Leg ulcer in hydroxyurea-treated patients.

A cutaneous ulcer is a lesser known complication of hydroxyurea treatment. Out of 39 patients [18 polycythaemia vera (PV), 13 essential thrombocythaemia (ET), 4 chronic myeloid leukemia (CML), 4 undefined myeloproliferative diseases (MPD)] treated with hydroxyurea, 6 (4ET, 1PV, 1CML) developed a cutaneous ulcer during a period of less that 2 years' treatment. In all but one of the patients the ulcers were situated in the ankle region. At the time of onset of ulceration, none of them had extreme values in their peripheral blood counts. All had one or more of the predisposing factors such as minor trauma or mild varicosity. None of the patients had any alteration in arterial or venous circulation when examined by non-invasive means. No hyperviscosity was found as measured by capillary viscosimeter. The ulcers were cured in three patients without discontinuation of the drug. One patient later developed an ulcer on the other leg. The ulcers healed in two patients only after having stopped the hydroxyurea medication. One patient still had the ulcer when she succumbed to the underlying CML in transformation. In conclusion, cutaneous, ulceration of the leg is relatively common during hydroxyurea therapy. Predisposing factors are also involved in its development. Its healing does not necessarily require the discontinuation of the drug.

Lymphocyte subset reconstitution after allogeneic bone marrow transplantation using radiation-free conditioning regimen for patients with chronic granulocytic leukemia.

Conditioning regimens for BMT are important in determining transplant outcome. A radiation-free protocol containing Mitobronitol (DBM), Cytarabine (Ara-C) and Cyclophosphamide (Cy) was used for conditioning of patients with chronic granulocytic leukemia (CGL). Using this conditioning treatment, fewer transplant related complications, including acute GVHD, VOD and severe infections, were observed. Acute GVHD did not develop, but chronic GVHD, accompanied with graft-versus leukemia, was present in half of the cases. To determine the clinical effect of the DBM/Ara-C/Cy conditioning, the recovery of peripheral blood lymphocytes was examined after allogeneic BMT for patients with CGL in comparison with TBI/Cy conditioning. The lymphocyte subsets of 11 DBM patients were followed and analyzed periodically (30-90 days, 4-12 months and > 13 months) using ten monoclonal antibodies and flow cytometry. Decreased percentage of total T cells as well as CD4+ and CD8+ subpopulations, significantly decreased T cell activation and increased proportion of TCR gamma delta + cells were found to be characteristic in the early post-transplant period in the DBM group. Early recovery and consistently higher percentage of B cells were observed for the whole follow-up period of patients receiving DBM conditioning. A high proportion of NK cells was observed in all transplant recipients. These findings suggest that the characteristic pattern of recovering lymphocytes is associated with the lack of severe transplant-related clinical complications following DBM/Ara-C/Cy conditioning.

Clinical significance of longitudinal complement measurements in recipients of bone marrow transplant.

Overall activity of classical complement pathway, C3 and C4 levels, level of circulating immune complexes and concentration of serum immunoglobulins were measured in 38 patients transplanted with HLA-identical bone marrow before and after transplantation for at least 4 years. Changes of complement parameters and their association with acute and chronic GVHD and with infections were analysed. A strong association was found between the development of chronic GVHD and hypercomplementaemia measured in 16 long-term survivors. Low pre-transplantation C4 activity was found to predict the development of severe acute GVHD. These findings indicate that longitudinal complement measurements may have clinical value in BMT patients.

[Experience with the molecular genetical detection of the chimera gene of the Philadelphia chromosome]. / Philadelphia chromosoma chimera génje molekuláris genetikai kimutatásának tapasztalatai.

Molecular genetical techniques could be developed for detection of the chimera gene of Philadelphia chromosome or that of its gene product, due to the relatively conserved structure of the chimera gene. The authors successfully analysed 123 blood/bone marrow samples from 106 patients using these molecular techniques adapted from the literature. Patients were classified by the first diagnosis, 65 CML, 7 AML, 13 ALL patients were studied. 12 patients had the diagnosis of myeloproliferative syndrome, and 9 patients were after bone marrow transplantation. 57% of the total, and by diagnosis, 74% of CML, 28% of AML, 54% of ALL, and 33% of post-transplant samples have shown the chimera gene structure characteristic for Philadelphia chromosome. All patients of myeloproliferative syndrome were negative. In some cases the authors had the opportunity to study simultaneously the peripheral blood and the bone marrow sample of the same patient and of the same date. The ratio of the positivity of the two samples varied from one to infinite. The authors could follow the effect of interferon in one case, the change of clonality of the leukemic cell line in an other case. They had the opportunity to detect two different abnormal gene structures in the sample of an AML patient.

TCR gamma/delta bearing lymphocytes in peripheral blood of allogenic bone marrow transplanted patients.

The presence of two distinct T-cell receptors (TCR) alpha/beta and gamma/delta dimers as well as of the activated T cells was analysed in peripheral blood mononuclear cells from seventeen recipients of allogeneic bone marrow transplants for leukemia and for severe aplastic anemia. Nine of seventeen recipients expressed an elevated percentage of T cells bearing TCR gamma/delta receptors in their peripheral blood. Seven out of nine cases having elevated gamma/delta positive cells showed chronic graft-versus-host (GVH) disease; one patient was treated with Cyclosporin A, and one patient was asymptomatic. In the twelve patients with GVH or other clinical symptoms, activated T cells (CD3+/HLA-DR+) were elevated indicating an autoreactive or alloreactive cell population. Our results confirmed earlier in vitro data showing that TCR-gamma/delta-bearing lymphocytes may be an activated T-cell population, and this T cell subset might be involved in mediating GVH disease, or in prolonging immunodeficiency after transplantation.

[Importance of the detection of minimal residual disease in the management of acute leukemia]. / Minimális reziduális leukaemia vizsgálatának jelentösége az akut leukaemiás betegek kezelésében.

Between 1984-1988, 57 adult acute leukemic patients were treated with intensive combined chemotherapy in the National Institute of Haematology and Blood Transfusion. For the evaluation of response to therapy, 4 investigations were performed in parallel: bone marrow aspirate, bone marrow biopsy, cytogenetic analysis and bone marrow culture. Nonparametric test for samples taken for the evaluation of remission status showed that bone marrow biopsy was significantly the most sensitive method for the detection of residual disease. The bone marrow culture was also on the borderline of significance, but the low CFU-GM level did not always correlate with the further clinical course. Occasionally, karyotypic abnormality was the only sign of the residual disease. It would be of great importance to quantitate the minimal residual disease in order to evaluate and compare the various intensive postinduction therapeutic strategies.

Growth kinetics and blast-colony forming cell binding capacity of stromal cells in various haematological malignancies.

Growth kinetics of bone marrow stromal layers from normal, AML, ALL and CML patients was studied. Significantly reduced time for confluency was observed in AML patients in complete remission, in CML patients in chronic phase, or CML patients after allogenic bone marrow transplantation. The functional capacity of these stromal layers did not differ: they all bound similar amounts of blast colony forming cells (BL-CFC) from normal bone marrow. The stromal layers from bone marrow transplanted patients varied in their BL-CFC binding capacity: two CML patients (10.5 and 49 months after transplantation) showed normal values, while two ALL patients (1.5 and 3 months, respectively, after transplantation) as well as one patient transplanted for CML (19.5 months after transplantation) showed significantly reduced BL-CFC binding capacity.

Growth kinetics and blast-colony forming cell binding capacity of aplastic anaemic stromal cells.

The kinetics of bone marrow cell growth and a special function of stromal cells (the capability of binding blast colony forming cells) were studied in patients with aplastic anaemia (AA). All 10 patients studied showed faster growth of bone marrow stromal cells. The time for a confluent stromal layer formation was 24.5 days for AA bone marrow as opposed to 33.0 days for normal bone marrow. This faster growth rate could also be observed if normal bone marrow cells, depleted of plastic non-adherent fraction, were plated, suggesting that at least one of the reasons for altered stromal cell growth kinetics in AA is the changes in the ratio of plastic adherent/non-adherent cells. Functionally, i.e. in supporting the growth of normal bone marrow blast colonies, AA stromal layers did not differ from that of normal stromal layers, independently of the clinical state of the disease (AA or SAA; in one patient before or after ATG treatment; in two patients after successful allogenic bone marrow transplantation). Moreover, in some AA patients this blast colony forming cell binding function of AA stromal layers could also be detected in samples cultured without hydrocortisone (i.e. in the absence of fat cells), suggesting that AA stroma also differs qualitatively from normal stroma without inducing a defective microenvironment for stem cell homing.

Stromal cell growth and blast colony-forming cells in AML bone marrow.

Bone marrow cells (BMC) from normal donors and from patients with acute myeloid leukaemia (AML) were cultured. Growth kinetics and the efficiency of stromal layers in supporting the adhesion of normal blast-colony forming cells (BL-CFCs) were studied. BMC from treated AML patients formed confluent stromal layers faster than normal BMCs. BL-CFC binding capacity of normal and AML stromal layers did not differ: on normal stromal layers 67.3-147, on AML stromal layers 63-117 colonies per 5 x 10(5) plastic non-adherent BMC were formed. The amount and/or binding capacity of BL-CFCs was found to be normal in two AML patients in complete remission, while a significantly reduced number and/or binding capacity of BL-CFCs was found in AML non-treated patients and in patients within 4 weeks after the last cytostatic course.

[Cytochemical, immunologic and gene rearrangement studies in adult acute leukemia]. / Cytochemiai, immunológiai és génátrendezödés vizsgálatok felnöttkori akut leukaemiákban.

Results of morphological, cytochemical and immunological studies performed in adult acute leukaemias have been compared. Thirty one cases proved to be acute myeloid leukaemia, while 25 cases were shown to be acute lymphoid leukaemia. Based on our results we conclude that immunophenotyping with monoclonal antibodies does not help in distinguishing the subtypes of AML. For purposes of clinical diagnosis cytochemical methods are valuable. On the other hand the monoclonal antibodies are essential in distinguishing the very immature myeloid and lymphoid leukaemias and this is of great importance from the clinical point of view, in determining therapy. Moreover, the diagnosis of acute lymphoid leukaemias is not possible without the specific monoclonal antibodies. Their application is first of all in haematological centers caring for leukaemia patients nowadays already obligatory. Gene rearrangement studies make the diagnosis more accurate and help in the diagnosis of leukaemias of unknown immunological origin.

[Gene rearrangement in leukemia]. / Génátrendezödés leukaemiákban.

In this report we summarize our experiences based on the gene rearrangement study of 111 leukaemic patients of different kind. The lymphocyte DNA of the patients was studied for rearrangement of the immunoglobulin light chain constant-, the heavy chain joining- and the T cell receptor beta chain constant region. Our data have well supplemented the results of the monoclonal antibody experiments. In 33 cases the DNA study was in good agreement with the immunological data. In 42 our data helped in gave different results, immunological results. In 11 cases evaluating the DNA and immunological data indicating the necessity of further investigation. The results were inconclusive in 25 cases. As a conclusion we consider the gene rearrangement study to be useful for diagnostic purposes.

Clinical value of cytomorphologic, immunologic and cytogenetic investigations of acute leukaemias.

The valuability of immunophenotyping of acute myeloid and lymphoid leukaemias in comparison to morphological and cytochemical classification were approached in 56 cases. In the case of acute myeloid leukaemias the immunophenotyping by monoclonal antibodies CD14, CD13, CD33 was less informative concerning the subtypes of the disease. The clinical diagnosis can be achieved on the basis of cytochemical investigation alone. In contrast, the diagnosis of lymphoid leukaemias requires all information obtained by immunophenotyping by a series of monoclonal antibodies CD3, CD2, CD4, CD8, CD1, CD19, CD20, CD21 and CD10. On the other hand, the monoclonal antibodies are essential in differentiation of the very immature myeloid and lymphoid leukaemias. This is of great importance from the clinical point of view for determining the therapy. Molecular genetic studies based on the characterisation of the state of gene rearrangement of immunoglobulin and T-cell receptor beta chains have basic importance in the confirmation of the result of immunophenotyping and in the determination of leukaemias of unknown origin.

[Granulocyte-macrophage progenitor cells in allogenic bone marrow transplantation: correlation of progenitor cell content and regeneration in the graft]. / Granulocyta-makrophag össejtszint vizsgálata allogén csontvelöátültetésben: az oltvány össejttartalma és a regeneráció összefüggése.

The amount of granuloid macrophage progenitors (CFU-GM) was studied in 16 donor bone marrows used for allogenic bone marrow transplantation in the National Institute of Haematology and Blood Transfusion between January, 1984 and January, 1988. In 10 bone marrow transplanted patients long-term follow up of bone marrow CFU-GM regeneration was carried out. Graft sizes were the following: 2.91 +/- 0.62 X 10(8)/kg body weight nucleated cells and 19.2 +/- 14 X 10(4)/kg body weight (CFU-GM. Preinfusion procedures (centrifugation and resuspension) did not alter CFU-GM content of the grafts. Separation of nucleated cells with hydroxyethylstarch, applied for ABO mismatched donor bone marrow, however, resulted in a 30 per cent loss in CFU-GM. Since higher than threshold graft-sizes for successful engraftment were used, no linear correlation between graft size and speed of granulocyte and platelet recovery was found. Significant difference between regeneration kinetics of bone marrow CFU-GM of patients transplanted for CML or AML and ALL was observed: in AML and ALL patients normal bone marrow CFU-GM level was found 4 to 6 months after transplantation, while in CML patients CFU-GM level approached the lower limit of the normal value only 10 to 14 months after transplantation. Granulocyte and thrombocyte recovery of CML patients showed a significant delay when compared to transplanted AML and ALL patients.

Results of immunosuppression in 170 cases of severe aplastic anaemia. Report of the European Group of Bone Marrow Transplant (EGBMT).

This is a report of the European Group of Bone Marrow Transplant (EGBMT) on the influence of immunosuppression (IS) on survival of patients with severe aplastic anaemia (SAA). Fourteen teams participated in this survey involving 170 cases treated from 1974 to December 1980. The 1 year survival was 62.7%. Three types of treatment were used: (1) anti-thymocyte globulin (ATG) alone, (2) anti-thymocyte globulin and haplo-identical related bone marrow infusion, and (3) high dose bolus-6-methyl-prednisolone (b-6 MePr). There was no statistical difference in survival between these three groups. Androgens did not modify survival. Blood counts before treatment had a significant prognostic value. Patients with less than 0.2 X 10(9)/l granulocytes and less than 10 X 10(9)/l reticulocytes had 40% 1 year survival; the others had more than 70% 1 year survival. Patients with complete or partial reconstitution had the same good prognosis. In contrast, patients with no improvement after IS had a 27% 1 year survival. Several successive courses of IS improved the prognosis of non-responding patients. This survey confirms that IS improves the survival of patients with SAA. A prospective study will be performed to define the best and safest form of IS and to correlate clinical results with in vitro tests.