Salvatore Bertolone

Mutations in proto-oncogene GFI1 cause human neutropenia and target ELA2

Mice lacking the transcriptional repressor oncoprotein Gfi1 are unexpectedly neutropenic. We therefore screened GFI1 as a candidate for association with neutropenia in affected individuals without mutations in ELA2 (encoding neutrophil elastase), the most common cause of severe congenital neutropenia (SCN; ref. 3). We found dominant negative zinc finger mutations that disable transcriptional repressor activity. The phenotype also includes immunodeficient lymphocytes and production of a circulating population of myeloid cells that appear immature. We show by chromatin immunoprecipitation, gel shift, reporter assays and elevated expression of ELA2 in vivo in neutropenic individuals that GFI1 represses ELA2, linking these two genes in a common pathway involved in myeloid differentiation.

Treatment of childhood acute immune thrombocytopenic purpura with anti-D immune globulin or pooled immune globulin

OBJECTIVE To evaluate the effectiveness of initial treatment of children with acute immune thrombocytopenic purpura (ITP) with anti-D immune globulin (anti-D) or pooled IgG immune globulin (IVIg). STUDY DESIGN The medical charts of 33 children diagnosed with acute ITP from May 1995 to October 1997 were reviewed. Patient data were eligible for analysis if, for the new diagnosis of acute ITP, the patient had received either anti-D at 45 to 50 microg/kg (WinRho SD, NABI) or IVIg at 0.8 to 1 g/kg (Gammagard SD, Baxter-Highland). The platelet response time for each treatment group was compared by the Mann-Whitney U test. RESULTS Time to achieve a platelet count >/=20 x 10(9 )/L (20,000/mm3 ) was 1.54 +/- 0.51 days in the IVIg group (n = 13) and 1.26 +/- 0.82 days in the anti-D group (n = 14) (P =.34). Time to achieve a platelet count >/=40 x 10(9 )/L (40,000/mm3 ) was 1.77 +/- 0.74 and 1.49 +/- 1.01 days for the IVIg and anti-D groups, respectively (P =.32). Children given IVIg were hospitalized for 2.1 +/- 0.87 days, whereas those given anti-D were hospitalized for 1.94 +/- 1.08 days. A net decrease in hemoglobin concentration was observed after receipt of IVIg (9.1 +/- 7.3 g/L [0.91 +/- 0.73 g/dL]) and after anti-D therapy (4.5 +/- 10.3 g/L [0.45 +/- 1.03 g/dL], P =.23). No patient required intervention for hemolysis. CONCLUSIONS In this retrospective analysis anti-D was as effective as IVIg for the treatment of acute ITP in children. However, randomized, controlled trials are needed to establish the role of anti-D in the treatment of acute ITP in children.

A phase II study of cisplatin therapy in recurrent childhood brain tumors

Thirty-six children with brain tumors were treated with surgery, radiation and/or adjuvant chemotherapy. After tumor recurrence, cisplatin (60 mg/m2/day IV×2) was given every three to four weeks. CT scans were used to measure drug response prior to the first, third and fifth courses. Complete and partial responses were demonstrated in nine of 31 evaluable patients. Dose limiting toxicities were renal and auditory. Seven patients developed the syndrome of inappropriate antidiuretic horm one secretion. This study confirms that cisplatin is active in a spectrum of brain tumors.

Mutations in Growth Factor Independent-1 Associated with Human Neutropenia Block Murine Granulopoiesis through Colony Stimulating Factor-1

Severe congenital neutropenia (SCN) is characterized by a deficiency of mature neutrophils, leading to recurrent bacterial and fungal infections. Although mutations in Elastase-2, neutrophil (ELA2) predominate in human SCN, mutation of Ela2 in mice does not recapitulate SCN. The growth factor independent-1 (GFI1) transcription factor regulates ELA2. Mutations in GFI1 are associated with human SCN, and genetic deletion of Gfi1 results in murine neutropenia. We examined whether human SCN-associated GFI1N382S mutant proteins are causal in SCN and found that GFI1 functions as a rate-limiting granulopoietic molecular switch. The N382S mutation inhibited GFI1 DNA binding and resulted in a dominant-negative block to murine granulopoiesis. Moreover, Gfi1N382S selectively derepressed the monopoietic cytokine CSF1 and its receptor. Gfi1N382S-expressing Csf1-/- cells formed neutrophils. These results reveal a common transcriptional program that underlies both human and murine myelopoiesis, and that is central to the pathogenesis of SCN associated with mutations in GFI1. This shared transcriptional pathway may provide new avenues for understanding SCN caused by mutations in other genes and for clinical intervention into human neutropenias.

Neonatal immune neutropenia following the administration of intravenous immune globulin

Background : In adults—but not neonates—neutropenia has been reported to complicate treatment with intravenous immunoglobulin, but the mechanism is unknown. Purpose : To describe for the first time the case of a newborn infant who, after intravenous immunoglobulin, demonstrated serum antineutrophil antibodies and neutropenia. Patients and Methods : The 1,425-g, 36-week-gestation boy was healthy except for intrauterine growth retardation. Intravenous immunoglobulin (lg/dose × 3) was administered to treat alloimmune thrombocytopenia. Neutrophil-specific antibodies were detected by a granulocyte immunofluorescence assay. Results : After the intravenous immunoglobulin, the platelet count normalized but the neutrophil count declined to 450/mm3. Neutrophil-specific antibodies were detected in the serum of the infant but not in the maternal serum. Furthermore, cross-matching revealed that the maternal serum did not react with the infants granulocytes. Two of three random lots of intravenous immunoglobulin contained detectible anti-neutrophil antibodies. Conclusions : After intravenous immunoglobulin, the infants serum contained one or more anti-neutrophil antibodies that were not maternal in origin. We speculate that the neutropenia resulted from the administration of intravenous immunoglobulin containing antineutrophil antibodies.

Assessment of cerebral tissue oxygenation in patients with sickle cell disease: effect of transfusion therapy.

This study used spatially resolved transcranial near-infrared spectroscopy (NIRS) to compare brain tissue oxygenation in sickle cell disease (SCD) patients with that of healthy children. In addition, NIRS was used to measure the dynamic response of cerebral oxygen balance to erythrocytapheresis. Transcranial NIRS measurements were obtained from 25 children with SCD who were not receiving transfusion or hydroxyurea therapy (NT-SCD). These patients were divided into two subgroups, those with mild (n = 10) or severe (n = 15) SCD symptoms. In addition, NIRS measurements were performed in 16 SCD patients with severe disease maintained on long-term erythrocytapheresis (T-SCD) and in 35 control children. The lowest mean brain tissue oxygen saturation occurred in the NT-SCD subgroup with severe symptoms (48 ± 9%; P < 0.001 vs. control). NT-SCD patients with mild symptoms had higher saturation (62 ± 8%; P < 0.001 vs. control), while the highest appeared in the control group (72 ± 7%). In T-SCD patients, however, brain tissue oxygen saturations were higher than severely symptomatic NT-SCD children and similar to mildly symptomatic NT-SCD children (65 ± 7%). Non-invasive measurements of brain tissue oxygenation with NIRS revealed that abnormal oxygen saturation levels in SCD patients correlated with the severity of their clinical manifestations. Additionally, cerebral oxygen balance seems to be favorably affected by erythro-cytapheresis.

Antibody development in pediatric sickle cell patients undergoing erythrocytapheresis

Erythrocytapheresis, or red blood cell exchange transfusion (RBCX), with donor red blood cell (RBC) units is now increasingly used in the treatment of acute and chronic complications of sickle cell disease (SCD). As in all transfusions, RCBX carries a risk of immunization against foreign antigen on transfused cells. However, by selecting donor units with RBC phenotypes similar to the patient, the risk of allo‐ and autoimmunization can be reduced.

Successful treatment of refractory autoimmune hemolytic anemia with monthly rituximab following nonmyeloablative stem cell transplantation for sickle cell disease.

Autoimmune hemolytic anemia (AIHA) can occur following hematopoietic stem cell transplantation (HSCT) and may be associated with other cytopenias. It can also occur in the context of chronic red cell transfusion in patients maintained on hypertransfusion regimens. There are an increasing number of reports on the successful treatment of autoimmune cytopenias with the monoclonal anti-CD20 antibody rituximab, including a few patients in a post-HSCT setting. The authors report the successful treatment with rituximab of refractory AIHA following allogeneic nonmyeloablative bone marrow transplantation in a child with sickle cell disease.

Pediatric dosing of rituximab revisited: serum concentrations in opsoclonus-myoclonus syndrome.

To longitudinally assess serum concentrations of rituximab, it was administered intravenously to 25 children with opsoclonus-myoclonus syndrome at 375 mg/m2 on each of 4 consecutive weeks with (Group I and II) or without (Group III) conventional immunotherapy. Serum rituximab levels, drawn before and after each infusion and at later intervals, were analyzed by enzyme-linked immunosorbent assay. Rituximab concentration increased stepwise with each infusion, dropping by the next infusion, thereby forming 4 discrete peaks (Cmax) and troughs (Cmin). It then fell precipitously to trace levels at 4 months. However, Cmax and Cmin curves differed significantly between groups. Compared with the youngest children (Group I), the oldest (Group III) had a 34% lower rituximab concentration at the fourth infusion, 45% less IgM depletion 1 month later, and received 20% less rituximab when the dose was recalculated as mg/kg. Serum IgM and rituximab levels were negatively correlated. Peak rituximab concentration did not correlate with adrenocorticotropic hormone dose. These results indicate that the degree of serum IgM depletion is a useful indicator for rituximab dose equivalency in children of different ages. They also suggest that pediatric rituximab dosing should be based on body weight, not surface area. (ClinicalTrials.gov NCT00244361).

Cathlink® 20: A subcutaneous implanted central venous access device used in children with sickle cell disease on long‐term erythrocytapheresis—a report of low complication rates

Experience with the use of central venous access device (CVAD) in children with sickle cell disease (SCD) on hypertransfusion is limited and published studies report wide variability in the rates of CVAD‐associated complications.