bluuebunny/crossref_metadata_2025 huggingface dataset:
Detection of donor cell derived acute myelogenous leukaemia in a patient transplanted for chronic myelogenous leukaemia using fluorescence in situ hybridization
Authors: Lowsky Robert , Fyles Gillian , Minden Mark , Lipton Jeffrey , Meharchand Jacinta , Tejpar Ismet , Zipursky Alvin , Messner Hans
Published: October 2003

The recurrence of leukaemia following allogeneic bone marrow transplantation appears to develop rarely in donor cells. However, the standard method for assigning the origin of recurrence, metaphase analysis, can be unreliable. We have applied the technique of fluorescence in situ hybridization (FISH) directly on archival Wright stained bone marrow slides obtained from a patient who developed acute myelogenous leukaemia (AML) following allogeneic bone marrow transplantation (BMT) for chronic myelogenous leukaemia (CML). Using a chromosome‐specific DNA probe we linked a chromosomal aberration, previously detected by conventional metaphase analysis, directly to morphologically identifiable blast cells. In this way we were able to assess cell‐lineage involvement of the secondary leukaemia and assign a donor origin.
Discordant clinical presentation and outcome in infant twins sharing a common clonal leukaemia
Authors: Campbell Myriam , Cabrera Maria Elena , Legues Maria Eugenia , Ridge Susan , Greaves Mel
Published: October 2003

We report the different presentation features and clinical outcome between two identical infant twins with acute lymphoblastic leukaemia with a shared clonal disease and MLL gene rearrangement. One twin relapsed and died, but the other is in complete remission >4 years after diagnosis. These data, and similar observations on other twin infants with leukaemia, suggest that despite a common clonal in utero, post‐natally these leukaemias can evolve independently, at different rates, in the twinned individuals, and that the usually fatal leukaemia associated with t(4;11) MLL gene rearrangement can be effectively treated when the leukaemic burden is small.
Expression of multidrug resistance P‐glycoprotein in myeloid progenitor cells of different phenotype: comparison between normal bone marrow cells and leukaemia cells
Authors: TAKESHITA AKIHIRO , SHINJO KAORI , OHNISHI KAZUNORI , OHNO RYUZO
Published: October 2003

We examined the multidrug resistant P‐glycoprotein (P‐gp) on normal bone marrow (BM) cells and acute myeloid leukaemia (AML) cells, using newly devised flow cytometric multi‐parameter analysis with CD33, CD34 and MRK16 monoclonal antibodies. In both normal BM cells and AML cells, CD34+CD33− cells expressed P‐gp strongly, CD34+CD33+ cells moderately, and CD34−CD33+ cells weakly. Acute promyelocytic leukaemia, mainly expressing CD34−CD33+ but not CD34+CD33− at diagnosis, expressed less P‐gp. P‐gp expression of AML cells at diagnosis was increased as compared with normal cells of the same phenotype. P‐gp expression was more increased in relapsed cases, especially in immature subpopulations.
Changes in the pattern of distribution of von Willebrand factor in rat aortic endothelial cells following thrombin generation in vivo
Authors: Senis Yotis A. , Richardson Mary , Tinlin Shawn , Maurice Donald H. , Giles Alan R.
Published: October 2003

The pattern of distribution of von Willebrand factor (VWF) in relatively large sheets of rat aortic endothelial cells (EC) obtained by the Häutchen technique were analysed by immunocytochemistry and light microscopy. EC were examined pre and post administration of a procoagulant mixture of factor Xa (F.Xa) and phosphotidylcholine/phosphotidylserine (PCPS) vesicles which was demonstrated to result in the selective loss of high molecular weight multimers (HMWM) of plasma VWF in the rat. In placebo animals the pattern was heterogenous both in overall distribution and in individual cells which showed both a diffuse and granular pattern. Groups of intensely stained EC were oriented parallel to the longitudinal axis of the aorta and staining was particularly prominent around the orifices of the intercostal arteries, implicating shear‐stress as a possible factor in VWF expression by EC. Changes in the pattern of distribution of staining were observed at various time points post‐infusion of F.Xa/PCPS, suggesting the immediate release of VWF from EC stores followed by the recruitment of EC to synthesize and store VWF. These changes are consistent with the decrease in EC Weibel‐Palade body (WPB) content observed by EM in previously reported studies using this model.
Serum p53 accumulation and altered antibody responses to Epstein‐Barr virus proteins precede diagnosis of haemopoietic malignancies of lymphoid origin
Authors: Lehtinen T. , Luostarinen T. , Dillner J. , Aromaa A. , Hakama M. , Hakulinen T. , Knekt P. , Leinikki P. , Lumio J. , LÄhdeaho M.‐L. , Maatela J. , Teppo L. , Lehtinen M.
Published: October 2003

We analysed antibodies to Epstein‐Barr virus nuclear antigens (EBNAs 1, 2, 5 and 6) and the presence of serum p53 in 100 individuals, 37 of whom had developed a haemopoietic malignancy during a 12‐year follow‐up of 39 000 Finnish adults. Serum p53 was detectable in six of the 63 (10%) matched controls and in 13/31 (42%) patients who developed a malignancy of lymphoid origin approximately 7 years after serum withdrawal. Six patients who developed a malignancy of myeloid origin were negative for p53. The presence of p53 alone was associated with a highly significant increased risk of lymphoid malignancies (relative risk (RR)p53 = 6.7, 95% confidence limits (CL) 1.9, 24) whereas high levels of antibody to EBNA2 seemed to be inversely related to the risk (RREBNA2 = 0.1, CL 0.0, 1.1). Among lymphoid malignancies, a combination of serum p53 and high EBNA1 antibody levels gave a greater than expected risk (RRp53  and  EBNA1 = 14, CL 1.4, 130; RRexpected = 4.4), whereas interaction with high levels of EBNA5 antibody gave an expected risk (RRp53 and EBNA5 = 19, CL 1.7, 220; RRexpected = 17). Thus detectable levels of p53 appear early in the development of lymphoid malignancies, and high EBNA1 antibody levels, and accumulated p53 may both be synergistic risk indicators for lymphoid malignancies, whereas high EBNA5 antibody levels and accumulation of p53 seem to raise the RR independently of each other.
Treatment of homozygous protein C deficiency with subcutaneous protein C concentrate
Authors: MINFORD A. M. B. , PARAPIA L. A. , STAINFORTH C. , LEE D.
Published: October 2003

Subcutaneous protein C concentrate (Immuno, Vienna) was used to treat a child with homozygous protein C deficiency who was formerly treated with intravenous protein C concentrate. After 3000 units subcutaneous protein C concentrate (250 iu/kg), protective protein C levels were maintained for 48h after infusion, with peak levels at 12h. Subcutaneous protein C concentrate is given every third day and is well tolerated by the patient. No thrombotic events have occurred. We conclude that subcutaneous administration of protein C concentrate is a valuable therapeutic option in the long‐term management of homozygous protein C deficiency and avoids the potential hazards of long‐term central venous lines.
Soluble interleukin‐6 receptor (sIL‐6R), a new prognostic factor in multiple myeloma
Authors: KYRTSONIS MARIA‐CHRISTINA , DEDOUSSIS GEORGES , ZERVAS CONSTANTIN , PERIFANIS VASSILIS , BAXEVANIS CONSTANTIN , STAMATELOU MARINA , MANIATIS ALICE
Published: October 2003

sIL‐6R is a 55 kD soluble molecule mediating the interleukin‐6 (IL‐6) signal through the IL‐6 receptor‐associated transmembrane signal transducer, gp130. It has recently been suggested that sIL‐6R serum levels may reflect disease severity in multiple myeloma (MM). We determined sIL‐6R serum levels in 25 normal controls (NC) and in 80 MM patients at diagnosis and during the course of the disease. Measurements were done by ELISA. In NC, sIL‐6R levels ranged from 14 to 40 ng/ml (median 28 ng/ml) whereas in MM patients the range was 10–200 ng/ml (median 38 ng/ml) (P < 0.01). 61 patients entered remission and 19 were resistant. Median sIL‐6R value at diagnosis was 36 ng/ml (10–120) in responding patients, and 82 ng/ml (20–200) in non‐responding patients (P < 0.001). During a follow‐up from 12 to 89 months, sIL‐6R values remained more or less stable in most patients. High sIL‐6R levels correlated with poor survival.
The effect of the GM‐CSF/IL‐3 fusion protein PIXY321 on bone marrow and circulating haemopoietic cells of previously untreated patients with cancer
Authors: Ghielmini Michele , Pettengell Ruth , Coutinho Lucia H. , Testa Nydia , Crowther Derek
Published: October 2003

This is a phase I/II study of the GM‐CSF/IL‐3 fusion protein PIXY321. Patients were treated with PIXY321 at a daily subcutaneous dose of 500, 750 and 1000 μg/m2 for 14 d. Side‐effects were mild and consisted mainly of injection‐site reactions and constitutional symptoms. A biphasic modest increase of white blood count (2.5‐fold) and platelets (1–1.5‐fold) was seen, accompanied by an increased bone marrow cellularity and an increase in circulating progenitors. Colony‐forming cells in the blood rose to a median of 184 granulocyte/macrophage‐colony forming cells (GM‐CFC)/ml, eight Mix‐CFC/ml, 250 burst forming units‐erythroid (BFU‐E)/ml and 140 CFU‐megakaryocytes/ml, corresponding to a 10‐, 2.5‐, 8‐ and 30‐fold increase respectively. When seeded for long‐term culture on irradiated bone marrow stroma, the mobilized cells were not able to sustain haemopoiesis in vitro to the same degree as bone marrow. Taken together, these results indicate that PIXY321 has a biological effect in humans more similar to that of IL‐3 than to that of GM‐CSF.
Macrophages are the major target cell for HIV infection in long‐term bone marrow culture and demonstrate dual susceptibility to lymphocytotropic and monocytotropic strains of HIV‐1
Authors: Gill Vikki , Shattock Robin J. , Freedman Andrew R. , Robinson Grant , Griffin George E. , Gordon‐Smith Edward C. , Gibson Frances M.
Published: October 2003

Haematological abnormalities are often seen in patients infected with HIV. A number of mechanisms are thought to contribute to this bone marrow suppression, including impaired stromal function and direct infection of progenitor cells. Evidence suggests that both bone marrow progenitor cells and perhaps stromal cells are open to infection by HIV, which raises the possibility that bone marrow stromal cells may serve as a reservoir for HIV. This study investigated the cellular targets and kinetics of in vitro infection of stroma in long‐term bone marrow culture (LTBMC) using both mono‐ and lymphocytotropic strains of HIV‐1. p24 ELISA and reverse transcriptase (RT) assay demonstrated that stroma could be infected with HIV and release infectious virions. The target cells for infection were shown to be macrophages by immunohistochemistry (APAAP), dual immunofluorescence staining (using CD68 and p24) and electron microscopy.

The data show that it was possible to infect stroma in LTBMC with HIV and that such infection was productive. The main target for infection was bone marrow macrophages. In contrast to peripheral blood derived macrophages, these cells were susceptible to both lymphocytotropic and monocytotropic strains of HIV‐1. The data suggests that these bone marrow macrophages may act as a reservoir for HIV. Infection of bone marrow macrophages may affect haemopoiesis either by transmission of HIV infection to developing progenitor cells through direct cell‐to‐cell contact or by altering the ability of the stroma to support normal development.
Long‐term salvage treatment by cyclosporin in refractory autoimmune haematological disorders
Authors: Emilia Giovanni , Messora Carlo , Longo Giuseppe , Bertesi Marcello
Published: October 2003

We report on long‐term treatment (13–62 months) with cyclosporin A (CyA) in eight patients with autoimmune haematological disorders, resistant to all usual therapies. Three patients had an autoimmune haemolytic anaemia (AHA); four an idiopathic thrombocytopenic purpura (ITP), and one an Evans syndrome. All patients were responsive: six achieved complete remission and two partial remission. The side‐effects were moderate and transient. The majority of surviving patients remain dependent on continued drug administration. The CyA appears to be recommendable as a salvage treatment in life‐threatening, resistant autoimmune haematological diseases.
THROMBO‐EMBOLIC DISEASE AFTER SPLENECTOMY FOR HEREDITARY STOMATOCYTOSIS
Authors: Stewart Gordon W. , Amess John A. L. , Eber Stefan W. , Kingswood Christopher , Lane Peter A. , Smith Brian D. , Mentzer William C.
Published: October 2003

Nine cases of hereditary stomatocytosis (HSt) are presented which show documented thrombotic complications after splenectomy. In three cases, patients became severely ill with pulmonary hypertension and a fourth developed portal hypertension. One unsplenectomized affected adult relative had suspected but unconfirmed thrombotic pathology; the six other affected unsplenectomized adults did not. Since splenectomy is of only limited therapeutic benefit in stomatocytosis, it should be not be performed without careful consideration. A tendency to iron overload, even without hypertransfusion and irrespective of splenectomy, is evident in many of these patients.
Graft‐versus‐leukaemia activity can be predicted by natural cytotoxicity against leukaemia cells
Authors: Glass Bertram , Uharek Lutz , Zeis Matthias , Loeffler Helmut , Mueller‐Ruchholtz Wolfgang , Gassmann Winifried
Published: October 2003

We have investigated graft‐versus‐leukaemia (GVL) effects after allogeneic bone marrow transplantation (BMT), using three murine leukaemia models, A20 (B lymphocytic), WEHI‐3 (myelomonocytic) and PU5‐1R (myeloid). Injection of leukaemia cells in a high number (106 cells) into syngeneic Balb/c mice (H‐2d) invariably led to death with a median survival time of 22 d (A20), 18 d (WEHI‐3) and 45 d (PU5‐1R). A lower tumour load of A20 (5 × 105 cells) was used in some experiments resulting in a leukaemic death rate of 94%. Lethal total‐body irradiation followed by syngeneic BMT prolonged survival (P < 0.05) for animals bearing the leukaemia A20 and WEHI‐3 but was unsuccessful for animals injected with cells from the monocytic leukaemia PU5‐1R. Graft‐versus‐host (GVH)‐nonreactive marrow of (C57 × Balb/c)F1 mice (H2bxd) exerted a significant GVL‐effect with reduced relapse rate and improved survival in mice receiving the leukaemia cell line A20. In animals with low tumour load a significant reduction of the relapse rate from 82% following syngeneic BMT to 47% following allogeneic, GVH‐nonreactive BMT could be achieved. Depletion of natural killer (NK) cells from the GVL‐reactive semi‐allogenic bone marrow graft enhances the relapse rate of the leukaemia A20 to 65%. In mice bearing the leukaemias WEHI‐3 or PU5‐1R allogeneic GVH‐nonreactive BMT did not improve survival compared to syngeneic BMT. Transplantation of GVH‐reactive bone marrow from DBA mice (MHC identical to Balb/c, minor difference) caused only a limited and insignificant reduction of relapse rate for animals with the leukaemia A20. These in vivo data are in close correlation with in vitro natural killer cell (NK) activity of the donor strains against the respective leukaemia targets. Depletion of NK cells from the GVL‐reactive (C57 × Balb/c)F1 bone marrow resulted in a significant loss of GVL activity. We conclude that NK cells are involved in graft‐versus‐leukaemia effects independent of graft‐versus‐host disease (GVHD).
Generation of activated natural killer (A‐NK) cells in patients with chronic myelogenous leukaemia and their role in the in vitro disappearance of BCR/abl‐positive targets
Authors: Silla Lucia M. R. , Pincus Steven M. , Locker Joseph D. , Glover Janice , Elder Elaine M. , Donnenberg Albert D. , Nardi N. B. , Bryant John , Ball Edward D. , Whiteside Theresa L.
Published: October 2003

Activated natural killer (A‐NK) cells, a subset of CD56dimCD3– lymphocytes, are obtained from PBMC of normal donors by adherence to plastic and culture in the presence of IL2. In this study we tested the feasibility of generating A‐NK cells in patients with Ph+ chronic myeloid leukaemia (CML). Cultures obtained from patients with early chronic phase (ECP; n=7) contained a mean (±SD) of 83 ± 7% of CD56+CD3– cells, and those from patients with advanced chronic phase (ACP; n=7) contained 27 ± 33% CD56+CD3– cells. In three patients with leukaemia in a blastic phase (BP) it was only possible to obtain one culture enriched in CD56+CD3– cells (81%). Cellular aggregates of myeloid cells and large granular lymphocytes were observed in early A‐NK cell cultures. Paired freshly‐adherent and cultured A‐NK cells were tested for the presence of BCR/abl mRNA by RT‐PCR. The BCR/abl+ cells were detected in all 12 preparations of the freshly adherent A‐NK cells tested. In 6/12 the BCR/abl+ cells were no longer detectable by RT‐PCR on day 14 of culture. Both proliferation and antileukaemic cytotoxicity were significantly higher (P = 0.002 and P = 0.029, respectively) in the BCR/abl– cultures than those in the six BCR/abl+ cultures. 5/6 BCR/abl– cultures were highly enriched in A‐NK cells on day 14, and 1/6 contained predominantly CD56+CD3+ cells. Only 2/6 BCR/abl+ cultures were enriched in A‐NK cells on day 14, but they had poor cytotoxicity and a low proliferative index. Myeloid cells (CD33+) were more frequently detected in the BCR/abl+ than BCR/abl– A‐NK cell cultures (P = 0.028). These observations suggest that: (1) populations of benign A‐NK cells can be generated from the peripheral blood of CML patients; (2) the ability to generate A‐NK cells is impaired in patients with advanced CML; and (3) the ability to generate A‐NK cells with antileukaemic activity correlates with the disappearance of BCR/abl+ cells from these cultures.
Idiopathic myelofibrosis in children
Authors: SEKHAR M. , PRENTICE H. G. , POPAT U. , ANDERSON D. , JANMOHAMMED R. , ROBERTS I. , BRITT R. P.
Published: October 2003

Childhood myelofibrosis (Mf ) is rare with variable outcome reported in the literature. We present clinical and investigative details of three children who presented with idiopathic Mf in early childhood. Two of these children were identical twins and have been haematologically stable over the past 7 years since their diagnosis. The third patient underwent an allogeneic bone marrow transplant (BMT) procedure as her clinical status was deteriorating. She remains engrafted at 13 months post BMT. None of the children had hepatosplenomegaly although extramedullary haemopoiesis was demonstrated on 52Fe studies in two patients. Circulating progenitors in these patients were increased in the face of reduced marrow precursors. The aetiology of childhood Mf is unclear and its natural history seems different from the adult disease. Allogeneic BMT can be an option for definitive treatment.
Cyclosporine A alleviates severe anaemia associated with refractory large granular lymphocytic leukaemia and chronic natural killer cell lymphocytosis
Authors: Bible Keith C. , Tefferi Ayalew
Published: October 2003

Large granular lymphocytic (LGL) leukaemia and chronic natural killer cell lymphocytosis (CNKL) are chronic indolent disorders often associated with neutropenia and constitutional symptoms. Severe anaemia occurs in about 20% of patients and is currently treated with corticosteroids followed by oral cyclophosphamide in non‐responders. 30% of patients fail initial measures, and salvage therapy is inadequate. We describe three transfusion‐dependent patients (two with T‐LGL leukaemia, one with CNKL) refractory to corticosteroids, cyclophosphamide, and in one case fludarabine. Cyclosporine A (CSA) initiation resulted in prompt transfusion‐independence and was well tolerated in all patients, making it an attractive alternative therapy for this disorder.
Leucocyte filterability: comparing diluted blood with purified cell suspensions
Authors: Evans Shelley‐Ann
Published: October 2003

Current rheological trends in filtration studies involve purification of cell subpopulations in order to study near‐homogenous populations of cells, and this means that time‐consuming and possibly cell‐damaging procedures are used. Filtration of diluted blood offers the possibility of determining the properties of subpopulations of cells with minimal cell manipulation. The filtration properties of granulocytes and lymphocytes result in their being detected as one kinetic population, and therefore a combined granulocyte/lymphocyte pore transit time is calculated. There was no significant change seen in the combined granulocyte/lymphocyte transit time over the normal range of differential count. Filtration of diluted blood is a more reproducible way of measuring granulocyte and lymphocyte filterability, because purification of these cells gives rise to more scatter and larger ranges in the measured pore transit time, and significant day‐to‐day variation. A combined granulocyte/lymphocyte transit time is therefore an acceptable and reproducible way of assessing leucocyte rheology in diluted blood, when the differential count lies within the normal range. If changes in the granulocyte/lymphocyte transit time are detected, then cell purification may yield additional information on the subpopulations. Therefore, particularly in pathological samples, a combination of these approaches will yield more information than either test alone.
ACUTE MYELOBLASTIC LEUKAEMIA PRESENTING WITH HERPES SIMPLEX TYPE‐1 VIRAEMIA AND PNEUMONIA
Authors: James E. , Robinson L. , Griffiths P. D. , Prentice H. G.
Published: October 2003

We report a patient with acute myeloblastic leukaemia who presented with a pneumonia and herpes simplex viraemia associated with primary herpes simplex virus‐1 infection. The importance of detecting and treating viral infections in haematology patients is discussed.
STEM CELL FACTOR AND THE REGULATION OF DENDRITIC CELL PRODUCTION FROM CD34+ PROGENITORS IN BONE MARROW AND CORD BLOOD
Authors: Saraya K. , Reid C. D. L.
Published: October 2003

Dendritic cells (DC) are essential for the presentation of antigen in primary immune responses and they develop from CD34+ cells in the bone marrow. Although both granulocyte macrophage colony stimulating factor (GM‐CSF) and tumour necrosis factor (TNF) are known to stimulate the development of mature DC from their progenitor (CFU‐DL), the function of stem cell factor (SCF) in this pathway remains to be determined. The interactions of SCF with GM‐CSF, TNF, interleukin‐3 (IL‐3) and macrophage colony stimulating factor (M‐CSF) in promoting CFU‐DL development have now been studied in serum‐free cultures of unfractionated as well as progenitor enriched cells from either bone marrow or cord blood. Although SCF alone is without effect on colony formation, it enhances both the numbers and size of DC colonies generated in vitro by GM‐CSF and TNF. It acts directly on progenitors and in the presence of GM‐CSF can also induce suboptimal DC growth even in the absence of TNF. SCF appears to recruit very early progenitors of a high proliferative potential with the capacity to differentiate into erythroid and myeloid as well as dendritic cell progeny. In combination with other cytokines it may therefore be useful for the ex vivo generation of large numbers of DC for clinical purposes.
Nerve growth factor enhances survival and cytotoxic activity of human eosinophils
Authors: Hamada Atsuo , Watanabe Naohiro , Ohtomo Hiroshi , Matsuda Hiroshi
Published: October 2003

Phenotypic change of blood‐type eosinophils to tissue‐type eosinophils is induced by various cytokines. We examined the effect of nerve growth factor (NGF) as a candidate for a constitutive cytokine which is able to induce the phenotypic change of eosinophils. The viability of human peripheral blood eosinophils cultured for 4 d was increased from a mean of 26% to a mean of 50% (P < 0.001) by the addition of NGF (50 ng/ml). Cytotoxic activity of eosinophils determined by larvicidal activity was increased 2–3‐fold by the addition of NGF (50 ng/ml) (P < 0.05). Furthermore, eosinophil chemotactic activity of NGF was demonstrated by the blind well chamber method. Since NGF is produced constitutively from various kinds of cells in local tissues, it is suggested that NGF might be a cytokine responsible for phenotypic change to tissue type eosinophils in conditions without immune stimuli.
Transmission of symptomatic parvovirus B19 infection by clotting factor concentrate
Authors: Yee T. T. , Cohen B. J. , Pasi K. J. , Lee C. A.
Published: October 2003

The risk of acquiring diseases from transfusion of blood and blood products is well recognized and the issue of parvovirus in haemophiliacs is not a new one. We report two patients with haemophilia acquiring iatrogenic parvovirus B19 infection, resulting in life‐threatening sepsis in one, an immunocompetent adult. Over the last 10 years there has been great progress in manufacturing safer products with regard to enveloped viruses such as HIV, hepatitis B and C. A recent outbreak across Europe of hepatitis A in haemophiliacs treated with plasma‐derived factor VIII concentrates has made haemophilic treaters concerned about the known (parvovirus B19 and hepatitis A) and the unknown non‐lipid enveloped viruses that may be contained in the clotting factor concentrates, because these are resistant to the existing viral inactivating techniques. The possibility of HIV itself mutating into a non‐lipid enveloped virus emphasizes the need to seek and use safer products.
Presenting Features and Prognosis in 72 Patients With Multiple Myeloma Who Were Younger Than 40 Years
Authors: Bladé Joan , Kyle Robert A. , Greipp Philip R.
Published: October 2003

The purpose of this study was to analyse the presenting clinical and laboratory features and the outcome of 72 patients with multiple myeloma (MM) who were younger than 40 years. The records of all Mayo Clinic patients with MM younger than 40 years who were seen between 1 January 1956 and 31 December 1992 were reviewed. Survival was measured from the date when treatment was required to the date of last follow‐up or death. The frequency of MM in patients younger than 40 and 30 years in 3278 Mayo Clinic patients was 2.2% and 0.3%, respectively. The main presenting clinical features were bone pain (66%), fatigue (26%), extramedullary plasmacytomas (19%) and bacterial infection (11%). Renal function impairment (creatinine level ≥ 177 μmol/l) and hypercalcaemia (serum calcium value ≥2.75 mmol/l) occurred in 29% and 30% of patients, respectively. Among the 57 patients evaluable for response the objective response rate was 54%. 14/35 patients treated with a single alkylating agent achieved an objective response, whereas 17/22 patients given combination chemotherapy had an objective response (P = 0.013). However, this higher response rate did not result in a significantly longer survival. The median survival for the 72 patients was 54 months. Patients with good prognostic features (normal renal function or low β2‐microglobulin level) had a median survival of 8 years. The actuarial survival at 5 and 10 years after initiation of therapy was 43% and 13%, respectively. In summary, survival in very young patients with myeloma is longer than that observed in series of patients of all ages, especially in those with good prognostic factors.
Low incidence of specific anti‐platelet antibodies detected by the MAIPA assay in the serum of thrombocytopenic MDS patients and lack of correlation between platelet autoantibodies, platelet lifespan and response to danazol therapy
Authors: HEBBAR M. , KAPLAN C. , CAULIER M. T. , WATTEL E. , MOREL P. , WETTERWALD M. , BAUTERS F. , FENAUX P.
Published: October 2003

We prospectively studied the mechanism of thrombocytopenia in 30 patients with myelodysplastic syndrome (MDS), who had a platelet count ≤70×109/l , and marrow blasts ≤10% by analysis of platelet lifespan, platelet‐bound IgG antibodies by platelet radioactive antiglobulin test (PRAT), and circulating and platelet‐bound autoantibodies specifically directed against platelet glycoproteins by indirect and direct MAIPA test.  PRAT analysis was positive in 21/30 patients (70%), but antiplatelet antibodies were found in only eight by the MAIPA test. Platelet lifespan was shortened in five cases, only one of whom had a positive MAIPA test. Patients were treated by danazol (600 mg/d), and 9/26 evaluable patients (35%) responded, including three of the five patients with shortened platelet lifespan.  No correlation between platelet lifespan, results of PRAT and MAIPA tests, and response to danazol was found.  In conclusion, specific antiplatelet autoantibodies, reduced platelet lifespan, and efficacy of androgen therapy on platelets were seen in 20%, 17% and 35% of MDS with thrombocytopenia, respectively, but no correlation between these findings was found in this study.
Multiplex PCR for rapid detection of T‐cell receptor‐gamma chain gene rearrangements in patients with lymphoproliferative diseases
Authors: Födinger Manuela , Buchmayer Heidi , Schwarzinger Ilse , Simonitsch Ingrid , Winkler Karin , Jäger Ulrich , Knobler Robert , Mannhalter Christine
Published: October 2003

We describe a multiplex polymerase chain reaction (PCR) method suitable for the detection of all T‐cell receptor (TCR) γ‐chain gene rearrangements in patients with lymphoproliferative diseases. 40 patients with various lymphoproliferative disorders and 40 healthy individuals were tested. Clonal TCRγ rearrangements were identified in all patients with malignant disorders, and in one of 10 cases with established reactive lymphocytosis but not in normal controls. In all individuals testing positive, the patient’s specific V and J segment involved in the rearrangement could be determined by simply splitting the multiplex primer mix. Our data show that the multiplex PCR technique enables rapid, simple and sensitive screening for clonal TCRγ chain gene rearrangements.
p16INK4A gene homozygous deletions in human acute leukaemias with alterations of chromosome 9
Authors: Faienza Maria Felicia , Ragione Fulviodella , Basso Guiseppe , Coppola Brigida , Miraglia del Giudice Emanuele , Schettini Francesco , Iolascon Achille
Published: October 2003

Acute leukaemias are characterized by non‐random chromosomal aberrations which are often strictly related to the inactivation of tumour suppressor genes (TSGs). Alterations at the short arm of chromosome 9 have been reported in a remarkable percentage of acute lymphoblastic leukaemias (ALL) and have been suggested to cause the loss of activity of the putative TSG, p16INK4A (MTS1/CDKN2) gene. In order to evaluate the correlation between this gene inactivation and visible cytogenetic abnormalities, we have investigated p16INK4A homozygous gene deletions in 10 paediatric acute leukaemias of different cell lineages which demonstrated karyotype aberrations involving chromosome 9. Moreover, the dimension of the genetic alteration was evaluated by studying the loss of heterozygosity of two highly polymorphic markers of chromosome 9p, namely α‐interferon (IFNA) and D9S104, and the deletion of 5′‐methylthioadenosine phosphorylase (MTAPase) gene. Finally, the deletion of a gene belonging to p16INK4A family, the p18 gene, was analysed in these acute leukaemias. Our results demonstrated that: (i) the biallelic loss of p16INK4A gene is strictly related to a specific immunophenotype, namely ALL of T‐cell lineage; (ii) no significant correlation exists between alterations at chromosome 9p level and the homozygous deletions of p16INK4A gene; and (iii) p18 gene was not deleted in the examined cases. These findings suggest a possible correlation between the T‐lymphocyte phenotype and the expression of p16INK4A gene. Moreover, the absence of MTAPase activity seems to be a valuable marker of p16INK4A gene inactivation, thus indicating that the deleted chromosomal area on 9p21 very frequently involves the MTAPase gene.
The detection of rhodamine 123 efflux at low levels of drug resistance
Authors: Webb Malcolm , Raphael Cary L. , Asbahr Heinke , Erber Wendy N. , Meyer Brian F.
Published: October 2003

Although many cell models of multidrug resistance (MDR) have been developed, most have been high‐resistance models which generate up to 100‐fold increases in drug resistance. However, the drug concentrations required to achieve these levels of resistance are much higher than those found in vivo. In this paper we describe the development of a cell model that reflects the resistance levels that are likely to be found clinically. We then investigated the methods used to detect MDR1 expression at these low levels of drug resistance. We demonstrated that the immunological and PCR‐based methods are unable to detect increased MDR1 expression in cells with a <5.2‐and 6.5‐fold increase in vinblastine resistance, respectively, in our drug‐resistant sublines. The rhodamine 123 (Rh 123) efflux assay was able to discriminate the vinblastine‐sensitive parent cells from all the vinblastine‐resistant sublines, including cells with a 1.7‐fold increase in resistance. However, this assay is non‐specific to the MDR1 gene and may detect the activity of other drug efflux proteins such as the MDR‐associated protein (MRP). Our results show that the Rh 123 efflux assay is able to detect the activity of drug efflux proteins such as the P‐glycoprotein, MRP or other efflux systems at the low levels of drug resistance that are likely to be attained in vivo
Isochromosome 12p in two cases of acute myeloid leukaemia without evidence of germ cell tumour
Authors: HEINONEN Kristiina , NAGESH RAO P. , SLACK JAMES L. , CRUZ JULIA , BLOOMFIELD CLARA D. , MRÓZEK KRZYSZTOF
Published: October 2003

An isochromosome 12p [i(12p)], typical of germ cell tumours (GCT), has, to date, been observed in 10 cases of acute myeloid leukaemia (AML) or myelodysplastic syndrome, nine of which had concurrent or preceding GCT. We report two i(12p)‐positive AML cases without clinical evidence of GCT. One patient with AML‐M1 had two i(12p) as the only cytogenetic anomalies. In the other case of AML‐M3 with t(15;17)(q22;q11‐12) at diagnosis, the i(12p) was clearly a secondary rearrangement since it first appeared at relapse and always accompanied the t(15;17). Our results suggest that i(12p) does not always indicate neoplastic disease of germ cell origin.
Establishment of cell lines derived from chronic lymphocytic leukaemic cells by transfection with myc and ras
Authors: ZHENG CHANG YU , PABELLO PAUL , MAKSYMIUK ANDREW W. , SKINNIDER LEO F.
Published: October 2003

Seventy‐one samples from 37 cases of chronic lymphocytic leukaemia (CLL) were transfected with human c‐myc and/or N‐ras. In only three cases did the CLL cells further transform and four cell lines were established following transfection with myc plus ras. Surface marker profiles and immunoglobulin gene rearrangement studies confirm that all cell lines have arisen from the original CLL cells. The cell lines are EBNA positive but negative for EBV latent membrane protein (LMP). Karyotyping of the cell lines is as follows: SA4, 46,XY; SA4‐2, 45,XY,−20; RK4, 48,XY,+5,+12; EHC4, 47,XY, t(14;18)(q32;q21),+12. Although the majority of CLL cells appear resistant to transformation in vitro with myc and ras with the methods used, it does occur in a small percentage (about 8%). There appeared to be no correlation between the ability to transform in vitro and clinical behaviour.
Cytogenetic clonality analysis: typical patterns in myelodysplastic syndrome and acute myeloid leukaemia
Authors: VAN LOM K. , HAGEMEIJER A. , VANDEKERCKHOVE F. , SMIT E. M. E. , LÖWENBERG B.
Published: October 2003

The cell morphology and karyotype of bone marrow samples from 24 patients with myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML) were studied simultaneously with a combined technique of May‐Grünwald‐Giemsa (MGG) staining and fluorescence in situ hybridization (FISH) with chromosome‐specific DNA probes. This enabled us to investigate cell lineage involvement in three malignant conditions: MDS (n = 12), leukaemia‐transformed MDS (LT‐MDS) (n = 5) and de novo AML (n=7). In MDS we found blasts and often significant proportions of mature granulocytic and erythroid cells to be cytogenetically abnormal. Percentages of granulocytic and erythroid cells with cytogenetic aberrations were generally less than those of blasts. These data support the involvement of a transformed pluripotent stem cell that has retained maturation abilities. In two patients with chronic myelomonocytic leukaemia (CMMoL) the clonal involvement of monocytes was predominant. Results in the five patients with LT‐MDS were similar to those in MDS. In the bone marrow of five of the seven de novo AML patients the cytogenetic abnormalities were restricted to the blasts and did not include the more mature granulocytic or erythroid populations. In the other two patients with AML, both with a t(8;21) and a loss of the Y chromosome, high percentages of mature neutrophils were cytogenetically abnormal. These patterns of clonal lineage involvement in MDS, LT‐MDS, t(8;21) AML and AML appear typical and may be of clinical use, for example, for distinguishing LT‐MDS from de novo AML in newly presenting patients.
Analysis of granulocyte colony stimulating factor receptor isoforms, polymorphisms and mutations in normal haemopoietic cells and acute myeloid leukaemia blasts
Authors: Bernard Tanya , Gale Rosemary E. , Linch David C.
Published: October 2003

Deletion mutants of the intracytoplasmic domain of the granulocyte colony stimulating factor receptor (G‐CSFR) have shown that it contains a membrane‐proximal region which must be conserved to allow the receptor to transduce a mitotic signal, and a C‐terminal region necessary for transduction of cell differentiation. Changes in the intracytoplasmic domain may result in the uncoupling of these two processes, as in acute leukaemia, and such alterations could occur either as isoforms or mutations. We have studied the transmembrane domain and intracytoplasmic tail of the G‐CSFR in RNA from blood or bone marrow of 11 haematologically normal controls and 40 patients with acute myeloid leukaemia (AML). Two novel transcripts of the receptor were identified, both were minor components and are unlikely to be of major physiological significance. We could find no evidence for altered levels of expression of these transcripts in the AML patients. In addition, only one point mutation was detected in the 40 patients screened by RT‐PCR‐SSCP, a C → A substitution at nucleotide 2088 which changes a threonine to asparagine in the transmembrane domain and is probably a polymorphism. These results suggest that abnormalities in the G‐CSFR are uncommon in AML.
Transfusion‐related acute lung injury due to HLA‐A2‐specific antibodies in recipient and NB1‐specific antibodies in donor blood
Authors: Bux Jürgen , Becker Frank , Seeger Werner , Kilpatrick David , Chapman Judith , Waters Alan
Published: October 2003

Transfusion‐related acute lung injury (TRALI) is a hazardous but little‐known complication of blood transfusion, characterized by non‐cardiogenic lung oedema after blood transfusion. Leucoagglutinating antibodies in the donor plasma are considered to play a central role in the pathogenesis of TRALI but no recommended procedure currently exists for their detection, and most of them have not yet been well characterized. Serum samples of two patients who have developed TRALI within 30 min of blood transfusion and the sera of the involved blood donors were investigated for leucocyte antibodies by granulocyte immunofluorescence, granulocyte agglutination and lymphocytotoxicity assays using typed test cells. Suspected specificities of the detected antibodies were confirmed by a luminoimmunoblot assay and the antigen capture assay MAIGA. One case was associated with granulocyte agglutinating anti‐HLA‐A2 antibodies in the recipient’s (i.e. patient's) own blood and the other with donor‐related non‐agglutinating antibodies directed against the granulocyte‐specific antigen NB1. Leucocyte incompatibility between donor and recipient was shown in both cases by crossmatching and typing of the incompatible cells for the appropriate antigen. The results show that TRALI is associated not only with donor‐ but also with recipient‐related leucocyte antibodies. In addition to leucoagglutinating antibodies, non‐agglutinating granulocyte‐specific antibodies can be also involved. For immunodiagnosis, sera from both must be investigated by a combination of granulocyte and lymphocyte (HLA) antibody screening tests and leucocyte incompatibility verified by crossmatching.
An aggressive subtype of B‐CLL is characterized by strong CD44 expression and lack of CD11c
Authors: Eisterer Wolfgang , Hilbe Wolfgang , Stauder Reinhard , Bechter Oliver , Fend Falko , Thaler Josef
Published: October 2003

In a retrospective study based on 107 B‐CLL patients, the expression of the adhesion molecules CD44, CD11a, CD11b, CD11c, CD18, CD25 and CD54 was analysed in bone marrow cryostat sections by immunohistochemistry. CD44 expression clearly identified two subgroups of B‐CLL patients with different clinical course. In particular, CD44‐positive patients presented with advanced disease, more often displayed a diffuse pattern of bone marrow infiltration, and had a worse prognosis. 33/61 patients positive for CD44 died within the observation period compared to 7/46 patients negative for CD44 (P = 0.0012). Multivariate analysis emphasized the independent prognostic value of CD44 expression for overall survival (P = 0.022). In contrast, patients positive for CD11c showed a longer survival, with 9/40 patients dying within the observation period compared to 31/67 negative for CD11c (P = 0.0013). Patients lacking CD11c were in advanced Rai and Binet stage. Multivariate analysis confirmed CD11c as a relevant independent prognostic marker (P = 0.033). Moreover, CD11c was able to separate patients with significantly different prognosis in the subgroup of CD44‐positive cases. 4/18 patients positive for CD44 and CD11c died before median survival time was reached. Patients positive for CD44 but negative for CD11c had an adverse prognosis: 29/43 patients died, median survival time was 33.4 months.  Our results indicate that CD44 positivity and CD11c negativity are associated with more advanced disease and worse prognosis in B‐CLL and suggest CD44‐positive/CD11c‐negative cases represent a more aggressive form of the disease.
Successful treatment of virus‐associated haemophagocytic syndrome in adults by cyclosporin A supported by granulocyte colony‐stimulating factor
Authors: TSUDA HIROYUKI , SHIRONO KENJI
Published: October 2003

Two young adult patients with typical virus‐associated haemophagocytic syndrome (VAHS) were treated with cyclosporin A and granulocyte colony‐stimulating factor (G‐CSF). Clinical symptoms such as high fever and malaise disappeared rapidly with concurrent haematological improvement in both patients. The serum levels of interleukin‐6 (IL‐6), soluble IL‐2 receptor, tumour necrosis factor and macrophage‐CSF were all elevated before treatment but that of G‐CSF was not. The dramatic effect of cyclosporin A observed implies that it efficiently and rapidly suppresses the cytokine storm caused by dysregulated T cells in VAHS. In addition, G‐CSF may promote haematological recovery without syndrome regression. We believe that the combination of cyclosporin A and G‐CSF may be effective, at least in selected patients with VAHS. Further studies are required to confirm its role as first‐line therapy for adult patients with VAHS.
A phase I/II trial of Z‐Dex (oral idarubicin and dexamethasone), an oral equivalent of VAD, as initial therapy at diagnosis or progression in multiple myeloma
Authors: Cook G. , Sharp R. A. , Tansey P. , Franklin I. M.
Published: October 2003

We designed an oral equivalent regime to mimic VAD and its hybrids, using idarubicin and dexamethasone (Z‐Dex) given in four cycles to induce cytoreduction prior to dose intensification in multiple myeloma cases. 20 patients (de novon=15, replaced VAD n=2, relapsed n=2, and resistant n=1), 13 males and seven females with a median age of 54 years (range 40–65 years) received Z‐Dex therapy. The overall response rate was 70% (14/20), with one patient (5%) achieving complete remission (CR). The response rate for previously untreated patients was 80% (12/15), with a CR rate of 6.7% (1/15). Both patients who received Z‐Dex in place of VAD continued to respond. Myelosuppression was seen in 14/20 patients (70%); 4/20 (20%) developing severe neutropenia with one death from neutropenic sepsis. Gastrointestinal toxicity and alopecia were infrequently reported. Satisfactory responses can be obtained using an oral regime equivalent to VAD with tolerable toxicity and morbidity.
Differential expression of CD3 and CD7 in T‐cell malignancies: a quantitative study by flow cytometry
Authors: Ginaldi Lia , Matutes Estella , Farahat Nahla , De Martinis Massimo , Morilla Ricardo , Catovsky Daniel
Published: October 2003

Most T‐cell antigens are expressed on normal and neoplastic T lymphocytes and for this reason it is not easy to distinguish between the immunophenotype of normal and malignant T cells. We have addressed this problem by comparing the levels of expression of CD3 and CD7 on T lymphocytes from 18 healthy donors with those of 61 cases of T‐cell leukaemia using quantitative flow cytometry with a method that converts fluorescence intensity into number of antigen molecules per cell. Normal T lymphocytes expressed 124 ± 25 CD3 and 20 ± 3 × 103 CD7 molecules per cell. The mean CD3 values were significantly lower in all types of T‐cell leukaemia than in normal T cells ( P < 0.05), with the exception of Sezary syndrome. The lowest CD3 values were found in T‐lymphoblastic leukaemia (T‐ALL), 30 ± 21 × 103, and adult T‐cell leukaemia/lymphoma (ATLL), 38 ± 31 × 103, followed by T‐prolymphocytic leukaemia (T‐PLL), 92 ± 47 × 103, and granular lymphocyte leukaemia (GLL), 95 ± 21 × 103. In contrast, the number of CD7 molecules was significantly higher in T‐ALL, 35 ± 7 × 103 ( P < 0.01), and T‐PLL, 29 ± 12 × 103, than the normal controls ( P < 0.01), whereas ATLL and GLL showed a low CD7 expression, 13 ± 3 and 12 ± 3 × 103, respectively. Our results show that the quantitative analysis of CD3 and CD7 and their combined evaluation may enable a distinction between normal and leukaemic T cells and could facilitate the monitoring of minimal residual disease. This study has also defined the T prolymphocyte as a cell of intermediate maturity between thymic derived and peripheral T lymphocytes.
A heteroplasmic point mutation of mitochondrial tRNALeu(CUN) in non‐lymphoid haemopoietic cell lineages from a patient with acquired idiopathic sideroblastic anaemia
Authors: Gattermann Norbert , Retzlaff Stefan , Wang Yan‐Ling , Berneburg Mark , H einisch Jürgen , Wlaschek Meinhard , Aul Carlo , Schneider Wolfgang
Published: October 2003

Acquired idiopathic sideroblastic anaemia (AISA) has been proposed to be a disorder of mitochondrial DNA (mtDNA). The hallmark of mitochondrial iron overload may be attributable to a respiratory chain defect leading to impaired reduction of ferric iron (Fe3 +) to ferrous iron (Fe2 +), which is essential to the last step of mitochondrial haem biosynthesis. In a 71‐year‐old patient we identified a point mutation in one of the two mitochondrial transfer‐RNAs coding for leucine (tRNAleu(CUN)). The mutation involves a G → A transition in the anticodon loop, immediately adjacent to the anticodon triplet (mtDNA position 12301). The mutated guanine is highly conserved in a wide range of species. The mutation is heteroplasmic, i.e. there is a mixture of normal and mutated mitochondrial genomes (ratio c. 50:50). Heteroplasmy of mtDNA is not found in normal individuals, but is a typical feature of mitochondrial cytopathies. The point mutation was present in the patient's bone marrow and whole blood samples, in purified platelets, and in the granulocyte/erythrocyte pellet after mononuclear cell separation by density gradient centrifugation. The mutation was not found in T‐ and B‐lymphocytes isolated by immunomagnetic bead separation. It was also absent from buccal mucosa cells and cultured skin fibroblasts. This pattern of involvement suggests that the mutation occurred in a self‐renewing myeloid stem cell of the CFU‐GEMM type.