Biomedical subjects

Frank Westermann

Publications and source records attributed to Frank Westermann.

17 recordsLinked to original sources

PubMed2007

Allelic variants of CAMTA1 and FLJ10737 within a commonly deleted region at 1p36 in neuroblastoma.

Deletion of a distal portion of 1p is seen in a wide range of human malignancies, including neuroblastoma. Here, a 1p36.3 commonly deleted region of 216 kb has been defined encompassing two genes, CAMTA1 and FLJ10737. Low expression of CAMTA1 has been recently shown to be an independent predictor of poor outcome in neuroblastoma patients. The present study surveys CAMTA1 and FLJ10737 for genetic alterations by fluorescence-based single strand conformation polymorphism (SSCP) using a panel of DNAs from 88 neuroblastomas, their matching blood samples and 97 unaffected individuals. Nucleotide variants encoding amino acid substitutions were found in both genes. One CAMTA1 variant (T1336I) was not detected in 97 unaffected individuals, another (N1177K) resides in a conserved domain of the CAMTA1 protein and was found hemizygous in six neuroblastomas. We found no evidence for somatic mutations in FLJ10737 or CAMTA1. Further investigations are needed to address the functional impact of the identified variants and their possible significance for neuroblastoma.

Alleles↗

PubMed2006

Targeted MYCN expression affects cytotoxic potential of chemotherapeutic drugs in neuroblastoma cells.

Neuroblastoma (NB) is a solid childhood tumour that exhibits heterogeneous biological and clinical phenotypes. Multiple drug resistance marks a major complication especially in high-risk patients with advanced tumour stages and specific genetic aberrations, such as MYCN amplification and lp deletion. As an approach to further address the mechanisms of chemotherapeutic responsiveness of NB, we used a MYCN-inducible in vitro system and tested the susceptibility of NB cells to anti-tumour drugs currently included in NB treatment protocols dependent on MYCN expression. We observed cytotoxic effects using drug concentrations corresponding to blood plasma levels achieved in NB patients. The most potent drugs were microtubule inhibitors vindesin, paclitaxel and vincristin. Less effective were doxorubicine, arsenic trioxide, cisplatin, etoposide and carboplatin. Exposed to anti-tumour agents, NB cells with induced MYCN expression exhibited higher specific apoptosis than NB cells lacking MYCN expression. Anti-tumour drugs in MYCN-on cells accelerated G1-S phase transition, led to enhanced accumulation of cell populations in G2/M phase, and increased levels of apoptosis. In contrast, MYCN-off cell populations arrested in G1 and, to a smaller extent, in G2/M and exhibited delayed onset of apoptosis. In summary, apoptosis profiles and anti-proliferative potential of chemotherapeutic drugs, used at in vivo tolerable doses, are affected by MYCN overexpression and deregulated cell cycle in SH-EP(MYCN) cells.

Antineoplastic Agents↗

PubMed2006

Classification of neuroblastoma patients by published gene-expression markers reveals a low sensitivity for unfavorable courses of MYCN non-amplified disease.

Currently, Pubmed lists 385 marker genes for neuroblastoma outcome. Using a customized neuroblastoma-microarray, we evaluated the prognostic impact of the gene-expression pattern of 349 of these candidates (90.6%) in 127 neuroblastoma patients with divergent outcome. By significance analysis of microarrays (SAM) and both uncorrected and Bonferroni-corrected ANOVA, 166/349 (47.5%), 218/349 (62.5%) and 128/349 (36.4%) candidates showed significant differential expression between patients with contrasting outcome. By Prediction Analysis for Microarrays (PAM), a 38-gene-classifier was derived from all markers, which classified patients outcome with an overall accuracy of 78.5%. However, patients with unfavorable outcome of MYCN non-amplified disease were largely misclassified (accuracy: 35%), suggesting that these courses are not identified by current marker genes.

Analysis of Variance↗

PubMed2006

Customized oligonucleotide microarray gene expression-based classification of neuroblastoma patients outperforms current clinical risk stratification.

PURPOSE: To develop a gene expression-based classifier for neuroblastoma patients that reliably predicts courses of the disease. PATIENTS AND METHODS: Two hundred fifty-one neuroblastoma specimens were analyzed using a customized oligonucleotide microarray comprising 10,163 probes for transcripts with differential expression in clinical subgroups of the disease. Subsequently, the prediction analysis for microarrays (PAM) was applied to a first set of patients with maximally divergent clinical courses (n = 77). The classification accuracy was estimated by a complete 10-times-repeated 10-fold cross validation, and a 144-gene predictor was constructed from this set. This classifier's predictive power was evaluated in an independent second set (n = 174) by comparing results of the gene expression-based classification with those of risk stratification systems of current trials from Germany, Japan, and the United States. RESULTS: The first set of patients was accurately predicted by PAM (cross-validated accuracy, 99%). Within the second set, the PAM classifier significantly separated cohorts with distinct courses (3-year event-free survival [EFS] 0.86 +/- 0.03 [favorable; n = 115] v 0.52 +/- 0.07 [unfavorable; n = 59] and 3-year overall survival 0.99 +/- 0.01 v 0.84 +/- 0.05; both P < .0001) and separated risk groups of current neuroblastoma trials into subgroups with divergent outcome (NB2004: low-risk 3-year EFS 0.86 +/- 0.04 v 0.25 +/- 0.15, P < .0001; intermediate-risk 1.00 v 0.57 +/- 0.19, P = .018; high-risk 0.81 +/- 0.10 v 0.56 +/- 0.08, P = .06). In a multivariate Cox regression model, the PAM predictor classified patients of the second set more accurately than risk stratification of current trials from Germany, Japan, and the United States (P < .001; hazard ratio, 4.756 [95% CI, 2.544 to 8.893]). CONCLUSION: Integration of gene expression-based class prediction of neuroblastoma patients may improve risk estimation of current neuroblastoma trials.

Biomarkers, Tumor↗

PubMed2006

Marked and independent prognostic significance of the CpG island methylator phenotype in neuroblastomas.

The CpG island methylator phenotype (CIMP) was closely associated with poor overall survival (OS) in Japanese neuroblastoma (NBL) cases in our previous study. Here, in German NBL cases, CIMP(+) cases (n=95) showed markedly poorer OS (hazard ratio (HR)=9.5; P<0.0001) and disease-free survival (DFS) (HR=5.4; P<0.0001) than CIMP(-) cases (n=50). All the 23 cases with N-myc amplification had CIMP. Among the remaining cases without N-myc amplification, CIMP(+) cases (n=27) had a poorer OS (HR=4.5; P=0.02) and DFS (HR=5.2; P<0.0001) than CIMP(-) cases (n=95). In multivariate analysis, CIMP and N-myc amplification had an influence on OS and DFS independent of age and disease stage. CIMP had a stronger influence on DFS than N-myc amplification while N-myc had a stronger influence on OS.

Adolescent↗

PubMed2006

Reduced expression of CAMTA1 correlates with adverse outcome in neuroblastoma patients.

PURPOSE: A distal portion of 1p is frequently deleted in human neuroblastomas, and it is generally assumed that this region harbors at least one gene relevant for neuroblastoma development. A 1p36.3 commonly deleted region, bordered by D1S2731 and D1S214 has been defined. The present study surveys whether expression of genes mapping to this region is associated with tumor behavior. EXPERIMENTAL DESIGN: Candidate genes localized within the deleted region were identified by sequence data analysis. Their expression was assessed in a cohort of 49 primary neuroblastomas using cDNA microarray analysis. Gene expression patterns associated with known prognostic markers and patient outcome were further evaluated by quantitative real-time reverse transcription-PCR in a cohort of 102 neuroblastomas. RESULTS: The commonly deleted region spans 261 kb and encompasses two genes, FLJ10737 and CAMTA1. We found no evidence for an association of FLJ10737 expression with established prognostic variables or outcome. In contrast, low CAMTA1 expression characterized tumors with 1p deletion, MYCN amplification, and advanced tumor stages 3 and 4. Moreover, low CAMTA1 expression was significantly associated with poor outcome (P < 0.001). In multivariate analysis of event-free survival, the prognostic information of low CAMTA1 expression was independent of 1p status, MYCN status, tumor stage, and age of the patient at diagnosis (hazard ratio, 3.52; 95% confidence interval, 1.21-10.28; P = 0.02). CONCLUSIONS: Our data suggest that assessment of CAMTA1 expression may improve the prognostic models for neuroblastoma and that it will be important to define the biological function of CAMTA1 in this disease.

Age Factors↗

PubMed2006

Increased WSB1 copy number correlates with its over-expression which associates with increased survival in neuroblastoma.

Gain of chromosome 17 is the most prevalent genetic abnormality identified in neuroblastoma (NB) and distal 17q gain has prognostic significance in NB. In this report, we have combined array-based comparative genomic hybridization (A-CGH) and gene expression analysis to investigate gene copy number changes and its impact on the gene expression level as well as their association with prognosis genes located on chromosome 17 in NB tumors. We observed differential gains of chromosome 17 between Stages 4- and 4S tumors. We found that WSB1, mapping to 17q11.1, which was frequently gained in 4S- tumors and not changed in 4- tumors, showed strong correlations between expression level and copy number. Furthermore, the increase of WSB1 gene expression is associated with good outcome in patients with NB of all stages. WSB1 also enhances the prognostic prediction when combined with other current prognostic factors in NB. Our results demonstrate that WSB1 copy number correlates with its expression level and that its high expression associates with good prognosis suggesting a possible role of this gene in the biology of favorable outcome NB. This article contains Supplementary Material available at http://www.interscience.wiley.com/jpages/1045-2257/suppmat.

Cell Line, Tumor↗

PubMed2005

Characterization of a complex genomic alteration on chromosome 2p that leads to four alternatively spliced fusion transcripts in the neuroblastoma cell lines IMR-5, IMR-5/75 and IMR-32.

Genetic aberrations in neuroblastoma (NB) have been extensively characterized over the last years. Alterations of the short arm of chromosome 2 (2p) have been of particular interest, since amplification of the MYCN oncogene on 2p24 is associated with an adverse outcome in NB patients. Here, we report on the characterization of a novel genomic rearrangement involving genetic material from 2p13 and 2p24 in NB cell lines that was discovered based on a serial analysis of gene expression (SAGE) profile of the MYCN-amplified NB cell line IMR-5. By analysis of a highly expressed SAGE tag not matching a Unigene cluster we identified four alternatively spliced corresponding transcripts, each of which consisted of the first 14 exons of the anthrax toxin receptor 1 gene (2p13.1) and varying combinations of exons of an unidentified gene located 1.3 Mb telomeric of MYCN (2p24.3) that was termed novel neuroblastoma gene 1. By Southern Blotting, Fluorescent In Situ Hybridization and Long Distance Inverse-PCR we disclosed that these transcripts result from a genomic alteration including material from distinct regions of chromosome 2p and four genomic breakpoints that are joined by short sequences of unknown origin. Furthermore, we show that this rearrangement lies within the homogeneous staining regions (HSR) in IMR-32 cells and is prevalent in both IMR-32 cells and their sub-clones IMR-5 and IMR-5/75, but not in a panel of 70 primary NB tumors. Our work is the first study discovering a fusion transcript based on a SAGE profile and for the first time precisely describes the DNA sequence of amplified breakpoint regions in NB.

Alternative Splicing↗

PubMed2005

Altered expression of cell cycle genes distinguishes aggressive neuroblastoma.

In this study, gene expression profiling was performed on 103 neuroblastoma (NB) tumors, stages 1-4 with and without MYCN amplification, using cDNA microarrays containing 42,578 elements. Using principal component analysis (PCA) to analyse the relationships among these samples, we confirm that the global patterns of gene expression reflect the phenotype of the tumors. To explore the biological processes that may contribute to increasing aggressive phenotype of the tumors, we utilized a statistical approach based on PCA. We identified a specific subset of the cell cycle and/or chromosome segregation genes that distinguish stage 4 NB tumors from all lower stage tumors, including stage 3. Furthermore, the control of the kinetochore assembly emerges from the Gene Ontology analysis as one of the key biological processes associated with an aggressive NB phenotype. Finally, we establish that these genes are further upregulated in the most aggressive MYCN-amplified tumors.

Cell Cycle↗

PubMed2004

Prediction of clinical outcome using gene expression profiling and artificial neural networks for patients with neuroblastoma.

Currently, patients with neuroblastoma are classified into risk groups (e.g., according to the Children's Oncology Group risk-stratification) to guide physicians in the choice of the most appropriate therapy. Despite this careful stratification, the survival rate for patients with high-risk neuroblastoma remains <30%, and it is not possible to predict which of these high-risk patients will survive or succumb to the disease. Therefore, we have performed gene expression profiling using cDNA microarrays containing 42,578 clones and used artificial neural networks to develop an accurate predictor of survival for each individual patient with neuroblastoma. Using principal component analysis we found that neuroblastoma tumors exhibited inherent prognostic specific gene expression profiles. Subsequent artificial neural network-based prognosis prediction using expression levels of all 37,920 good-quality clones achieved 88% accuracy. Moreover, using an artificial neural network-based gene minimization strategy in a separate analysis we identified 19 genes, including 2 prognostic markers reported previously, MYCN and CD44, which correctly predicted outcome for 98% of these patients. In addition, these 19 predictor genes were able to additionally partition Children's Oncology Group-stratified high-risk patients into two subgroups according to their survival status (P = 0.0005). Our findings provide evidence of a gene expression signature that can predict prognosis independent of currently known risk factors and could assist physicians in the individual management of patients with high-risk neuroblastoma.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed2004

cDNA array-CGH profiling identifies genomic alterations specific to stage and MYCN-amplification in neuroblastoma.

BACKGROUND: Recurrent non-random genomic alterations are the hallmarks of cancer and the characterization of these imbalances is critical to our understanding of tumorigenesis and cancer progression. RESULTS: We performed array-comparative genomic hybridization (A-CGH) on cDNA microarrays containing 42,000 elements in neuroblastoma (NB). We found that only two chromosomes (2p and 12q) had gene amplifications and all were in the MYCN amplified samples. There were 6 independent non-contiguous amplicons (10.4-69.4 Mb) on chromosome 2, and the largest contiguous region was 1.7 Mb bounded by NAG and an EST (clone: 757451); the smallest region was 27 Kb including an EST (clone: 241343), NCYM, and MYCN. Using a probabilistic approach to identify single copy number changes, we systemically investigated the genomic alterations occurring in Stage 1 and Stage 4 NBs with and without MYCN amplification (stage 1-, 4-, and 4+). We have not found genomic alterations universally present in all (100%) three subgroups of NBs. However we identified both common and unique patterns of genomic imbalance in NB including gain of 7q32, 17q21, 17q23-24 and loss of 3p21 were common to all three categories. Finally we confirm that the most frequent specific changes in Stage 4+ tumors were the loss of 1p36 with gain of 2p24-25 and they had fewer genomic alterations compared to either stage 1 or 4-, indicating that for this subgroup of poor risk NB requires a smaller number of genomic changes are required to develop the malignant phenotype. CONCLUSIONS: cDNA A-CGH analysis is an efficient method for the detection and characterization of amplicons. Furthermore we were able to detect single copy number changes using our probabilistic approach and identified genomic alterations specific to stage and MYCN amplification.

Cell Line, Tumor↗

PubMed2004

Increased rates of spontaneous sister chromatid exchange in lymphocytes of BRCA2+/- carriers of familial breast cancer clusters.

Heterozygous carriers of germ-line mutations of the BRCA2 breast cancer susceptibility gene are predisposed to breast, ovarian, pancreatic and other cancers. The BRCA2 protein is implicated in the maintenance of chromosome stability through its essential function in double-strand DNA repair and recombination. Our previous studies had revealed multiple intrachromosomal rearrangements, duplications, inversions and deletions on 9p23-24 in lymphocytes and fibroblasts of BRCA2+/- members from independently ascertained familial breast cancer clusters. In pursuit of evaluating if there is a subtle genomic instability in BRCA2+/- individuals, we have determined frequencies of spontaneous sister chromatid exchanges (SCEs) in BRCA2 wild-types and BRCA2 mutation carriers of two familial breast cancer clusters. Here, we demonstrate an average increase of 65% of spontaneous SCEs in BRCA2+/- versus BRCA2+/+ family members. In one cluster, the number of metaphases with multiple SCEs was 5-times higher in BRCA2+/- compared to wild-type members, while in the second cluster BRCA2+/- members had 8.9% of metaphases with multiple SCEs compared to a level below detection in BRCA2 wild types. To investigate the correlation between SCE and genomic instability in 9p, we performed fluorescence detection of SCEs and FISH analysis with 9p probes. The frequency of SCE in 9p of BRCA2 mutation carriers was 3-4 fold (P = 0.005) higher compared to BRCA2 wild-types. Collectively, the increased rates of SCE in BRCA2 heterozygous mutation carriers indicate a BRCA2 haploinsufficiency, which might be an important factor for the accumulation of structural chromosomal alterations with the consequence of damage in as yet unidentified genes.

Breast Neoplasms↗

PubMed2003

Neuroblastoma-derived sulfhydryl oxidase, a new member of the sulfhydryl oxidase/Quiescin6 family, regulates sensitization to interferon gamma-induced cell death in human neuroblastoma cells.

In neuroblastoma cells, apoptotic programs can be activated by cytokines and cytostatic drugs. Apoptotic dysfunction confers resistance against therapeutic drugs and is a major complication for achieving optimal therapy response. Deregulated expression of the MYCN gene is a critical determinant in neuroblastoma progression, and one of the pleiotropic functions of the MYCN protein is cellular sensitization to cytokine-induced and drug-induced apoptosis. By using the functional approach of technical knockout (TKO), we have identified five genes that regulate sensitization for IFN-gamma-induced cell death. Most efficient among them is the newly identified SOXN (neuroblastoma-derived sulfhydryl oxidase), which comprises 12 exons and maps to 9q34.3. SOXN encodes a putative protein of 698 amino acids that contains a signal sequence, a protein-disulfide-isomerase-type thioredoxin and a yeast ERV1 domain and is highly homologous to members of the sulfhydryl oxidase/Quiescin6 family. The SOXN protein is predominantly located in the plasma and in the nuclear membrane. Antisense SOXN confers resistance to IFN-gamma-induced apoptosis. In contrast, ectopic overexpression of sense-SOXN sensitizes the cells to induced cell death. These results identify SOXN as a major player in regulating the sensitization of neuroblastoma cells for IFN-gamma-induced apoptosis.

Amino Acid Sequence↗

PubMed2003

Ataxin-2 promotes apoptosis of human neuroblastoma cells.

Neuroblastoma is a highly heterogeneous tumor of young children. Although many advances have been made towards understanding the molecular mechanisms dictating the phenotypic heterogeneity, the prognosis of children with neuroblastoma, particularly of progressively growing variants, has remained dire. About 10% of neuroblastomas regress spontaneously, probably by apoptosis, while another 20% have amplified the MYCN gene resulting in a poor prognosis. In pursuit of identifying cell death-associated genes in neuroblastoma, we encountered the SCA2 gene, coding for ataxin-2, as an important player. Here, we report that enforced expression of wild-type ataxin-2, but not of mutant ataxin-2, sensitizes neuroblastoma cells for apoptosis. In line with this, higher levels of ataxin-2 were detected in apoptotic cells compared to nonapoptotic cells. Neuroblastoma tumors with amplified MYCN contain significantly less ataxin-2 protein than tumors without amplified MYCN. Collectively, our data suggest that ataxin-2 has an important role in regulating the susceptibility of neuroblastoma cells to apoptotic stimuli in vitro and in vivo.

Amino Acid Chloromethyl Ketones↗

PubMed2003

Neuroblastoma: biology and molecular and chromosomal pathology.

Neuroblastoma is the most frequently occurring solid tumour in children, with an incidence of 1.3 cases per 100000 children aged 0-14 years. Despite many advances during the past three decades, neuroblastoma has remained an enigmatic challenge to clinical and basic scientists. 20 years ago, the MYCN gene was found to be amplified in neuroblastomas, and research since then has focused on the search for other genetic markers. It has emerged that neuroblastoma cells, like cells of many other tumour types, often suffer from extensive, non-random genetic damage at multiple genetic loci. Elucidation of the exact molecular make-up of neuroblastomas will enable researchers to analyse how much specific markers, alone or in combination, can help to stratify disease in prospective studies; at present, stratification is based on age, stage, MYCN, and Shimada pathology. Neuroblastoma may be one of the first examples of the use of genetic tumour markers as a tool for defining tumour behaviour and to aid clinical staging.

Child↗

PubMed2002

Genetic parameters of neuroblastomas.

Neuroblastoma is a malignant childhood tumor of migrating neuroectodermal cells derived from the neural crest and destined for the adrenal medulla and the sympathetic nervous system. The biological behavior of neuroblastomas is extremely variable and in some respects unique. Neuroblastomas tend to regress spontaneously in a portion of infants or to differentiate into a benign ganglioneuroma in some older patients. Unfortunately, in the majority of patients neuroblastoma is metastatic at the time of diagnosis, and it usually undergoes rapid progression with a fatal outcome. The mechanisms leading to this diverse clinical behavior of neuroblastomas are largely unclear. From the analysis of tumors at the cytogenetic and molecular level non-random genetic changes have been identified, including ploidy changes, amplification of the oncogene MYCN, deletions of chromosome 1p, gains of chromosome arm 17q, and deletions of 11q as well as of other genomic regions that allow tumors to be classified into subsets with distinct biological features and clinical behavior. MYCN status is widely accepted for therapy stratification. Additional genetic parameters are currently under investigation to refine risk assessment, but so far the molecular monitoring tools for prediction of therapy response and disease outcome are still incomplete. This should lead to more risk-adapted therapies according to the clinical-genetic parameters by which individual tumors are characterized. This review aims at discussing the role of genomic changes in neuroblastomas of diverse biological and clinical types.

Child↗

PubMed2002

Fluorescence in situ hybridization analyses of chromosome band 1p36 in neuroblastoma detect two classes of alterations.

Chromosomal alterations in 1p36 were investigated in 196 neuroblastoma tumors using fluorescence in situ hybridization. Additionally, by using the same technique, it was determined whether MYCN was amplified in 149 of these. The most frequent finding was a deletion in 1p36, leading to monosomy of this region (29 cases, 15%). Furthermore, we found tumors with at least two intact copies of chromosome 1 and additional 1p36-deleted copies. Altogether, 21 tumors (11%) displayed this imbalance of 1p36. Similar to the cases with deletion, imbalances were predominantly found in stage 4 tumors (81%), and they were significantly associated with an increased patient age (P = 0.01). Nearly all 1p-deleted tumors showed amplification of MYCN (24/27 analyzed samples, 89%), whereas only 8 of 21 (38%) with imbalance did. Eight cases with imbalance were investigated for loss of heterozygosity (LOH) using microsatellite markers in 1p35-36. Only 4 displayed 1p36 LOH, whereas the remaining 4 were heterozygous. Both patients with deletion of 1p and with imbalance had a poor outcome [3-year rate of event-free-survival (EFS): 33 +/- 15% and 41 +/- 15%], which was significantly worse compared to the outcome of patients without 1p alterations (3-year EFS: 70 +/- 5%; P = 0.01 and P = 0.0059). We conclude that besides monosomic short arm deletions, imbalance of 1p36 is a strong marker of a poor prognosis in neuroblastoma and not necessarily associated with MYCN amplification and LOH.

Adult↗