J Vandesompele

Emerging patterns of cryptic chromosomal imbalance in patients with idiopathic mental retardation and multiple congenital anomalies: a new series of 140 patients and review of published reports

Background: Chromosomal abnormalities are a major cause of mental retardation and multiple congenital anomalies (MCA/MR). Screening for these chromosomal imbalances has mainly been done by standard karyotyping. Previous array CGH studies on selected patients with chromosomal phenotypes and normal karyotypes suggested an incidence of 10–15% of previously unnoticed de novo chromosomal imbalances. Objective: To report array CGH screening of a series of 140 patients (the largest published so far) with idiopathic MCA/MR but normal karyotype. Results: Submicroscopic chromosomal imbalances were detected in 28 of the 140 patients (20%) and included 18 deletions, seven duplications, and three unbalanced translocations. Seventeen of 24 imbalances were confirmed de novo and 19 were assumed to be causal. Excluding subtelomeric imbalances, our study identified 11 clinically relevant interstitial submicroscopic imbalances (8%). Taking this and previously reported studies into consideration, array CGH screening with a resolution of at least 1 Mb has been undertaken on 432 patients with MCA/MR. Most imbalances are non-recurrent and spread across the genome. In at least 8.8% of these patients (38 of 432) de novo intrachromosomal alterations have been identified. Conclusions: Array CGH should be considered an essential aspect of the genetic analysis of patients with MCA/MR. In addition, in the present study three patients were mosaic for a structural chromosome rearrangement. One of these patients had monosomy 7 in as few as 8% of the cells, showing that array CGH allows detection of low grade mosaicisims.

A new recurrent inversion, inv(7)(p15q34), leads to transcriptional activation of HOXA10 and HOXA11 in a subset of T-cell acute lymphoblastic leukemias

Chromosomal translocations with breakpoints in T-cell receptor (TCR) genes are recurrent in T-cell malignancies. These translocations involve the TCRαδ gene (14q11), the TCRβ gene (7q34) and to a lesser extent the TCRγ gene at chromosomal band 7p14 and juxtapose T-cell oncogenes next to TCR regulatory sequences leading to deregulated expression of those oncogenes. Here, we describe a new recurrent chromosomal inversion of chromosome 7, inv(7)(p15q34), in a subset of patients with T-cell acute lymphoblastic leukemia characterized by CD2 negative and CD4 positive, CD8 negative blasts. This rearrangement juxtaposes the distal part of the HOXA gene cluster on 7p15 to the TCRβ locus on 7q34. Real time quantitative PCR analysis for all HOXA genes revealed high levels of HOXA10 and HOXA11 expression in all inv(7) positive cases. This is the first report of a recurrent chromosome rearrangement targeting the HOXA gene cluster in T-cell malignancies resulting in deregulated HOXA gene expression (particularly HOXA10 and HOXA11) and is in keeping with a previous report suggesting HOXA deregulation in MLL-rearranged T- and B cell lymphoblastic leukemia as the key factor in leukaemic transformation. Finally, our observation also supports the previous suggested role of HOXA10 and HOXA11 in normal thymocyte development.

Comparative genomic hybridization (CGH) analysis of stage 4 neuroblastoma reveals high frequency of 11q deletion in tumors lacking MYCN amplification

We have studied the occurrence and association of 11q deletions with other chromosomal imbalances in Stage 4 neuroblastomas. To this purpose we have performed comparative genomic hybridization (CGH) analysis on 50 Stage 4 neuroblastomas and these data were analyzed together with those from 33 previously published cases. We observed a high incidence of 11q deletion in Stage 4 neuroblastoma without MYCN amplification (59%) whereas 11q loss was only observed in 15% of neuroblastomas with MYCN‐amplification (p = 0.0002) or 11% of cases with 1p deletion detected by CGH (p = 0.0001). In addition, 11q loss showed significant positive correlation with 3p loss (p = 0.0002). Event‐free survival was poor and not significantly different for patients with or without 11q deletion. Our study provides further evidence that Stage 4 neuroblastomas with 11q deletions represent a distinct genetic subgroup that typically shows no MYCN‐amplification nor 1p deletion. Moreover, it shows that neuroblastomas with 11q deletion also often present 3p deletion. This genetic subgroup shows a similar poor prognosis as MYCN amplified 4 neuroblastomas.

Molecular analysis of 1p36 breakpoints in two Merkel cell carcinomas

Merkel cell carcinoma (MCC) is a rare aggressive neuroendocrine tumor of the skin. Only little information is available on the genetic alterations occurring in this tumor. Cytogenetic studies thus far have not shown recurrent chromosomal changes, although various structural chromosome 1 rearrangements, including deletions, often leading to loss of distal 1p material appear to be frequent. We report on fluorescence in situ hybridization and loss of heterozygosity analyses of an MCC tumor and MCC cell line UISO. The present study has shown that two distinct regions in the most distal band 1p36 on the short arm of chromosome 1 can be implicated in MCC. One region at 1p36.3 was delineated by a distal deletion in the MCC tumor as a result of an unbalanced translocation, resulting in loss of all markers distal to ENO1. This region was previously shown to be deleted in different tumor types including neuroblastoma. In cell line UISO an insertion in 1p36.2 was identified. The insertion breakpoint indicates a second, more proximal, region on 1p involved in MCC. The insertion breakpoint was mapped within a cluster of repetitive tRNA and snRNA genes and thus could coincide with the constitutional 1p36 breakpoint previously reported in a patient with neuroblastoma. Genes Chromosomes Cancer 23:67–71, 1998.

Escape from p53-mediated tumor surveillance in neuroblastoma: switching off the p14 ARF -MDM2-p53 axis

A primary failsafe program against unrestrained proliferation and oncogenesis is provided by the p53 tumor suppressor protein, inactivation of which is considered as a hallmark of cancer. Intriguingly, mutations of the TP53 gene are rarely encountered in neuroblastoma tumors, suggesting that alternative p53-inactivating lesions account for escape from p53 control in this childhood malignancy. Several recent studies have shed light on the mechanisms by which neuroblastoma cells circumvent the p53-driven antitumor barrier. We review here these mechanisms for evasion of p53-mediated growth control and conclude that deregulation of the p14ARF-MDM2-p53 axis seems to be the principal mode of p53 inactivation in neuroblastoma, opening new perspectives for targeted therapeutic intervention.

Genome wide measurement of DNA copy number changes in neuroblastoma: dissecting amplicons and mapping losses, gains and breakpoints

In the past few years high throughput methods for assessment of DNA copy number alterations have witnessed rapid progress. Both ‘in house’ developed BAC, cDNA, oligonucleotide and commercial arrays are now available and widely applied in the study of the human genome, particularly in the context of disease. Cancer cells are known to exhibit DNA losses, gains and amplifications affecting tumor suppressor genes and proto-oncogenes. Moreover, these patterns of genomic imbalances may be associated with particular tumor types or subtypes and may have prognostic value. Here we summarize recent array CGH findings in neuroblastoma, a pediatric tumor of the sympathetic nervous system. A total of 176 primary tumors and 53 cell lines have been analyzed on different platforms. Through these studies the genomic content and boundaries of deletions, gains and amplifications were characterized with unprecedented accuracy. Furthermore, in conjunction with cytogenetic findings, array CGH allows the mapping of breakpoints of unbalanced translocations at a very high resolution.

EVI1-mediated down regulation of MIR449A is essential for the survival of EVI1 positive leukaemic cells

Chromosomal rearrangements involving the MECOM (MDS1 and EVI1 complex) locus are recurrent genetic events in myeloid leukaemia and are associated with poor prognosis. In this study, we assessed the role of MECOM locus protein EVI1 in the transcriptional regulation of microRNAs (miRNAs) involved in the leukaemic phenotype. For this, we profiled expression of 366 miRNAs in 38 MECOM‐rearranged patient samples, normal bone marrow controls and MECOM (EVI1) knock down/re‐expression models. Cross‐comparison of these miRNA expression profiling data showed that MECOM rearranged leukaemias are characterized by down regulation of MIR449A. Reconstitution of MIR449A expression in MECOM‐rearranged cell line models induced apoptosis resulting in a strong decrease in cell viability. These effects might be mediated in part by MIR449A regulation of NOTCH1 and BCL2, which are shown here to be bona fide MIR449A targets. Finally, we confirmed that MIR449A repression is mediated through direct promoter occupation of the EVI1 transcriptional repressor. In conclusion, this study reveals MIR449A as a crucial direct target of the MECOM locus protein EVI1 involved in the pathogenesis of MECOM‐rearranged leukaemias and unravels NOTCH1 and BCL2 as important novel targets of MIR449A. This EVI1‐MIR449A‐NOTCH1/BCL2 regulatory axis might open new possibilities for the development of therapeutic strategies in this poor prognostic leukaemia subgroup.

Ectopic microRNA-150-5p transcription sensitizes glucocorticoid therapy response in MM1S multiple myeloma cells but fails to overcome hormone therapy resistance in MM1R cells.

Glucocorticoids (GCs) selectively trigger cell death in the multiple myeloma cell line MM1S which express NR3C1/Glucocorticoid Receptor (GR) protein, but fail to kill MM1R cells which lack GR protein. Given recent demonstrations of altered microRNA profiles in a diverse range of haematological malignancies and drug resistance, we characterized GC inducible mRNA and microRNA transcription profiles in GC sensitive MM1S as compared to GC resistant MM1R cells. Transcriptome analysis revealed that GCs regulate expression of multiple genes involved in cell cycle control, cell organization, cell death and immunological disease in MM1S cells, which remain unaffected in MM1R cells. With respect to microRNAs, mir-150-5p was identified as the most time persistent GC regulated microRNA, out of 5 QPCR validated microRNAs (mir-26b, mir-125a-5p, mir-146-5p, mir-150-5p, and mir-184), which are GC inducible in MM1S but not in MM1R cells. Functional studies further revealed that ectopic transfection of a synthetic mir-150-5p mimics GR dependent gene expression changes involved in cell death and cell proliferation pathways. Remarkably, despite the gene expression changes observed, overexpression of mir-150-5p in absence of GCs did not trigger significant cytotoxicity in MM1S or MM1R cells. This suggests the requirement of additional steps in GC induced cell death, which can not be mimicked by mir-150-5p overexpression alone. Interestingly, a combination of mir-150-5p transfection with low doses GC in MM1S cells was found to sensitize therapy response, whereas opposite effects could be observed with a mir-150-5p specific antagomir. Although mir-150-5p overexpression did not substantially change GR expression levels, it was found that mir-150-5p evokes GR specific effects through indirect mRNA regulation of GR interacting transcription factors and hormone receptors, GR chaperones, as well as various effectors of unfolded protein stress and chemokine signalling. Altogether GC-inducible mir-150-5p adds another level of regulation to GC specific therapeutic responses in multiple myeloma.