Biomedical subjects

T Eckschlager

Publications and source records attributed to T Eckschlager.

At least 19 recordsLinked to original sources

PubMed2006

Expression of glutamate carboxypeptidase II in human brain.

Glutamate carboxypeptidase II (GCPII) is a transmembrane glycoprotein expressed in various tissues. When expressed in the brain it cleaves the neurotransmitter N-acetylaspartylglutamate (NAAG), yielding free glutamate. In jejunum it hydrolyzes folylpoly-gamma-glutamate, thus facilitating folate absorption. The prostate form of GCPII, known as prostate specific membrane antigen (PSMA), is an established cancer marker. The NAAG-hydrolyzing activity of GCPII has been implicated in a number of pathological conditions in which glutamate is neurotoxic (e.g. amyotrophic lateral sclerosis, Huntington's disease, Alzheimer's disease, epilepsy, schizophrenia, and stroke). Inhibition of GCPII was shown to be neuroprotective in tissue culture and in animal models. GCPII is therefore an interesting putative therapeutic target. However, only very limited and controversial data on the expression and localization of GCPII in human brain are available. Therefore, we set out to analyze the activity and expression of GCPII in various compartments of the human brain using a radiolabeled substrate of the enzyme and the novel monoclonal antibody GCP-04, which recognizes an epitope on the extracellular portion of the enzyme and is more sensitive to GCPII than to the homologous GCPIII. We show that this antibody is more sensitive in immunoblots than the widely used antibody 7E11. By Western blot, we show that there are approximately 50-300 ng of GCPII/mg of total protein in human brain, depending on the specific area. Immunohistochemical analysis revealed that astrocytes specifically express GCPII in all parts of the brain. GCPII is enzymatically active and the level of activity follows the expression pattern. Using pure recombinant GCPII and homologous GCPIII, we conclude that GCPII is responsible for the majority of overall NAAG-hydrolyzing activity in the human brain.

Aged↗

PubMed2006

Reovirus - possible therapy of cancer.

Oncolytic viruses infect, replicate in, and eventually lyse tumor cells but spare normal ones. In addition to direct lysis, a result of viral replicative cycle, viruses also mediate tumor cell destruction by inducing nonspecific and specific antitumor immunity. Some viruses express proteins that are cytotoxic to tumor cells. Viruses recognized as oncolytic agents can therefore be divided into three categories: 1/ naturally occurring viruses (e.g. Newcastle disease virus, vesicular stomatitis virus, autonomous parvoviruses, some measles virus strains, reovirus) that selectively replicate in tumor cells, in some instances owing to their relative resistance to interferon action; 2/ virus mutants in which some genes essential for replication in normal cells but evitable in cancer cells have been deleted (e.g.adenovirus ONYX 015 that replicates only in cells with defected p53 or herpes virus G207 which exacts the presence of ribonucleotide reductase); 3/ virus mutants modified by the introduction of tissue-specific transcriptional elements that drive viral genes (e.g.adenovirus CV706 that has PSA restricted expression of E1A and E1B and adenovirus adMycTK that binds selectively on myc protein). Reovirus is prevalent in the human population but not associated with any known human disease. Studies have shown that reovirus multiplicate preferentially in tumor cells with activated gene of ras family or ras-signaling pathway while sparing normal cells. Activated ras or its pathway could be found in as many as 60-80% of human malignancies. In our studies we used cell lines that demonstrably express activated ras. We showed the cytopathic effect of reovirus (serotype 3 strain Dearing) on medulloblastoma cell lines and compared it with its acting on normal human fibroblasts. Oncolytics Biotech Inc. is currently guiding three Phase I or Phase I/II Reolysin studies, and has completed two clinical studies and concluded enrolment in a third one.

Animals↗

PubMed2005

Characterization of drug-resistant neuroblastoma cell lines by comparative genomic hybridization.

Three parental neuroblastoma cell lines and nine derived lines resistant to Vincristin, Doxorubicin and Cisplatin, respectively, using CGH were studied. CGH profiles of all three parental cell lines were obtained using DNA from a healthy volunteer as reference DNA. Labeled DNA from each of the drug resistant daughter cell lines and labeled DNA from their parental sensitive cell lines were hybridized to obtain a comparison of gains and losses that accompanied the development of resistance for that particular drug. All three parental cell lines were characterized by typical findings for high risk neuroblastoma: N-myc amplification, gain of 17q, and loss of 1p36.2-36.3. Acquired drug resistance in the neuroblastoma cell lines appeared to be accompanied by a large array of DNA sequence copy number changes. The regions frequently affected in chemo-resistant cell lines included gains of 13q14.1-32, and 7q11.2-31.3, 4 q. Amplifications were seen at 7q 21.1 consistent with MDR1 amplification in UKF-NB-2 VCR, UKF-NB-3 DOXO, UKF-NB-4 VCR, and UKF-NB-4 DOXO, but not in any Cisplatin resistant line. All Cisplatin and Doxorubicin and two Vincristin resistant line (UKF-NB-2 VCR and UKF-NB-4 VCR) had a deletion of part of 19q or the whole 19 chromosome. All lines resistant to Vincristin or Doxorubicin and two Cisplatin resistant lines (UKF-NB-2 CDDP and UKF-NB-4 CDDP) had a deletion of at least part of 17q, UKF-NB-4 DOXO had deletion of the whole chromosome 17. The loss of 17q may cause chemoresistance by deletion of topoisomerase IIalpha gene. Deletion of 19 q in all but one chemo-resistant lines may influence of cytochromes P450 genes which are located on 19q13.2. Also gains of 15q 22, which were detected in UKF-NB-4 VCR, UKF-NB-2 DOXO and UKF-NB-4 DO X O, may affect other cytochromes P450 genes.

Antineoplastic Agents↗

PubMed2004

Lymphocyte subpopulations and immunoglobulin levels in Hodgkin's disease survivors.

We investigated the frequency of lymphocyte populations (CD3+ /T lymphocytes/, CD4+ /helpers/, CD8+ /suppressor and cytotoxic/, CD3- CD16+ /NK cells/, CD3+ HLA-DR+ /activated T lymphocytes/, and CD20+ /B lymphocytes/) and immunoglobulin G, A, M, and E levels in a group of two hundred twenty nine Hodgkińs disease long term survivors. The most frequent pathological findings were increased IgE levels, decreased CD3+ and CD4+ proportions, an increased CD20+ proportion and especially a low CD4/CD8 proportion. Decreased CD3+ and CD4+ and increased CD20+ proportions were more frequently found in the group with recurrent infections. IgM and IgA levels were positively correlated with plasmatic cholesterol and triacylglycerols levels. We suppose that immunological defects (increase of IgE levels, decreased T and helper lymphocytes) in Hodgkińs disease survivors are inherent and are not related to atopy. Examination of lymphocyte subpopulations may be helpful in the prediction of an increased risk of recurrent infections.

Adult↗

PubMed2003

[CD nomenclature].

Explore the source record for details and available documents.

Antigens, CD↗

PubMed2002

[Manipulation of hematopoietic stem cell grafts and their use in clinical practice].

Autologous stem cell transplantation has been successfully used in treatment of various hematological malignancies and solid tumors in children and adults. Published data have confirmed that bone marrow harvests and peripheral blood stem cell collections frequently contain a significant number of tumor cells. Contaminating tumor cells can contribute to the disease relapse in posttransplant period, so attempts are made to eliminate contaminating tumor cells from autografts. In the case of allogenic transplantation, T-lymphocytes depletion from graft decreases the risk of the graft versus host disease after transplantation. In this article we comment techniques available and method used for elimination of tumor cells. Commentary is aimed on potential benefits and risks of every method.

Bone Marrow Purging↗

PubMed2001

Detection of minimal bone marrow infiltration in patients with localized and metastatic Ewing sarcoma using RT-PCR.

Ewing sarcoma and related neoplasias are characterized by the presence of specific chromosomal translocations resulting in EWS/ETS gene rearrangements. Created EWS/ETS-oncogene fusion transcripts can be detected in up to 98% of ESFT and provide tumour-specific markers useful in diagnostics. Using RT-PCR for detection of this aberration we can reveal minimal amounts of tumour cells contaminating BM, blood or apheresis products. We have examined BM samples from 22 patients (21 newly diagnosed and one recurrent disease) with histologically confirmed ESFT for the presence of contaminating tumour cells in BM at the time of diagnosis. Sixteen patients presented with localized disease, six had distant metastases at the first presentation. Ewing sarcoma cells were detected in the BM of 5/16 (31%) patients with localized disease and 3/6 (50%) with clinically detectable metastases at diagnosis. BM smears prepared from the same aspirates evaluated by light microscopy were all negative, even in two patients with multiple bone disease. We have confirmed the high sensitivity of the RT-PCR assay for detection of minimal BM infiltration in localized and metastatic ESFT. We have found that more than a quarter of patients with localized ESFT have minimal BM infiltration. Although the clinical significance of the minimal disease detected at the molecular level remains unknown, RT-PCR evaluation may enable better stratification of patients into risk groups in the future.

Adolescent↗

PubMed2001

Significance of P-glycoprotein expression in childhood malignant tumors.

Resistance to chemotherapy significantly affects the treatment results in various cancers. Multidrug resistance caused by P-glycoprotein expression is now widely studied in human malignancies. We present the results of P-glycoprotein expression examination in 91 tumor tissue samples obtained from children treated for different malignant tumors in the Dept. of Pediatric Oncology, Prague. The correlation between the level of P-glycoprotein expression and tumor histology, clinical outcome, use of therapy, relapse rate and metastatic disease was made. P-glycoprotein expression was found significantly more frequent in soft tissue sarcomas, neuroblastomas, and hepatoblastomas, and generally in disseminated disease. On the contrary, a high expression of P-glycoprotein was not found in malignant brain tumors and nephroblastomas. The data strongly support the possibility that the percentage of P-glycoprotein expressing cells in selected tumors (soft tissue sarcomas, neuroblastomas), may have a clinical importance.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2000

Cyclin A down-regulation in TGFbeta1-arrested follicular lymphoma cells.

Transforming growth factor beta1 (TGFbeta1) induces growth arrest in many cell types, including B lymphocytes. We examined the effect of TGF on cell cycle progression of a non-Hodgkin lymphoma cell line of follicular lymphoma subtype (FL). After 48 h of TGFbeta1 (10 ng/ml) treatment, a significantly increased number of DoHH2 cells was retained in G(0)/G(1) phase. We examined the level of cell cycle components, cyclins, cyclin-dependent kinases (cdk), and their inhibitors. We found that the expression of cyclin A and p21(WAF1) molecules was primarily modulated by TGFbeta1 treatment while the expression of other regulatory components, like cyclins D, cyclin E, cdk2, cdk4, and cdk6 or p15(INK4B), p16(INK4A), and p27(KIP1) was not significantly affected. We further examined expression and activity of CREB/ATF family members to examine their roles in cyclin A inhibition. The binding activity of CREB-1 and ATF-2 to the CRE region of the cyclin A promoter was almost completely abolished due to the treatment. The total level of CREB-1, ATF-2, and ATF-3 was notably reduced. Moreover, CREB-1 was dephosphorylated due to the treatment as revealed by immunoblotting. We assume that down-regulation of cyclin A was mediated by the absence of CREB/ATF activation dimers. The profound effect on the ATF family of transcription factors indicates the complexity of TGFbeta1 action on FL B malignant cells.

Cell Cycle↗

PubMed2000

[Detection of minimal residual disease in Ewing's sarcoma using RT-PCR].

BACKGROUND: More than 90% of Ewing's sarcomas (ES) contain a fusion of the EWS and FLI-1 genes, due to the t(11;22)(q23;q12) translocation. At the molecular level, the EWS-FLI-1 rearrangement shows great diversity. Specifically, many different combinations of exons from EWS-FLI-1 encode in-frame fusion transcripts and result in differences in length and composition of the chimeric protein, which function as an oncogenic aberrant transcription factor. The finding of this translocation gives evidence for the presence of ES cells. The aim of this prospective study was to verify applicability of the RT-PCR method for the detection of minimal residual disease in patients with ES. METHODS AND RESULTS: Conditions for the detection of Ewing's sarcoma cells by means of the reverse-transcriptase polymerase chain reaction (RT-PCR) at fusion transcripts in peripheral blood, bone marrow (BM) and autologous hematopoietic stem cell grafts in patients with ES were appointed. 31 samples of BM, 5 samples of blood and 7 peripheral blood grafts obtained from 23 patients were investigated. Presence of tumor cells was identified in 7 BM samples from 7 different patients (31 samples from 16 patients), all the peripheral blood and graft samples were negative. CONCLUSIONS: The high sensitivity of RT-PCR method in detection of cells bearing t(11;22)(q23;q12) was demonstrated in the experimental model and clinical samples. Likewise the literary statements, the RT-PCR method was found to be more sensitive than cytology.

Adolescent↗

PubMed2000

Gene aberrations in childhood brain tumors.

We present the results of the examination of prognostic markers in 40 children suffering from brain tumors. Prognostic markers such as amplification of the N-myc and c-myc, deletion of the 17p, and DNA ploidy are indispensable factors for the determination of diagnosis. An increased number of c-myc gene copies was found in malignant brain tumors, especially embryonal, more often than reported in the literature. N-myc amplification occurs in our group seldom, but it seems to be a sign of worse prognosis in glial and embryonal brain tumors. DNA aneuploidy was not found very frequently, but in high-grade tumors only.

Adolescent↗

PubMed1999

Acyclic nucleotide analogues suppress growth and induce apoptosis in human leukemia cell lines.

Acyclic nucleotide analogues perturb DNA replication by terminating the growing DNA chain. The analogues selected for testing on human leukemia cell lines, namely 9-[2-(phosphonomethoxy)ethyl]adenine (PMEA), 9-[2-(phosphonomethoxy)ethyl]-2,6-diaminopurine (PMEDAP), and 9-[2-(phosphonomethoxy)ethyl]guanine (PMEG) exhibited growth-inhibiting activity at low concentrations, and apoptosis-inducing activity at high concentrations. A common feature was a reduction of the proportion of G1 cell cycle phase. Activities of the analogues increased in the order PMEA<PMEDAP<PMEG. The lymphoid cell line MOLT-4 was more susceptible to the agents than the myelogenous cell lines HL-60 and ML-1. In semicontinuous cultures in the presence of low-concentration PMEG the steady-state viable cell concentration was lower and the proportion of G1 phase cells was suppressed. Upon gradual removal of PMEG from the medium, the cell concentration and the DNA profile returned to values characteristic for the control culture. It is concluded that low concentrations of the analogues cause reversible slowdown of growth, due to continuous repairing of damaged DNA, while high-concentrations induce apoptosis in irreparably damaged cells.

Adenine↗

PubMed1996

Congenital-infantile fibrosarcoma: a clinicopathological study of five patients entered on the Prague children's tumor registry.

Five children with congenital-infantile fibrosarcoma are analyzed. The tumor was found at birth in four children: in one patient it was recognized at the age of 7 months. In three children the tumor affected the lower extremity. In one patient the inguinal region was the primary site, in another the abdominal wall. The morphology was that of a highly cellular spindle cell sarcoma with cells arranged in a fascicular pattern. Variations of this common pattern such as a cartwheel arrangement, and foci of small oval cells were observed. The immunohistochemistry revealed positivity of vimentin in four investigated tumors and muscle specific actin in three. Desmin, sarcomeric actin and myoglobin were all negative. There were scattered cells positive with KP1 (CD68), MAC 387, and in one case, with factor XIIIa antibodies which were considered to be reactive rather than tumor cells. The flow cytometry study showed DNA content in three tumors within diploid range; one tumor was hyperdiploid with the DNA index 1.2. Three children are disease-free from nine to 21 years after the diagnosis. One of them had the tumor preoperatively irradiated, and the subsequent histological examination revealed an almost complete tumor necrosis. In one patient there were six recurrences (treated by surgery only), and the child is well 25 months after the last recurrence. In one child the disease had an unusually aggressive course, and the patient died of widespread metastases to the lungs, lymph nodes and bones.

Abdominal Neoplasms↗

PubMed1996

Platelet membrane receptors during short cardiopulmonary bypass--a flow cytometric study.

To elucidate a mechanism of platelet dysfunction during extracorporeal circulation, we performed a study on the surface expression of platelet adhesive receptors (GPIb, GPIIb-IIIa) and activation markers (GMP140, GP53) during short cardiopulmonary bypass (CPB). Ten paediatric patients, age 6-13 years, with atrial or atrioventricular septal defects were studied. The mean CPB time was 52 min (21-110 min). During CPB, a significant drop in platelet count was observed, but not below 130 x 10(3)/microliter. The expression of platelet GPIb decreased slightly during CPB and the decrease was not significant. The decrease of GPIIb-IIIa was significant, but only in samples collected either at the end of CPB (89 +/- 13%, p < 0.05) or before leaving the operating room (74 +/- 14%, p < 0.05). The value of surface expression of platelet activation markers (GMP140, GP53) during CPB was in the range of values for resting platelets. Our results suggest that generalized CPB-induced defects of primary haemostasis are not directly connected to circulation of activated degranulated platelets or to loss of platelet adhesive receptors GPIb-IX and GPIIb-IIIa.

Adolescent↗

PubMed1996

DNA ploidy in neuroblastoma.

Neuroblastoma is perhaps the most heterogeneous childhood cancer in terms of clinical behavior. Stage of disease, age at diagnosis, levels of urinary catecholamine excretion, N-myc amplification, and DNA ploidy have been found to be significant prognostic factors. The aims of this combined retrospective-prospective study are to verify the prognostic significance of DNA ploidy and to show its correlation with other prognostic signs. Thirty six fresh and thirty three paraffin embedded samples from patients with histologically confirmed neuroblastoma (41 prior to receiving any chemotherapy) were available for flow cytometry DNA analysis. Our results showed that the maturation induced during chemotherapy could give rise to aneuploidy therefore we analyzed the associations between the DNA ploidy and other prognostic markers only in patients examined before chemotherapy. There were no significant correlations between DNA ploidy and urinary catecholamine metabolites levels or tumor localization. DNA aneuploidy was significantly more frequent in patients with lower clinical stage, lower age at diagnosis, and without N-myc gene amplification. Patients with DNA aneuploid neuroblastomas died less frequently than patients with DNA diploid tumors. There were no significant associations among the S-phase or proliferation fraction and other prognostic factors.

Catecholamines↗

PubMed1996

Comparison of fluorescent in situ hybridization (FISH) and the polymerase chain reaction (PCR) for detection of residual neuroblastoma cells.

Residual neuroblastoma (NB) cells in bone marrow or peripheral progenitor hematopoietic cells harvests may be a source of relapse after autologous transplantation in patients with high stage neuroblastoma. Therefore a sensitive method for detecting minimal residual disease (MRD) in the harvested product is important so that an appropriate purging techniques can be applied and optimized. We report on the detection of NB cells by fluorescence in situ hybridization (FISH) for N-myc amplification and compare the sensitivity of FISH with a semiquantitative polymerase chain reaction (PCR) assay. As a model of MRD we used the neuroblastoma cell line IMR-32 diluted with normal peripheral blood lymphocytes. We were able to detect a single NB cell in 1000 normal mononuclear cells by FISH. The PCR method, using ethidium-bromide-stained gels, required at least 10% NB cells to be present for detection of an amplified band of the N-myc oncogene. Thus, FISH is ten to one hundred times more sensitive in detection of N-myc amplification than a differential PCR and thus it is the method of choice for detection of MRD in NB patients.

DNA, Neoplasm↗

PubMed1995

[DNA cytometry analysis in childhood tumors].

BACKGROUND: DNA contents in cells may be determined by flow cytometry. The relationship between malignant cell aneuploidy and prognosis is known in many types of neoplasms in adults and in children. In some situations, demonstration of an aneuploid clone verifies presence of malignant cells. Aneuploidy is rare in benign diseases. This report summarizes our first experiences with cytometric DNA analysis and shows the method's abilities to other potential users. METHODS AND RESULTS: We investigated DNA contents in blood and bone marrow (BM) specimens of 25 children with leukemia, in 41 unfixed solid tumors after biopsy and in 24 specimens of paraffin embedded neuroblastoma tissue. We also investigated 5 specimens of cerebrospinal fluid (CSF) of patients with medulloblastoma, 18 specimens of CSF from patients with leukemia or lymphoma, 4 pleural exudates suspected from malignancies, and 45 specimens of possibly infiltrated BM from primary solid tumors. As the purpose of this study was to test the method on a relatively small number of specimens, we did not perform statistical analysis of our data. As reported previously, aneuploidy was frequent in CALLA + acute lymphoblastic leukemia and in types of neuroblastoma with favorable prognosis (lower clinical stages and less than 2 years of age). CONCLUSIONS: Our results show that DNA ploidy may be tested by flow cytometry in an easy and fast way. The source of the material may be unfixed tumors, deparaffinized tumors, BM, blood, CSF and pleural exudates.

Adolescent↗

PubMed1994

Renal cell carcinoma in children: a single institution's experience.

Seven patients with renal cell carcinomas (RCC) were treated at the Department of Pediatric Oncology in Prague over a period of 15 years. The average age at diagnosis was 8 years 9 months. Histology review showed five tumors to be of clear cell type with mixed tubular, papillary, and solid patterns; one tumor was purely papillary eosinophilic type and one was composed of clear and eosinophilic cells. Five patients received chemotherapy and four of them also received radiotherapy. Four patients are without evidence of disease with a minimal follow-up period of 2 years. These four were all clinical stage I or II. Two stage III patients died of metastatic disease. One patient died of tumor emboli to the lungs at operation. Tumor diagnosis and treatment of the patients in the lower clinical stages seem to have a favorable prognostic impact in childhood RCC. Combined chemotherapy with vinblastine as one of the agents, and possibly additional immunotherapy, appears to have promising results, although the number of patients treated thus far is limited.

Adolescent↗