Biomedical subjects
Publications and source records attributed to K D Zang.
In very rare patients tetrasomy of chromosome 8 is observed as the only karyotypic event. We present a 74-year-old man with acute monocytic leukemia (FAB M5) displaying 46,XY/48,XY, +8, +8 in unstimulated bone marrow cells. The patient died 2 weeks after diagnosis. Tetrasomy 8 appears to identify a subgroup of acute nonlymphocytic leukemia. It may indicate a poor prognosis.
The met proto-oncogene was found to be amplified in a human glioblastoma cell line (T3095) established from a glioblastoma multiform WHO grade IV. Amplification of epidermal growth factor receptor, transforming growth factor alpha and N-myc which have been described previously in glioblastoma were not observed in T3095. There was, however, an 8-fold met amplification. Giemsa-stained metaphases of T3095 cells revealed multiple (> 5) double minutes (dmins) in the majority of cells. Following xenografting in nude mice there was a significant increase in the number and frequency of dmins. The increase in dmins correlates with the level of met amplification (50-fold), suggesting localisation of the amplified met on dmins. Here we report the first case of met amplification in glioblastoma. Correlation between met amplification and extrachromosomal elements (dmins) has not been reported previously.
Recently, methods have been developed for the isolation of expressed sequences from particular human chromosomes. Using Alu consensus sequences as primers, cDNA synthesis has been initiated from interspecies hybrid cell lines that contain single human chromosomes. Alu consensus sequences have also been utilized to amplify human genomic sequences via polymerase chain reaction (PCR). Here, we describe the use of Alu-PCR to isolate expressed sequences from human chromosomes selectively. Heteronuclear (hn) RNA is transcribed into cDNA by using poly-(dT)15 primer sequences. Subsequently, human specific cDNA sequences are amplified by Alu-PCR and cloned into pBluescript. To verify the chromosomal assignment, cloned PCR products are sequenced, converted into STS markers, and tested on a different somatic hybrid that contains human chromosome 22. The method provides a fast, reliable way to identify expressed sequence tagged sites from selected human chromosomes.
The applicability of fluorescence in situ DNA hybridization to hair root preparations for the detection of numerical chromosome anomalies is demonstrated. This simple method yields a high number of scorable nuclei, allowing rapid identification of epithelial cell chromosome composition. In clinical genetics, this technique may become useful in cases where other types of cells are difficult to obtain.
Meningiomas are common tumours derived from the thin membrane that surrounds the brain and spinal cord. Currently, the molecular mechanisms responsible for the initiation and progression of these tumors are largely unknown. Toward the elucidation of such mechanisms, we have formulated an experimental design utilizing the technique of subtractive hybridization that is aimed at identifying the changes in gene expression between intracranial meningiomas and their normal precursor cells, leptomeningeal cells. We report here the identification and initial characterization of three genes whose expression is altered or aberrant in meningioma cell lines and tumours relative to cultures of normal leptomeningeal cells. Complementary DNA probes from one of these genes detect transcripts of altered size in several meningiomas relative to normal leptomeningeal cells. Another of these genes demonstrates decreased expression in meningiomas and in tumours associated with the disorder neurofibromatosis 2. A third gene isolated by this procedure is differentially expressed in both meningiomas and breast carcinomas. Therefore, the decreased expression of these genes may play roles in growth-regulatory pathways that are abrogated not only in meningiomas, but in other tumor types as well.
We describe a boy with monosomy for the distal part of the short arm of chromosome 3. He had a congenital heart defect, tetramelic hexadactyly, and typical craniofacial anomalies. Comparison with previously reported cases confirms that the phenotype consists of an identifiable pattern of malformation.
By extending non-isotopic in situ hybridization of DNA probes (targeted to metaphase chromosomes or interphase nuclei) to hybridization using tissue sections, additional topological information on the DNA structure and specific alterations can be obtained. We have established a method for the application of two different, chromosome-specific probes labelled with two colour dyes allowing simultaneous detection of two-colour signals. This method was tested in and is applicable to tissue sections of various origins. To demonstrate its sensitivity, prostate carcinomas (either as cryosections or as sections from paraffin blocks) were investigated for the presence or absence of chromosomes 1 and Y. The technique presented here, comparable to immunohistochemical staining, is particularly useful for routine application in diagnostic laboratories and testing of fresh or archival material.
Cytogenetic and molecular studies of various solid tumors have indicated that a series of different chromosomal regions may be deleted in the tumor genome. Usually, losses of heterozygosity are observed and, from this finding, the presence of specific genes acting as tumor suppressors has been deduced. In particular tumors, however, only a single chromosome site appears to be affected. Therefore, we have carried out a study of human meningioma, investigating 7 such putative suppressor regions by applying twelve site-specific DNA markers. In 6 out of 19 tumors, we exclusively found loss of heterozygosity for markers of the long arm of chromosome 22; none of the tumors showed statistically significant additional allelic losses for the regions 1p, 3p, 5p, 5q, 11p, 13q, 17p. Our data support the long-standing observation that only losses of or within chromosome 22 are associated with the development of meningiomas. Other suppressor regions are apparently not involved.
Although prostatic cancer is the most common malignant disease in men in western countries, only limited data on chromosomal changes are available. We report on our cytogenetic findings in cell cultures from four primary prostatic carcinomas indicating the existence of intratumoral focal heterogeneity concerning chromosomal anomalies in prostatic cancer.
A classical minisatellite family, the M13 tandem repeat, is shown here to be spread over all human chromosomes. M13 shows an R-band-like pattern, in contrast to the 33.15 family of Jefferys, which preferentially clusters in the telomeric regions.
The human c-myb gene which encodes a DNA binding protein and which is rarely amplified in neoplastic cells was found to be altered in four human glioblastoma cell lines. It exists in multiple copies in 2 out of 4 cases studied. The degree of amplification as determined by densitometry was about 10-fold, a rearrangement within the coding region and an enhanced gene activity of c-myb were noted. The observation of c-myb oncogene amplification and activity in glioblastoma cell lines presents the first report of this effect in human brain tumor cells.
Immunophenotyping of cultured cancer cells requires intact antigenic structures; these are mostly destroyed by conventional chromosome preparation techniques. Thus, the simultaneous cytogenetic and immunocytochemical characterization of solid tumor cells appears unfeasible. Here, we describe a novel method that allows in situ chromosome preparation from monolayer cultures of solid tumor cells without affecting their immunological features. Using this technique, it is possible to achieve detailed cytogenetic data including chromosome banding together with the demonstration of cytoplasmic and nuclear antigens within the same tumor cells.
DNA bending has been suggested to play a role in the regulation of gene expression, initiation of DNA-replication, site specific recombination, and DNA packaging. In the human mitochondrial DNA we have found a DNA curvature structure within the 3'-region of ther URF2 sequence in front of the L-strand origin of replication. This structure interacts specifically with a protein factor isolated from mitochondria. Based on the localization of this DNA curvature structure and the known function of such structures the data suggest a model in which this DNA signal sequence and its specific protein binding is involved in the regulatory initiation event of L-strand replication.
Expression of 16 oncogenes was investigated in a series of human meningiomas showing a normal chromosome complement or the characteristic monosomy 22 but no structural aberrations detectable by banding analysis. By dot hybridization, the only expressed sequence detected was KRAS. The expression was elevated approximately 6--8-fold in comparison to matrix tissue (meninges) and to fibroblasts of the corresponding patient. Northern blot analysis displayed the typical banding pattern and an 8--10-fold overexpression. DNA analysis did not reveal gene amplification or major rearrangements in the KRAS gene structure.
Using a biochemical approach (evaluation of esterase D activity) and recombinant DNA techniques (in situ and filter hybridization with specific DNA probes) a glioblastoma cell line with karyotypical nullisomy 13 was shown to contain several chromosome #13-specific sequences. They were assigned to a marker chromosome. This finding suggests that cytogenetic descriptions of deletions or chromosome losses will gain considerable power of the statement when supplemented by molecular analysis, particularly since, by now, DNA probes have been accumulated for all chromosomes and all their subfragments.
Differences in the structure and number of mitochondria in tumor cells were found. Using isoelectrofocusing two-dimensional polyacrylamide gel electrophoresis which allows detection of alterations in the protein pattern of tumor mitochondria, we studied both quantitative and qualitative changes in the mitochondrial protein pattern of human gastrointestinal tumors and corresponding normal matrix tissues. One low molecular protein spot was found to be quantitatively changed in the tumors. The approximate molecular weight was 21 x 10(3) daltons and the pI value 5.7.
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