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M Fiegl

Publications and source records attributed to M Fiegl.

34 records · Page 2Linked to original sources

Presence of a p53 gene deletion in patients with multiple myeloma predicts for short survival after conventional-dose chemotherapy.

In multiple myeloma (MM), previous studies showed that mutations of the p53 gene are rare events in patients with newly diagnosed disease, but it is not known whether deletions of p53 are of any significance in MM. To address this question, we used interphase fluorescence in situ hybridization (FISH) with a DNA probe specific for the p53 locus at 17p13 and investigated bone marrow plasma cells from 72 patients with MM (59 patients = 81.9% before therapy). By FISH, deletions of p53, which were found to be predominantly monoallelic, were detected in 32.8% and 54.5% of patients with newly diagnosed and relapsed MM, respectively. Karyotypes from six of the patients with a p53 deletion by FISH showed a structural abnormality of 17p in only one of them. Additional FISH studies including a distal-17p probe (specific for the D17S34 locus) provided evidence for an interstitial deletion on 17p resulting in loss of p53 hybridization signals in myeloma cells. Among all 59 patients with newly diagnosed MM, presence of a p53 deletion was associated with stage III (P = .054), but not with other laboratory and clinical parameters. Patients with a p53 deletion had significantly shorter survival time compared with those without a deletion, both from the time of diagnosis (median 13.9 v 38.7 months; P < .0001) and from the time of initiation of induction treatment consisting of conventional dose chemotherapy (median 15.9 months v median not reached at 38 months; P < .0002). On stepwise multivariate regression analysis, presence of a p53 deletion was the most significant independent parameter predicting for shortened survival (P = .002). We conclude that a p53 gene deletion, which can be identified by interphase FISH in almost a third of patients with newly diagnosed MM, is a novel prognostic factor predicting for short survival of MM patients treated with conventional-dose chemotherapy.

Adult↗

Alveolar rhabdomyosarcoma in a 68-year-old patient identified by cytogenetic analysis of bone marrow.

Pancytopenia and fulminant disseminated intravascular coagulation in a 68-year-old woman suggested an acute hematologic malignancy. However, cytogenetic analysis on a bone marrow sample revealed a near-tetraploid karyotype with an isochromosome 1q and a translocation (2;13) (q35;q14), which was suggestive of an alveolar rhabdomyosarcoma (ARMS). This diagnosis was subsequently confirmed by indirect immunohistochemistry. ARMS has not yet been observed in a patient of this age. Thus, our case underlines the importance of cytogenetics, to establish an a priori unexpected tumor diagnosis.

Aged↗

Aneuploidy of chromosome 8 as detected by interphase fluorescence in situ hybridization is a recurrent finding in primary and metastatic breast cancer.

Previous work from our laboratory demonstrated aneuploidy for several chromosomes by interphase fluorescence in situ hybridization (FISH) in a high proportion of breast cancer specimens. In the literature, only limited data are available concerning chromosome 8 anomalies in breast cancer. To determine chromosome 8 ploidy status in primary and metastatic specimens from 81 breast cancer patients, FISH analysis with a DNA probe recognizing chromosome 8 centromeres was performed. In all primary tumor specimens (n = 30), significant proportions of cells were aneuploid exhibiting gain of chromosome 8 copy numbers; in 75% of effusion specimens previously classified as malignant by cytology and/or FISH for various chromosomes (n = 40), cell populations aneuploid for chromosome 8 were detected; effusions previously classified non-malignant (n = 11) were diploid in 10 cases, whereas one specimen contained rare hyperdiploid cells. Among these cells complex chromosomal aneuploidy could be demonstrated by two-color FISH, suggesting malignancy. Trisomic and tetrasomic clones were predominant in the majority of samples, but a marked intratumor cytogenetic heterogeneity was observed in most cases. Primary tumors and corresponding positive axillary lymph nodes revealed similar distributions of chromosome 8 copy numbers, analogous to previous findings with other chromosomes. This implies that, by using suitable FISH probes after examination of the respective primary tumor, an efficient search for (micro)metastasis might be feasible.

Adult↗

Chromosomal imbalances in primary and metastatic pancreatic carcinoma as detected by interphase cytogenetics: basic findings and clinical aspects.

To date, cytogenetic studies on pancreatic carcinoma are rare, and little is known about the frequency of cytogenetic aberrations in primary carcinomas compared with metastatic tumour cells. We therefore evaluated the frequency of chromosomal aberrations in 12 primary pancreatic carcinomas and in effusion specimens from 25 patients with pancreatic cancer by using interphase fluorescence in situ hybridization (FISH) and a panel of four centromeric probes. Hyperdiploidy and chromosomal imbalances, predominantly affecting chromosome 8, were a constant finding in metastatic effusion cells, whereas concordant gain of chromosomes or relative loss of chromosome 18 characterized primary pancreatic carcinomas. The potential role of oncogenes located on chromosome 8 for pancreatic cancer progression was further investigated by double-hybridization studies of aneuploid effusion cells with a probe to 8q24 (MYC) and a centromeric probe to chromosome 8, which demonstrated amplification of the MYC oncogene in two of ten cases (20%). Finally, a potential application of basic findings in the clinical setting was tested by searching for micrometastatic cells in effusions from pancreatic cancer patients primarily negative by FISH. Two-colour FISH in combination with extensive screening (>10,000 nuclei) seems to be a useful tool to unequivocally identify micrometastatic cells by demonstrating hyperdiploidy and intranuclear chromosomal heterogeneity.

Ascites↗

Interphase fluorescence in situ hybridization improves the detection of malignant cells in effusions from breast cancer patients.

In diagnostic evaluation of effusions, difficulties are encountered when atypical reactive mesothelial cells have to be differentiated from malignant cells. We tested the impact of fluorescence in situ hybridization (FISH) to identify metastatic cells in breast cancer effusions by detection of numerical chromosomal changes. Pleural and ascitic fluid samples (n=57) from 41 breast cancer patients were concomitantly evaluated by routine cytology and FISH, using centromere-specific probes representing chromosomes 7, 11, 12, 17 and 18. After setting stringent cut-off levels deduced from non-malignant control effusions (n=9), the rates of cells with true aneuploidy were determined in each effusion sample from breast cancer patients. The occurrence of aneuploid cells, as detected by FISH and indicative of malignancy, was correlated with the cytological findings. Routine cytology revealed malignancy in 60% of effusions. Using FISH, aneuploid cell populations could be observed in 94% of cytologically positive and in 48% of cytologically negative effusions, thus reverting diagnosis to malignancy. To confirm malignancy in cases with a low frequency of aneuploid cells, two-colour FISH was additionally performed and indeed showed heterogeneous chromosomal aneuploidy within single nuclei. We conclude that FISH is a valuable tool in the diagnosis of malignancy and may serve as an adjunct to routine cytological examination, as demonstrated here for breast cancer effusions.

Adult↗

Involvement of P-glycoprotein in the transmembrane transport of interleukin-2 (IL-2), IL-4, and interferon-gamma in normal human T lymphocytes.

The physiological role of the multidrug resistance P-glycoprotein (P-gp), which is expressed by normal human T lymphocytes, is still largely unknown. To investigate whether or not P-gp is involved in the transport of cytokines, peripheral blood lymphocytes were stimulated with phytohemagglutinin (PHA) in the absence or presence of P-gp inhibitors, and concentrations of cytokines (interleukin-2 [IL-2], IL-4, IL-6, interferon-gamma [IFN-gamma]) in the supernatants of these cultures were quantitated by enzyme-linked immunosorbent assay. P-gp inhibitors included verapamil (Ver), tamoxifen (Tmx), and the P-gp specific monoclonal antibody UIC2. Release of IL-2 was significantly suppressed by these inhibitors at concentrations that were also effective in blocking efflux of Rhodamine-123 from normal T lymphocytes. IL-2 mRNA expression in lymphocytes was not different between PHA control and the cultures with P-gp inhibitors. Ver and Tmx did not interfere with T-cell activation as determined by CD25 and CD69 expression. In a nonhematological model, the P-gp expressing HCT-8 adenocarcinoma cell line, exogenously added IL-2 was shown to exert an inhibitory effect on P-gp mediated Rhodamine-123 efflux. In addition, transepithelial transport of IL-2 by electrophysiologically tight and polarized HCT-8 monolayers was examined. A time-dependent flux of IL-2 across dense monolayers, which was partially inhibited by Ver, was observed. We also investigated whether or not P-gp inhibitors suppressed release of other cytokines produced by activated T cells (IL-4, IL-6, IFN-gamma). Release of IL-4 and IFN-gamma was significantly inhibited by Ver, Tmx, and UIC2; however, release of IL-6 remained unaffected. These data show P-gp mediated transmembrane flux of IL-2 in T lymphocytes and HCT-8 cells. We conclude that P-gp participates in the transport of cytokines (IL-2, IL-4, and IFN-gamma) in normal peripheral T lymphocytes.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

[Molecular biology of tobacco smoke associated neoplasia].

Adducts, formed by carcinogens of tobacco smoke with DNA, can be detected by means of molecular techniques and are used as marker of internal exposure. Carcinogen-DNA adducts produce specific mutations in tumor-suppressor genes (e.g. p53) and oncogenes (e.g. ras), which can be involved in tumor initiation or in later stages of tumor progression (e.g. evolution of an invasive phenotype). Benzo(a)-pyrene, an important carcinogen of tobacco smoke, induces GT transversions, as demonstrated in in vitro systems and animal models. Mutations in the p53- or ras-gene are more common in human tumors of the lung, head and neck, bladder and pancreas in smokers than in non-smokers. Molecular biology of cancer gains increasing significance in clinical practice since 1.) the presence of certain mutations confers an unfavorable prognosis to malignant disease (e.g. ras mutations in lung cancer), 2.) ras and p53 mutations often occur early during tumor development and can thus facilitate diagnosis of malignant disease, and 3.) minimal residual disease can be detected using molecular techniques. After resection of cancer of the head and neck, tumor recurred more frequently in patients with no evidence of residual disease as assessed by pathohistologic criteria than in patients with no evidence of residual disease as evaluated by p53 immunostaining.

Carcinogens↗

Interphase fluorescence in situ hybridization identifies chromosomal abnormalities in plasma cells from patients with monoclonal gammopathy of undetermined significance.

Karyotypic studies in patients with monoclonal gammopathy of undetermined significance (MGUS) have been hampered by a low percentage of bone marrow plasma cells (BMPC), which are predominantly nonproliferating. By combining cytomorphology and interphase fluorescence in situ hybridization (FISH) we investigated whether or not chromosomal abnormalities occur in BMPC from patients with MGUS. Studying chromosomes 3, 7, 11, and 18, which we found to be frequently aneuploid by FISH in multiple myeloma (MM), we observed three hybridization signals for one of these chromosomes 3 were most common, occurring in 38.9% of patients, followed by gains of chromosomes 11 (25%), 7 (16.7%), and 18 (5.6%) Among BMPC, the frequency of aneuploid cells was 18.9% +/- 13.9% (mean +/- SD) for chromosome 3, 22.3% +/- 9.2% for chromosome 11, 23.2% +/- 22.0% for chromosome 7, and 6.1% +/- 2.3% for chromosome 18. In five patients, chromosomal abnormalities were shown to be restricted to BMPC expressing cytoplasmic immunoglobulins corresponding to the serum paraprotein. No gain of hybridization signals was observed in normal and reactive plasma cells. In one patient with MGUS, metaphase cytogenetics revealed one abnormal metaphase with 47, XY, +4, and trisomy 4 was also demonstrated in a subpopulation of BMPC by interphase FISH. FISH results from patients with MGUS and newly diagnosed MM at stage IA (n = 14) indicated that aberrations involving > or = 2 chromosomes occurred significantly more often in early stage MM (P < .01). With respect to clinical and laboratory features, MGUS patients with and without chromosomal abnormalities were indistinguishable. Our results indicate that MGUS already has the chromosomal characteristics of a plasma cell malignancy.

Adult↗

Multiple myeloma: high incidence of chromosomal aneuploidy as detected by interphase fluorescence in situ hybridization.

Because metaphase cytogenetic studies in multiple myeloma (MM) are hampered by a low proliferative activity of myeloma cells in vitro, interphase cytogenetics by means of fluorescence in situ hybridization (FISH) should improve the detection of chromosomal abnormalities in MM. We therefore investigated chromosomal aneuploidy in 36 patients with MM using interphase FISH and alpha-satellite DNA probes for chromosomes 1, 3, 7, 8, 11, 12, 16, 17, 18, and X. By FISH, myeloma cells from 32 patients (88.9%) were aneuploid for at least one of the chromosomes examined. In 24 patients (66%), aberrations of > or = 3 chromosomes were observed. Aneuploidy was predominantly characterized by a gain of chromosome numbers, with involvement of chromosomes 3, 7, and 11 occurring in > 50% of patients. Loss of a centromeric signal suggesting monosomy was most frequently observed for chromosomes 17 (22.2% of patients) and X (monosomic in 42.3% of female patients, but loss of chromosome X was never observed in males, P < 0.05). Dual-color FISH studies provided evidence for marked heterogeneity of aneuploid cells in 8 patients (22.8%). Occurrence of chromosomal aneuploidy was independent of stage and pretreatment status. Gain of chromosome 3 was significantly correlated with an IgA paraprotein (P < 0.05). In 12 patients, the direct comparison of metaphase cytogenetics and FISH showed that FISH detected aneuploidy of chromosomes in 9 patients that was missed by metaphase analysis. In conclusion, interphase FISH, by which chromosomal aneuploidy was detected in almost 90% of patients with MM, represents an approach for evaluating the clinical significance of specific chromosomal abnormalities in MM.

Adult↗

Splice site mutation in the glucocorticoid receptor gene causes resistance to glucocorticoid-induced apoptosis in a human acute leukemic cell line.

Induction of apoptosis is the molecular basis for the therapeutic application of glucocorticoids (GC) in human leukemia. The beneficial effect of endocrine therapy is, however, hampered by the occurrence of resistant clones evolving under selective GC pressure. To delineate molecular mechanisms of GC resistance, we PCR amplified, cloned, and sequenced GC receptor (GR) transcripts and gene segments from a GC-resistant subclone of the human acute leukemic cell line CCRF-CEM, termed CEM-R6. Our analyses revealed that one GR gene allele harbored a point mutation (L753F) previously shown to compromise GR functions in other CCRF-CEM derivatives. On the second allele, we identified an A to G point mutation in the 3'-splice junction of intron G. As a consequence, a cryptic splice site 8 base pairs downstream within exon 8 is recognized, which leads to an 8-base deletion in the GR mRNA, resulting in reading frame shift and 2 consecutive in-frame preterminal stop codons. Translation of this mutant mRNA would produce a truncated GR protein missing 93 amino acids of the ligand-binding domain and expressing 9 altered residues at its new COOH terminus. In concert with the L753F mutation on the other allele, this molecular defect explains the GC-resistant phenotype and provides further evidence for mutational GR gene inactivation as a mechanism for human leukemic cells to escape GC-induced apoptosis.

Amino Acid Sequence↗

Interphase cytogenetics reveals a high incidence of aneuploidy and intra-tumour heterogeneity in breast cancer.

The occurrence of aberrations involving chromosomes 11 and 17 in malignant tissues of breast cancer patients has not yet been studied systematically. Using fluorescence in situ hybridisation (FISH) with centromere-specific probes, we determined chromosome 11 and 17 status in interphase nuclei from primary and/or metastatic breast cancer cells. In all cancerous specimens obtained from 30 patients, FISH identified cells with clonal chromosomal abnormalities, with aneuploidy rates ranging from 6% to 92% (median 59%). There was a gain of centromeric signals for chromosome 11, most likely corresponding to hyperploidy; aberrations of chromosome 17 in specimens from 26 patients (87%) were hyperploid as well; however, four cases (13%) showed loss of chromosome 17 centromeres. All specimens contained heterogeneous aneuploid cell populations with excessive gain of signals in some cases. The pattern of aneuploidy did not appear to correlate with tumour grade/stage and was comparable in primary tumours and corresponding metastatic axillary lymph nodes, indicative of genetic instability early in tumour development. Screening with a panel of FISH probes may lead to enhanced sensitivity and specificity in detecting malignant cells, as demonstrated in this study with effusions which could not be conclusively interpreted as being malignant by cytological criteria.

Adult↗

Glucocorticoid-mediated immunomodulation: hydrocortisone enhances immunosuppressive endogenous retroviral protein (p15E) expression in mouse immune cells.

To define glucocorticoid (GC)-regulated genes contributing to the anti-inflammatory and immunosuppressive effects of GC, previous work from our laboratory revealed up-regulation of transcripts from endogenous type B mouse mammary tumour virus (Mtv) and type C murine leukaemia virus (Emv) loci by high dose GC treatment of P388D1 macrophage-like cells. This study demonstrates enhancement of expression from Mtv and Emv loci in P388D1 cells by more physiological hydrocortisone concentrations (1 microM), and shows direct transcriptional mode of regulation by blocking GC-mediated signal transduction at different levels. Furthermore, we found up-regulation of Emv mRNA steady-state levels in murine lymphoid lineage cells (T-like EL4 and BW5147 cells; B-like X63 cells) upon GC treatment. The Emv transcripts shown by us to be GC-up-regulated encode for the transmembrane envelope protein TM/p15E which is highly conserved in several retroviruses. TM/p15E and the p15E-like products found in humans exert immunosuppressive effects in different test systems. Thus, our findings raise the possibility that immunomodulation by GC might be mediated in part by enhanced expression of p15E(-like) products.

Adjuvants, Immunologic↗

Ulcerative breast cancer: case report and review of management.

In the literature, there is an astonishingly small amount of information on specific treatment modalities of locally advanced, ulcerated breast cancer. Here we present a case report and a literature review on ulceration in breast cancer. An older, so far untreated woman with complete ulcerative destruction of her right breast was inoperable because of the extent of the tumor. Primary anthracycline-based chemotherapy and hormonal therapy led to major tumor shrinkage with complete ulcer healing, which was maintained for nearly 2 years. There is increasing evidence from the literature that primary chemotherapy may particularly benefit patients with inoperable ulceration, as also illustrated by our case.

Aged↗