The effect of treatment with Campath-1H in patients with autoimmune cytopenias

We describe 21 patients with severe and life-threatening autoimmune cytopenias resistant to standard immunosuppression who were treated with the monoclonal antibody Campath-1H. Four patients had autoimmune neutropenia, four had autoimmune haemolytic anaemia, four had pure red cell aplasia, one had immune thrombocytopenia purpura (ITP), three had autoimmune haemolytic anaemia and ITP (Evan's syndrome), three had autoimmune pancytopenia (ITP, autoimmune neutropenia and autoimmune haemolytic anaemia), one had ITP (associated with acquired Glanzmann's disease) and autoimmune neutropenia, and one had ITP and red cell aplasia. Campath-1H was administered at a dose of 10 mg/d as an intravenous infusion for 10 d. Responses were seen in 15 patients, which were sustained in six. Relapse occurred in eight patients after Campath-1H treatment. Patients entering the study later, received cyclosporine after Campath-1H in an attempt to reduce the incidence of relapse. Three patients received a second course of Campath-1H; all responded but later relapsed. Fourteen patients are alive at a median of 12 months (range 4-61) after Campath-1H. Campath-1H represents an alternative therapeutic option for severe, refractory autoimmune cytopenias.     

Fibrinogen levels are associated with bleeding in patients with primary immune thrombocytopenia

Abstract

Bleeding is the most common clinical symptom and the leading cause of death in patients with primary immune thrombocytopenia (ITP). Our research intends to verify the role of fibrinogen levels as independent determinants of bleeding. We retrospectively analyzed the relationship between fibrinogen levels and bleeding events in 463 patients. Additionally, we confirmed the impact of fibrinogen level on clot firmness in 25 patients via thrombelastography (TEG). Fibrinogen levels (median and inter-quartile range, IQR) were significantly different (p < .001) between bleeding and non-bleeding patients [258(207–314) mg/dL vs. 315(262–407) mg/dL, respectively]. Further analyzes in three subgroups based on platelet (PLT) count showed that non-bleeding patients still had higher fibrinogen levels than bleeding patients. The optimal discriminant threshold of fibrinogen in bleeding was 288.5 mg/dL according to receiver operating characteristic (ROC) curves. Patients were divided into low (LF, 230[193–258] mg/dL) and high (HF, 349[313–424] mg/dL) fibrinogen groups based on this threshold. Bleeding event rates were significantly different (LF: 84.6% vs. HF: 60.4%, P < .001) between the two groups. Multivariable analyses further confirmed these differences. Moreover, TEG parameters showed elevated clot firmness in the HF group. Our data suggest that high fibrinogen levels are associated with reduced bleeding events.     

GM-CSF in the treatment of Fanconi's anaemia

We have used recombinant human (rh) GM-CSF in two 12-year-old Fanconi's aplastic anaemia patients. They had not received any previous therapy except blood transfusions. Each patient was given three 21 d courses of rh-GM-CSF, the first two at a dose of 3.5μg/kg/d and the third at 7 μg/kg/d s.c. There were significant increases in WBC and absolute neutrophil counts after the first week of rh-GM-CSF which lasted as long as the treatment was continued. Following the cessation of treatment, WBC and ANC dropped rapidly. We conclude that rh-GM-CSF can be used in FAA, especially in severely neutropenic cases.     

Mycophenolate mofetil for the treatment of refractory auto-immune haemolytic anaemia and auto-immune thrombocytopenia purpura

The treatment of both auto-immune haemolytic anaemia (AIHA) and auto-immune thromobocytopenic purpura (AITP) remains unsatisfactory in those refractory to first-line management. Mycophenolate mofetil (MMF) is an immunosuppressive agent originally used to prevent acute rejection of solid organ transplants and used more recently in the management of auto-immune conditions. We report its use in four patients with AIHA and six patients with AITP. All four patients with AIHA and five of the six patients with AITP showed a complete or good partial response to treatment with MMF, confirming a possible role in the treatment of these conditions.     

Plasma levels of endothelial-derived haemostatic factors in autoimmune thrombocytopenia and haemolytic anaemia

The endothelial cell-synthesized haemostatic factors tissue plasminogen activator (tPA), plasminogen activator inhibitor (PAI) and von Willebrand factor (vWF) were assayed in a cross-sectional study of patients with immune thrombocytopenia (ITP) (n = 12) and autoimmune haemolytic anaemia (AIHA) (n = 3), and compared with a simultaneously selected contrast group of other hospitalized patients with obscure blood cytopenia at the time of sampling. All three factors were grossly elevated in both the study group and the contrast group. In the autoimmune patient group, the three haemostatic variables were significantly correlated with orosomucoid levels, demonstrating the acute-phase nature of the increased levels of vWF, tPA and PAI. These findings support the view that haemostatic factors of the vessel wall are implicated in the pathophysiology of a wide spectrum of diseases.     

Use of mycophenolate mofetil for chronic, refractory immune cytopenias in children with autoimmune lymphoproliferative syndrome

Autoimmune lymphoproliferative syndrome (ALPS) is a disorder of apoptosis associated most often with heritable FAS mutations leading to lymphadenopathy, hypersplenism and chronic refractory autoimmune cytopenias. Mycophenolate mofetil (MMF) was used to treat cytopenias in 13 ALPS patients aged 9 months to 17 years from a cohort of 118 children (aged < 18 years) and 82 adults. Twelve responded for a median follow-up of 49 weeks (range 38-240 weeks), defined by maintenance of adequate blood counts and reduction in dosage or cessation of other immunosuppressive agents. This preliminary experience suggests that MMF may spare steroid usage in patients with ALPS-associated cytopenias.     

Serum G-CSF levels are not increased in patients with antibody-induced neutropenia unless they are suffering from infectious diseases

By the use of a G-CSF-specific ELISA we determined the serum granulocyte-colony stimulating factor (G-CSF) levels in 63 patients with antibody-induced neutropenia including neonatal immune neutropenia, autoimmune neutropenia, and drug-induced immune neutropenia. In the sera of 20 patients, elevated G-CSF levels of 60-1006 pg/ml (normal <39 pg/ml) were observed. These patients suffered from infectious diseases at the time of blood collection. G-CSF levels normalized after successful antibiotic treatment, indicating that increased G-CSF production in patients with immune neutropenia may be primarily the result of infection and not of neutropenia.     

Pregnancy-induced thrombocytopenia and TTP, and the risk of fetal death, in Upshaw–Schulman syndrome: a series of 15 pregnancies in 9 genotyped patients

Upshaw–Schulman syndrome (USS) is a congenital thrombotic thrombocytopenic purpura (TTP) due to mutations in the gene that encodes for ADAMTS13 ( ADAMTS13 ), but its clinical signs may be mild or absent during childhood. We have identified 37 patients with USS (24 females, 13 males) belonging to 32 families. The nine women from six families who were diagnosed during their first pregnancy are the focus of this report. Six of the nine women had episodes of thrombocytopenia during childhood misdiagnosed as idiopathic thrombocytopenic purpura. Thrombocytopenia occurred during the second–third trimesters in each of their 15 pregnancies, with 16 babies (one twin pregnancy), often followed by TTP. Of 15 pregnancies, eight babies were stillborn or died soon after birth, and the remaining seven were all premature except one, who was born naturally following plasma infusions to the mother that had started at 8 weeks' gestation. All nine USS women had severely deficient ADAMTS13 activity. ADAMTS13 analyses demonstrated that eight women were compound heterozygotes of Y304C/G525D (2 siblings), R125VfsX6/Q1302X (2 siblings), R193W/R349C (2 siblings), I178T/Q929X, and R193W/A606P; one woman was homozygous for R193W. Only the R193W mutation has been previously reported. These observations emphasize the importance of measuring ADAMTS13 activity in the evaluation of thrombocytopenia during childhood and pregnancy.     

GM-CSF in the treatment of Felty syndrome

Many therapeutic agents have been tried with variable success in the treatment of Felty neutropenia, but the reports are anecdotal. We now describe the second trial of recombinant granulocyte-macrophage colony-stimulating factor (GM-CSF), in a splenectomized, infected patient with Felty syndrome.     

Effect of thrombopoietin-receptor agonists on circulating cytokine and chemokine levels in patients with primary immune thrombocytopenia (ITP)

Background: Thrombopoietin-receptor-agonists (TPO-RAs) increase platelet production in Immune Thrombocytopenia (ITP) by stimulating Mpl. The effect of TPO-RAs on inflammatory cytokine production in ITP patients has not been well investigated. Methods: Plasma samples from 48 ITP patients treated with TPO-RAs (median age 50 years (inter-quartile range; IQR 20–69), median platelet counts 24 × 10⁹/L (IQR 15–47 × 10⁹/L), 28 females) and 16 healthy controls (nine females, median age 37 years, IQR 22–51 years) were collected before and during treatment, and analyzed for a panel of cytokines and chemokines by enzyme-linked immunosorbent assay and immuno-bead-based multiplex assay. Results: Elevated levels of C-X-C motif chemokine 10 (CXCL10; p < 0.001) and osteoprotegerin (OPG; p < 0.05) were observed in pretreatment samples compared to controls; these levels decreased during 6 months of treatment. Pretreatment levels of transforming growth factor (TGF)-β were lower than in healthy controls and increased after 6 months of treatment (p < 0.05). Levels of sCD40L increased after 6 months of treatment (p < 0.05), but decreased thereafter to pretreatment values. The increase in TGF-β and sCD40L may reflect increased platelet turnover. Levels of tumor necrosis factor (TNF)-α, interferon (IFN)-γ and interleukin (IL)-10 did not change during treatment. Conclusion: These findings suggest that treatment with TPO-RA creates a more balanced steady-state of immune activation.     

Metabolomic profile in patients with primary warm autoimmune haemolytic anaemia

Autoimmune haemolytic anaemia (AIHA) is a rare clinical condition with immunoglobulin fixation on the surface of erythrocytes, with or without complement activation. The pathophysiology of AIHA is complex and multifactorial, presenting functional abnormalities of T and B lymphocytes that generate an imbalance between lymphocyte activation, immunotolerance and cytokine production that culminates in autoimmune haemolysis. In AIHA, further laboratory data are needed to predict relapse and refractoriness of therapy, and thus, prevent adverse side-effects and treatment-induced toxicity. The metabolomic profile of AIHA has not yet been described. Our group developed a cross-sectional study with follow-up to assess the metabolomic profile in these patients, as well as to compare the metabolites found depending on the activity and intensity of haemolysis. We analysed the plasma of 26 patients with primary warm AIHA compared to 150 healthy individuals by mass spectrometry. Of the 95 metabolites found in the patients with AIHA, four acylcarnitines, two phosphatidylcholines (PC), asymmetric dimethylarginine (ADMA) and three sphingomyelins were significantly increased. There was an increase in PC, spermine and spermidine in the AIHA group with haemolytic activity. The PC ae 34:3/PC ae 40:2 ratio, seen only in the 12-month relapse group, was a predictor of relapse with 81% specificity and 100% sensitivity. Increased sphingomyelin, ADMA, PC and polyamines in patients with warm AIHA can interfere in autoantigen and autoimmune recognition mechanisms in a number of ways (deficient action of regulatory T lymphocytes on erythrocyte recognition as self, negative regulation of macrophage nuclear factor kappa beta activity, perpetuation of effector T lymphocyte and antibody production against erythrocyte antigens). The presence of PC ae 34:3/PC ae 40:2 ratio as a relapse predictor can help in identifying cases that require more frequent follow-up or early second-line therapies.     

Model analysis of the contrasting effects of GM-CSF and G-CSF treatment on peripheral blood neutrophils observed in three patients with childhood-onset cyclic neutropenia

Human cyclic neutropenia (CN) is a rare haematological disorder characterized by regular fluctuation in the serial count of blood neutrophils. The oscillations occur at subnormal levels with a stable 3-week period. To reduce the risk of serious infections during the severe neutropenic nadir phases (<0.25 ×10⁹ neutrophils/l) patients are usually treated with recombinant growth factors. Compared with G-CSF, which has been shown to enhance the amplitudes substantially, the response to GM-CSF is poor: neutrophil numbers are not amplified, the cycles remain unchanged or are dampened. However, two cases with a modest neutrophil increase have been reported in the literature. In a recently published clinical study the different effects of GM-CSF and G-CSF application have been investigated in the same patients. Based on a mathematical model of CN we previously proposed, the detailed neutrophil data measured in this study are analysed by simulation. The contrasting clinical results can be quantitatively explained by the model concept of regulatory control together with possible individual feedback defects, i.e. abnormally reduced mitotic responsiveness of granulopoietic progenitor cells to GM-CSF and G-CSF.     

Effect in vivo of recombinant GM-CSF on neutropenia and survival in mice treated by high-dose melphalan

High doses of melphalan cause severe neutropenia and may irreversibly damage hematopoietic stem cells. Treatment of mice with recombinant murine GM-CSF (GM-CSF) for 5 days immediately after 400 micrograms of melphalan did not prevent the severe neutropenia. However, GM-CSF accelerated the neutrophil recovery and reduced the mortality rate during the neutropenic period compared to melphalan-only treated mice. CFU-GM levels measured 6 d after melphalan treatment without GM-CSF were markedly reduced in the bone marrow while being elevated in the spleen. In comparison, GM-CSF further reduced the total CFU-GM population in melphalan-treated mice including the levels in the bone marrow and in the spleen. On d 14 after melphalan, the spleen regained its active CFU-GM production. By d 90, the number of circulating neutrophils, the number of bone marrow CFU-GM and splenic CFU-GM were the same in GM-CSF-treated and -untreated mice. The results suggest that GM-CSF could be used to shorten the neutropenic period and reduce mortality caused by a high dose of melphalan. Though this effect could be at the expense of a temporary reduction in CFU-GM population, GM-CSF did not induce more long-term damage to myelopoiesis than that already caused by melphalan alone.     

Effect of recombinant human thrombopoietin in nonhuman primates with chemotherapy-induced thrombocytopenia

We examined the effects of recombinant human thrombopoietin (rhTPO) on myelosuppressive chemotherapy-induced thrombocytopenia in cynomolgus monkeys. After treatment with nimustine (ACNU) on day 0, the monkeys intravenously received rhTPO at a dose of 0.04, 0.2 or 1 μg/kg/d or monkey's serum once each day from day 1 to day 28. Administration of rhTPO reduced the severity of thrombocytopenia and accelerated the rate of platelet recovery in a dose-dependent fashion. Treatment with the highest rhTPO dose completely prevented thrombocytopenia and stimulated a marked increase in platelet counts over the normal values. Animals treated with ACNU also became neutropenic and slightly anaemic. Administration of rhTPO following ACNU treatment significantly improved neutropenia with increasing doses of rhTPO, but had no effect on anaemia. Compared to the control animals, rhTPO-treated animals exhibited no significant changes in several serum parameters, C-reactive protein concentration and some blood coagulation profiles within the study period. These results suggest a therapeutic efficacy of rhTPO in improving chemotherapy-induced thrombocytopenia.     

Serum thrombopoietin levels in patients with aplastic anaemia

Endogenous serum thrombopoietin (TPO) levels were measured in 31 patients with aplastic anaemia (AA) using an enzyme immunoassay with a sensitivity of 20 pg/ml. The median platelet count for all AA patients was 30 ± 29 × 10⁹/l (range 5–102) compared with a median of 284 ± 59 × 10⁹/l (range 148–538) for normal controls. Serum TPO levels were significantly elevated in all patients compared with normals (1706 ± 1114.2, range 375–5000 v 78 ± 54, range 16.5–312.9, P  < 0.0001). There was no correlation between serum TPO levels and the degree of thrombocytopenia in AA patients, but TPO levels were significantly higher in patients who were platelet transfusion dependent than in patients who were transfusion independent ( P  < 0.01). There was a trend for higher TPO levels in patients with severe AA compared with non-severe AA patients. Clinical trials of TPO and a related truncated, pegylated molecule, megakaryocyte growth and development factor (PEG-rHuMGDF), are awaited to determine whether treatment with these drugs will result in increased platelet counts in patients with AA.     

A case of severe pancytopenia caused by ibuprofen

We here present the case of a patient with severe neutropenia, haemolytic anaemia and thrombocytopenia associated with long-term use of ibuprofen. The blood parameters rapidly normalized when the drug was discontinued, and no further treatment, except for a short course of antibiotics, was required.     

Effect of low-dose granulocyte-macrophage colony-stimulating factor (LD-GM-CSF) on platelet transfusion-dependent thrombocytopenia

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a hematopoietic growth factor known to promote the proliferation and differentiation of precursors of granulocytes and monocytes. GM-CSF at standard doses (125–500 μg/m²) alleviates neutropenia secondary to cytotoxic chemotherapy, myelodysplastic syndromes, and aplastic anemia, but has minimal effect on anemia or thrombocytopenia. GM-CSF at doses <30 μg/m² has been reported to improve platelet counts in some patients exhibiting cytopenia related to hematologic disorders such as aplastic anemia and myelodysplastic syndrome. Lowdose GM-CSF (10–20 μg/m²) was evaluated in 20 patients with transfusion-dependent thrombocytopenia persisting after myeloablative cytotoxic chemotherapy or with disease-related cytopenia. Seven patients (35%) responded as defined by a reduction in the platelet transfusion requirements by at least 75%. Low-dose GM-CSF did not significantly increase neutrophil counts or decrease red blood cell transfusion requirements. These results indicate that low-dose GM-CSF has a thrombopoietic effect in about one-third of patients with platelet transfusion-dependent thrombocytopenia which has not been observed at higher doses. © 1994 Wiley-Liss, Inc.