Mutlu Arat

NBEAL2 is mutated in gray platelet syndrome and is required for biogenesis of platelet α-granules

Gray platelet syndrome (GPS) is an autosomal recessive bleeding disorder that is characterized by large platelets that lack α-granules. Here we show that mutations in NBEAL2 (neurobeachin-like 2), which encodes a BEACH/ARM/WD40 domain protein, cause GPS and that megakaryocytes and platelets from individuals with GPS express a unique combination of NBEAL2 transcripts. Proteomic analysis of sucrose-gradient subcellular fractions of platelets indicated that NBEAL2 localizes to the dense tubular system (endoplasmic reticulum) in platelets.

Late cardiovascular events after allogeneic hematopoietic stem cell transplantation : a retrospective multicenter study of the Late Effects Working Party of the European Group for Blood and Marrow Transplantation

Long-term outcome after hematopoietic stem cell transplantation including late transplant-related events is of increasing interest. This study shows that long-term survivors after allogeneic hematopoietic stem cell transplantation are likely to have an increased risk of premature cardiovascular accidents. See related perspective article on page 1132. Background Long-term outcome after hematopoietic stem cell transplantation including late transplant-related events is of increasing interest. The aim of this study was to evaluate the incidence of cardiovascular events after allogeneic HSCT and to search for their risk factors. Design and Methods This is a retrospective mutlicenter European Group of Blood and Marrow Transplantation (EBMT) analysis, including 548 long-term survivors treated in ten EBMT transplant centers, who underwent hematopoietic stem cell transplantation between 1990 and 1995 and survived ≥1 year after the transplant. All arterial events occurring after hematopoietic stem cell transplantation (cerebral, coronary, peripheral) were reported. Results Twenty (3.6%) out of 548 patients had a cardiovascular event in at least one arterial territory. The median age at occurence of cardiovascular events was 54 years (range, 41–70). The cumulative incidence of a first arterial event 15 years after hematopoietic stem cell transplantation was 6% (95% CI, 3%–10%). The cumulative incidence for patients with a high global cardiovascular risk score, defined as having ≥50% of the risk factors (arterial hypertension, diabetes, dys-lipidemia, increased body-mass index, physical inactivity, smoking) was 17%, as compared to 4% in those with a low risk score. In multivariate analysis age older than 30 years at last follow-up, and a high global cardiovascular risk score were associated with, respectively, 6.4-fold and 9.8-fold increases in the risk of an arterial event. Conclusions Long-term survivors after allogeneic hematopoietic stem cell transplantation are likely to have an increased risk of premature cardiovascular accidents.

Lamivudine prophylaxis for prevention of chemotherapy-induced hepatitis B virus reactivation in hepatitis B virus carriers with malignancies.

Summary.  Although hepatitis B virus (HBV) reactivation in HBV carriers undergoing immunosuppressive therapy is clearly documented, the role of antiviral prophylaxis in such individuals is still controversial. The aim of this study was to determine the efficacy of lamivudine prophylaxis in HBV carriers with haemato/oncological malignancies, who receive chemotherapy. Eighteen HBV carriers with malignancy, who were candidates for chemotherapy, were enrolled. Eight subjects (three with leukaemia, four with lymphoma and one with multiple myeloma) were enrolled for prophylactic lamivudine therapy. The remaining 10 patients (six with leukaemia, three with lymphoma and one with breast cancer) were not treated with lamivudine and were used as a control. Lamivudine was administered beginning on the same day as the chemotherapy and was maintained for a year after chemotherapy was discontinued. No HBV‐related mortality was observed in either group. In the lamivudine‐treated group, none of the subjects had clinical, biochemical or serological evidence of HBV reactivation during the time they were receiving chemotherapy and after their chemotherapy was discontinued. In contrast, five of the 10 HBV carriers not receiving lamivudine therapy experienced a reactivation of HBV infection. This reactivation of HBV was observed during the chemotherapy in four with one individual experiencing a HBV activation 12 months after chemotherapy was discontinued. No lamivudine‐related major adverse effects were observed. Hence prophylactic lamivudine treatment in HBV carriers with haemato/oncological malignancy receiving chemotherapy prevents chemotherapy‐induced HBV reactivation.

Gray platelet syndrome: natural history of a large patient cohort and locus assignment to chromosome 3p

Gray platelet syndrome (GPS) is an inherited bleeding disorder characterized by macrothrombocytopenia and absence of platelet α-granules resulting in typical gray platelets on peripheral smears. GPS is associated with a bleeding tendency, myelofibrosis, and splenomegaly. Reports on GPS are limited to case presentations. The causative gene and underlying pathophysiology are largely unknown. We present the results of molecular genetic analysis of 116 individuals including 25 GPS patients from 14 independent families as well as novel clinical data on the natural history of the disease. The mode of inheritance was autosomal recessive (AR) in 11 and indeterminate in 3 families. Using genome-wide linkage analysis, we mapped the AR-GPS gene to a 9.4-Mb interval on 3p21.1-3p22.1, containing 197 protein-coding genes. Sequencing of 1423 (69%) of the 2075 exons in the interval did not identify the GPS gene. Long-term follow-up data demonstrated the progressive nature of the thrombocytopenia and myelofibrosis of GPS resulting in fatal hemorrhages in some patients. We identified high serum vitamin B(12) as a consistent, novel finding in GPS. Chromosome 3p21.1-3p22.1 has not been previously linked to a platelet disorder; identification of the GPS gene will likely lead to the discovery of novel components of platelet organelle biogenesis. This study is registered at www.clinicaltrials.gov as NCT00069680 and NCT00369421.

Late complications after hematopoietic stem cell transplantation

Hematopoietic stem cell transplantation (HSCT) offers the opportunity for cure to patients with leukemia, lymphoma and severe non-malignant diseases. More than 40,000 HSCTs are performed annually worldwide. Therefore, the number of long-term survivors, free of the disease for which they were transplanted is continuously increasing. Despite the improved prognosis of HSCT, long-term outcome may be impaired by transplant-associated morbidity and mortality. Long-term survivors can present a variety of malignant and non-malignant complications, impairing physical and psychological performance, normal integration in family and social life, and quality of life. Conditioning regimens, particularly when including total-body irradiation as well as graft-versus-host disease, play a key role in the development of late effects. However, with increasing time since transplantation new types of late effects may emerge. Awareness on long-term effects after HSCT is crucial to provide adapted pretransplant counseling, and recommendations for post-transplant screening, prevention and early treatment.

Thrombopoietic cytokines in patients with iron deficiency anemia with or without thrombocytosis.

Iron deficiency anemia is a cause of reactive thrombocytosis. A moderate increase in platelet numbers is common but sometimes counts may exceed 1,000 × 109/l. The mechanisms causing reactive thrombocytosis are unclear. In this study, we evaluated 15 women with iron deficiency anemia and thrombocytosis (platelets >450 × 109/l) and 16 women with iron deficiency anemia with normal platelet counts. Serum samples were taken before oral iron replacement therapy, after 1 and 3 months and at the end of replacement therapy. Thrombopoietin, erythropoietin (EPO), leukemia inhibitory factor, interleukin-6 and interleukin-11 levels were assayed. There was no change in the levels of thrombopoietic cytokines except for EPO. The correlation between high EPO levels and high platelet counts may suggest that EPO increases platelet counts, but the same EPO level changes can also be demonstrated in women with iron deficiency anemia but normal initial platelet counts. The fact that the levels of other cytokines remained unchanged during treatment suggests that either these cytokines have no effect on reactive thrombocytosis or the change in platelet counts in our patients is in a narrow range and is thus not affected by the cytokine levels.

Administration of granulocyte-colony-stimulating factor for allogeneic hematopoietic cell collection may induce the tissue factor-dependent pathway in healthy donors.

Summary:The hypercoagulable state caused by the use of recombinant human granulocyte colony-stimulating factor (rhG-CSF) has been cited in anecdotal reports. Since tissue factor (TF) is the main initiator of the coagulation cascade, we examined if rhG-CSF had an inductive effect on the TF-dependent pathway in 18 healthy donors receiving rhG-CSF (10 μg/kg/day × 5 days) for peripheral blood progenitor cell mobilization. After rhG-CSF, there were increases both in TF antigen (TF:Ag) (P=0.01) and TF procoagulant activity (TF:PCA) (P=0.06) plasma levels and in TF:Ag cytofluorimetric expression on CD33 (+) cells (P=0.04). Mean activities of FVIII and vWF also increased significantly. Thrombin time was slightly prolonged (P=0.06) due to significant increases in plasma D-dimer levels. In addition, while FIX activity remained stable, there were marked reductions in mean plasma FX and FII activities and a slight decrease in FVII activity that resulted in a significant prolongation of prothrombin time within normal ranges. In conclusion, the administration of rhG-CSF led to a ‘prothrombotic state’ via stimulation of TF and increased endothelial markers, such as F VIII and vWF. In the light of these findings, the use of rhG-CSF for stem cell mobilization should be undertaken cautiously in healthy donors with underlying thrombotic risk factors.

Bone marrow microvessel density (MVD) in adult acute myeloid leukemia (AML): therapy induced changes and effects on survival.

Based on the strong evidence in favor of an increase in microvessel density (MVD) in hematological malignancies, we evaluated VEGF immunoreactivity and MVD measurement in bone marrow biopsies of 36 AML patients at diagnosis and following therapy. MVD assessment was based on CD31, CD34 expressing vessels. The values were calculated for only one marker if the other vascular marker was positive on blasts, otherwise both markers were used. VEGF immunoreactivity was also scored. Comparison of MVD values of 36 AML patients with 18 non-malignant controls showed a significantly higher MVD in AML (CD31: P = 0.004, CD34: P < 0.001), which is independent of other variables such as cellularity or blast percentage. Following induction chemotherapy, the responders showed a significant decrease in blast counts (P < 0.001), cellularity (P = 0.001) and MVD (P = 0.050) quantification with CD31. Higher baseline MVD (CD34) values were associated with shorter overall survival (P = 0.0027). These results are encouraging for inclusion of MVD enumeration in bone marrow examinations of AML patients at diagnosis as an additional prognostic factor.

Autoimmune thrombocytopenia in a patient with small cell lung cancer developing after chemotherapy and resolving following autologous peripheral blood stem cell transplantation.

A 46-year-old white male with small cell lung cancer (SCLC) limited to the thorax developed autoimmune thrombocytopenic purpura (AITP), following a cyclophosphamide, paclitaxel and G-CSF-containing regimen for peripheral blood stem cell (PBSC) mobilization. AITP associated with small or non-small cell lung cancer has been reported. We considered that the AITP in this case may be a part of paraneoplastic syndrome, which is frequently seen in patients with SCLC. The patient received HDC and autologous PBSC transplantation (APBSCT) for SCLC and the AITP resolved following transplantation, thus supporting the concept of HDC + APBSCT for the treatment of autoimmune diseases.

Febrile neutropenia in allogeneic and autologous peripheral blood stem cell transplantation and conventional chemotherapy for malignancies.

The risk and outcome of infection in febrile neutropenic patients is mainly determined by the duration of neutropenia, the underlying disease or the treatment. This study was undertaken to compare infections and the outcome after conventional chemotherapy (CCT), allogeneic PBSC transplantation (alloPBSCT) or autologous PBSC transplantation (autoPBSCT), during the period of neutropenia, in a single center. A total of 145 patients (50 in CCT group, 50 in alloPBSCT and 45 in autoPBSCT) were evaluated. In the alloPBSCT group, 86% of the patients (43/50), in the autoPBSCT group 93% of the patients (42/45) and in the CCT group 92% (46/50) of the patients had at least one febrile episode during their neutropenic period (P > 0.05). Microbiologically and/or clinically documented infection rates were 50% (25/50), 42% (19/45) and 48% (24/50) respectively. Gram-positive pathogens, mostly coagulase-negative staphylococci were the most frequent cause of bacteremias in all groups. The frequency of CNS infections was significantly higher in the alloPBSCT and autoPBSCT groups compared to the CCT group (P < 0.008 and P < 0.04, respectively). Catheter infections were frequent in the pbsct groups and pulmonary infections were more frequent in the cct group (P < 0.05). The CCT group needed longer antibiotic usage compared to the alloPBSCT group (P < 0.006). The duration of neutropenia and the type of treatment given, does not affect the rate of febrile episodes, but affects the type of infections in febrile neutropenic patients. Bone Marrow Transplantation (2000) 26, 211–214.