Biomedical subjects

B Ylstra

Publications and source records attributed to B Ylstra.

14 recordsLinked to original sources

PubMed2006

Cross-platform array comparative genomic hybridization meta-analysis separates hematopoietic and mesenchymal from epithelial tumors.

A series of studies have been published that evaluate the chromosomal copy number changes of different tumor classes using array comparative genomic hybridization (array CGH); however, the chromosomal aberrations that distinguish the different tumor classes have not been fully characterized. Therefore, we performed a meta-analysis of different array CGH data sets in an attempt to classify samples tested across different platforms. As opposed to RNA expression, a common reference is used in dual channel CGH arrays: normal human DNA, theoretically facilitating cross-platform analysis. To this aim, cell line and primary cancer data sets from three different dual channel array CGH platforms obtained by four different institutes were integrated. The cell line data were used to develop preprocessing methods, which performed noise reduction and transformed samples into a common format. The transformed array CGH profiles allowed perfect clustering by cell line, but importantly not by platform or institute. The same preprocessing procedures used for the cell line data were applied to data from 373 primary tumors profiled by array CGH, including controls. Results indicated that there is no apparent feature related to the institute or platform and that array CGH allows for unambiguous cross-platform meta-analysis. Major clusters with common tissue origin were identified. Interestingly, tumors of hematopoietic and mesenchymal origins cluster separately from tumors of epithelial origin. Therefore, it can be concluded that chromosomal aberrations of tumors from hematopoietic and mesenchymal origin versus tumors of epithelial origin are distinct, and these differences can be picked up by meta-analysis of array CGH data. This suggests the possibility of prospectively using combined analysis of diverse copy number data sets for cancer subtype classification.

Chromosome Aberrations↗

PubMed2006

Expression profiling and prediction of distant metastases in head and neck squamous cell carcinoma.

BACKGROUND: For breast and prostate cancer, a gene expression signature of the tumour is associated with the development of distant metastases. Regarding head and neck squamous cell carcinoma (HNSCC), the only known risk factor is the presence of > or =3 tumour-positive lymph nodes. AIM: To evaluate whether a HNSCC gene expression signature can discriminate between the patients with and without distant metastases. METHODS: Patients with HNSCC with and without distant metastases had >3 tumour-positive lymph nodes, and did not differ with respect to other risk factors. Statistical analysis was carried out using Student's t test, as well as statistical analysis of microarrays (SAM), to assess the false discovery rate for each gene. These analyses were supplemented with a newly developed method that computed deviations from gaussian-order statistics (DEGOS). To validate the platform, normal mucosa of the head and neck was included as control. RESULTS: 2963 genes were differently expressed between HNSCC and normal mucosa (t test; p<0.01). More rigorous statistical analysis with SAM confirmed the differential expression of most genes. The comparison of genes in HNSCC with and without metastases showed 150 differently expressed genes (t test; p<0.01), none of which, however, could be confirmed using SAM or DEGOS. CONCLUSIONS: No evidence for a metastasis signature is found, and gene expression profiling of HNSCC has seemingly no value in determining the risk of developing distant metastases. The absence of such a signature can be understood when it is realised that, for HNSCC in contrast with breast cancer, the lymph nodes are a necessary in-between station for haematogenous spread.

Adult↗

PubMed2006

Genome-wide DNA copy number alterations in head and neck squamous cell carcinomas with or without oncogene-expressing human papillomavirus.

Oncogene-expressing human papillomavirus type 16 (HPV16) is found in a subset of head and neck squamous cell carcinomas (HNSCC). HPV16 drives carcinogenesis by inactivating p53 and pRb with the viral oncoproteins E6 and E7, paralleled by a low level of mutations in TP53 and allelic loss at 3p, 9p, and 17p, genetic changes frequently found in HNSCCs of nonviral etiology. We hypothesize that two pathways to HNSCC exist: one determined by HPV16 and the other by environmental carcinogens. To define the critical genetic events in these two pathways, we now present a detailed genome analysis of HNSCC with and without HPV16 involvement by employing high-resolution microarray comparative genomic hybridization. Four regions showed alterations in HPV-negative tumors that were absent in HPV-positive tumors: losses at 3p11.2-26.3, 5q11.2-35.2, and 9p21.1-24, and gains/amplifications at 11q12.1-13.4. Also, HPV16-negative tumors demonstrated loss at 18q12.1-23, in contrast to gain in HPV16-positive tumors. Seven regions were altered at high frequency (>33%) in both groups: gains at 3q22.2-qter, 5p15.2-pter, 8p11.2-qter, 9q22-34.1, and 20p-20q, and losses at 11q14.1-qter and 13q11-33. These data show that HNSCC arising by environmental carcinogens are characterized by genetic alterations that differ from those observed in HPV16-induced HNSCC, and most likely occur early in carcinogenesis. A number of genetic changes are shared in both tumor groups and can be considered crucial in the later stages of HNSCC progression.

Adult↗

PubMed2006

Increased gene copy numbers at chromosome 20q are frequent in both squamous cell carcinomas and adenocarcinomas of the cervix.

Genome-wide microarray-based comparative genomic hybridization (array CGH) was used to identify common chromosomal alterations involved in cervical carcinogenesis as a first step towards the discovery of novel biomarkers. The genomic profiles of nine squamous cell carcinomas (SCCs) and seven adenocarcinomas (AdCAs), as well as four human papillomavirus (HPV)-immortalized keratinocyte cell lines, were assessed. On a genome-wide scale, SCCs showed significantly more gains than AdCAs. More specifically, there was a striking and highly significant difference between the two histological types for gain at 3q12.1-28, which was predominantly observed in SCC. Other frequent alterations included gains of 1q21.1-31.1 and 20q11.21-13.33, and losses of 11q22.3-25 and 13q14.3-21.33. Subsequent FISH analysis for hTR, located at 3q26, confirmed the presence of 3q gain in SCCs and HPV-immortalized cell lines. Fine mapping of chromosome 20q using multiplex ligation-dependent probe amplification (MLPA) showed copy number increases for a number of genes located at 20q11-q12, including DNMT3B and TOP1. For DNMT3B, this correlated with elevated mRNA expression in 79% of cases. In conclusion, the assessment of frequent genomic alterations resulted in the identification of potential novel biomarkers, which may ultimately enable a better risk stratification of high-risk (hr)-HPV-positive women.

Adenocarcinoma↗

PubMed2005

DNA copy number changes at 8q11-24 in metastasized colorectal cancer.

BACKGROUND: C-Myc, a well-known oncogene located on 8q24.12-q24.23, is often amplified and over-expressed in both primary and metastasizing colorectal cancer. In addition, PRL-3 (also known as PTP4A3), a tyrosine phosphatase located on 8q24.3, is amplified in colorectal cancer metastasis. Beside PRL-3 and c-myc, other oncogenes located on the 8q23-24 region might be involved in this process. Therefore, the present study aims to correlate DNA copy number status of a series of genes at 8q23-24 in colorectal cancer at high resolution in correlation to metastatic disease. MATERIALS AND METHODS: Thirty-two cases of colorectal cancer, 10 stage B1, 10 B2 and 12 D (Astler-Coller) with their corresponding liver metastasis and one colorectal cell line (colo205, previously analyzed by array-CGH), were included in this study. A chromosome 8 specific MLPA probe mixture was used to analyze the presence of DNA copy number changes. The probe mixture contained 29 probes covering 25 genes on chromosome 8, as well as 6 control probes on other chromosomes. RESULTS AND DISCUSSION: MLPA results obtained of the colo205 colorectal cell line were comparable with previous array-CGH results, thus validating the MLPA probe mixture. Astler-Coller B1 and B2 colorectal cancers differed significantly in DNA copy number of the genes, MOS (p=0.04), MYC (p=0.007), DDEF1 (p=0.004), PTK2 (p=0.02) and PTP4A3 (p=0.04). When comparing these with Astler-Coller D primary tumors, significant differences were seen for several genes as well (MYC (p<0.000), DDEF1 (p<0.000), SLA (p<0.000), PTK2 (p<0.000), PTP4A3 (p=0.002), and RECQL4 (p=0.01)). When comparing primary Astler-Coller D tumors and their corresponding liver metastases, a similar pattern of gains and losses was observed. Most of the liver metastases showed higher DNA copy number ratios than the corresponding primary tumors, but this difference was only significant for TPD52 (p=0.02) and EIF3S6 (p=0.007). CONCLUSION: In addition to c-myc, multiple genes on chromosome 8 differed significantly between primary colorectal cancers with and without liver metastases. This observation is consistent with the concept that clinical behaviour, like risk of liver metastasis, is determined by the genomic profile that is already present in the primary tumor.

Base Sequence↗

PubMed2004

Microarray-based comparative genomic hybridization and its applications in human genetics.

Through the years, several techniques capable of detecting DNA copy number changes have been developed. A number of those, such as karyotyping and fluorescence in situ hybridization (FISH), have proven to be valuable tools in both research and diagnostics. Recently, a new technique, called microarray-based comparative genomic hybridization (array CGH), has been introduced. Array CGH has proven to be a specific, sensitive, and fast technique, with considerable advantages compared to other methods used for the analysis of DNA copy number changes. Array CGH enables analysis of the whole genome in a single experiment. Until now, its applications have been mainly directed at detecting genomic abnormalities in cancer. However, array CGH is also suitable for the analysis of DNA copy number aberrations that cause human genetic disorders. This review gives an overview of array CGH and its applications in human genetics. Advantages, limitations, and future perspectives of array CGH are discussed.

Gene Dosage↗

PubMed2004

High resolution microarray comparative genomic hybridisation analysis using spotted oligonucleotides.

BACKGROUND: Currently, comparative genomic hybridisation array (array CGH) is the method of choice for studying genome wide DNA copy number changes. To date, either amplified representations of bacterial artificial chromosomes (BACs)/phage artificial chromosomes (PACs) or cDNAs have been spotted as probes. The production of BAC/PAC and cDNA arrays is time consuming and expensive. AIM: To evaluate the use of spotted 60 mer oligonucleotides (oligos) for array CGH. METHODS: The hybridisation of tumour cell lines with known chromosomal aberrations on to either BAC or oligoarrrays that are mapped to the human genome. RESULTS: Oligo CGH was able to detect amplifications with high accuracy and greater spatial resolution than other currently used array CGH platforms. In addition, single copy number changes could be detected with a resolution comparable to conventional CGH. CONCLUSIONS: Oligos are easy to handle and flexible, because they can be designed for any part of the genome without the need for laborious amplification procedures. The full genome array, containing around 30000 oligos of all genes in the human genome, will represent a big step forward in the analysis of chromosomal copy number changes. Finally, oligoarray CGH can easily be used for any organism with a fully sequenced genome.

Breast Neoplasms↗

PubMed2001

Assembly of microarrays for genome-wide measurement of DNA copy number.

We have assembled arrays of approximately 2,400 BAC clones for measurement of DNA copy number across the human genome. The arrays provide precise measurement (s.d. of log2 ratios=0.05-0.10) in cell lines and clinical material, so that we can reliably detect and quantify high-level amplifications and single-copy alterations in diploid, polyploid and heterogeneous backgrounds.

Aneuploidy↗

PubMed2001

Comprehensive genome sequence analysis of a breast cancer amplicon.

Gene amplification occurs in most solid tumors and is associated with poor prognosis. Amplification of 20q13.2 is common to several tumor types including breast cancer. The 1 Mb of sequence spanning the 20q13.2 breast cancer amplicon is one of the most exhaustively studied segments of the human genome. These studies have included amplicon mapping by comparative genomic hybridization (CGH), fluorescent in-situ hybridization (FISH), array-CGH, quantitative microsatellite analysis (QUMA), and functional genomic studies. Together these studies revealed a complex amplicon structure suggesting the presence of at least two driver genes in some tumors. One of these, ZNF217, is capable of immortalizing human mammary epithelial cells (HMEC) when overexpressed. In addition, we now report the sequencing of this region in human and mouse, and on quantitative expression studies in tumors. Amplicon localization now is straightforward and the availability of human and mouse genomic sequence facilitates their functional analysis. However, comprehensive annotation of megabase-scale regions requires integration of vast amounts of information. We present a system for integrative analysis and demonstrate its utility on 1.2 Mb of sequence spanning the 20q13.2 breast cancer amplicon and 865 kb of syntenic murine sequence. We integrate tumor genome copy number measurements with exhaustive genome landscape mapping, showing that amplicon boundaries are associated with maxima in repetitive element density and a region of evolutionary instability. This integration of comprehensive sequence annotation, quantitative expression analysis, and tumor amplicon boundaries provide evidence for an additional driver gene prefoldin 4 (PFDN4), coregulated genes, conserved noncoding regions, and associate repetitive elements with regions of genomic instability at this locus.

Animals↗

PubMed2000

Quantitative mapping of amplicon structure by array CGH identifies CYP24 as a candidate oncogene.

We show here that quantitative measurement of DNA copy number across amplified regions using array comparative genomic hybridization (CGH) may facilitate oncogene identification by providing precise information on the locations of both amplicon boundaries and amplification maxima. Using this analytical capability, we resolved two regions of amplification within an approximately 2-Mb region of recurrent aberration at 20q13.2 in breast cancer. The putative oncogene ZNF217 (ref. 5) mapped to one peak, and CYP24 (encoding vitamin D 24 hydroxylase), whose overexpression is likely to lead to abrogation of growth control mediated by vitamin D, mapped to the other.

Breast Neoplasms↗

PubMed1999

Analysis of mRNA stabilities during pollen development and in BY2 cells

RNA stability is important in post-transcriptional gene expression. However, its role during plant male gametophyte development has not been studied in detail, probably because of technical difficulties and because earlier work suggested that, in general, mRNAs expressed during late pollen development would be stable. We determined mRNA stabilities of 12 pollen-expressed genes. We measured pollen mRNA decay during in planta development and in vitro maturation after transcriptional inhibition with ActinomycinD (ActD). Because five of these genes were also expressed in somatic cells we also measured RNA decay in BY2 cells after transcriptional inhibition with ActD. Ten of the mRNAs were highly stable during pollen development (t1/2 > 30 h), including all seven pollen-specific mRNAs. However, two mRNAs showed a logarithmic decrease as pollen development proceeded, both in planta and in vitro. In fact, for one of these two, GUT8-2b, the mRNA level decreased significantly within 3 h. Our results show that many pollen-expressed mRNAs are indeed long-lived but that at least some mRNAs undergo decay during pollen maturation. Thus, there is no overall cytoplasmic condition in pollen that stabilizes mRNAs and specific mRNA turnover mechanisms must exist. We detected reciprocal mRNA stabilities with two genes that are expressed in both BY2 cells and pollen. GUT15 mRNA, known to be unstable (t1/2 < 3 h) in BY2 cells, was highly stable in pollen. In contrast, GRP2 mRNA decayed in pollen but was highly stable in BY2 cells. Therefore, we conclude that mRNA stability can be tissue-specific.

Journal Article↗

PubMed1998

Hexose transport in growing petunia pollen tubes and characterization of a pollen-specific, putative monosaccharide transporter.

We investigated the molecular and physiological processes of sugar uptake and metabolism during pollen tube growth and plant fertilization. In vitro germination assays showed that petunia (Petunia hybrida) pollen can germinate and grow not only in medium containing sucrose (Suc) as a carbon source, but also in medium containing the monosaccharides glucose (Glc) or fructose (Fru). Furthermore, high-performance liquid chromatography analysis demonstrated a rapid and complete conversion of Suc into equimolar amounts of Glc and Fru when pollen was cultured in a medium containing 2% Suc. This indicates the presence of wall-bound invertase activity and uptake of sugars in the form of monosaccharides by the growing pollen tube. A cDNA designated pmt1 (petunia monosaccharide transporter 1), which is highly homologous to plant monosaccharide transporters, was isolated from petunia. Pmt1 belongs to a small gene family and is expressed specifically in the male gametophyte, but not in any other vegetative or floral tissues. Pmt1 is activated after the first pollen mitosis, and high levels of mRNA accumulate in mature and germinating pollen. A model describing the transport of sugars to the style, the conversion of Suc into Glc and Fru, and the active uptake by a monosaccharide transporter into the pollen tube is presented.

Amino Acid Sequence↗

PubMed1996

Flavonols are not essential for fertilization in Arabidopsis thaliana.

Flavonols are plant metabolites suggested to serve a vital role in fertilization of higher plants. Petunia and maize plants mutated in their flavonol biosynthesis are not able to set seed after self-pollination. We have investigated the role of these compounds in Arabidopsis thaliana. Like in all other plant species, high levels of flavonols could be detected in pollen of wild-type A. thaliana. No flavonols were detected in reproductive organs of the A. thaliana tt4 mutant in which the chs gene is mutated. Surprisingly, this mutant did set seed after self-fertilization and no pollen tube growth aberrations were observed in vivo. The role of flavonols during fertilization of Arabidopsis is discussed.

Acyltransferases↗

PubMed1992

Flavonols stimulate development, germination, and tube growth of tobacco pollen.

The effect of anther-derived substances on pollen function was studied using pollen produced by in vitro culture of immature pollen of tobacco (Nicotiana tabacum L.) and petunia (Petunia hybrida). Addition of conditioned medium consisting of diffusates from in situ matured pollen strongly increased pollen germination frequency and pollen tube growth, as well as seed set after in situ pollination. Thin-layer chromatography and depletion of phenolic substances by Dowex treatment indicated that flavonols are present in the diffusate and may be the active compounds. When added to the germination medium, flavonols (quercetin, kaempferol, myricetin) but not other flavonoids strongly promoted pollen germination frequency and pollen tube growth in vitro. The best results were obtained at very low concentrations of the flavonols (0.15-1.5 mum), indicating a signaling function. The same compounds were also effective when added during pollen development in vitro.

Journal Article↗