A Hrubá

University Hospital Motol, Praha, Praha, Czech Republic

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Publications (44)51.78 Total impact

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    ABSTRACT: Objectives: Transplantation of allogeneic stem cells from healthy donor can cure several malignant and non-malignant disorders in children. In this paper authors evaluate results of allogeneic SCT (stem cell transplantation) in children performed at transplant center from November, 1989, to December, 2005. At certain time points they determine survival rates, death rates due to toxicity or relapse of malignancy, changes in donors and grafts composition. Patients and methods: From 1989 till 2005 altogether 265 patients at the age of 0.2-20.5 years (median 9.7) underwent first allogeneic SCT at transplant unit (Teaching Hospital Motol, Prague). 71% (n=189) were transplanted for malignant disease (acute leukemia, myelodysplastic syndrome, chronic myelogenous leukemia, lymphoma, etc.), other for life-threatening non-malignant disorder (congenital or acquired bone marrow failure, severe immunodeficiency, osteopetrosis, mucopolysaccharidosis type I., adrenoleukodystrophy, etc.). At the beginning matched sibling donors were predominantly used (81%). Bone marrow was used as a source of stem cells in majority of patients (93%). Over last period authors use more frequently graft obtained from unrelated donor (68%), bone marrow is currently used in only 58% of transplants. Results: Out of 265 transplanted patients 165 (62%) survive. The survival is higher in the group of children transplanted for non-malignant disorder (80% versus 55%). Over the time the authors successfully reduced overall transplant related mortality from 29% to 9%. Survival rate improved markedly from 46% (1989-2000) to 80% (2001-2005). Conclusions: Allogeneic SCT at the transplant unit over the passed years became a standard procedure in the therapy of advanced stages of malignant diseases or some life-threatening non-malignant disorders for children from all the Czech Republic. For such children transplantation represents the last chance to be cured. Even with majority of transplants being done using grafts from alternative donors, the authors achieve the same excellent results as they do in outstanding European pediatric transplant centers.
    No preview · Article · Jan 2007
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    ABSTRACT: 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.
    No preview · Article · Aug 2002 · Casopís lékar̆ů c̆eských
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    ABSTRACT: Cyclophosphamide (4 g/m2) and paclitaxel (Taxol) (175, 200 or 250 mg/m2) therapy with subsequent administration of G-CSF (10 micrograms/kg) has been used as intensification and as mobilization therapy for patients with breast cancer. This regimen was used in 19 patients, as part of adjuvant therapy in 14 and as part of therapy of metastatic disease in five. Median number of collected CD34+ cells was 17.5 x 10(6)/kg (2.9-48.1). All patients except one (94.7%) reached minimal required number of CD34+ cells (> or = 3 x 10(6)/kg). Median number of leukapheresis was two. The required number of cells (> or = 3 x 10(6)/kg) was collected in one leukapheresis in 17 out of 19 patients (89.5%) and more than five and 10 x 10(6)/kg CD34+ cells respectively were collected in 14 (73.7%) and 11 (57.9%) patients respectively. No factor significantly influencing the amount of collected cells (except the trend in favour of later year of therapy and large-volume leukapheresis) was identified. Leukopenia gr. 4 was observed in 88.9% of treated patients and febrile neutropenia developed in 46.2% patients. Although the antitumour activity of this chemotherapy was not possible to assess it seems that this intensification could be successfully used as a therapy and as very potent mobilization regimen.
    No preview · Article · Jan 2002 · Sbornik lekarsky
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    ABSTRACT: The aim of this study was to establish a suitable method for in vitro T cell depletion in peripheral blood stem cell grafts for mismatched/haploidentical transplantation in children and adults with severe hematological disorders and for autologous transplantation in patients with autoimmune diseases refractory to conventional immunosuppressive treatment. Two different selection techniques have been used: CD34+ selection using immunoaffinity columns (CellPro Ceprate) followed by T cell depletion by E-rosetting or CD34+ selection using submicroscopic paramagnetic beads (CliniMACS device) with T cell depletion in a one step procedure. The mean purity and recovery of CD34+ cells and efficiency of T cell removal in the final product were compared. From March 1995 to December 1998 we prepared twelve allografts using Cell Pro system for eight children with high-risk hematological malignancies and six autografts for six patients with severe autoimmune diseases. From January 1999 to October 2000 we prepared fifteen allografts using CliniMACS system for ten children with high-risk hematological diseases and inborn metabolic disorders or primary immunodeficiences, five allografts for three adult patients with high-risk hematological malignancies and two autografts for two patients with autoimmune diseases. In allogeneic transplantation the median purity of CD34+ cells in the final products after CellPro and E-rosetting was 85.6% (55.3%-95.7%); median recovery was 24.8% (17%-35%), median transplanted doses of T cells per kilogram of body weight were 0.66x10(4) (0-2.8); in autologous transplantation the median purity of CD34+ was 92.6% (55.6%-96%), median recovery was 28% (22%-46.2%), median transplanted doses of T cells per kilogram of body weight were 0.39x10(4) (0.0-3.6). After CliniMACS technique the median purity of CD34+ cells was 94.87% (69.15%-99%),medianrecoverywas 58% (30%-79.6%), median transplanted doses of T cells per kg of body weight were 0.254x10(4) (0-14.15); in autologous transplantation the median purity of CD34+ was 94% (94%-94%, median recovery was 97.4% (95%-99.8%), median transplanted doses of T cells per kilogram of body weight were 0.87x10(4) (0.49-1.24). We consider both methods of CD34+ selection and T cell depletion suitable for peripheral blood stem cell processing before mismatched hemopoietic stem cell transplantation in patients without identical donor or before autologous transplantation for severe autoimmune diseases. However, magnetic separation using CliniMACS system results in higher levels of purity and recovery with efficient T cell depletion.
    No preview · Article · Feb 2001 · Neoplasma
  • Z Gasová · Iuri Marinov · Alena Hrubá · K Benesová · Petr Turek
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    ABSTRACT: Transplantations of peripheral blood progenitor cells (PBPC) are able to assure a complete haematopoietic and immunologic reconstitution. The efficient mobilization of progenitor cells into peripheral blood is the main factor responsible for quality of the graft as well as timing and technique of collections. The aim of the present paper was to find the optimum time for starting PBPC collections and consequently to minimize the number of procedures required. The study was performed in patients with haematological malignancies using an autologous collection regimen. We attempted to determine a relationship between the concentration of CD 34+ cells in peripheral blood at the beginning of the collection and the number of CD 34+ cells in the leukapheresis product prepared in the standard mode processing 2-3 total blood volumes. We assessed the significance of the CD 34+ cells concentration in peripheral blood for the adequate collection of CD 34+ cells. We also evaluated the time of engraftment in patients after autologous PBPC transplantation whenever possible. The study was performed in 70 patients. Two groups were defined: Group I patients were well mobilized, whereas Group II patients were weakly mobilized. CD 34+ counts, using flow cytometry were found to be useful in predicting the optimal time for collections.
    No preview · Article · Jul 1999 · Transfusion Science
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    ABSTRACT: Transplantations of haematopoietic progenitor cells from peripheral blood (PBPC) are able to ensure haematopoietic and immunological reconstitution as well as stable long term engraftment. Autologous PBPC are administered after previous myeloablative chemotherapy to patients with haematological and non-haematological malignancies. The objective of the submitted study was to follow-up the results of autologous separations of PBPC in patients with a good effect of mobilisation therapy. The authors evaluated in PBPC concentrates the content of cell parameters needed for transplantation. In the subsequent part of the trial they mention the times of engraftment after autologous transplantation. The authors evaluated parameters of 26 separations of PBPC in 11 haematooncological patients with a good effect of mobilisation therapy and with concentration of CD 34+ cells higher than 20 x 10(3)/ml of peripheral blood. The separations of PBPC were implemented on the Cobe Spectra and Baxter CS 3000 Plus equipment under a standard regime with processing of 12 l blood, i.e. 2.7 total blood volumes of the patients. In the mentioned group of patients already from one separation an adequate amount of CD 34+ cells for their transplantation was obtained. Transplantation doses were prepared on average from two separations and amounted as regards MNC/kg, CD 34+ cells/kg and CFU-GM/kg to 4.3 x 10(8), 17.1 x 10(6) and 2.5 x 10(4). The assembled parameters correspond to, or in some parameters exceed, recommended amounts for their transplantation. In well mobilised patients under the regime of standard separations adequate amounts of progenitor cells for their autologous transplantation were obtained. Transplantation doses were prepared from two collections. Investigation of pre-separation concentration of CD 34+ cells in the peripheral blood is a reliable indicator for starting PBPC separation. Early post-transplantation results indicate the time of engraftment 10 days on average and minor need of substitution therapy with blood products.
    No preview · Article · Jul 1999 · Casopís lékar̆ů c̆eských
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    ABSTRACT: 13 patients have been transplanted at Institute of Hematology and Blood Transfusion since 1995 using allogeneic PBPC either alone or with bone marrow as a source of progenitor cells. All donors were G-CSF mobilised HLA identical family members. PBPC harvests were performed on D 4,5, (6) of G-CSF administration. The medium content of TNC, CD34+, CD3+, CD4+and CD8+cells/kg b.w. of the recipients in the grafts were: 13,1x10(8)(TNC), 11,4x10(6)(CD34+), 393x10(6)(CD3+) 243x10(6)(CD4+), 125x10(6)(CD8+) The patients received either BuCy2 or CyTBI preparative regimen and Cyclosporin A + short course of Methotrexate for GVHD prophylaxis. Engraftment of ANC >500 was achieved by D+16 and PLT >20.000 by D+19. Three of ten evaluable patients developed acute and three of nine chronic GVHD. 8 patients survive with the longest follow up 776 days.
    No preview · Article · Jan 1999 · Bone Marrow Transplantation
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    ABSTRACT: Most children with acute lymphoblastic leukemia (ALL) and increasing number of children with acute myelogenous leukemia (AML) are currently cured with conventional chemotherapy. Despite of this success there is a subset of patients with high-risk features at diagnosis who are predisposed to a very high risk of relapse. Relapse of AML and early bone marrow relapse of ALL can not be cured by conventional chemotherapy. Allogeneic hematopoietic stem cell transplantation (HSCT) is therapeutic option in these children with very high-risk acute leukemia. Between XI/1989-XII/1996 33 children with acute leukemia (ALL: 22, AML: 11) underwent an allogeneic HSCT from HLA identical related donors (HLA-identical sibling: 30, twin: 1, other HLA-identical relative: 2) at the 2nd Dept. of Pediatrics, University Hospital Motol. Median age of our group was 9 years (1.5-19 y.), boys (n = 23) clearly dominated over the girls (n = 10). The resource of stem cells was bone marrow in 31 children, bone marrow and peripheral blood progenitor cells (PBPC) and PBPC in one child respectively. Myeloablative conditioning regimen varied, consisting of total body irradiation and chemotherapy in 21 children and chemotherapy in 12 children. HSCT was performed in first complete remission of acute leukemia in 9 children (AML: 7, ALL: 2), in second remission in 14 children (AML: 2, ALL: 12), in third remission in 4 children (ALL: 4). Six children underwent HSCT in first partial remission (n = 1) and in second (n = 4) or third (n = 1) chemoresistant relapse. Seven (21%) children died due to post-transplant complications. Nine (28%) children suffered from clinically significant acute graft-versus-host reaction (GVH) and 15% (4/27) children who survived 100 days post-transplant suffered from chronic GVH disease. Relapse of leukemia was diagnosed in 39% (12/31) children. Fourteen (42%) children are alive and well in continuous remission with median follow-up 42 months. Allogeneic HSCT can cure children with very high-risk acute leukemia in the situations where conventional chemotherapy fails. Relapse of leukemia and GVH reaction are most important causes of post-transplant morbidity and mortality.
    No preview · Article · Jul 1998 · Casopís lékar̆ů c̆eských
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    ABSTRACT: The authors describe the results of purification of bone marrow and peripheral progenitor cells (PBPC) for clinical transplantations. Vepeside was used to purify in 1990-1995 a total of 41 bone marrows of adults and children. Of these 23 were transplanted. Maphosphamide was used bone marrow purging in two patients; transplantation was performed in one case. By a combination of Vepeside with methylprednisolone haematopoietic cells of 24 patients were purged, transplantations were performed in 10. Three-day cultivation of haematopoietic cells in the presence of Desferal was used for purging cells of 22 patients with neuroblastoma; transplantations were performed in 10 patients. The authors give the values of nucleated cells, haematopoietic colonies of CFU-GM and CD34 positive cells obtained after purification calculated per kg body weight of the patient and the percentage yields.
    No preview · Article · Apr 1997 · Casopís lékar̆ů c̆eských
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    ABSTRACT: Bone marrow transplantation has become the therapeutic method in some forms of malignant haemotopoietic diseases, malignant tumours, inborn errors of metabolism and immunodeficiency states. The objective of the presented work is the analysis of 40 allogenic bone marrow transplantations in children made in 1989-1994. Bone marrow transplantation was made in 40 children (26 boys and 14 girls), mean age 10.5 years (range 1.5-17.5 years). Indications were acute lymphoblastic leukaemia in 11, acute myeloid leukaemia in 10, chronic myeloid leukaemia in 6, myelodysplastic syndrome in 2, aplastic anaemia and Fanconi's anaemia in 7, non-Hodgkin lymphoma in 2 and inborn errors in 2 children. The donor was in 33 patients in HLA identical sibling and in seven instances a monozygotic twin, HLA non-identical sibling or relative or unrelated matched donor. Bone marrow engraftment was achieved in 35 (87.5%) patients, in one instance the bone marrow was rejected (2.5%) and in four patients (10%) early death occurred before the bone marrow engraftment. On Aug. 15, 1995 20 patients (50%) survived, a relapse developed in 7 (17.5%) and 13 patients died in conjunction with the transplantation (32.5%). The most frequent cause of death were infectious complications (9 children) either in conjunction with the development of graft versus host reaction (6x) or without signs of this reaction (3x). As a prophylaxis of graft versus host disease 24x Cyclosporine A with corticosteroids was used, 16x with methotrexate. A chronic graft versus host disease developed in 6 of 28 children surviving 100 days after transplantation. The greatest problem are infectious (bacterial and mycotic) complications in the phase of bone marrow aplasia before engraftment of the transplanted bone marrow or in conjunction with a graft versus host reaction which cannot be completely avoided by preventive measures. Bone marrow transplantation is also in children an effective therapeutic method of some forms of malignant haematopoietic diseases, malignant tumours and immunodeficiency states. The correct indication, suitable donor, preventive measures against the graft versus host reaction and protection against infectious complications are essential for the success of this pretentious treatment.
    No preview · Article · Jan 1996 · Casopís lékar̆ů c̆eských
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    ABSTRACT: In the submitted paper the authors summarize their experience with changes of lymphocyte subpopulations after transplantations of haematopoietic progenitor cells on account of malignant tumours in children. After transplantation the percentage of T lymphocytes increases, the CD4/CD8 ratio declines in the second month and remains reduced up to the second year, and the percentage of B lymphocytes is very low after transplantation and gradually rises. At the end of the first year after transplantation the ratio of B lymphocytes in blood and bone marrow is elevated, and the finding in the bone marrow reminds of lymphoblastic leukaemia. Reconstitution of immunity does not differ in relation to the type of graft, not even after the use of an ex vivo purified graft.
    No preview · Article · Jan 1996
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    ABSTRACT: The authors are using the autologous PBPC transplantations since 1992. For mobilisation has been used primarily cyclophosphamide alone, later cyclophosphamide and G-CSF of intensive chemotherapy and G-CSF. 104 separations in 28 children were made using the Cobe Spectra device with 17 transplantations, two children died on peritransplantation complications. By use of the MCS 3p Haemonetics device were done 46 separations in 16 children, the graft was used in five cases, always successfully.
    No preview · Article · Jan 1995
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    ABSTRACT: The first allogenic bone marrow transplantation (TKD), when for the preparation whole body irradiation was used, was implemented in the Institute of Haematology and Blood Transfusion (UHKT) in Prague in 1986. Before June 1992 36 TKD were performed incl. 28 allogenic, 2 syngenic and 6 autologous. For the first time bone marrow from a non-related donor was transplanted. Of 30 allogenic and syngenic TKD to the present time 17 patients survive, i.e. 56.6% of the whole group. According to individual diagnoses 8 patients with the diagnosis of chronic myeloid leukaemia (CML) survive, 5 of 10 patients with the diagnosis of acute leukaemia (AL) and 3 of 4 patients with the diagnosis of severe aplastic anaemia (SAA) or with Fancon's anaemia (FA) resp. The survival period of the whole group is from 1-62 months since the transplantation. The main cause of death of 8 from 13 patients who died were infections associated with acute or chronic disease of the graft against the host (GVHD). In autologous TKD the bone marrow was treated with etoposide. Of the six transplanted patients with AL five survive 1.5-30 months after transplantation. The authors present some general information of pretransplantation preparation, prevention of GVHD, its incidence and results of TKD.
    No preview · Article · Jan 1993 · Casopís lékar̆ů c̆eských
  • J Soucek · V Chudomel · A Hrubá · G Lindnerová
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    ABSTRACT: The stimulating effect of cytosol fraction (F3) isolated from human myeloblasts (m.w. ranging from 30 to 100 kDa) and monoclonal antibody CD 3 (MEM 57) was tested on NK and LAK cell activities in peripheral mononuclear cells (PMNC) of normal donors and leukemic patients. The F3 fraction added in 10% of the total volume of RPMI 1640 medium induced significantly increased lytic activity of normal lymphocytes or IL-2 activated lymphocytes against K 562 cells in 3-day culture. Similarly, the proliferation of both cell cultures was enhanced by F3. The effect of F3 on NK and LAK cell activities in culture of PMNC from leukemic patients was less pronounced and synergic action of F3 did not occur, whereas the proliferation of leukemic cells was significantly enhanced. MEM 57 increased the cytotoxicity and cell proliferation in 3-day culture of normal lymphocytes at concentrations ranging from 10 to 250 ng/ml. MEM 57 stimulated also NK cytotoxicity estimated in freshly isolated normal lymphocytes. The deficient NK cell activity observed in PMNC of leukemic patients was induced by MEM 57 or its combination with IL-2 in 3-day culture. These observations indicate that combination of more immunomodulating agents can lead to shortening of the incubation time necessary for correction of the NK cell defective activity in PMNC of leukemic patients.
    No preview · Article · Feb 1991 · Neoplasma
  • J. Soucek · V. Chudomel · A. Hruba · G. Lindnerova

    No preview · Article · Jan 1991
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    ABSTRACT: Normal or leukemic human lymphocytes treated with direct current (DC) showed enhanced antileukemic cytotoxicity. The enhancing effect of DC-treated lymphocytes was dependent on current density and time exposure. A desirable effect was achieved with current densities ranging from 5 to 10 mA/cm2 at a short exposition time (5--10s). Enhanced lymphocyte cytotoxicity occurred after a 48 h cultivation at 37 degrees C in a humidified atmosphere containing 5% CO2 and was proved by the increased number of trypan blue stained target cells, tumor-binding cells, and lymphocytes with activated nucleoli. Lymphocyte cytotoxicity measured immediately after DC-treatment was not enhanced. Furthermore, the cytotoxic effect was potentiated using media conditioned with interleukin-2 (IL-2) or cytosol fraction (F3) isolated from human leukemic cells. Such in vitro stimulated cytotoxic cells displayed reactivity against K 562 cells as well as fresh leukemic cells of allogeneic origin. Of considerable clinical interest is the observation that lymphocytes treated with DC in IL-2 or F3 conditioned media may enhance antileukemic cytotoxicity in peripheral lymphocytes of patients with hematological malignancies.
    No preview · Article · Feb 1989 · Neoplasma
  • K Koubek · P Korínkovà · A Hrubà · P Hausner
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    ABSTRACT: We report on the characterization of four monoclonal antibodies which were prepared against membrane markers of human myeloid lineage. Fusion, isolation of hybridoma cells and their cloning and testing of the monoclonal antibodies by indirect immunofluorescence and FACS 440 analysis were performed by means of standard procedures. The results indicate that the monoclonal antibodies have a specificity against membrane markers of human myeloid lineage (exactly promyelo-granulocytes). These monoclonal antibodies do not react with human T and B lymphocytes, monocytes, erythrocytes and thrombocytes of peripheral blood. In normal bone marrow reactivity to matured myeloid cells was found to occur in promyelocytes and expressed on all granulocytes. These monoclonal antibodies also react with cells of myeloid cell lines and with other precursor cell lines represented by NALM-1, NALM-16, HEL, K-562, REH no reactivity was detected. The produced antibodies react with some leukaemic cells from patients with more mature myeloid cells (AML with promyelocytes, myelocytes and CML) they do not react with pathological cells from patients with CLL, AML (with myeloblasts), ALL, hairy cell leukaemia, erythroleukaemia and several types lymphoma. All antibodies have a IgM class and express granulocytotoxic and granuloagglutination activity. Using flow cytometry comparative analysis with other monoclonal antibodies was performed to detect the membrane structure (X-haptene) included in standard International classification as CD 15 group.
    No preview · Article · Feb 1988 · Folia haematologica (Leipzig, Germany: 1928)
  • A Hrubá · J Soucek · V Chudomel

    No preview · Article · Aug 1986 · Casopís lékar̆ů c̆eských
  • E Paluska · A Hrubá · O Stĕrba · J Kopecek
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    ABSTRACT: A synthetic polymer, Duxon, was developed and tested as a substitute of blood plasma for transfusion purposes. Tests of this preparation included a test for its influence on the haemopoietic stem cells and the graft-versus-host reaction (GVHR). A single dose or repeated doses of Duxon did not reduce the number of pluripotent haemopoietic colonies (CFU-S) in mice and short-term incubation of cells from haemopoietic organs of mice with Duxon resulted in a slight, yet significant, increase in the number of CFU-S. Injection of Duxon non-significantly diminished the manifestations of the regional GVHR. Incubation of spleen cells with Duxon significantly reduced their GVH reactivity in the regional test. The systemic GVHR was moderately inhibited by repeated doses of Duxon in sublethally irradiated mice. A significant delay in deaths of animals from GVHR was seen in lethally irradiated mice receiving a single dose of Duxon on day 4 after GVHR induction. Survival of lethally and sublethally irradiated mice was prolonged following injection of cells incubated with Duxon. Possible uses of Duxon, a preparation with mild immunosuppressive properties, not impairing haemopoiesis, are discussed.
    No preview · Article · Feb 1986 · Folia biologica
  • A Hrubá · E Paluska · V Chudomel
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    ABSTRACT: Mouse spleen and bone marrow cells were incubated for 2 h with ZnCl2 and Li2SO4 at different concentrations and tested for the ability to evoke the graft-versus-host reactions (GVHR) and to form pluripotent haemopoietic colonies. ZnCl2 at concentrations 5 X 10(-6) to 5 X 10(-4) M inhibited the regional GVHR. At a concentration of 5 X 10(-6) ZnCl2 also inhibited the ability to elicit the systemic GVHR in irradiated mice. Haemopoiesis was stimulated in cells incubated with ZnCl2 at concentration 5 X 10(-6) M, it was inhibited after incubation of A/01a cells with 5 X 10(-4) M ZnCl2, whereas cells from non-inbred ICR mice were stimulated by the latter concentration of ZnCl2 for haemopoiesis. Li2SO4 inhibited the ability to induce the regional GVHR at concentrations of 5 X 10(-3) and 10(-2) M, had no effect on survival of mice during the systemic GVHR and exerted rather an inhibitory effect on haemopoiesis.
    No preview · Article · Feb 1986 · Folia biologica