Biomedical subjects
Publications and source records attributed to K D Zang.
Of 85 patients with ALS, the authors identified 3 patients with balanced translocations and 2 patients with pericentric inversions, all affecting distinct chromosomal loci. The high rate of constitutional aberrations (5.9%) suggests that ALS is, in part, associated with recombination-based rearrangements of genomic sequences.
In a recent study, 23 microdissected areas of 10 glioblastoma multiforme (GBM) were investigated for quantitative genomic aberrations using comparative genomic hybridization (CGH). To validate the chromosomal aberrations, as revealed by CGH after microdissection, parallel tissue sections were stained immunohistochemically with an antibody that detects both wild-type epidermal growth factor receptor (EGFR) and the deletion mutant form of the receptor (EGFRvIII). Immunostaining was correlated with CGH data of chromosome 7, because chromosome 7 is the most frequently aberrant chromosome in GBM (here four of 10 tumors), and this aberration often indicates an abnormality of EGFR. Nine of nine areas that showed gain in or amplification (2 areas) of chromosome 7 with CGH contained EGFR-immunoreactive cells. Only three of 14 areas without abnormality of chromosome 7 in CGH contained EGFR-immunoreactive cells; eleven of 14 areas were immunonegative. Our findings demonstrate a strong correlation between immunohistochemistry of EGFR and the copy numbers of chromosome 7, as revealed by CGH after microdissection in glioblastoma multiforme.
Glioblastoma multiforme (GBM) is characterized by intratumoral heterogeneity as to both histomorphology and genetic changes, displaying a wide variety of numerical chromosome aberrations the most common of which are monosomy 10 and trisomy 7. Moreover, GBM in vitro are known to have variable karyotypes within a given tumor cell culture leading to rapid karyotype evolution through a high incidence of secondary numerical chromosome aberrations. The aim of our study was to investigate to what extent this mitotic instability of glioblastoma cells is also present in vivo. We assessed the spatial distribution patterns of numerical chromosome aberrations in vivo in a series of 24 GBM using two-color in situ hybridization for chromosomes 7/10, 8/17, and 12/18 on consecutive 6-microm paraffin-embedded tissue slides. The chromosome aberration patterns were compared with the histomorphology of the investigated tumor assessed from a consecutive HE-stained section, and with the in vitro karyotype of cell cultures established from the tumors. All investigated chromosomes showed mitotic instability, i.e., numerical aberrations within significant amounts of tumor cells in a scattered distribution through the tumor tissue. As to chromosomes 10 and 17, only monosomy occurred, as to chromosome 7 only trisomy/polysomy, apparently as a result of selection in favor of the respective aberration. Conversely, chromosomes 8, 12, and 18 displayed scattered patterns of monosomy as well as trisomy within a given tumor reflecting a high mitotic error rate without selective effects. The karyotypes of the tumor cell cultures showed less variability of numerical aberrations apparently due to clonal adaptation to in vitro conditions. We conclude that glioblastoma cells in vivo are characterized by an extensive tendency to mitotic errors. The resulting clonal diversity of chromosomally aberrant cells may be an important biological constituent of the well-known ability of glioblastomas to preserve viable tumor cell clones under adaptive stress in vivo, in clinical terms to rapidly recur after antitumoral therapy including radio- or chemotherapy.
Meningioma is the most frequent tumor of neuroectodermal origin in humans. It is usually benign. Only a minority of cases shows progression to an anaplastic tumor (WHO grade II and III). Meningioma is generally a sporadic tumor. Multiple and familial cases are rare and mostly associated with (hereditary) neurofibromatosis 2 (NF2). Meningiomas show an unexpectedly high recurrence rate. Also, completely removed low-grade tumors can recur. Recurrence and multiplicity are correlated with the formation of a peritumoral edema. On the cytogenetic level, meningioma is the best-studied tumor in humans. Grade I tumors show either uniform monosomy 22 or a diploid karyotype. The majority of high-grade, but only a minority of low-grade, meningiomas show loss of merlin, a cytoskeleton-cytoplasm-linker protein. Merlin is the product of the NF2 gene located on chromosome 22. A second tumor suppressor gene on chromosome 22 has not yet been detected. In contrast to other solid tumors, progression of meningiomas is correlated with increasing hypodiploidy, showing characteristic clonal evolutions that mostly include chromosomes 14, 18, and 19 and, more rarely, 6 and 10. Structural aberrations are infrequent, except for the loss of the short arm of chromosome 1, which appears to be the decisive step for anaplastic growth. Comparative histochemical and molecular cytogenetic studies point to the alkaline phosphatase gene (ALPL, liver-bone-kidney type) located on 1p36.1-->p34 as a candidate tumor suppressor gene. A model is proposed that tries to explain - with a minimum number of essential steps - the origin, progression, infiltration, and recurrence of meningiomas.
OBJECT: The goal of this study was to determine whether in meningiomas cytogenetic findings are suitable as a predictive parameter relevant to prognosis. METHODS: Between 1992 and 1998 at the Department of Neurosurgery, Saarland University, 198 patients underwent surgery to resect meningiomas. The meningiomas were investigated cytogenetically and the patients were followed up for a mean period of 33 months. On the basis of the cytogenetic findings, the meningiomas were subdivided into four groups: Group 0 meningiomas displayed a normal diploid chromosome set; Group 1 tumors were found to have monosomy 22 as the sole cytogenetic aberration; Group 2 tumors were markedly hypodiploid meningiomas with loss of additional autosomes in addition to monosomy 22; and Group 3 meningiomas had deletions of the short arm of a chromosome 1, as well as additional chromosomal aberrations including loss of one chromosome 22. One hundred ninety-eight patients in whom tumor resections were determined to be Simpson Grade I or II could be followed up after complete tumor extirpation. In 20 patients, one or several recurrences were documented during the period of observation. The tumors were classified according to their different, but mostly uniform chromosomal aberrations. Recurrences were found in six (4.3%) of 139 tumors in Groups 0 and 1 and in two (10.5%) of 19 tumors in Group 2; the highest rate of recurrence was found in 12 (30%) of 40 tumors in Group 3. This supports the notion that the deletion of the short arm of one chromosome 1 is an important prognostic factor in meningiomas. The results of this study document a significant correlation between histological grade (p < 0.0001), location (p < 0.0001), and recurrences of meningiomas (p < 0.0001) (significance determined using chi-square tests). CONCLUSIONS: The cytogenetic classification of meningiomas provides a significant contribution to the predictability of tumor recurrence and is, therefore, a valuable criterion for the neurosurgeon's postoperative management protocol.
The tumor suppresser protein p53 is critical for guarding the genome from incorporation of damaged DNA (Lane, D. P. (1992) Nature 358, 15-16). A relevant stress that activates p53 function is UV light (Noda, A., Toma-Aiba, Y., and Fujiwara, Y. (2000) Oncogene 19, 21-31). Another well known component of the mammalian UV response is the transcription factor c-Jun (Angel, P., and Karin, M. (1991) Biochim. Biophys. Acta 1072, 129-157). We show here that upon UV irradiation p53 activates transcription of the human mismatch repair gene MSH2. Interestingly, this up-regulation critically depends on functional interaction with c-Jun. Hence, the synergistic interaction of a proto-oncogene with a tumor suppresser gene is required for the regulation of the mammalian stress response through activation of expression of MSH2.
We examined homogenized tissue samples of biopsies from 19 astrocytomas of different grades for genetic imbalances using comparative genomic hybridization (CGH): three astrocytomas grade II, and 16 astrocytomas grade IV (glioblastoma multiforme), one of the glioblastomas representing the recurrence of a benign oligoastrocytoma. In two of three cases of astrocytoma grade II, a gain of chromosome 7 was found. The alterations in the glioblastomas were complex, and most frequently showed the characteristic gain of chromosome 7 and loss of chromosome 10. The single analyzed case of recurrence of an oligoastrocytoma was characterized by a unique CGH pattern. This tumor showed two distinct alterations: apart from an amplification on 15q24q26, we found a distinct amplification of a small region on 20p11.2p12, which has not been previously described in brain tumors. Partial or complete gains of chromosome 20 arose in six other tumors; we conclude that chromosome 20 in particular 20p11. 2p12, may harbor relevant genes for glioma progression.
Calpain, also named CANP (for calcium-activated neutral protease), is an intracellular cytoplasmatic non-lysosomal cysteine endopeptidase that requires calcium ions for activity. Many substrates of the calpain isoenzymes, such as the transcription factors c-Fos and c-Jun, the tumor supressor protein p53, protein kinase C, pp60c-src and the adhesion molecule integrin, have been implicated in the pathogenesis of different human tumors, suggesting an important role of the calpains in malignant diseases. We now report differential expression of the calpain I gene (CL I) in a variety of tumors, extending our study to a larger series of renal cell carcinomas. Using Northern-blot analysis, we studied calpain I expression in 30 renal cell carcinomas as compared with matched healthy tissues. Tumor samples were classified according to their histological type: 21 clear cell carcinomas, 4 chromophobe carcinomas, 3 papillary carcinomas and 2 oncocytomas. In renal tumor samples, calpain I gene mRNA was expressed at highly variable levels, significantly depending on the different histological types. Moreover, there was a correlation of higher calpain I expression with increased malignancy: within the clear cell carcinoma subset, tumor samples with advanced nodal status (N1 and N2) showed a significantly higher calpain I expression than tumors without metastasis to regional lymph nodes. Our data suggest an important role of calpain isoenzymes in carcinogenesis and tumor progression.
SV40 and/or DNA sequences indistinguishable from SV40 have been detected in several types of human tumours. The oncoprotein of Simian virus 40, SV40 large T-antigen (Tag), is known to bind and inactivate tumour suppressor proteins, such as members of the retinoblastoma family and p53, thereby promoting cell transformation. In this study, we used the yeast two-hybrid system to investigate whether the Simian virus 40 (SV40) large T-antigen is able to interact with p73, a noval discovered putative tumour suppressor, that is homologous both structurally and functionally to p53. The yeast two-hybrid system is a genetic method to detect protein-protein-interactions in vivo. Our results suggest that the SV40 large T-antigen is not able to bind p73 in yeast although both proteins are expressed in the transformed yeast strain as was shown by western blot analysis.
Smear preparations of 23 fresh astrocytoma biopsies were analyzed by two-color fluorescence in situ hybridization with cosmids specific for the P16 and the TP53 genes. Additionally, tissue sections of the same tumors were immunostained with the use of a monoclonal antibody that recognizes both wild-type and mutant TP53 protein. In 21 astrocytomas, loss of P16 was observed in a significant proportion of cells. Cells with homozygous P16 loss were present in 13 astrocytomas; 14 astrocytomas showed cells with heterozygous loss of P16. Remarkably, 5 astrocytomas showed a scattered mosaic pattern of cells with homozygous and, respectively, heterozygous p16 loss. Homozygous deletion of TP53 was not observed. Cells with heterozygous TP53 loss were detected in 12 tumors, in 7 of them in association with P16 loss. One tumor showed aberrant cells for neither TP53 nor P16 but strong immunostaining for TP53. Positive TP53 immunostaining was found in 16 astrocytomas. Heterozygous loss of TP53 was significantly correlated with TP53 protein expression. We conclude that, unlike typical tumor suppressor genes, P16 might enhance cellular proliferation after heterozygous loss through a dosage effect and that the distribution of cells with homozygous loss of P16 speaks in favor of a polyclonal loss of the second copy of this gene.
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The term "multiforme" in glioblastoma multiforme (GBM) indicates the highly variable histomorphology that cannot be addressed by studies on homogenized tissue probes. In order to relate genetic findings with histomorphologically distinct areas we used microdissection to procure defined cell populations from microscopic tissue sections under direct visualization. Formalin-fixed and paraffin-embedded tissue sections of 10 GBM were evaluated for intratumoral genetic heterogeneity by microdissection of multiple areas of 20-50 tumor cells and DOP-PCR of DNA isolated from the dissected cell groups, followed by comparative genomic hybridization (CGH). Microdissected cells from histomorphologically normal extratumoral blood vessels from the same slides served as controls. The individual tumors showed variable combinations of primary chromosomal gains and losses common to all studied areas of a given case along with secondary, area-specific additional aberrations. CGH displayed a wider variety of chromosomal aberrations than metaphase cytogenetics of cell cultures from the same tumors. The most frequent aberrations observed were previously unperceived gains on chromosomes 4q (8/10) and 5q (5/10). Other nonrandom aberrations were gains on 12q (6/10), 13q (6/10), and 7 (5/10), and losses of 22 (5/10). Amplifications on 7p were intratumorally heterogeneous and only found in single areas of 2 tumors. In contrast to normal extratumoral vessels, vascular proliferates in most cases demonstrated chromosomal aberrations (CGH) which were partially different from the aberrations observed in the tumor itself. The described method gives evidence of considerable intratumoral genetic heterogeneity in GBM and provides a sensitive tool for the detection of quantitative chromosomal changes that are present only regionally within a given tumor.
Chordoid meningioma is a rare histological subtype of meningiomas. Cytogenetic analysis of three cases revealed the unique feature of an unbalanced translocation der(1)t(1;3)(p12-13;q11) that was ascertained by chromosome microdissection and reverse fluorescence in situ hybridization. As the t(1;3) has not been observed in other subtypes of meningioma, it may represent a specific cytogenetic marker of chordoid meningiomas. It is not yet clear whether a fusion gene or the combined loss of genes from chromosome arms 1p and 3p is the pathogenetically important outcome of the translocation.
Meningiomas are cytogenetically characterized by loss of one chromosome 22 as a typical primary aberration and progression-associated secondary chromosome changes, of which monosomy 1p is the most common. The aim of this study was to evaluate the significance of monosomy 1p and enzyme activity loss of tissue nonspecific alkaline phosphatase (ALPL), whose gene maps to chromosome 1p36.1-p34, as parameters for the diagnosis of progression-prone meningiomas. We analyzed smear preparations of 56 meningiomas and additional paraffin sections of 17 of the cases by two-color fluorescence in situ hybridization (FISH) using the D1Z1 and D1Z2 probes and by a metaphase cytogenetic analysis of 30 of these tumors. The results were compared to clinical and morphological parameters and the expression of ALPL. Smear preparations showed deletion of 1p36 in 27% of common-type, 70% of atypical (intermediate-type), and 100% of anaplastic meningiomas. Monosomy 1p, as detected by FISH or the karyotype, was strongly associated with complete loss of ALPL activity. Intermediate-type and anaplastic meningiomas of younger patients displayed an increasing rate of cells with trisomy 1q and relative loss of 1p. The highly significant correlation of FISH results and ALPL histochemistry with clinical parameters gives evidence of their strong prognostic relevance. The complete activity loss of ALPL and the immunologically detected loss of ALPL protein in areas of meningiomas with monosomy 1p indicate a cytogenetically undetectable inactivation of the homologous Alpl allele. The apparently homozygous loss of expression of ALPL supports the notion that Alpl is a candidate tumor suppressor gene in meningiomas.
By screening a meningioma expression library with autologous serum we identified four cDNA clones representing a novel gene with striking homology to Caenorhabditis elegans hyaluronidase as indicated by BLASTP analysis. In humans hyaluronidase has been implicated in cancer development and three human genes are known to encode proteins with hyaluronidase activity. None of the human genes, however, showed any homology at the nucleotide or amino acid sequence level to the newly isolated antigen we termed meningioma expressed antigen 5 (MGEA5). Somatic cell hybrid mapping and fluorescence in situ hybridization mapped the gene for MGEA5 to chromosomal band 10q24.1-q24.3. Reverse transcription (RT)-PCR and northern blot hybridization revealed expression of the gene encoding MGEA5 in several meningioma and additional human tissues. Expression analysis also indicated an alternative splicing event giving rise to a shorter and altered transcript termed MGEA5s. The expression of MGEA5 and MGEA5s as fusion proteins revealed an approximate molecular weight of 92 and 54 kDa, respectively. Using heterologous sera we found antibodies against MGEA5s in five out of 23 meningioma patients, whereas no immune reaction was detected in 12 control sera from healthy individuals. Confirmation of hyaluronidase activity was independently achieved by turbidometric analysis and a gel matrix assay. A model for involvement of the novel hyaluronidase gene in meningioma development is proposed.
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Clonal chromosomal changes were detected in three of five sporadic angiomyolipomas of the kidney irrespective of a solitary or multifocal appearance of this benign tumor type. No specific chromosomal changes have been identified. Including the cytogenetic data of the four renal angiomyolipomas reported in the literature, trisomy 7 as the single clonal chromosomal abnormality was detected in two angiomyolipomas. Because trisomy 7 has been reported in both neoplastic and nonneoplastic kidney cells, it may be assumed that trisomy 7 is already physiologically resident in renal cells but undergoes positive selection in this tumor type.
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