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Background: Paediatric tumours are often characterised by the presence of recurrent DNA copy number alterations (CNAs). These DNA copy number profiles, obtained from a tissue biopsy, can aid in the correct prognostic classification and therapeutic stratification of several paediatric cancer entities (e.g. MYCN amplification in neuroblastoma) and are part of the routine diagnostic practice. Liquid biopsies (LQBs) offer a potentially safer alternative for such invasive tumour tissue biopsies and can provide deeper insight into tumour heterogeneity., Procedure: The robustness and reliability of LQB CNA analyses was evaluated. We performed retrospective CNA profiling using shallow whole-genome sequencing (sWGS) on paired plasma circulating cell-free DNA (cfDNA) and tissue DNA samples from routinely collected samples from paediatric patients (n = 128) representing different tumour entities, including osteosarcoma, Ewing sarcoma, rhabdomyosarcoma, Wilms tumour, brain tumours and neuroblastoma., Results: Overall, we observed a good concordance between CNAs in tissue DNA and cfDNA. The main cause of CNA discordance was found to be low cfDNA sample quality (i.e. the ratio of cfDNA (<700 bp) and high molecular weight DNA (>700 bp)). Furthermore, CNAs were observed that were present in cfDNA and not in tissue DNA, or vice-versa. In neuroblastoma samples, no false-positives or false-negatives were identified for the detection of the prognostic marker MYCN amplification., Conclusion: In future prospective studies, CNA analysis on LQBs that are of sufficient quality can serve as a complementary assay for CNA analysis on tissue biopsies, as either cfDNA or tissue DNA can contain CNAs that cannot be identified in the other biomaterial., Competing Interests: Conflict of interest statement The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

From June 1989 through to November 1998, 121 children with newly diagnosed T-cell lymphoblastic lymphoma (T-LBL) were included in the EORTC 58881 trial conducted by the Children's Leukaemia Group. The therapy regimen was based on a Berlin-Frankfurt-Munster protocol, for a total duration of 24 months. Cranial irradiation, prophylactic cranial and local, was omitted, even for patients with central nervous involvement at diagnosis. In total, 119 patients were evaluable. The median follow-up was 6.7 years. The overall event-free survival (EFS) rate at 6 years was 77.5% (standard error (SE)=4%). Median time of relapse was 1 year after complete remission (range 0.2-5.9 years). Only two (1.8%) patients had an isolated central nervous system relapse. For patients with complete response (n=16) to the 7-day prephase, the EFS rate at 6 years was 100% versus 14% (P<0.001) for patients with no response (n=7). Overall survival rate at 6 years was 86% (SE=3%). An intensive acute lymphoblastic leukaemia type chemotherapy regimen without irradiation leads to a high cure and survival rate in childhood T-LBL without an increased CNS recurrence. This suggests that prophylactic cranial irradiation can safely be omitted. Response to the prephase appeared to be a strong prognostic factor for EFS.

An important problem in the treatment of children with acute lymphoblastic leukaemia (ALL) is pre-existent or acquired resistance to structurally and functionally unrelated chemotherapeutic compounds. Various cellular mechanisms can give rise to multidrug resistance (MDR). Best studied is the transmembrane protein-mediated efflux of cytotoxic compounds that leads to decreased cellular drug accumulation and toxicity. Several MDR-related efflux pumps have been characterised, including P-glycoprotein (P-gp), multidrug resistance-associated protein 1 (MRP1), breast cancer resistance protein (BCRP) and lung resistance protein (LRP). P-gp expression and/or activity has been associated with unfavourable outcome in paediatric ALL patients, whereas MRP1 and BCRP do not seem to play a major role. LRP might contribute to drug resistance in B-lineage ALL, but larger studies are needed to confirm these results. The present review summarises the current knowledge concerning multidrug resistance-related proteins and focuses on the clinical relevance and prognostic value of these efflux pumps in childhood ALL.

Although neuroblastoma is the most common extracranial solid tumour of childhood, little is known about its aetiology. Together with MYCN amplification and chromosome 17q gain, chromosome 1p deletion is one of the most frequently occurring genetic abnormalities in neuroblastoma. Based upon mapping of deletion breakpoints, putative tumour suppressor gene loci have been assigned to the distal part of the short arm of chromosome 1. Recently, the EXTL1 gene was suggested as a candidate neuroblastoma-suppressor gene and to evaluate this hypothesis, we performed 1p deletion analysis and mutation screening of the EXTL1-coding region on DNA from 22 primary neuroblastomas and 21 neuroblastoma cell lines. Deletions of the chromosome region 1p36.1, including the EXTL1 gene, were detected in several neuroblastoma cell lines and primary tumours. EXTL1 mutation screening resulted in the detection of one unclassified variant (Ser28Cys) but could not provide additional evidence of EXTL1 being involved in the aetiology of neuroblastoma.

Deletions and translocations resulting in loss of distal 1p-material are known to occur frequently in advanced neuroblastomas. Fluorescence in situ hybridisation (FISH) showed that 17q was most frequently involved in chromosome 1p translocations. A review of the literature shows that 10 of 27 cell lines carry 1;17 translocations. Similar translocations were also observed in primary tumours. Together with the occurrence of a constitutional 1;17 translocation in a neuroblastoma patient, these observations suggest a particular role for these chromosome re-arrangements in the development of neuroblastoma. Apart from the loss of distal 1p-material, these translocations invariably lead to extra copies of 17q. This also suggested a possible role for genes on 17q in neuroblastoma tumorigenesis. Further support for this hypothesis comes from the observation that in those cell lines without 1;17 translocations, other chromosome 17q translocations were present. These too lead to extra chromosome 17q material. Molecular analysis of 1;17 translocation breakpoints revealed breakpoint heterogeneity both on 1p and 17q, which suggests the involvement of more than 2 single genes on 1p and 17q. The localisation of the different 1p-breakpoints occurring in 1;17 translocations in neuroblastoma are discussed with respect to the recently identified candidate tumor suppressor regions and genes on 1p. In this study, we focused on the molecular analysis of the 17q breakpoints in 1;17 translocations. Detailed physical mapping of the constitutional 17q breakpoint allowed for the construction of a YAC contig covering the breakpoint. Furthermore, a refined position was determined for a number of 17q breakpoints of 1;17 translocations found in neuroblastoma cell lines. The most distal 17q breakpoint was identified in cell line UHG-NP and mapped telomeric to cosmid cCI17-1049 (17q21). This suggests that genes involved in a dosage-dependent manner in the development of neuroblastoma map in the distal segment 17q22-qter. Future studies aim at the molecular cloning of 1;17 translocation breakpoints and at deciphering the mechanisms leading to 1;17 translocations and possibly to the identification of neuroblastoma genes at or in the vicinity of these breakpoints.

Deletions of the short arm of chromosome 1, extra copies of chromosome 17q and MYCN amplification are the most frequently encountered genetic changes in neuroblastomas. Standard techniques for detection of one or more of these genetic changes are karyotyping, FISH analysis and LOH analysis by Southern blot or PCR. Each of these techniques has its own particular limitations. More recently, comparative genomic hybridisation (CGH) was introduced for detection of genomic imbalances including deletions, duplications and gene amplification. We evaluated the sensitivity and reliability of CGH for detection of the most frequently encountered genetic changes in neuroblastoma. For this purpose a panel of well-characterised neuroblastoma cell lines as well as a series of 11 primary neuroblastomas was analysed. Our results show that CGH is a valuable tool for the genetic characterisation of neuroblastomas, both for the detection of frequently occurring genomic imbalances and for the identification of previously unnoticed genetic changes.

Cytogenetic and molecular studies in neuroblastoma suggest the presence of a tumour suppressor gene at the distal chromosome band 1p36. Previously, we hypothesised that a constitutional translocation involving the region 1p36 [t(1;17)(p36;q12-q21)] in a patient with neuroblastoma predisposed him to tumour development. Here we report the molecular delineation of the translocation breakpoints. Somatic cell hybrids containing the derivative chromosomes were used to determine the position of chromosome 1p and 17q DNA probes respective to the breakpoints using fluorescence in situ hybridisation. The 1p breakpoint was localised between the PND and D1S56 loci. The chromosome 17q breakpoint is flanked by NF1 and SCYA7, as proximal and distal marker, respectively. We redefined the translocation as t(1;17)(p36.31-13;q11.2-q12). The identification of flanking markers of the breakpoints is a prerequisite for breakpoint cloning and identification of a putative neuroblastoma suppressor gene.

Loss of chromosome 1 short arm material, resulting from terminal deletions or unbalanced translocations, is a frequent finding in advanced neuroblastoma. In translocations, often relatively small portions of a second chromosome are translocated to the chromosome 1 short arm. The chromosomal origin of this translocated material could often not be identified using banding analysis only. Recent studies, applying fluorescent in situ hybridisation, showed that in the majority of these translocations, chromosome 17 is involved. In this study, the nonrandom occurrence of unbalanced 1;17 translocations is further supported by their presence in 3/7 neuroblastoma cell lines. Analysis of the 1p breakpoints extends our earlier observation of breakpoint heterogeneity. A similar scattering of 17q breakpoints was observed. The 1p and 17q breakpoints of the constitutional 1;17 translocation did not coincide with any of the 1;17 translocation breakpoints found in neuroblastoma cell lines. Cell lines, not containing 1;17 translocations, contained other chromosome 17 rearrangements. As a result, extra copies of 17q are found in all cell lines, suggesting a role for genes on 17q in neuroblastoma development. The possible significance of 1;17 translocations in neuroblastoma is discussed.

A multicenter EORTC study was conducted in children with acute lymphocytic leukemia to determine whether 5 g/m2 of methotrexate (MTX) (24 h i.v. infusion, four cycles) is an appropriate dosage for obtaining CSF drug concentrations approaching the critical cytotoxic level of 10(-6) M. A total of 193 cycles were analyzed for 58 patients. At the end of the 24 h infusion, the mean MTX serum level was 65.27 +/- 33.11 microM; the mean CSF MTX level was 1.47 +/- 1.1 microM; no significant difference in CSF MTX levels was observed between patients with (n = 20) and those without i.v. Ara-C (n = 38). The mean CSF MTX/serum MTX ratio was 0.029 +/- 0.027. CSF drug concentrations greater than or equal to 10(-6) M were achieved in 81% of the courses. The highest level was 8.4 X 10(-6) M. Only 5% of patients failed to achieve this drug concentration in at least one cycle. No significant correlation was observed between blood and CSF MTX levels. Mean CSF MTX levels were comparable from one cycle to another.