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Biomedical subjects

W Fiedler

Publications and source records attributed to W Fiedler.

At least 55 records · Page 3Linked to original sources
PubMed1999

P53 genotyping - an effective concept for molecular testing of head and neck cancer?

P53 mutations are currently recognized as the most common genetic alteration in human tumors. The purpose of our study was to evaluate the significance and reliability of p53 genotyping in head and neck cancer as a possible marker permitting the prediction of tumor behavior and clinical outcome. P53 genotyping in our study refers to highly sensitive molecular screening in order to detect structural alterations in the nucleic acid sequence of the gene. Exons 2-11 and adjacent intronic regions were screened for mutations by direct genomic sequencing or by bi-directional dideoxyfingerprinting in 66 primary tumors of the larynx, pharynx and oral cavity. Alterations in the of the p53 gene were detected in 36% (24 of 66) of the analyzed tumors, no mutation was found in our cohort outside exons 5-8. The frequency of p53 mutation had no correlation to the tumor stage or tumor site. The recurrence rate in patients with a p53 alteration was not significantly higher compared to patients without a p53 mutation in their primary tumors. Summarizing the results of our study only limited reliability of p53 genotyping as an effective concept for molecular testing of head and neck cancer was found.

Biomarkers, Tumor↗
PubMed1999

Determination of telomerase activity in multifocal renal cell carcinoma.

In a consecutive series of 245 patients with renal cell carcinoma (RCC) all nephrectomy specimens were examined regarding the presence of additional multifocal lesions. Therefore, 3-mm step sections were performed and investigated by macroscopical and histopathological examination. Twenty-six multifocal tumors were found in 19 specimens by this procedure. In order to characterize the biological activity of these multifocal lesions in more detail, their telomerase activity was analyzed. Fourteen of the 23 investigated multifocal tumors (60.9%) displayed telomerase activity. In three cases the tissue probes were too small for further investigations. Telomerase activity was also detected in 10 out of 15 (66.7%) matched primary renal cell carcinomas. In contrast, all corresponding normal kidney cortex tissues used as control were telomerase negative. From our results we conclude, that small tumor lesions of RCC must be expected to have a malignant potential similar to that of primary carcinomas. Hence, nephron-sparing surgery cannot be routinely carried out even in patients with small tumors.

Carcinoma, Renal Cell↗
PubMed1999

Molecular cytogenetic characterization of a prenatally detected supernumerary minute marker chromosome 8.

The characterization of a prenatally detected very small (approximately half of 18p-(karyotype: 47,XX,+mar[16]/46,XX[7]) supernumerary marker chromosome (SMC) identified by GTG-banding analysis is described. The marker has been identified as derived from chromosome 8 centromeric material using a combination of different cytogenetic (GTG-, NOR-, CBG banding), molecular cytogenetic (24 colour-fluorescent in situ hybridization [FISH], three-colour FISH using centromeric probes for all human chromosomes) and molecular genetic techniques (microsatellite analysis). This is the first case described with such a minute SMC derived from chromosome 8 diagnosed prenatally, the 15th case reporting on a SMC originating from chromosome 8 and the third such case without any severe clinical features.

Adult↗
PubMed1998

Functional expression and localization of vascular endothelial growth factor and its receptors in the human epididymis.

Blood supply is essential for the maintenance of epididymal function. Since there is no considerable neovascularization in the epididymis, this tissue could represent a suitable model to study the vascular endothelial growth factor (VEGF) effect for vascular permeability. We studied the expression and function of VEGF and its receptors fms-like tyrosine kinase (Flt-1) and fetal liver kinase (designated as kinase insert domain-containing receptor, KDR in the human) in the human epididymis. VEGF and VEGF receptors mRNA were detected in the human epididymal tissue. VEGF protein was localized in peritubular and in ciliated cells of efferent ducts as well as in peritubular and basal cells of the epididymal duct. Vascular endothelial cells did not express VEGF. Flt-1 protein was localized in ciliated cells of efferent ducts and in lymphatic vessels. Vascular endothelial cells were negative for Flt-1 but positive for KDR. In vitro VEGF165 treatment of epididymal tissue induced endothelial fenestrations and opening of interendothelial junctions. Additionally, we observed for the first time that VEGF could induce transendothelial gaps. We conclude that these gaps might be of importance not only for molecular transport but also for cell passage across the vessel wall, which may be significant for tumor metastasis. VEGF may act as a paracrine effector to influence the permeability of lymphatic vessels via Flt-1, and of blood vessels via KDR.

Aged↗
PubMed1998

Gain of DNA copy number on chromosomes 3q26-qter and 5p14-pter is a frequent finding in head and neck squamous cell carcinomas.

Comparative genomic hybridization (CGH) was used to study genomic imbalances in 77 squamous cell carcinomas of the head and neck (HNSCC) and in four cell lines derived from oral carcinomas. Particular attention was paid to all tumors characterized by a gain of two specific chromosomal segments, i.e. 3q26-qter and 5p14-p15. The 57 tumors containing both or one of the two imbalances were compared with 20 tumors lacking both with regard to genomic complexity, as well as to associated genomic, histopathologic and clinical peculiarities. 60% of the oral, and 66% of the non-oral cancers exhibited a gain of 3q26-qter, while a gain of 5p14-p15 was found in 66% of the oral, but only in 48% of the non-oral tumors. 48% of all tumors were characterized by both gains together, 26% exhibited only one of the two alterations. It could be shown that presence of both, gain of 3q26-qter and 5p14-p15, was clearly associated with a significantly higher complexity of genomic changes which was not only expressed as a high frequency of DNA copy number alterations (CNAs) but was also connected with a considerable number of additional amplifications of various chromosomal segments and a high conformity of the patterns of genomic imbalances in these tumors. Clear differences of the extent and of the patterns of genomic imbalance could be observed between oral and non-oral tumors. With respect to histopathological parameters, no clear association could be found for specific imbalances to the grade of differentiation, nor the invasiveness or metastatic status of the tumors. However, a higher number of patients with tumors characterized by gain of 3q26-qter plus 5p14-p15 died within a short period (i.e. less than 15 months) after excision of the tumor compared to the group without these imbalances. The implications of these findings are discussed from the oncogenetic and clinical aspects and in comparison with other reports.

Journal Article↗
PubMed1998

Pattern of genomic imbalances in oral squamous cell carcinomas with and without an increased copy number of 11q13.

Among 23 squamous cell carcinomas (SCC) of the oral cavity which were screened for DNA copy number alterations (CNAs) using comparative genomic hybridization, 14 showed a gain of, and 5 of these 14 even an amplification of band 11q13. Amplification of 11q13 was also detected in three of the four studied SCC cell lines and was confirmed by interphase FISH. The number of CNAs in addition to 11q13 varied from 14 to 47 in these carcinomas. All these tumors had seven other specific CNAs in common, i.e. gain on 1p36.3-36.6, 5p15, 9q34, 12p12-13, 14q32, 19 and 20q, all but one showed also an increase of copy number in 7p22, 8q24, 10q26, 12q26, 15q24-25, 16p, 16q23-24, 17q and 22q12-qter. These imbalances were distinctly rarer in the tumors without CNA in 11q13. Loss of material apparently played a minor role in these tumors with gain of 11q13, the most frequent losses (3p12-14 and 5q21) being present in 10 of the 14 cases and loss of 9p13-21 in 5/14 tumors. The three tumors with the highest number of CNAs in addition to 11q13, were histologically classified as pT4, three of the five tumors with 11q13 amplification were highly node-positive (pN 2b-2c). Two of the pT4 tumors shared as many as 23 specific chromosomal segments affected by CNA. Thus, gain of 11q13, though being found at different stages of karyotypic evolution, is apparently associated with a rather specific pattern of other CNAs and involved in progressed stages of malignancy in oral squamous cell carcinoma. In addition, the proportion of patients deceased within one year after diagnosis was clearly higher in the group whose tumors showed an increased 11q13 copy number as compared to the group without this increase. This could point to an association of gain in 11q13 and aggressiveness of the respective tumor.

Aged↗
PubMed1997

Vascular endothelial growth factor and its receptors in normal human testicular tissue.

Expression of vascular endothelial growth factor (VEGF), also known as vascular permeability factor (VPF), and its receptors Flt-1 and KDR (Flk-1 in mouse) and their localization in the human testis were analyzed by means of reverse transcriptase-polymerase chain reaction (RT-PCR), Western blotting and immunohistochemistry. VEGF mRNA was detected in the human testicular tissue and in fragments of seminiferous tubules by means of RT-PCR, while fragments of blood vessels isolated from testes were negative. Western blotting procedure using a specific VEGF antibody, revealed two protein bands corresponding to 24 and 49 kDa in the extracts prepared from the whole testis and in the seminiferous tubules while no such bands were found in isolated fragments of human testicular blood vessels. Also immunohistochemically, human testicular blood vessels show no VEGF immunoreactivity, while Leydig cells and Sertoli cells were positive. The mRNA of the VEGF receptor Flt-1 was found to be expressed in human testicular tissue, in isolated fragments of testicular blood vessels and in seminiferous tubules as determined by RT-PCR procedure. In accordance with these results, the Flt-1 protein was immunohistochemically localized in Leydig, Sertoli and perivascular cells. Endothelial cells of certain segments of human testicular microvasculature also stained positive for Flt-1. Expression of VEGF receptor, KDR, could be demonstrated in human testicular tissue, in isolated seminiferous tubules and in isolated fragments of human testicular blood vessels by means of RT-PCR. Immunohistochemically, the KDR protein was localized in endothelial cells and perivascular cells of capillaries within the lamina propria of seminiferous tubules. Leydig cells and Sertoli cells show KDR immunoreactivity, too. Thus we demonstrate the presence of both types of VEGF receptors Flt-1 and KDR on Leydig as well as on Sertoli cells which are normal non-endothelial cells, suggesting hitherto unrecognized and novel functions for such receptors. The results obtained permit us to suggest VEGF as a paracrine mitogenic and angiogenic factor, responsible for modulating the capillarization of the human testicular tissue and maintaining the functions of testicular microvasculature. VEGF may also influence the permeability of capillaries passing through the groups of Leydig cells and those localized within the lamina propria of human seminiferous tubules. The differences in the expression pattern of the VEGF receptors in the human testicular tissue probably reflect different VEGF effects in different compartments of human testis.

Blotting, Western↗
PubMed1997

Vascular endothelial growth factor, a possible paracrine growth factor in human acute myeloid leukemia.

Vascular endothelial growth factor (VEGF) is a multifunctional cytokine involved in angiogenesis, inflammation, and wound healing. It is secreted by a variety of tumor cell lines, including hematopoietic lines. Therefore, we investigated expression of VEGF and its receptors on fresh leukemic blasts. VEGF-specific transcripts were detected by polymerase chain reaction (PCR) in 20 of 28 patients with de novo acute myeloid leukemia (AML) and in 3 of 5 patients with secondary AML. Using immunocytochemistry, we found VEGF protein in 2 leukemic cell lines and in 8 AML patients, in concordance with PCR results. Supernatants of fresh leukemic cells from 24 AML patients contained significantly more VEGF than supernatants from bone marrow cells of 9 normal donors or of CD34-enriched cells from 3 normal volunteer donors as determined by an enzyme-linked immunosorbent assay. VEGF possesses two high-affinity receptors, KDR and FLT1. Using a sensitive nested PCR assay, we detected expression of FLT1 in 10 of 20 patients with de novo AML and 3 of 5 patients with secondary AML. KDR was expressed in 4 of 22 patients with de novo AML and 1 of 4 with secondary AML. To study possible paracrine growth stimulation of AML blasts, endothelial cells from human umbilical cords were incubated with increasing concentrations of VEGF. A dose-dependent increase of granulocyte-macrophage colony-stimulating factor secretion from endothelial cells was identified.

Acute Disease↗
PubMed1997

[Telomeres and telomerase].

Complex mechanisms have evolved in mammalian cells for regulating cellular lifespan. Normal cells demonstrate a strictly limited growth potential and senescence after a defined number of cell divisions. In contrast, tumor cells often exhibit an apparently unlimited proliferation potential and are termed immortalized. It has been proposed that the progressive shortening of the tips of the eukaryotic chromosomes--the telomeres--is an important component of senescence and is involved in the control of cell cycle. The enzyme telomerase adds TTAGGG repeats onto mammalian telomeres, preventing their shortening. Telomerase is normally inactive in most somatic cells, but detectable in tumor cells. The activation of telomerase in malignant cancers seems to be an important step in tumorigenesis in order to gain the ability of indefinite proliferation and to become immortal. This review describes the present knowledge of telomeres and telomerase and their role in cellular senescence and human aging. It summarizes aspects of telomerase in cancer and its function as a diagnostic and prognostic tumor marker.

Animals↗
PubMed1997

Antimycotic therapy with liposomal amphotericin-B for patients undergoing bone marrow or peripheral blood stem cell transplantation.

Suspected deep or systemic mycosis in patients undergoing high-dose therapy and autologous or allogeneic bone marrow transplantation (BMT) requires an immediate systemic antimycotic therapy. Intravenous therapy with the standard drug conventional amphotericin-B is associated with severe adverse effects like nephrotoxicity and chills. Furthermore, BMT patients often receive other potential nephrotoxic drugs such as CsA or virustatics. In this study, we report 74 BMT-patients treated with liposomal amphotericin-B for culture-documented aspergillosis (n = 5) or candidiasis (n = 6), or for serologically (n = 35) or clinically suspected mycosis or as prophylaxis (n = 2). Therapy was initiated with a median dose of 2.8 (0.64-5.09) mg/kg body-weight and continued for 13 (1-55) days. The drug was excellently tolerated and only in one was therapy stopped due to severe chills and fever. Severe organ impairment was not observed under therapy with liposomal amphotericin-B. Creatinine decreased in five patients after an increase under preceding therapy with the conventional formulation. Influence of liposomal amphotericin-B on bilirubin and transaminases was difficult to evaluate due to therapy-related toxicity, veno-occlusive disease (VOD), and graft-versus-host disease (GvHD). 10/11 culture-positive patients died from aspergillosis (5/5) or candidiasis (5/6), but in 9/11 of these subjects the immunity was additionally compromised by GvHD, steroid therapy, and VOD. Liposomal amphotericin-B was effective in preventing relapse of systemic mycosis in 10/12 patients with a history of aspergillosis (n = 11) or candidiasis (n = 1). We conclude, that favourable toxicity of liposomal amphotericin-B should encourage dose escalation studies of liposomal amphotericin-B randomised against the conventional formulation and that the comparison of patients undergoing BMT with patients under standard chemotherapy might be difficult because of additional risk factors of the BMT-patients.

Adolescent↗
PubMed1997

High dose chemotherapy with busulfan, cyclophosphamide, and etoposide as conditioning regimen for allogeneic bone marrow transplantation for patients with acute myeloid leukemia in first complete remission.

We explored the combination of busulfan/cyclophosphamide/etoposide as conditioning regimen prior to bone marrow transplantation in 31 patients with acute myeloid leukemia (AML) in first complete remission. The preparative regimen consisted of 16 mg/kg busulfan, 30-60 mg/kg VP-16, and 120 mg/kg cyclophosphamide. With a median follow-up of 30.5 months (range, 5-60 months), 25 patients are alive in continuous complete remission. Estimated disease-free survival at 5 years is 80.5%. Death was due to transplant-related toxicity (graft-versus-host disease and cytomegalovirus infection, graft-versus-host disease and pneumonia, sepsis and mucositis, respectively). None of the patients have relapsed. As demonstrated by the results of this analysis, the conditioning regimen busulfan/cyclophosphamide/etoposide is effective and well tolerated in patients with AML in first complete remission. Main nonhematological toxicities were mucositis and hepatotoxicity. The low mortality and relapse rate appears to justify allogeneic bone marrow transplantation for patients with AML in first complete remission who have an HLA-identical donor. Whether this regimen offers a substantial improvement in disease-free and overall survival over presently used regimens warrants further investigation.

Acute Disease↗
PubMed1997

Telomeres and telomerase: biological and clinical importance.

We review the present knowledge of telomeres and telomerase with special attention to their role in cell proliferation, cellular senescence, and human aging. We summarize the functional aspects of telomerase in cancer, as well as its role as a useful diagnostic and prognostic tumor marker, and discuss possible approaches to telomerase inhibition as a target for cancer therapy.

Aging↗
PubMed1996

Mass spectrometric mapping of protein epitope structures of myocardial infarct markers myoglobin and troponin T.

Monoclonal antibodies are widely used analytical tools in biochemical research. The knowledge of their corresponding epitopes is of major interest. One possible approach for epitope characterization is the application of protein antigen proteolysis in combination with mass spectrometric peptide mapping analysis. Two complementary analytical strategies were applied: (a) limited proteolysis of antibody-bound antigen followed by removal of nonbound peptides and detachment of the antigenic peptides (epitope excision) and (b) enzymatic digest of the antigen followed by extraction of the antigenic peptides with the antibody and detachment of antigenic peptides after removal of nonbinding fragments (epitope extraction). In the few examples published so far, immobilized antibodies were used for these studies. In this study we present a method for characterization of the epitope sequences without prior immobilization of the monoclonal antibody. The separation of nonepitope peptides from antibody-bound peptides was carried out by ultrafiltration. The epitope and nonepitope fractions were analyzed by MALDI-MS without further purification, and the epitope sequences were identified. The method was developed using a model system consisting of the synthetic C-terminal cyanogen bromide fragment CB3 of myoglobin and the commercial monoclonal anti-myoglobin MG1. In further investigations the epitope sequence of a synthetic 32 amino acid peptide derived from heart muscle protein troponin T toward a monoclonal antibody MAb-M7, which was raised against the intact protein, was characterized. With this approach the epitope binding site of this antibody was determined, and selective shielding of potential cleavage sites in the immune complex could be observed. Furthermore, statements about the three-dimensional structure of the bound antigen were made.

Amino Acid Sequence↗
PubMed1996

[Immunosuppressive treatment of a spontaneous inhibitor hemophilia A using cyclophosphamide, vincristine and prednisone following prior Factor VIII stimulation].

A 59-year-old patient who presented with hematuria and recurrent soft tissue bleeding was found to have a factor VIII inhibitor level of 52 Bethesda units (BU)/ml and acquired hemophilia was diagnosed. After treatment with immunoglobulins (0.4 g IgG/kg per day for one week) the factor VIII inhibitor titer decreased to 12 BU/ml. Because of another episode of retroperitoneal hemorrhage, the patient was put on an immunosuppressive combination therapy which was first described by Lian et al. (1988). Our patient was infused with a factor VIII concentrate followed by cyclophosphamide, vincristine and prednisone. This regimen was repeated every 3-4 weeks. After 6 courses a further decline in the factor VIII inhibitor concentration, but no complete eradication of the autoantibody, was achieved. The factor VIII inhibitor level has remained at 2.5 BU/ml for more than 7 weeks without further bleeding episodes. The pathophysiology and treatment of acquired hemophilia are discussed.

Autoantibodies↗
PubMed1996

Lactic acidosis and hypoglycemia in a patient with high-grade non-Hodgkin's lymphoma and elevated circulating TNF-alpha.

A 71-year-old patient with high-grade non-Hodgkin's lymphoma stage IVB, severe lactic acidosis and tumor-associated hypoglycemia is described. Endocrine causes of hypoglycemic episodes were excluded because of low serum concentrations of insulin and "insulin-like growth factor 1", and normal concentrations of growth hormone and thyroid hormone. Clinical conditions associated with lactic acidosis such as diabetes mellitus, biguanide intoxication, septicemia, acute hypoxemia, or circulatory insufficiency were ruled out. Enhanced glucose metabolism within the tumor was visualized by positron emission tomography employing 2-fluro-2-deoxy-D-glucose (FDG) as a tracer. A markedly elevated tumor necrosis factor-alpha (TNF-alpha) level was found which decreased after cytoreductive therapy paralleling the normalization of serum lactate. In contrast to the majority of cases of lymphoma-associated lactic acidoses reviewed to date, in our case lactate elimination was not reduced.

Acidosis, Lactic↗
PubMed1996

Philadelphia chromosome-positive secondary acute myeloid leukemia following high-dose chemotherapy with peripheral blood progenitor cell support for relapsed low-grade non-Hodgkin's lymphoma.

We report the case of a 47-year-old patient who developed acute myelogenous leukemia (AML) 18 months after receiving high-dose chemotherapy with peripheral blood progenitor cell support (PBPCT) for relapsed low-grade follicular non-Hodgkin's lymphoma (NHL). Cytogenetic analysis of the leukemic cells showed the translocation (9;22)(q34;q11). In three mitoses, an additional Philadelphia chromosome (Ph1) was present. In the literature, Philadelphia chromosome-positive secondary AML has been described only once before in a patient with multiple myeloma.

Acute Disease↗