescence in situ hybridization. The MYNC gene was found in single copy in both tumors. Loss of heterozygosity (LOH) and restriction fragment length polymorphism analyses were performed by using DNA from frozen tumor cells and from microdissected tumor areas excised from paraffin-embedded sections. We detected somatic LOH at locus D1S468 (1p36) in a tumor-cell population with a trisomy 1 of the stage-2 patient. Neuroblastoma cells of the stage-4 patient were diploid and showed allelic loss at the following loci: D1S172, D1S80, D1S94, D1S243, D1S468, D1S214, D1S241, and D1S164. Haplotype study showed that the siblings inherited the same paternal 1p36-->pter chromosome region by homologous recombination and that, in the two tumors, arm 1p of different chromosomes of maternal origin was damaged. Our results suggest that the siblings inherited the predisposition to neuroblastoma associated with paternal 1p36 region and that tumors developed as a consequence of somatic loss of the maternal 1p36 allele.
Abdominal Neoplasms↗
131I-metaiodobenzylguanidine (131I-MIBG) therapy for residual neuroblastoma: a mono-institutional experience with 43 patients.
Incomplete response to therapy may compromise the outcome of children with advanced neuroblastoma. In an attempt to improve tumour response we incorporated 131I-metaiodobenzylguanidine (131I-MIBG) in the treatment regimens of selected stage 3 and stage 4 patients. Between 1986 and 1997, 43 neuroblastoma patients older than 1 year at diagnosis, 13 with stage 3 (group A) and 30 with stage 4 disease (group B) who had completed the first-line protocol without achieving complete response entered in this study. 131I-MIBG dose/course ranged from 2.5 to 5.5 Gbq (median, 3.7). The number of courses ranged from 1 to 5 (median 3) depending on the tumour response and toxicity. The most common acute side-effect was thrombocytopenia. Later side-effects included severe interstitial pneumonia in one patient, acute myeloid leukaemia in two, reduced thyroid reserve in 21. Complete response was documented in one stage 4 patient, partial response in 12 (two stage 3, 10 stage 4), mixed or no response in 25 (ten stage 3, 15 stage 4) and disease progression in five (one stage 3, four stage 4) Twenty-four patients (12/13 stage 3, 12/30 stage 4) are alive at 22-153 months (median, 59) from diagnosis. 131I-MIBG therapy may increase the cure rate of stage 3 and improve the response of stage 4 neuroblastoma patients with residual disease after first-line therapy. A larger number of patients should be treated to confirm these results but logistic problems hamper prospective and coordinated studies. Long-term toxicity can be severe.
3-Iodobenzylguanidine↗
Real-time quantitative PCR for the measurement of MYCN amplification in human neuroblastoma with the TaqMan detection system.
BACKGROUND: Neuroblastoma is the most common extracranial malignant solid tumor in children under 5 years and is characterized by a wide clinical and biological heterogeneity, from spontaneously regressive forms to cancers with a rapid and fatal progression. MYCN oncogene amplification is considered the most important prognostic factor to evaluate survival and therapeutic choices in these patients. METHODS: Here we present a new assay for rapid and accurate measurement of MYCN amplification, based on real-time quantitative PCR with the TaqMan(TM) reaction. The degree of MYCN amplification was derived from the ratio of the MYCN oncogene and the single-copy reference gene, beta-actin. The absolute abundance of these two genes in tumor sample DNA was obtained by extrapolation on external calibration curves generated with reference DNA. RESULTS: We found a variable degree of MYCN amplification, from 2 to 29, in 26 of 49 (53%) neuroblastomas. These results were well correlated to those obtained with a competitive PCR assay in the same samples (r = 0. 987). MYCN amplification was associated mainly with advanced cancer stages, and the analysis of overall survival confirmed that the measurement of MYCN amplification is a predictor of patient outcome in neuroblastoma. Patients without MYCN amplification had a cumulative survival significantly higher than patients with low (<9; P = 0.02) and high (>/=9; P = 0.03) oncogene amplification. CONCLUSION: The assay is rapid and reproducible and does not require any post-PCR analytical procedure.
Calibration↗