http://repub.eur.nl/res/aut/5238/ List of Publications en http://repub.eur.nl/static-eur/img/logo.png http://repub.eur.nl http://repub.eur.nl/res/pub/34670/ 2011-06-01T00:00:00Z http://repub.eur.nl/res/pub/28487/ 2010-11-18T00:00:00Z Background: Tissue MicroArray technology aims to perform immunohistochemical staining on hundreds of different tissue samples simultaneously. It allows faster analysis, considerably reducing costs incurred in staining. A time consuming phase of the methodology is the selection of tissue areas within paraffin blocks: no utilities have been developed for the identification of areas to be punched from the donor block and assembled in the recipient block.Results: The presented work supports, in the specific case of a primary subtype of breast cancer (tubular breast cancer), the semi-automatic discrimination and localization between normal and pathological regions within the tissues. The diagnosis is performed by analysing specific morphological features of the sample such as the absence of a double layer of cells around the lumen and the decay of a regular glands-and-lobules structure. These features are analysed using an algorithm which performs the extraction of morphological parameters from images and compares them to experimentally validated threshold values. Results are satisfactory since in most of the cases the automatic diagnosis matches the response of the pathologists. In particular, on a total of 1296 sub-images showing normal and pathological areas of breast specimens, algorithm accuracy, sensitivity and specificity are respectively 89%, 84% and 94%.Conclusions: The proposed work is a first attempt to demonstrate that automation in the Tissue MicroArray field is feasible and it can represent an important tool for scientists to cope with this high-throughput technique. http://repub.eur.nl/res/pub/32777/ 2010-09-01T00:00:00Z http://repub.eur.nl/res/pub/36551/ 2007-12-01T00:00:00Z In recent years, biobanks have evolved into professional infrastructures that acquire, validate, process, store, manage and distribute biological material of human origin to public or private end-users/researchers. This article (a) highlights the importance of quality assurance for both the biobank basic processes and sample annotation in order to ensure reliable results of research based on these samples, (b) suggests that certification according to international standards can contribute to the organization of the biobanking processes while accreditation can contribute to the organization of sample characterization/validation, and (c) provides a compilation of all existing guidelines against an International Organization for Standardization (ISO) format. Copyright http://repub.eur.nl/res/pub/36495/ 2007-03-01T00:00:00Z Studies using fresh-frozen tissue samples originating from different centres, as is often the case in EORTC related translational research, can show conflicting research results due to heterogeneity in the quality of samples and associated data from each centre. The development of infrastructure for the European Human Frozen Tumour Tissue Bank (TuBaFrost) anticipated this problem and Standard Operating Procedures (SOPs) have been developed to ensure samples collected are of consistent high quality and variation in research results is minimised. The SOPs drew on the best practice standard workflows and operating procedures employed by members of the TuBaFrost Consortium and key tissue bank initiatives worldwide. It was essential to provide workable solutions that reflect the variety in infrastructure and resources at the potential collecting centres and also the fact that it is not necessary to standardise every step of the collection and storage process in order to collect high quality tissue. Hence, the TuBaFrost SOPs detail the compulsory measures that must be implemented in order to become a TuBaFrost collecting centre and also make advisory recommendations regarding the less critical factors. Accordingly, the TuBaFrost SOPs are very flexible and to illustrate this the complete SOP for collecting, freezing and storing tissue at the Erasmus MC Tissue Bank is included. These TuBaFrost SOPs could equally be applicable to centres collecting samples for EORTC related translational research studies in order to standardise sample quality and produce reliable and reproducible research results. http://repub.eur.nl/res/pub/13800/ 2005-05-01T00:00:00Z Fusion of the MN1 gene to TEL (ETV6) results in myeloid leukemia. The fusion protein combines the transcription activating domain of MN1 and the DNA binding domain of TEL and is thought to act as a deranged transcription factor. In addition, disruption of the large first exon of the MN1 gene is thought to inactivate MN1 function in a meningioma. To further investigate the role of MN1 in cancer, we generated Mn1 knockout mice. Mn1(+/-) animals were followed for 30 months, but they had no higher incidence of tumor formation than wild-type littermates. Mn1 null mice, however, were found to die at birth or shortly thereafter as the result of a cleft palate. Investigation of newborn or embryonic day 15.5 (E15.5) to E17.5 null mice revealed that the development of several bones in the skull was abnormal. The affected bones are almost exclusively formed by intramembranous ossification. They are either completely agenic at birth (alisphenoid and squamosal bones and vomer), hypoplastic, deformed (basisphenoid, pterygoid, and presphenoid), or substantially thinner (frontal, parietal, and interparietal bones). In heterozygous mice hypoplastic membranous bones and incomplete penetrance of the cleft palate were observed. We conclude that Mn1 is an important factor in development of membranous bones. http://repub.eur.nl/res/pub/9871/ 2002-01-01T00:00:00Z Malignant transformation of Barrett's esophagus is characterized by three distinct premalignant stages: intestinal metaplasia (MET), low- (LGD), and high-grade dysplasia (HGD). We reported recently an increase in the frequency of loss of 7q33-q35 between LGD and HGD as determined by comparative genomic hybridization (P. H. J. Riegman et al., Cancer Res., 61: 3164-3170, 2001). Now the 7q32.3-q36.1 region was additionally characterized by allelotype analysis with 11 polymorphic markers in 15 METs, 20 LGDs, 20 HGDs, and 20 Barrett's adenocarcinomas from different patients. Low percentages of imbalance were determined in METs and LGDs, 7% and 10%, respectively, whereas HGDs and Barrett's adenocarcinomas revealed high percentages of loss, 75% and 65%, respectively. This difference in frequency between LGDs and HGDs appeared highly significant: P = 0.00007. The majority of imbalances were found at D7S2439 and D7S483, located on 7q36.1. These data suggest that markers from this area can be used as a diagnostic tool in Barrett's esophagus, i.e., to distinguish between watchful waiting and active treatment. http://repub.eur.nl/res/pub/9624/ 2001-01-01T00:00:00Z The incidence of adenocarcinoma in Barrett's esophagus has been increasing rapidly over the past decades. Neoplastic progression is characterized by three well-defined premalignant stages: metaplasia, low-grade dysplasia, and high-grade dysplasia. A genome-wide overview, based on comparative genomic hybridization, was performed, evaluating 30 Barrett's adenocarcinomas and 25 adjacent precursors, i.e., 6 metaplasias, 9 low-grade dysplasias, and 10 high-grade dysplasias. The frequency of losses and gains significantly increased in the subsequent stages of malignant transformation. Losses of 5q21-q23, 9p21, 17p12-13.1, 18q21, and Y were revealed in low-grade dysplasias. This was followed by loss of 7q33-q35 and gains of 7p12-p15, 7q21-q22, and 17q21 in high-grade dysplasias along with high-level amplification (HLA) of 7q21 and 17q21. In the invasive cancers, additional losses of 3p14-p21, 4p, 4q, 8p21, 13q14-q31, 14q24.3-q31, 16q21-q22, and 22q as well as gains of 3q25-q27, 8q23-24.1, 12p11.2-12, 15q22-q24, and 20q11.2-q13.1 were distinguished along with HLAs of 8p12-p22 and 20q11.2-q13.1. Approximately one-third of the alterations in the dysplasias were also found in the adjacent adenocarcinomas, illustrating that multiple clonal lineages can be present in Barrett's esophagus. Novel findings include loss on 7q, gain on 12p, and the observation of several HLAs in high-grade dysplasias. Furthermore, loss of 7q33-q35 was found to represent a significant distinction between low-grade and high-grade dysplasia (P = 0.01), whereas loss of 16q21-q22 and gain of 20q11.2-q13.1 were disclosed to significantly discriminate between high-grade dysplasia and adenocarcinoma (P = 0.02 and P = 0.03, respectively). This inventory of genetic aberrations increases our understanding of malignant transformation in Barrett's esophagus and might provide useful biomarkers for disease progression. http://repub.eur.nl/res/pub/9650/ 2001-01-01T00:00:00Z Analyses of cancer incidence data in the United States and Western Europe revealed steadily rising rates over the past decades of adenocarcinomas of the esophagus and gastric cardia. Genetic information on gastric cardia adenocarcinoma and its preneoplasias is sparse. We have used comparative genomic hybridization to obtain a genome-wide overview of 20 archival gastric cardia adenocarcinomas and 10 adjacent preneoplastic lesions (4 metaplasias, 1 low-grade dysplasia, 5 high-grade dysplasias). Multiple genetic alterations were discriminated in all adenocarcinomas. Frequent loss (> or =25% of all tumors) was detected, in decreasing order of frequency, on 5q, 18q, 4q, 3p, 9p, 2q, 11q, 14q, 21q, 4p, 9q, 16q, 1p, and 8p. Frequent gain (> or =25% of all tumors) was disclosed, in decreasing order of frequency, on 20q, 7p, 8q, 1q, 7q, 20p, 17q, 13q, Xp, 6q, 8p, 19q, 5p, 6p, and Xq. Loss of the Y chromosome was found in 60% of male cases. High level amplification was frequently (>10% of all tumors) detected on 7q21, 8p22, 12p11.2, 17q12-q21, and 19q13.1-q13.2. The precursor lesions showed multiple aberrations in all high-grade dysplasias, whereas few genetic changes were discerned in LGD and metaplasias. High level amplifications were also found in high-grade dysplasias, ie, on 7q21, 8p22, and 17q12-q21. Moreover, the percentage of aberrations was not significantly different for invasive carcinomas or high-grade dysplasias. Approximately 70% of the precursor aberrations were also present in the adjacent carcinoma. Minimal overlapping regions in the preneoplasias included loss on 18q12-q21 and gains on 8q23 and 17q12-q21, suggesting involvement of genes residing in these regions. In conclusion, we have (i) created a map of genetic alterations in gastric cardia adenocarcinomas and (ii) provided evidence for the presence of a metaplasia-dysplasia-carcinoma sequence in this poorly understood type of cancer. http://repub.eur.nl/res/pub/8773/ 1998-01-01T00:00:00Z We describe a G-->A transition within intron 5 of the NF2 gene. This mutation creates a consensus splice branch point sequence. To our knowledge this is the first report of a mutation that creates a functional branch point sequence in a human hereditary disorder. The new branch point sequence is located 18 bp upstream of a consensus splice acceptor site. A consensus splice donor site is found 106 bp 3' of the acceptor site. Asa consequence the G-->A transition results in an alternatively spliced mRNA containing an additional exon 5a of 106 bp derived from intron sequences. We cloned the mutant cDNA and show that due to an in-frame stop codon the cDNA codes for a truncated NF2 protein. The mutation was observed in three affected members of an NF2 family. In a tumour of one of the family members both alternatively spliced and wild-type mRNA were found, although the wild-type allele of the gene is absent due to an interstitial deletion on chromosome 22. We also show that immunoprecipitations reveal the presence of full-length wild-type NF2 protein in the tumour lysate. These data support the hypothesis that some degree of normal splicing of the mutant precursor RNA is taking place. It is therefore likely that this residual activity of the mutant allele explains the relatively mild phenotype in the family. These data also indicate that complete inactivation of the gene is not required for tumour formation. http://repub.eur.nl/res/pub/40328/ 1992-11-18T00:00:00Z As a start of our study of prostate-specific and androgen-dependent expression of PA, various PA cDNAs (Chapter II) and the PA gene (Chapter Ill) were isolated and characterized. The PA gene turned out to be a member of a small kaHikrein family, encompassing two other closely related genes Ithe human Glandular Kal!ikrein-1 gene (hGK-1), and the tissue kallikrein gene {KLK1)]. Genetic characterization of the human kallikrein genes showed clustering in a 60 kb segment on chromosome 19q13.2-13.4 (Chapters IV and V). The hGK-1 gene shows a strong homology to PA and is, similarly to PA, exclusively expressed in prostate tissue. KLK1 is mainly expressed in kidney, pancreas and salivary glands. In addition to PA eDNA, hGK-1 cDNAs were isolated and characterized. This allowed the comparison of PA and hGK-1 mRNA expression. Using hGK-1 and ?A-specific eDNA probes, androgen-stimulated mRNA expression of PA and hGK-1 could be determined (Chapter VI). Further, an androgen responsive element in the promoter region of the PA gene was defined and tested for its functional activity (Chapter Vl!). The homology of the PA, hGK-1 and KLK1 genes does not only include the open reading frame, but extends into the promoter regions, although the genes are at least partially expressed in different tissues and at different levels. The above mentioned aspects, resulted in the development of a model system for the study of tissue-specific and hormone-responsive gene expression in the human prostate.