Biomedical subjects

W Siegert

Publications and source records attributed to W Siegert.

At least 55 records · Page 3Linked to original sources

PubMed2000

Successful autologous bone marrow rescue in patients who failed peripheral blood stem cell mobilization.

We assessed autologous bone marrow (BM) harvest and hematologic recovery after high-dose chemotherapy (HDCT) in patients who failed to achieve peripheral blood stem cell (PBSC) mobilization. One hundred and ninety-three patients with germ cell tumor, malignant lymphoma, sarcoma or medulloblastoma were scheduled for HDCT. In 123 patients, PBSC were mobilized by disease-specific chemotherapy plus granulocyte colony-stimulating factor (G-CSF). In 110/ 123 patients (89%) with circulating CD34+ cell counts 2 > or = 10/microl, sufficient hematopoietic autografts were collected (group A). In 13/123 patients (11%) with peripheral CD34 + cell counts < 10/microl, PBSC harvesting was not performed (group B). These latter patients were classified as "poor mobilizers" and underwent second-line BM harvest at a median of 46 (range 10-99) days after mobilization failure. Seventy patients with first-line BM harvest (group C) acted as historical controls. Ten patients from group B proceeded to HDCT and nine were evaluable for hematopoietic reconstitution. Recovery to neutrophils >0.5 x 10(9)/l was comparable with group C patients: 16 (range 9-34) days vs 13 (range 8-98) days. However, platelet (PLT) reconstitution >20 x 10(9)/l was significantly slower, with a median of 35 (range 13-50) days as compared with 19 (range 9-148) days (P = 0.0106) for control patients. Supportive care requirements, febrile days and length of hospital stay were not significantly different between the two groups of patients. We conclude that patients who fail to mobilize PBSC should be evaluated for second-line BM harvest. This approach may preserve the therapeutic option of HDCT for these patients.

Adolescent↗

PubMed2000

Presence of human beta- and gamma-herpes virus DNA in Hodgkin's disease.

Herpes viruses have been implicated in the etiology of Hodgkin's disease (HD). We studied the prevalence of human cytomegalovirus (CMV), human herpes viruses type-6 (HHV-6), type-7 (HHV-7) and type 8 (HHV-8) DNA in up to 88 Hodgkin's disease biopsies in comparison to Epstein-Barr virus (EBV) DNA by polymerase chain reaction (PCR). Non-Hodgkin lymphomas (NHL) and reactive lesions served as controls. CMV and HHV-6 were found in 8/86 (9%) and 11/88 (13%) HD cases, respectively, by nested primer PCR. Except for three cases harbouring HHV-6 type-B, only HHV-6 type-A was detected in HD. HHV-7 was observed by nested PCR in 33/88 (38%) HD cases and was already detectable in 15/88 (17%) HD cases by a single-round PCR indicating elevated virus copy numbers. Seven of these cases showed co-infection with HHV-6, and 11 cases were found to contain EBV DNA. 7/8 CMV-positive HD cases also harboured EBV DNA. HHV-8 DNA was not detected by single round or nested PCR in any HD case investigated. Thus, CMV, HHV-6, and HHV-7 were present in small proportions of HD cases, with frequent co-infection of HHV-6 and HHV-7, and frequent association with EBV. In contrast to EBV, beta-herpes viruses are therefore unlikely to have a role in the aetiology of HD. Rather, the presence of these viruses seems to reflect impaired immunological surveillance.

Betaherpesvirinae↗

PubMed2000

Diagnosis of toxoplasmosis in bone marrow transplant recipients: comparison of PCR-based results and immunohistochemistry.

Toxoplasmosis in bone marrow transplant recipients is a rare but serious complication and if untreated, almost uniformly fatal. The diagnosis, however, remains difficult. We therefore compared serial determination of antibody titers specific for T. gondii before and after transplantation, serial PCR for T. gondii DNA in serum, PCR and nested PCR for T. gondii DNA in various tissues, conventional histology and immunohistochemistry for detection of parasites in three patients with autopsy-confirmed toxoplasmosis after bone marrow transplantation. Immunohistochemistry demonstrated the presence of parasites in 13 out of 20 organs investigated (65%), whereas PCR detected T. gondii-specific DNA in 15 out of 20 organs (75%). Immunohistochemistry revealed concordant results to PCR data in 60% of the specimens. With the use of a nested PCR protocol, eight out of nine samples (89%) were positive for T. gondii-specific DNA. The combination of both methods detected the presence of parasites in 90% of the specimens. Serial PCR in serum did not yield positive results. Neither PCR nor immunohistochemistry was able to detect parasites in all organs investigated, but both methods together improved sensitivity to 90% and consequently, should be used jointly to maximize diagnostic precision. Bone Marrow Transplantation (2000) 25, 1257-1262.

Adult↗

PubMed2000

Treatment of BK virus-associated hemorrhagic cystitis and simultaneous CMV reactivation with cidofovir.

Hemorrhagic cystitis (HC) is a common complication following high-dose chemotherapy and bone marrow transplantation, and the treatment of virus-associated HC remains to be optimized. This is the first report on the successful use of cidofovir in a patient with HC and polyoma viruria concomitant with CMV reactivation after allogeneic BMT. Treatment led to a significant decrease in viruria and to sustained suppression of CMV reactivation. Administered with probenecid and hydration, cidofovir was well tolerated, and there were no side-effects.

Antiviral Agents↗

PubMed2000

Rapid quantification and differentiation of human polyomavirus DNA in undiluted urine from patients after bone marrow transplantation.

A combined PCR assay was developed for the detection and typing of human polyomavirus (huPoV) in clinical samples, consisting of (i) a qualitative seminested PCR assay (snPCR) to discriminate between huPoV BK and JC and (ii) a high-throughput, quantitative TaqMan PCR assay (TM-PCR) for the general detection of huPoV. The TM-PCR detects huPoV DNA in a linear range from 10(7) to 10(1) copies per assay. In reproducibility runs, the inter- and intra-assay variabilities were < or =60 and < or =50%, respectively. The snPCR assay uses a set of four primers for the same region of the BK and JC viral genomes. In the first round of amplification, two general primers were used; in the second round, one of these general primers and two additional, BK- or JC-specific primers were used simultaneously to produce amplicons of different sizes specific for BK virus (246 bp) and JC virus (199 bp), respectively. We tested different urine dilutions in order to determine the inhibitory effects of urine on PCR amplification. Furthermore, we compared the use of native urine with DNA purified by different preparation procedures. Our results show, that a 1:10 dilution of the urine led to complete reduction of the amplification inhibition found with 6% of undiluted urine samples. In a clinical study including 600 urine specimens, our assay turned out to be fast, cheap, and reliable in both qualitative and quantitative aspects.

BK Virus↗

PubMed2000

Detection of human cytomegalovirus DNA by real-time quantitative PCR.

A real-time PCR assay was developed to quantify human cytomegalovirus (CMV) DNA. This assay was used to demonstrate a higher CMV DNA load in plasma of bone marrow transplant patients than in that of blood donors. The CMV load was higher in CMV antigen-positive patients than in antigen-negative patients.

Blood Donors↗

PubMed2000

Detection of germ-cell tumor cells in peripheral blood progenitor cell harvests: impact on clinical outcome.

Our study was conducted to evaluate the impact of tumor cell contamination in peripheral blood progenitor cell (PBPC) harvests on the clinical outcome of patients with germ-cell tumors undergoing high-dose chemotherapy (HDCT) and autologous PBPC reinfusion. Samples of mononuclear cells from progenitor cell harvests of 57 patients with advanced or recurrent germ-cell tumors were retrospectively screened for contaminating tumor cells using immunocytochemical staining for cytokeratin filaments and reverse transcription-PCR (RT-PCR) testing for germ-cell alkaline phosphatase mRNA. The results were correlated to clinical prognostic variables as well as to the overall and event-free survival of these patients. Tumor cell contamination was detected in PBPC harvests of 16 of 57 enrolled patients (28%), and, among these, in 14 of 51 (27%) who underwent HDCT. The presence of contaminating tumor cells as detected by either immunocytochemical staining, RT-PCR, or both was strongly associated with a reduced overall survival (43% versus 71%, P = 0.0037) and event-free survival (0% versus 52%, P = 0.0005) after 1 year. In multivariate analysis, the demonstration of contaminating tumor cells had a higher predictive value for a poor event-free survival than other known prognostic variables. The presence of contaminating tumor cells in PBPC harvests of patients with germ-cell tumors seems to predict a poor overall and event-free survival in patients undergoing HDCT and autologous PBPC reinfusion.

Adult↗

PubMed1999

High-dose salvage chemotherapy. Germ-cell tumor treatment results in Germany.

During the past 10 years, high-dose chemotherapy (HDCT) has emerged as a feasible and curative treatment option for patients with multiply relapsed and/or refractory germ-cell tumors. Increasing experience and the results from clinical trials have helped to optimize the application of HDCT and to define its role in the salvage concepts for germ-cell tumors.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed1999

Interferon-alpha before allogeneic bone marrow transplantation in chronic myelogenous leukemia does not affect outcome adversely, provided it is discontinued at least 90 days before the procedure.

The influence of interferon-alpha (IFN) pretreatment on the outcome after allogeneic bone marrow transplantation (BMT) in chronic myelogenous leukemia (CML) is controversial. One goal of the German randomized CML Studies I and II, which compare IFN +/- chemotherapy versus chemotherapy alone, was the analysis of whether treatment with IFN as compared to chemotherapy had an influence on the outcome after BMT. One hundred ninety-seven (23%) of 856 Ph/bcr-abl-positive CML patients were transplanted. One hundred fifty-two patients transplanted in first chronic phase were analyzed: 86 had received IFN, 46 hydroxyurea, and 20 busulfan. Forty-eight patients (32%) had received transplants from unrelated donors. Median observation time after BMT was 4.7 (0.7 to 13.5) years. IFN and chemotherapy cohorts were compared with regard to transplantation risks, duration of treatments, interval from discontinuation of pretransplant treatment to BMT, conditioning therapy, graft-versus-host disease prophylaxis and risk profiles at diagnosis and transplantation, and IFN cohorts also with regard to performance and resistance to IFN. Outcome of patients receiving related or unrelated transplants pretreated with IFN, hydroxyurea, or busulfan was not significantly different. Five-year survival after transplantation was 58% for all patients (57% for IFN, 60% for hydroxyurea and busulfan patients). The outcome within the IFN group was not different by duration of prior IFN therapy more or less than 5 months, 1 year, or 2 years. In contrast, a different impact was observed in IFN-pretreated patients depending on the time of discontinuation of IFN before transplantation. Five-year survival was 46% for the 50 patients who received IFN within the last 90 days before BMT and 71% for the 36 patients who did not (P =.0057). Total IFN dosage had no impact on survival after BMT. We conclude that outcome after BMT is not compromised by pretreatment with IFN if it is discontinued at least 3 months before transplantation. Clear candidates for early transplantation should not be pretreated with IFN.

Adolescent↗

PubMed1999

Aerosolized amphotericin B inhalations as prophylaxis of invasive aspergillus infections during prolonged neutropenia: results of a prospective randomized multicenter trial.

We performed a prospective, randomized, multicenter trial to evaluate the effectiveness of prophylactic inhalations with aerosolized amphotericin B (aeroAmB) to reduce the incidence of invasive aspergillus (IA) infections in patients after chemotherapy or autologous bone marrow transplantation and an expected duration of neutropenia of at least 10 days. From March 1993 until April 1996, 382 patients with leukemias, relapsed high-grade non-Hodgkin lymphomas, or solid tumors were randomized with a 13:10 ratio to receive either prophylactic aeroAmB inhalations at a dose of 10 mg twice daily or no inhalation prophylaxis in an unblinded fashion. The incidence of proven, probable, or possible IA infections was 10 of 227 (4%) in patients who received prophylactic aeroAmB. This did not differ significantly from the 11 of 155 (7%) incidence in patients who received no inhalation prophylaxis (P =.37). Moreover, no differences in the overall mortality (13% v 10%; P =.37) or in the infection-related mortality (8% v 7%; P =.79) were found. In contrast to other nonrandomized trials, we observed no benefit from prophylactic aeroAmB inhalations, but the overall incidence of IA infections was low.

Administration, Inhalation↗

PubMed1999

Identification of human cytomegalovirus variants by analysis of single strand conformation polymorphism and DNA sequencing of the envelope glycoprotein B gene region-distribution frequency in liver transplant recipients.

Single strand conformation polymorphism analysis (SSCP) of PCR-amplified DNA and subsequent DNA sequencing of human cytomegalovirus (HCMV) glycoprotein B (gB) gene were applied to identify known HCMV strains and to detect new virus variants. 61 HCMV PCR positive patients were studied out of a cohort of 410 patients after liver transplantation (LTX). SSCP was able to distinguish between strains Davis, AD169, and Towne, and in addition could identify five new virus variants (Berlin B, C, E, F, and H). Their frequency, gB and gH types were determined. Simultaneous infections with two or three strains or variants, as well as a switch from one virus to another virus were observed during long-term follow-up. No correlation between the occurrence of certain virus strains or gB types and defined clinical manifestations of HCMV infection after LTX was drawn.

Base Sequence↗

PubMed1999

Determination of platinum complexes in clinical samples by a rapid flameless atomic absorption spectrometry assay.

The frequent use of platinum (Pt) complexes in cancer chemotherapy and the application of new therapeutic options and dosing strategies have increased the need for rapid analytic procedures to determine Pt concentrations in the biologic fluids of patients. Therefore a flameless atomic absorption spectrometry method for the quantification of Pt in plasma and ultrafiltrate was developed and validated. A simple sample preparation of only one dilution step was established. Only 400 microL of whole blood was required for duplicate analysis of Pt in both matrices. The matrix-specific temperature programs took less than 75 seconds. The lower limit of quantification was 40 ng Pt/mL and 20 ng Pt/mL for plasma and ultrafiltrate, respectively. Suitable linearity could be reached using separate calibration curves for the high and low Pt concentration ranges. Recovery of Pt was complete, and there were no major stability problems. The accuracy and precision of the new method met the international criteria for the validation of bioanalytic methods. In addition, the use of different anticoagulants for clinical sampling, ultrafiltration systems, and ultrafiltration conditions were investigated. The assay has already been extensively applied to pharmacokinetic studies. In conclusion, the new Pt assay proved to be rapid, simple, sensitive, and suitable for clinical use.

Anticoagulants↗

PubMed1999

Different real-time PCR formats compared for the quantitative detection of human cytomegalovirus DNA.

BACKGROUND: The aim of this study was to compare the ABI PRISM 7700 Sequence Detection System and the LightCycler to develop a quantitative real-time PCR assay for the detection of human cytomegalovirus (HCMV) DNA suitable for routine hospital application. METHODS: We used one exonuclease probe and five different hybridization probe sets as sequence-specific fluorescence detection formats. For the exonuclease assay and two hybridization probe sets, reproducibility and the detection limit were determined. To keep the total assay time to a minimum, we gradually shortened individual reaction steps on both instruments. RESULTS: The exonuclease assay can be interchangeably performed on the 7700 and the LightCycler. No change of reaction conditions is required, except for the addition of bovine serum albumin to the LightCycler reaction. The shortest possible total assay time is 80 min for the ABI PRISM 7700 Sequence Detection System and 20 min for the LightCycler. When the LightCycler is used, the exonuclease probe can be replaced by a set of hybridization probes. All assays presented here detected HCMV DNA in a linear range from 10(1) to 10(7) HCMV genome equivalents/assay (r >0.995) with low intraassay (<5%) and interassay (<10%) variation. CONCLUSIONS: The ABI PRISM 7700 Sequence Detection System as well as the LightCycler are useful instruments for rapid and precise online PCR detection. Moreover, the two principles of fluorescence signal production allow HCMV quantification with the same accuracy.

Cytomegalovirus↗

PubMed1998

Kinetics of the graft-versus-leukemia response after donor leukocyte infusions for relapsed chronic myeloid leukemia after allogeneic bone marrow transplantation.

Little is known about the mechanisms and the kinetics of the so-called graft-versus-leukemia (GVL) response induced by donor lymphocyte infusions (DLI) in patients with leukemic relapse after allogeneic bone marrow transplantation (BMT). We sought to elucidate this problem by sequentially studying three patients with relapsed chronic myeloid leukemia after sex-mismatched BMT from time before donor leukocyte infusion until achievement of complete molecular remission. Lineage-specific chimerism was assessed longitudinally by a combined fluorescent immunophenotyping and sex chromosome-specific in situ hybridization approach. Results were related to quantitative detection of bcr-abl transcripts by competitive differential reverse transcriptase-polymerase chain reaction (RT-PCR), qualitative bcr-abl RT-PCR, and multiplex PCR-based DNA donor/recipient chimerism. All patients had predominant donor lymphopoiesis at the time of DLI, suggesting a state of tolerance to recipient leukemic and/or normal cells. In contrast, granulopoiesis and erythropoiesis were mainly recipient derived in both patients with hematologic relapse and partly recipient derived in the patient with molecular relapse. Eighty percent, 90%, and 8% of CD34(+) cells, respectively, were found to be of recipient origin at relapse, and few donor stem cells predicted for cytopenia post-DLI. Responses were seen after a time lag of 5 to 13 weeks after DLI and resulted in reversal to full donor chimerism within a critical switch period of 4 to 5 weeks. A sudden decrease in recipient cells was paralleled by a sharp decrease in bcr-abl transcript numbers detectable several weeks before achievement of molecular remission and onset of clinical graft-versus-host disease (GVHD). This response pattern was confirmed by retrospective RT-PCR analysis in an additional five patients. Prospective monitoring of stem cell chimerism and response may enable us to individually tailor adoptive immunotherapy in the future.

Adult↗

PubMed1998

High-dose chemotherapy with autologous stem cell support in poor-risk germ cell tumors.

High-dose chemotherapy (HDT) and stem cell transplantation is a newer treatment option widely applied in poor-risk germ cell tumor patients. Due to the increasing practical clinical experience and the availability of hematopoietic growth factors, this treatment approach has become a relatively safe procedure. Depending on the drugs used. the most prominent therapy-associated side effects are myelosuppression, infections, mucositis, moderate, mostly reversible neurotoxicity, and renal impairment. Because of their unique pharmacologic characteristics, carboplatin and etoposide have proved to be the most important drugs for HDT. It is not known whether the addition of ifosfamide or cyclophosphamide or other drugs to the combination of carboplatin and etoposide leads to superior results. It is not yet clear if HDT should already be instituted in first-line treatment of poor-risk patients, or later after relapse or incomplete response. Even if precise data on the therapeutic value of HDT are still missing because randomized trials are not yet ready for evaluation, there is good evidence for the effectiveness of HDT. The demonstration of remissions in cisplatin-refractory patients, the inversion of remission durations, and the induction of long-term freedom from disease in multiply relapsed patients who were deemed incurable with conventional-dose procedures argue in favor of HDT. Finally, the delineation of prognostic factors associated with a poor probability of survival after HDT contributes to the selection of patients who are likely to profit from HDT and those who should be spared from dose-intensive treatment.

Antineoplastic Agents↗

PubMed1998

Human herpesvirus type 6 variants identified by single-strand conformation polymorphism analysis.

Six human herpesvirus 6 (HHV-6) variants were analyzed for heterogeneity using the polymerase chain reaction (PCR) and single-strand conformation polymorphism (SSCP). Two independent DNA regions were selected: a fragment of the gene U11 (position 18966-21578) coding for a basic phosphoprotein, the major antigenic structural protein pp100; and a fragment from an open reading frame (ORF) area of the gene U67, previously referred to as 13R (position 102458-103519), coding for a product of unknown function. The two PCR systems based on the above DNA sequences yielded products of 187 bp and 223 bp, respectively. DNA obtained from three laboratory reference strains (U1102, R104 and St.W.) and from HHV-6 infected peripheral white blood cells of bone marrow transplant patients and blood donors was used to test the applicability of two different SSCP analysis systems for the identification of HHV-6 variants using amplicons derived by PCR from the two genomic regions described above (U11 [pp100], U67 [13R-ORF]). The generation of characteristic SSCP patterns enables the rapid differentiation of HHV-6 A and B strains for the classification of variants derived from clinical samples, reducing the need for expensive and time-consuming direct sequencing analyses.

Base Sequence↗

PubMed1998

High-dose chemotherapy in germ cell tumours: a large single centre experience.

High-dose chemotherapy (HDCT) has evolved as a strategy to improve the treatment outcome in patients with relapsed and/or refractory germ cell tumours. Between August 1989 and September 1995, 150 consecutive patients with relapsed and/or refractory germ cell tumours were treated with conventional-dose salvage chemotherapy followed by one cycle of HDCT with carboplatin 1500-2000 mg/m2, etoposide 1200-2400 mg/m2 and ifosfamide 0-10 g/m2 and were retrospectively analysed. With a median follow-up time of 55 months (range 21-88 months) 51/150 (34%) patients are alive and disease free. The projected event-free and overall survival are 29% (confidence interval 22-37%) and 39% (confidence interval 31-47%) respectively. The relevance of prognostic variables for long-term survival after HDCT were prospectively confirmed. Persisting toxicities occurred in approximately one third of the long-term survivors. Treatment intensification with HDCT resulted in a significant proportion of the long-term survivors in patients with relapsed and/or refractory germ cell tumours. Trials to prospectively evaluate HDCT as an early intervention in these patients seem justified.

Adolescent↗

PubMed1998

Treatment of relapse after allogeneic bone marrow transplantation with unmanipulated G-CSF-mobilized peripheral blood stem cell preparation.

Donor lymphocyte infusions (DLI) are an effective treatment of leukemia relapse after allogeneic bone marrow transplantation. Undesired side-effects are the development of graft-versus-host disease (GVHD) and the occurrence of pancytopenia in some patients. In a pilot study, we investigated if unmanipulated G-CSF-mobilized peripheral blood stem cells which naturally contain large numbers of T lymphocytes (D-PBSC/LI) would be equally effective or even superior than DLI in generating a graft-versus-leukemia reaction (GVL) but could mitigate or prevent the development of pancytopenia. We treated 12 patients with CML chronic phase (n = 5), CML blast crisis (n = 2), AML (n = 2), ALL (n = 1), CLL (n = 1) and multiple myeloma (n = 1). In five patients with acute leukemia or CML blast crisis D-PBSC/LI followed intensive chemotherapy (group A), in seven patients D-PBSC/LI were given without any prior chemotherapy (group B). In group A two patients were evaluable for hematologic toxicity. Leukopenia <1000/microl lasted for 10 and 19 days, and thrombocytopenia <20,000/microl for 11 and 13 days, respectively. In group B leukopenia <1000/microl and thrombocytopenia <20,000/microl was observed in only one patient. Moderate cytopenia developed in four of five evaluable patients. A complete remission could be achieved in all seven patients with CML who all developed acute and/or chronic GVHD. None of the remaining five patients achieved a complete remission despite acute and/or chronic GVHD in two of them. Four patients died from disease progression, one patient from a secondary lymphoma, and one patient as a result of uncontrolled GVHD. In conclusion, D-PBSC/LI is effective in inducing GVL reaction but it does not prevent pancytopenia in each case. It remains unclear if it mitigates the incidence and severity of pancytopenia.

Adult↗