Freimut H. Schilling

Myeloablative megatherapy with autologous stem-cell rescue versus oral maintenance chemotherapy as consolidation treatment in patients with high-risk neuroblastoma: a randomised controlled trial

BACKGROUND Myeloablative megatherapy is commonly used to improve the poor outlook of children with high-risk neuroblastoma, yet its role is poorly defined. We aimed to assess whether megatherapy with autologous stem-cell transplantation could increase event-free survival and overall survival compared with maintenance chemotherapy. METHODS 295 patients with high-risk neuroblastoma (ie, patients with stage 4 disease aged older than 1 year or those with MYCN-amplified tumours and stage 1, 2, 3, or 4S disease or stage 4 disease and <1 year old) were randomly assigned to myeloablative megatherapy (melphalan, etoposide, and carboplatin) with autologous stem-cell transplantation (n=149) or to oral maintenance chemotherapy with cyclophosphamide (n=146). The primary endpoint was event-free survival. Secondary endpoints were overall survival and the number of treatment-related deaths. Analyses were done by intent to treat, as treated, and treated as randomised. FINDINGS Intention-to-treat analysis showed that patients allocated megatherapy had increased 3-year event-free survival compared with those allocated maintenance therapy (47% [95% CI 38-55] vs 31% [95% CI 23-39]; hazard ratio 1.404 [95% CI 1.048-1.881], p=0.0221), but did not have significantly increased 3-year overall survival (62% [95% CI 54-70] vs 53% [95% CI 45-62]; 1.329 [0.958-1.843], p=0.0875). Improved 3-year event-free survival and 3-year overall survival were also recorded for patients given megatherapy in the as-treated group (n=212) and in the treated-as-randomised group (n=145). Two patients died from therapy-related complications during induction treatment. No patients given maintenance therapy died from acute treatment-related toxic effects. Five patients given megatherapy died from acute complications related to megatherapy. INTERPRETATION Myeloablative chemotherapy with autologous stem-cell transplantation improves the outcome for children with high-risk neuroblastoma despite the raised risk of treatment-associated death.

Localized infant neuroblastomas often show spontaneous regression: results of the prospective trials NB95-S and NB97.

PURPOSE The excellent prognosis of localized neuroblastoma in infants, the overdiagnosis observed in neuroblastoma screening studies, and several case reports of regression of localized neuroblastoma prompted us to initiate a prospective cooperative trial on observation of localized neuroblastoma without cytotoxic treatment. PATIENTS AND METHODS For infants with localized neuroblastoma without MYCN amplification, chemotherapy was scheduled only in cases with threatening symptoms; otherwise, the tumor was either resected or observed by ultrasound and magnetic resonance imaging (MRI). RESULTS Of 340 eligible participants, 190 underwent resection, 57 were treated with chemotherapy, and 93 were observed with gross residual tumor. Of those 93 patients with unresected tumors, spontaneous regression was seen in 44, local progression in 28, progression to stage 4S in seven, and progression to stage 4 in four. Time to regression was quite variable, with first signs of regression noted 1 to 18 months after diagnosis and in 15 of 44 patients even after the first year of life. So far, complete regression was observed in 17 of 44 patients 4 to 20 months after diagnosis. Known clinical risk factors were not able to differentiate between patients with regression and regional or metastatic progression. Overall survival (OS; 3-year OS, 0.99 +/- 0.01) and metastases-free survival (rate at 3 years, 0.94 +/- 0.03) for patients with unresected tumors was excellent and was not different from patients treated with surgery or chemotherapy. CONCLUSION Spontaneous regression is regularly seen in infants with localized neuroblastoma and is not limited to the first year of life. A wait-and-see strategy is justified in those patients.

Long-term results and risk profiles of patients in five consecutive trials (1979–1997) with stage 4 neuroblastoma over 1 year of age

During the last two decades new diagnostic and therapeutic tools have been utilized to improve the poor survival chances of children with stage 4 neuroblastoma. This study reviews the risk profiles and the long-term outcome of patients from five consecutive German neuroblastoma trials. A total of 96% of all German patients registered at the German childhood cancer registry with neuroblastoma stage 4 over 1 year of age at diagnosis entered one of the trials during 1979-2001. Eight hundred and twenty-eight consecutive children were analyzed retrospectively. In spite of having significantly improved diagnostic tools like bone marrow superstaging and mIBG scintigraphy the stage 4 incidence did not increase after reaching completeness of the registry (5.4 cases/100,000 children at 1-14 years of age; P=0.52). The distribution of the primary tumors and of metastases was constant over the periods. The amount of bone marrow infiltration did not change with time. The risk factors lactate dehydrogenase, ferritin and MYCN, and the clinical risk groups 4A, 4B, 4C also remained constant over the trials with a few exceptions for NB97. The 5-year event free survival increased from 0.01+/-0.01 (NB79) to 0.14+/-0.03 (NB85), 0.16+/-0.04 (NB82), 0.27+/-0.02 (NB90), and 0.33+/-0.04 (NB97). The overall survival rates improved similarly from 0.04 (NB79) to 0.44 (NB97). In conclusion, the improved survival was associated with better treatment and not caused by lower risk profiles in stage 4 neuroblastoma patients.

Expression of mRNA for the neurotrophin receptor trkC in neuroblastomas with favourable tumour stage and good prognosis

Childhood neuroblastoma tumours of the sympathetic nervous system show a remarkable clinical heterogeneity ranging from spontaneous regression to unfavourable outcome despite intensive therapy. Favourable neuroblastomas often express high levels of trkA mRNA, encoding the tyrosine kinase receptor for nerve growth factor. We have investigated mRNA expression for the neurotrophin receptor trkC in 23 primary neuroblastomas using a sensitive RNAase protection assay. TrkC expression was detected in 19 of these tumours at highly variable levels with a 300-fold difference between the highest and lowest values. Significantly higher levels of trkC mRNA were found in tumours from patients with favourable features such as low age (P < 0.012), favourable tumour stage (P < 0.012) and favourable prognosis (P < 0.05). Children with intermediate or high trkC mRNA expression had better prognosis compared with those with low or undetectable levels (83.3% vs 20%, P = 0.005). Further characterisation of trkC mRNA expression by reverse transcriptase-polymerase chain reaction (RT-PCR) showed that mRNA encoding the full-length cytoplasmic tyrosine kinase domain of the receptor was only expressed in a subset of favourable tumours. These data show that favourable neuroblastomas may express the full trkC receptor while advanced tumours, in particular MYCN-amplified neuroblastoma, seem to either express no trkC or truncated trkC receptors of as yet unknown biological function. These data are suggestive of a role for trkC and its preferred ligand neutotrophin-3, NT-3, in neuroblastoma differentiation and/or regression.

Gangliocytes in neuroblastic tumors express alarin, a novel peptide derived by differential splicing of the galanin-like peptide gene

In neuroblastic tumors a relationship of differentiation of the tumor to galanin receptor expression and antip roliferative and apoptotic effects upon activation of galanin receptors in neuroblastoma cells was reported. To elucidate the expression of other components of the galanin peptide family in neuroblastic tumors, RT-PCR analysis of a variety of human neuroblastic tumor tissues was performed. Ganglioneuroma tissues revealed the presence of a splice variant of the galanin-like peptide (GALP) mRNA, which results in exclusion of exon 3 and a frame shift after the signal peptide sequence of GALP. This generates a peptide of 25 amino acids, which we have termed alarin because of the N-terminal alanine and the C-terminal serine. The novel neuropeptide alarin does not reveal significant homology to other peptides. Immunohistochemistry with antibodies directed against synthetic alarin peptide detected specific cytoplasmic granular staining in ganglia of human ganglioneuroma and ganglioneuroblastoma, as well as differentiated tumor cells of neuroblastoma tissues. Undifferentiated neuroblasts of these tumor tissues did not show alarin-like immunoreactivity and alarin-specific mRNA. Our findings indicate that alarin expression is a feature of ganglionic differentiation in neuroblastic tumor tissues.

Low aerobic mitochondrial energy metabolism in poorly- or undifferentiated neuroblastoma

BackgroundSuccinate dehydrogenase (SDH) has been associated with carcinogenesis in pheochromocytoma and paraganglioma. In the present study we investigated components of the oxidative phosphorylation system in human neuroblastoma tissue samples.MethodsSpectrophotometric measurements, immunohistochemical analysis and Western blot analysis were used to characterize the aerobic mitochondrial energy metabolism in neuroblastomas (NB).ResultsCompared to mitochondrial citrate synthase, SDH activity was severely reduced in NB (n = 14) versus kidney tissue. However no pathogenic mutations could be identified in any of the four subunits of SDH. Furthermore, no genetic alterations could be identified in the two novel SDH assembly factors SDHAF1 and SDH5. Alterations in genes encoding nfs-1, frataxin and isd-11 that could lead to a diminished SDH activity have not been detected in NB.ConclusionBecause downregulation of other complexes of the oxidative phosphorylation system was also observed, a more generalized reduction of mitochondrial respiration seems to be present in neuroblastoma in contrast to the single enzyme defect found in hereditary pheochromocytomas.

Galanin and galanin receptors in human cancers

The increasing interest in peptides and peptide receptors in cancer is based on the possibility of receptor targeting, because peptide receptors are often expressed in different human tumors. The neuropeptide galanin has also been suggested to be involved in the development of neuroendocrine tumors based on the development of estrogen-induced tumors in estrogen-sensitive rats. This study summarizes our current knowledge on the expression of galanin peptide and galanin receptors in different human neuroendocrine tumors. The expression of both, peptide and corresponding receptor, seems to be a common feature of human gliomas, pheochromocytomas, pituitary and neuroblastic tumors. The co-expression of galanin and its receptors supports a role for galanin in tumor cell pathology via autocrine/paracrine mechanisms.

Malignancy‐associated haemophagocytic lymphohistiocytosis in children and adolescents

Haemophagocytic lymphohistiocytosis (HLH) in the context of malignancy is mainly considered a challenge of adult haematology. While this association is also observed in children, little is known regarding inciting factors, appropriate treatment and prognosis. We retrospectively analysed 29 paediatric and adolescent patients for presenting features, type of neoplasm or preceding chemotherapy, treatment and outcome. Haemophagocytic lymphohistiocytosis was considered triggered by the malignancy (M‐HLH) in 21 patients, most of whom had T‐ (n = 12) or B‐cell neoplasms (n = 7), with Epstein–Barr virus as a co‐trigger in five patients. In eight patients, HLH occurred during chemotherapy (Ch‐HLH) for malignancy, mainly acute leukaemias (n = 7); an infectious trigger was found in seven. In M‐ and Ch‐HLH, median overall survival was 1·2 and 0·9 years, and the 6 month survival rates were 67% and 63%, respectively. Seven of 11 deceased M‐HLH patients exhibited active malignancy and HLH at the time of death, while only two out of five deceased Ch‐HLH patients had evidence of active HLH. To overcome HLH, malignancy‐ and HLH‐directed treatments were administered in the M‐HLH cohort; however, it was not possible to determine superiority of one approach over the other. For Ch‐HLH, treatment ranged from postponement of chemotherapy to the use of etoposide‐containing regimens.

Combined111In-pentetreotide scintigraphy and123I-mIBG scintigraphy in neuroblastoma provides prognostic information

BACKGROUND High-affinity somatostatin receptors (SRs) have been characterized in neuroblastomas and may be used as target structures for in vivo detection of SR. PROCEDURE Eighty-eight children with histologically proven neuroblastoma were investigated at diagnosis or relapse by (123)I-mIBG and (111)In-pentetreotide scintigraphy. All tumors were investigated for MYCN copy number, chromosome 1p36 status, and 68/88 also for DNA content, followed for a median follow-up of 35 months (range 1-88 months). RESULTS SR expression was detected in 56/88 tumors and (123)I-mIBG showed positivity in 83/88. (111)In-pentetreotide was less sensitive in detecting tumor tissue than was (123)I-mIBG (64% vs. 94%, P = 0.005). Survival (SUR) and event-free survival probability (EFS) according to Kaplan-Meier was significantly better for children with positive SR scintigraphy than for the children with a negative SR scan (SUR: 90% vs. 48% at 4 years log rank P < 0.003, EFS: 83% vs. 39% at 4 years, log rank P < 0.0002). CONCLUSIONS (123)I-mIBG scintigraphy remains the best scintigraphic method for detecting neuroblastoma tumor tissue, whereas additional SR scintigraphy is able to provide significant prognostic information with a minimum of invasiveness.

Heterogeneity of the MYCN Oncogene in Neuroblastoma

Purpose:MYCN amplification is an important therapy-stratifying marker in neuroblastoma. Fluorescence in situ hybridization with signal detection on the single-cell level allows a critical judgement of MYCN intratumoral heterogeneity. Experimental Design: The MYCN status was investigated by fluorescence in situ hybridization at diagnosis and relapse. Heterogeneity was defined as the simultaneous presence of amplified cells (≥5 cells per slide) and nonamplified cells within one tumor or sequential change of the amplification status during the course of the disease. Likewise, heterogeneity can be detected between primary tumor and metastasis. Results: From 1,341 patients analyzed, 1,071 showed no amplification, 250 showed homogeneous amplification, and 20 patients showed MYCN heterogeneity. Of the patients with heterogeneity, 12 of 20 had clusters of MYCN amplifications, 3 of 20 had amplified single cells, 3 of 20 showed MYCN amplifications in the bone marrow but not in the primary tumor, and 2 of 20 acquired MYCN amplification during the course of the disease. All stage 4 patients were treated according to high-risk protocols; 7 of 8 later progressed. Four patients with localized disease were treated according to high-risk protocol because of MYCN-amplified clusters; 1 of 4 later progressed. One patient treated with mild chemotherapy experienced progression. Seven patients with localized/4S disease underwent no chemotherapy: 4 of 5 patients with MYCN heterogeneity at diagnosis remained disease-free, and 1 of 5 experienced local progression. Two patients had normal MYCN status at diagnosis but acquired MYCN amplification during the course of the disease. Conclusion:MYCN heterogeneity is rare. Our results suggest that small amounts of MYCN-amplified cells are not correlated to adverse outcomes. More patients with heterogeneity are warranted to clarify the role of MYCN heterogeneity for risk classification.