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R Slater

Publications and source records attributed to R Slater.

At least 55 records · Page 3Linked to original sources
PubMed1997

Cloning of cold-inducible dehydrin-like genes from the blackcurrant (Ribes nigrum L.) using RT-PCR.

A method is described for the identification of conserved genes in one plant species by using sequence information on internal motifs from well-characterized clones from another species. This sequence information is used to design primers for reverse transcriptase polymerase chain reaction (RT-PCR) and to design oligonucleotide probes to identify genuine positive amplification products. The approach was successfully used to clone cDNAs encoding cold-inducible dehydrin-like genes from the woody perennial blackcurrant, Ribes nigrum L. The strategy described can accelerate the cloning of heterologous cDNAs and is a convenient alternative to direct screening of cDNA libraries.

Amino Acid Sequence↗
PubMed1997

Assessment of chromosomal gains and losses in oral squamous cell carcinoma by comparative genomic hybridisation.

Cytogenetic studies have demonstrated that oral squamous cell carcinomas (OSCCs) are usually characterised by complex karyotypes with many marker chromosomes. We analysed the genetic changes of six OSCC cell cultures by comparative genomic hybridisation (CGH). The CGH technique provides information on chromosomal gains and losses of the whole tumour genome in a single experiment and can therefore identify regions that harbour putative tumour suppressor genes (in the case of loss of chromosomal material) or oncogenes (in the case of gain or amplification of chromosomal material). Recurrent losses were detected at chromosome arms Xp and 3p (four cases). Gains consistently occurred at chromosome arms 8q and 9q (four cases) and at 1q, 3q, 5p, 7p, and 9p (three cases). The same six tumour cultures have previously been analysed by classical karyotyping. An important discrepancy between the two techniques was the number of losses detected: 55 with karyotyping versus 26 with CGH. On the basis of the cytogenetic complexity of these tumours and on FISH experiments that confirmed the CGH results, we conclude that genetic changes, particularly losses, can be more reliably detected by CGH analysis.

Carcinoma, Squamous Cell↗
PubMed1997

Comparative genomic hybridization analysis of Wilms tumors.

In this study we have applied the technique of comparative genomic hybridization (CGH) to a large series of sporadic Wilms tumors, including six samples of the associated nephroblastomatosis. The data obtained were compared with the findings of molecular studies carried out on the same material. The aims of the study were (1) to characterize the range of genetic variation in sporadic Wilms tumor and nephroblastomatosis, (2) to determine whether changes could be found that have not been detected by commonly used techniques, and (3) to compare the sensitivity of CGH with that of conventional molecular analysis. The chromosomes that showed gains and losses by CGH were similar to those previously found in molecular and cytogenetic studies, however loss of 4q was a new event identified in 2 out of 46 tumors. We did not detect amplified genetic material. Comparison of the data from the nephroblastomatosis and tumor samples from the same patient showed that loss of 7p may be associated with malignant transformation, and that losses in 1p, 11p, 4q and gains in 1q and 12q can be early events; whilst loss in 9p and gain of 8, 10q and 18 are possible secondary changes in tumor development. The combined CGH and molecular techniques used demonstrated involvement of two specific 1p regions in the etiology of Wilms tumor.

Chromosome Aberrations↗
PubMed1996

Allelic loss of chromosome 1p as a predictor of unfavorable outcome in patients with neuroblastoma.

BACKGROUND: Neuroblastoma is a childhood tumor derived from cells of the neural crest, with a widely variable outcome. Differences in the behavior and prognosis of the tumor suggest that neuroblastoma can be divided into several biologic subgroups. We evaluated the most frequent genetic abnormalities in neuroblastoma to determine their prognostic value. METHODS: We used Southern blot analysis to study the allelic loss of chromosomes 1p, 4p, 11q, and 14q, the duplication of chromosome 17q, and the amplification of the N-myc oncogene in 89 neuroblastomas. We also determined the nuclear DNA content of the tumor cells. RESULTS: Allelic loss of chromosome 1p, N-myc amplification, and extra copies of chromosome 17q were significantly associated with unfavorable outcome. In a multivariate analysis, loss of chromosome 1p was the most powerful prognostic factor. It provided strong prognostic information when it was included in multivariate models containing the prognostic factors of age and stage or serum ferritin level and stage. Among the patients with stage I, II, or IVS disease, the mean (+/- SD) three-year event-free survival was 100 percent in those without allelic loss of chromosome 1p and 34 +/- 15 percent in those with such loss; the rates of three-year event-free survival among the patients with stage III and stage IV disease were 53 +/- 10 percent and 0 percent, respectively. CONCLUSIONS: The loss of chromosome 1p is a strong prognostic factor in patients with neuroblastoma, independently of age and stage. It reliably identifies patients at high risk in stages I, II, and IVS, which are otherwise clinically favorable. More intensive therapy may be considered in these patients. Patients in stages III and IV with allelic loss of chromosome 1p have a very poor outlook, whereas those without such loss are at moderate risk.

Blotting, Southern↗
PubMed1996

Familial sideroblastic anemia with emergence of monosomy 5 and myelodysplastic syndrome.

The case history of two sisters with pyridoxine-refractory familial sideroblastic anemia (FSA) is presented in which one developed a myelodysplastic syndrome (MDS) with monosomy for chromosome 5. Bone marrow examination of both patients at diagnosis showed erythroid hyperplasia with more than 50% ring sideroblasts. Karyotypic analysis initially showed a normal 46, XX karyotype in both of the children. Therapeutic trials with pyridoxine, prednisone, and erythropoietin were unsuccessful. The first patient required regular transfusions and developed a significant hemosiderosis. At the age of 9 years, 7.5 years after the diagnosis of FSA, refractory anemia with excess of blasts (RAEB) was diagnosed. Bone marrow cytogenetic analysis revealed a clone with monosomy for chromosome 5. Her sister's illness was detected at the age of 12 years. She has a more benign course of disease, remains largely transfusion independent and until now shows no signs of myelodysplasia. To our knowledge this is the first observation of a transition of FSA to MDS accompanied by the appearance of a chromosomal abnormality. FSA might be another type of bone marrow failure syndrome, therefore close follow-up of these patients may be necessary.

Anemia, Refractory, with Excess of Blasts↗
PubMed1996

Positional cloning of genes involved in the Beckwith-Wiedemann syndrome, hemihypertrophy, and associated childhood tumors.

The Beckwith-Wiedemann syndrome (BWS) is an overgrowth malformation syndrome that occurs with an incidence of 1:13,700 births. There is a striking incidence of childhood tumors found in BWS patients. Various lines of investigation have localized "imprinted" genes involved in BWS and associated childhood tumors to 11p15. High resolution mapping of 8 rare balanced chromosomal BWS rearrangements enabled us to identify three distinct regions on chromosome 11p15 that might harbor genes involved in the above-mentioned disorders. These results suggest genetic heterogeneity that correlates with the clinical heterogeneity seen in the patients studied. Expressed candidate gene sequences from these regions have been cloned and partly sequenced. These transcripts are either disrupted by or are at least within a few kb of these BWS chromosome breakpoints. So far, zinc-finger sequences and one Kruppel-associated box (KRAB) domain were found in independent candidate genes which are compatible with a regulating function of growth promoting genes. The abundance of expression of these genes varies from low abundant in all adult and fetal tissues tested to detectable on Northern blots of adult tissues. In addition to our 11p15 studies we have analyzed additional chromosome regions, in particular 1p. Cytogenetic, loss of heterozygosity (LOH) and comparative genomic hybridization (CGH) studies have identified 1p35 as a region of interest. A positional cloning effort to identify a balanced 1p35 translocation found in a Wilms tumor has led to the isolation of a YAC, crossing this breakpoint.

Adult↗
PubMed1996

Centromeric breakage as a major cause of cytogenetic abnormalities in oral squamous cell carcinoma.

Cytogenetic analysis of short-term explant tumor cultures derived from 11 human oral squamous cell carcinomas (nine from primary tumors and two from nude mouse xenograft cultures) revealed clonal chromosomal aberrations with multiple numerical and structural changes in all tumors. Recurrent breakpoints were located at chromosomal bands 1p13 (five tumors), 11q13 (four tumors), 3q27-29 (three tumors), and 12q13 (three tumors). Four tumors had a homogeneously staining region at band 11q13. Consistent chromosomal losses included 3p, 9p13-pter, and 18q22-qter, each occurring in eight tumors. Gain of material was observed for chromosome arms 3q, 5p, 7p, and 8q. As many as 134 of a total of 218 chromosomal breakpoints (61%) occurred in centromeric regions, often resulting in isochromosomes and unbalanced whole-arm translocations. Using fluorescence in situ hybridization with chromosome-specific centromeric alphoid repeat probes, two whole-arm translocations, der(Xq;11q) and a der(3q;11q), each from a different tumor, were shown to contain juxtaposed centromeric sequences of both participating chromosomes, strongly suggesting that the breakpoints were within the centromeres. We propose that centromeric breakage is an important mechanism for the generation of genetic imbalance in the development of oral squamous cell carcinoma.

Animals↗
PubMed1996

Allelic loss of the short arm of chromosome 4 in neuroblastoma suggests a novel tumour suppressor gene locus.

Neuroblastoma is a childhood neural crest tumour, genetically characterized by frequent deletions of the short arm of chromosome 1 and amplification of N-myc. Here we report the first evidence for a neuroblastoma tumour suppressor locus on 4pter. Cytogenetically we demonstrated rearrangements of 4p in 7 out of 26 evaluable tumours (27%). Subsequent analysis of loss of heterozygosity (LOH) by Southern blotting revealed allelic loss of 4p in 16/82 (19.5%) informative neuroblastomas. Taken together cytogenetic and Southern blot analyses showed loss of 4p in 20/86 neuroblastomas analysed (23%). The common deleted region was bordered by the probe D4S123 and encompassed the distal 34 cM of 4p. We found no evidence for genomic imprinting of the 4p locus as the 4p alleles lost in the tumours were of random maternal and paternal origin. LOH4p was found at all disease stages and in every age group. Furthermore LOH4p was present both in cases with and without LOH1p and amplification of N-myc.

Alleles↗
PubMed1996

The application of microwave denaturation in comparative genomic hybridization.

Comparative genomic hybridization (CGH) is a powerful tool for analyzing unbalanced chromosomal rearrangements in a variety of tissues. However, reproducibility of the technique is poor. We have developed an alternative protocol involving microwave denaturation of the metaphase chromosome preparations prior to the hybridization step. The advantage of this method for CGH is the retention of the morphology of the chromosomes and hence an improved chromosome banding pattern. Furthermore, it results in a consistently strong hybridization which is not dependent on the batch of lymphocytes used to obtain the metaphase chromosome spreads. This procedure has also proved to be applicable to nucleic acid hybridizations in general. The protocol, its application and the results of this method in CGH is discussed. Furthermore preliminary results of this method in paint and DNA probe hybridizations to chromosome spreads and to RNA in tissue sections are presented.

DNA, Neoplasm↗
PubMed1995

DNA index and %S-phase cells determined in acute lymphoblastic leukemia of children: a report from studies ALL V, ALL VI, and ALL VII (1979-1991) of the Dutch Childhood Leukemia Study Group and The Netherlands Workgroup on Cancer Genetics and Cytogenetics.

DNA per cell content was routinely recorded by single-parameter flow cytometry in leukemic blasts from 473 children with acute lymphoblastic leukemia (ALL), enrolled in national studies ALL V, VI, and VII (1979-1991) of the Dutch Childhood Leukemia Study Group. The parameters bonemarrow %S-value and DNA Index were compared with clinical features, with chromosome number based on cytogenetic analyses and with treatment results in study ALL VI. %S-values, ranging between 1 and 36%, were unrelated to initial white blood cell count, immunophenotype, and DNA index but were lowest in blasts with L1 morphology. In study ALL VI (non-high risk), the survival of patients with < or = 6% S-phase cells was superior to that of patients with %S-values of > 6 (P = 0.030). Hyperdiploidy, defined by a DNA index > or = 1.16, was compared to the cytogenetic hyperdiploid classification of n > 50. Initially there were 25 discrepancies in 189 samples jointly analysed by flow cytometry and cytogenetics. After review only five discrepancies remained unresolved. Hyperdiploidy, independent of the method used, was found to be unrelated to blast morphology and %S-phase cells but closely associated with c-ALL and was absent in T-ALL. In study ALL VI, event-free survival at 8 years of hyperdiploid patients was 90.6% but was not significantly different from non-hyperdiploid patients (EFS = 82.1%; P = 0.08). Routine DNA flow cytometry appeared a valuable adjunct to cytogenetic analyses and allowed the identification of a large subset of non-high-risk ALL patients in study ALL VI with a DNA index > or = 1.16 or %S-value of < or = 6.0 with highest survival probability.

Bone Marrow↗
PubMed1995

Maintenance of vascular endothelial cell-specific properties after immortalization with an amphotrophic replication-deficient retrovirus containing human papilloma virus 16 E6/E7 DNA.

Primary human vascular endothelial cells were immortalized by the integration of a single DNA copy of an amphotrophic, replication-deficient retrovirus containing the E6/E7 genes of human papilloma virus. To date, the resulting cell lines, designated EC-RF7 and EC-RF24, have been cultured for more than 1 year. The cell lines have retained a diploid karyotype, display no abnormalities, and are able to grow in a polar mode. Analysis of the EC-RF cell lines by indirect immunofluorescence, using an extensive panel of monoclonal antibodies, showed expression of endothelial cell-specific soluble (von Willebrand factor) and surface-bound antigens (endoglin, PCAM-1) indistinguishable from that of primary cells. In addition, the expression of the markers CD9, 13, 14, 29, 36, 40, 51, and 55 that are not restricted to endothelial cells was also similar for the immortalized and the primary endothelial cells. Immortalization did not alter the expression of the surface adhesion molecules E-selectin, VCAM-1, and ICAM-1 nor transmigration of neutrophils. The regulation of extracellular proteolytic activity by EC-RF24 was established by measuring both the induction of functional tissue factor (promotion of Factor Xa generation) and the functional deposition of plasminogen activator inhibitor 1 in the subendothelial matrix (SDS-resistant complex formation with thrombin). Finally, the biosynthesis of the endothelial cell-specific von Willebrand factor was studied in detail in the EC-RF24 cell line and the results were compared with those of primary endothelial cells.

Antigens, CD↗
PubMed1995

A non-random chromosome abnormality found in precursor-B lineage acute lymphoblastic leukaemia: dic(9;20)(p1?3;q11).

A comparison of cytogenetical data on acute lymphoblastic leukaemia studied at four large European centres has revealed a non-random dicentric chromosome abnormality: dic(9;20) (p1?3;q11) in 10 patients, nine of whom were children. All had early precursor-B lineage ALL, and eight children had a non-standard risk clinical presentation. The origin of the dicentric chromosome was demonstrated using a range of chromosome banding techniques. This was confirmed by FISH using paints and centromeric probes for chromosomes 9 and 20, together with a number of cosmid probes. The follow-up time of these patients is presently too short and the number of patients too few to determine the prognostic significant of this chromosome abnormality.

Adolescent↗
PubMed1994

Recurrent 1;17 translocations in human neuroblastoma reveal nonhomologous mitotic recombination during the S/G2 phase as a novel mechanism for loss of heterozygosity.

Neuroblastomas often show loss of heterozygosity of the chromosomal region 1p36 (LOH 1p), probably reflecting loss of a tumor-suppressor gene. Here we describe three neuroblastoma tumors and two cell lines in which LOH 1p results from an unbalanced translocation between the p arm of chromosome 1 and the q arm of chromosome 17. Southern blot and cytogenetic analyses show that in all cases the chromosome 17 homologue from which the 1;17 translocation was derived is still present and intact. This suggests a model in which a translocation between the short arm of chromosome 1 and the long arm of chromosome 17 takes place in the S/G2 phase of the cell cycle and results in LOH 1p. Nonhomologous mitotic recombination in the S/G2 phase is a novel mechanism of LOH.

Alleles↗
PubMed1993

Acute leukemias of different lineages have similar MLL gene fusions encoding related chimeric proteins resulting from chromosomal translocation.

The MLL gene, on human chromosome 11q23, undergoes chromosomal translocation in acute leukemias, resulting in gene fusion with AF4 (chromosome 4) and ENL (chromosome 19). We report here translocation of MLL with nine different chromosomes and two paracentric chromosome 11 deletions in early B cell, B- or T-cell lineage, or nonlymphocytic acute leukemias. The mRNA translocation junction from 22 t(4;11) patients, including six adult leukemias, and nine t(11;19) tumors reveals a remarkable conservation of breakpoints within MLL, AF4, or ENL genes, irrespective of tumor phenotype. Typically, the breakpoints are upstream of the zinc-finger region of MLL, and deletion of this region can accompany translocation, supporting the der(11) chromosome as the important component in leukemogenesis. Partial sequence of a fusion between MLL and the AFX1 gene from chromosome X shows the latter to be rich in Ser/Pro codons, like the ENL mRNA. These data suggest that the heterogeneous 11q23 abnormalities might cause attachment of Ser/Pro-rich segments to the NH2 terminus of MLL, lacking the zinc-finger region, and that translocations occur in early hematopoietic cells, before commitment to distinct lineages.

Acute Disease↗