Miguel R. Abboud

Prevention of a First Stroke by Transfusions in Children with Sickle Cell Anemia and Abnormal Results on Transcranial Doppler Ultrasonography

BACKGROUND Blood transfusions prevent recurrent stroke in children with sickle cell anemia, but the value of transfusions in preventing a first stroke is unknown. We used transcranial Doppler ultrasonography to identify children with sickle cell anemia who were at high risk for stroke and then randomly assigned them to receive standard care or transfusions to prevent a first stroke. METHODS To enter the study, children with sickle cell anemia and no history of stroke had to have undergone two transcranial Doppler studies that showed that the time-averaged mean blood-flow velocity in the internal carotid or middle cerebral artery was 200 cm per second or higher. The patients were randomly assigned to receive standard care or transfusions to reduce the hemoglobin S concentration to less than 30 percent of the total hemoglobin concentration. The incidence of stroke (cerebral infarction or intracranial hemorrhage) was compared between the two groups. RESULTS A total of 130 children (mean [+/-SD] age, 8.3+/-3.3 years) were enrolled; 63 were randomly assigned to receive transfusions and 67 to receive standard care. At base line, the transfusion group had a slightly lower mean hemoglobin concentration (7.2 vs. 7.6 g per deciliter, P=0.001) and hematocrit (20.4 vs. 21.7 percent, P=0.002). Ten patients dropped out of the transfusion group, and two patients crossed over from the standard-care group to the transfusion group. There were 10 cerebral infarctions and 1 intracerebral hematoma in the standard-care group, as compared with 1 infarction in the transfusion group -- a 92 percent difference in the risk of stroke (P<0.001). This result led to the early termination of the trial. CONCLUSIONS Transfusion greatly reduces the risk of a first stroke in children with sickle cell anemia who have abnormal results on transcranial Doppler ultrasonography.

A Comparison of Conservative and Aggressive Transfusion Regimens in the Perioperative Management of Sickle Cell Disease

BACKGROUND Preoperative transfusions are frequently given to prevent perioperative morbidity in patients with sickle cell anemia. There is no consensus, however, on the best regimen of transfusions for this purpose. METHODS We conducted a multicenter study to compare the rates of perioperative complications among patients randomly assigned to receive either an aggressive transfusion regimen designed to decrease the hemoglobin S level to less than 30 percent (group 1) or a conservative regimen designed to increase the hemoglobin level to 10 g per deciliter (group 2). RESULTS Patients undergoing a total of 604 operations were randomly assigned to group 1 or group 2. The severity of the disease, compliance with the protocol, and the types of operations were similar in the two groups. The preoperative hemoglobin level was 11 g per deciliter in group 1 and 10.6 g per deciliter in group 2. The preoperative value for hemoglobin S was 31 percent in group 1 and 59 percent in group 2. The most frequent operations were cholecystectomies (232), head and neck surgery (156), and orthopedic surgery (72). With the exception of transfusion-related complications, which occurred in 14 percent of the operations in group 1 and in 7 percent of those in group 2, the frequency of serious complications was similar in the two groups (31 percent in group 1 and 35 percent in group 2). The acute chest syndrome developed in 10 percent of both groups and resulted in two deaths in group 1. A history of pulmonary disease and a higher risk associated with surgery were significant predictors of the acute chest syndrome. CONCLUSIONS A conservative transfusion regimen was as effective as an aggressive regimen in preventing perioperative complications in patients with sickle cell anemia, and the conservative approach resulted in only half as many transfusion-associated complications.

Effects of Recombinant Human Granulocyte Colony-Stimulating Factor on Neutropenia in Patients with Congenital Agranulocytosis

Congenital agranulocytosis is a disorder characterized by severe neutropenia and a profound deficiency of identifiable neutrophil progenitors in bone marrow. In an attempt to stimulate neutrophil production and thereby reduce the morbidity and mortality associated with this disease, we administered recombinant human granulocyte colony-stimulating factor (rhG-CSF) in doses of 3 to 60 micrograms per kilogram of body weight per day to five patients with congenital agranulocytosis. In all five patients, an increase in the number of neutrophils was noted eight to nine days after the initiation of the effective dosage (the dose at which the neutrophil count reached 1000 cells per microliter or more and the bone marrow showed granulocyte maturation beyond the myelocyte stage). The absolute neutrophil counts rose from less than 100 to between 1300 and 9500 cells per microliter. Marrow aspirates obtained after 14 days at the effective dosage showed maturation to the mature neutrophil stage. The side effects that were observed were medullary pain, splenomegaly, and an elevation of levels of leukocyte alkaline phosphatase. All five patients have had sustained neutrophil counts of 1000 cells per microliter or more for 9 to 13 months while receiving subcutaneous maintenance therapy. Preexisting chronic infections have resolved clinically, and the number of new infectious episodes and the requirement for intravenous antibiotics have decreased. We conclude that treatment with rhG-CSF can lead to a large increase in the numbers of functional neutrophils in patients with congenital agranulocytosis.

Risk of recurrent stroke in patients with sickle cell disease treated with erythrocyte transfusions

OBJECTIVE To determine the effect of a transfusion program on risk of stroke recurrence in children with sickle cell disease. DESIGN The clinical course and experience with transfusion therapy at eight centers were reviewed for subjects whose initial stroke occurred after January 1988. RESULTS Sixty subjects were observed for 191.7 patient-years. Eight had a single recurrent stroke (two intracranial hemorrhages and six infarctions) for a prevalence of 13.3%, or one recurrence for each 24 patient-years of observation. Thirteen subjects had 15 transient neurologic events; two of these had subsequent strokes, but the overall risk was similar for those who did and those did not have transient events. Hemoglobin S levels were greater than the desired maximum of 30% at the time of 7 of 16 transient events and five of six recurrent infarctions. The stroke recurrence rate was similar to those in previous reports of children receiving long-term transfusion therapy but significantly less than that reported for children who did not receive transfusions (p < 0.001). CONCLUSIONS We conclude that maintenance of hemoglobin S at a level less than 30% appears to be effective in reducing the rate of recurrent infarction but does not prevent transient neurologic events. Transient neurologic events are common but do not appear to be related to recurrent stroke.

Gene Therapy for Wiskott-Aldrich Syndrome—Long-Term Efficacy and Genotoxicity

Wiskott-Aldrich syndrome gene therapy is feasible, but γ-retroviral vectors contribute a substantial risk of leukemogenesis. Taking the Sting Out of Gene Therapy Wiskott-Aldrich syndrome (WAS) is a rare X-linked recessive disorder characterized by low platelet count, immune deficiency, autoimmunity, and high risk of cancer. WAS is primarily a disorder of blood cells, and hematopoietic stem cell transplantation (HSCT) has been the only hope of cure. However, HSCT is restricted to patients who can find matching donors. One way to overcome this limitation is through gene therapy that restores the function of the mutated protein in HSCs from the patient. Now, Braun et al. report correction of WAS protein (WASP) in 9 of 10 patients that underwent HSC gene therapy. The authors used a γ-retroviral vector to correct WASP expression in autologous HSCs. After transfer to patients, these cells engrafted and WASP was expressed in lymphoid and myeloid cells and platelets in 9 of 10 patients. What’s more, this therapy caused either partial or complete resolution of symptoms. However, seven patients developed acute leukemia, and further analysis revealed genetic alterations such as chromosomal translocations. These studies suggest that with improved vector design, gene therapy may be feasible and effective for patient with WAS. Wiskott-Aldrich syndrome (WAS) is characterized by microthrombocytopenia, immunodeficiency, autoimmunity, and susceptibility to malignancies. In our hematopoietic stem cell gene therapy (GT) trial using a γ-retroviral vector, 9 of 10 patients showed sustained engraftment and correction of WAS protein (WASP) expression in lymphoid and myeloid cells and platelets. GT resulted in partial or complete resolution of immunodeficiency, autoimmunity, and bleeding diathesis. Analysis of retroviral insertion sites revealed >140,000 unambiguous integration sites and a polyclonal pattern of hematopoiesis in all patients early after GT. Seven patients developed acute leukemia [one acute myeloid leukemia (AML), four T cell acute lymphoblastic leukemia (T-ALL), and two primary T-ALL with secondary AML associated with a dominant clone with vector integration at the LMO2 (six T-ALL), MDS1 (two AML), or MN1 (one AML) locus]. Cytogenetic analysis revealed additional genetic alterations such as chromosomal translocations. This study shows that hematopoietic stem cell GT for WAS is feasible and effective, but the use of γ-retroviral vectors is associated with a substantial risk of leukemogenesis.

Stroke Prevention Trial in Sickle Cell Anemia

Stroke occurs in 7-8% of children with Sickle Cell Disease (Hb SS) and is a major cause of morbidity. Rates of recurrence have been reduced from 46-90% to less than 10% through chronic blood transfusions. Prevention of first stroke, however, would be preferable because even one stroke can cause irreversible brain injury. Transcranial Doppler (TCD) ultrasound can detect arterial blood flow rates associated with subsequent stroke risk. By combining TCD screening and a potentially effective treatment, first stroke may be prevented. The Stroke Prevention Trial in Sickle Cell Anemia (STOP) is the first stroke prevention trial in Hb SS and the first randomized, controlled use of transfusion in Hb SS. This multi-center trial is designed to test whether reducing sickle hemoglobin to 30% or less with periodic blood transfusions will reduce first-time stroke by at least 70% compared to standard care. Primary endpoints will be clinically evident symptoms of cerebral infarction with consistent findings on Magnetic Resonance Imaging and Angiography (MRI/MRA) or symptomatic intracranial hemorrhage. Secondary endpoints will be asymptomatic brain lesions detected by MRI in brain areas not involved in primary endpoints. The design calls for a 6-month start-up interval, 18 months of TCD screening and randomization, and observation for stroke from entry through month 54. Key features of the trial are standardized TCD and MRI/MRA protocols interpreted blindly, and blinded adjudication of endpoints. The sample size (60 per treatment group) is based on prospective data relating TCD velocity to risk of stroke. A time-averaged mean velocity of > or = 200 cm/sec is associated with a 46% risk of cerebral infarction over 39 months. The sample size is sufficient to detect 70% reduction in the primary endpoint at 90% power. This trial will determine if transfusion is effective in the primary prevention of stroke. Secondary aims may further the understanding of the effects of transfusion on the brain and guide future research into cerebrovascular disease in Hb SS.

Granulocytosis causing sickle-cell crisis

mgikg), or topiramate (25 mg/kg) or a combination of the two by intraperitoneal injection. A control group received vehicle (0.9% saline) alone. An hour later, each animal was given 85 mgkg pentylenetetrazol subcutaneously; the time taken to the first generalised seizure was rec~rded .~ A cut-off point of 15 min was set. Although neither topiramate or lamotrigine exerted an effect, none of the animals pre-treated with the combination experienced a seizure throughout the observation period. It is not possible to be certain that the combination (and not topiramate alone) was the effective strategy in these patients. Nevertheless, although lamotrigine was administered to the limit of tolerability, doses of topiramate remained quite small: add-on regulatory trials with topiramate reported maximum seizure reduction with 400-800 mg daily.’ The study in mice also supports particular efficacy for the combination. This may be a consequence of the drugs’ wide range of mechanisms of action affecting Na’ and Ca” channels, and GABA-mediated inhibition and excitatory aminoacid neurotransmission.

Serum ferritin level changes in children with sickle cell disease on chronic blood transfusion are nonlinear and are associated with iron load and liver injury

Chronic blood transfusion is increasingly indicated in patients with sickle cell disease. Measuring resulting iron overload remains a challenge. Children without viral hepatitis enrolled in 2 trials for stroke prevention were examined for iron overload (STOP and STOP2; n = 271). Most received desferrioxamine chelation. Serum ferritin (SF) changes appeared nonlinear compared with prechelation estimated transfusion iron load (TIL) or with liver iron concentrations (LICs). Averaged correlation coefficient between SF and TIL (patients/observations, 26 of 164) was r = 0.70; between SF and LIC (patients/observations, 33 of 47) was r = 0.55. In mixed models, SF was associated with LIC (P = .006), alanine transaminase (P = .025), and weight (P = .026). Most patients with SF between 750 and 1500 ng/mL had a TIL between 25 and 100 mg/kg (72.8% +/- 5.9%; patients/observations, 24 of 50) or an LIC between 2.5 and 10 mg/g dry liver weight (75% +/- 0%; patients/observations, 8 of 9). Most patients with SF of 3000 ng/mL or greater had a TIL of 100 mg/kg or greater (95.3% +/- 6.7%; patients/observations, 7 of 16) or an LIC of 10 mg/g dry liver weight or greater (87.7% +/- 4.3%; patients/observations, 11 of 18). Although SF changes are nonlinear, levels less than 1500 ng/mL indicated mostly acceptable iron overload; levels of 3000 ng/mL or greater were specific for significant iron overload and were associated with liver injury. However, to determine accurately iron overload in patients with intermediately elevated SF levels, other methods are required. These trials are registered at www.clinicaltrials.gov as #NCT00000592 and #NCT00006182.

High levels of granulocyte and granulocyte-macrophage colony-stimulating factors in cord blood of normal full-term neonates

Because several human hematopoietic growth factors have been identified and shown to be effective for treatment of congenital or iatrogenic neutropenias, and cord blood contains stimulatory activities for blood-forming cells, we postulated that identification of these factors and analysis of their regulatory role in normal neonates would provide a rationale for their use in treating neonatal infections associated with neutropenia. We studied the plasma levels of granulocyte and granulocyte-macrophage colony-stimulating factors (G-CSF and GM-CSF, respectively) and the frequency of granulomonopoietic colony-forming cells (CFU-GM) in the umbilical cord blood of normal term neonates. Plasma growth factor levels were measured by a biologic assay. Circulating hematopoietic progenitors were assayed for colony formation with different recombinant growth factors used as exogenous growth stimulators. The cell cycle status of these progenitors was analyzed by the thymidine suicide technique. At birth the leukocyte count (mean +/- SD) was 11.0 +/- 3.9 x 10(9)L and the neutrophil count was 5.6 +/- 2.6 x 10(9)/L. The incidence of CFU-GM was significantly higher in umbilical cord blood than in normal adult peripheral blood (p less than 0.005) with up to 40% of the cells in S phase (less than 10% in normal adults). Plasma levels of G-CSF and GM-CSF at birth were 40.8 +/- 2.8 U/ml and 19.9 +/- 5.2 U/ml, respectively (normal adult plasma levels 2.5 +/- 1.5 U/ml for G-CSF and undetectable for GM-CSF). These high levels of G-CSF and GM-CSF in umbilical cord blood of normal neonates might play a role in maintaining adequate neutrophil production.

Diabetes mellitus, thiamine-dependentmegaloblastic anemia, and sensorineural deafness associated with deficient α-ketoglutarate dehydrogenase activity

Three brothers with diabetes mellitus, thiamine-responsive megaloblastic anemia, and sensorineural deafness are reported. Two had, in addition, congenital septal defects. The activities of thiamine-dependent enzymes were determined in one patient, revealing low alpha-ketoglutarate dehydrogenase activity, which could have caused a sideroblastic anemia with secondary megaloblastic changes. The anemia was thiamine dependent. The cause of the diabetes mellitus was not known, but it was not type 1.