Severe congenital neutropenia unresponsive to G-CSF

Summary Severe congenital neutropenia (SCN) is an inherited disorder characterized by severe neutropenia and recurrent infections from an early age, with bone marrow showing a maturational arrest of granulopoiesis at the promyelocyte stage. Since the introduction of G-CSF therapy the prognosis for affected children has improved dramatically. We describe two patients with SCN who were clinically unresponsive to G-CSF therapy. The results of in-vitro colony assays from these two patients are presented together with the results from the mother of one of these patients who also has a chronic neutropenia, and a further child with SCN who responded to treatment with G-CSF.     

Severe congenital neutropenia with unique features of dysgranulopoiesis

Congenital dysgranulopietic neutropenia (CDN) is a recently proposed entity that describes a small subgroup of children with clinically severe neutropenia. We followed and studied a 3-year-old girl with neutropenia (<500/mm³) and recurrent severe infections in whom repeated marrow evaluations revealed large (30-50 μm) multinucleated promyelocytes to polymorphonuclear cells with as many as 4 to 16 nuclei or nuclear lobes, respectively. In addition to the nuclear endoreduplication, ultrastructural and cytochemical evaluation of these cells demonstrated abnormalities in granule genesis and centriole structure. Concomitantly. immunoperoxidase staining indicated that many of the granules were devoid of lactoferrin but not lysozyme. In vitro proliferation studies revealed normal to increased thymidine labeling, normal numbers of colony-forming cells, and normal colony-stimulating activity from blood and marrow mononuclear cells, findings consistent with ineffective myelopoiesis. However, serum folate, B₁₂, and lysozyme levels were normal. The nuclear and cytoplasmic abnormalities in this patient result in an extreme example of CDN, distinct from previously described cases.     

Severe congenital neutropenia and the unfolded protein response

PURPOSE OF REVIEW: Severe congenital neutropenia is an inborn disorder of granulopoiesis. Mutations of ELA2, encoding neutrophil elastase, are present in approximately 50% of cases of severe congenital neutropenia and nearly all cases of cyclic neutropenia, a related disorder of granulopoiesis. Herein, we review recent studies of the molecular pathogenesis of severe congenital neutropenia, with an emphasis on those cases associated with mutations of ELA2. RECENT FINDINGS: Genetic evidence suggests that ELA2 mutations act in a dominant, cell-intrinsic fashion to disrupt granulopoiesis. A unifying model that accounts for the different clinical phenotypes (severe congenital neutropenia versus cyclic neutropenia) and the diversity of ELA2 mutations (over 50 have been identified), however, is lacking. Recent studies suggest that mutations of ELA2 may cause disease through induction of the unfolded protein response. In this model, the ELA2 mutations result in the production of misfolded neutrophil elastase protein, activation of the unfolded protein response, and ultimately apoptosis of granulocytic precursors. The propensity of individual neutrophil elastase mutants to misfold may determine the magnitude of unfolded protein response-induced apoptosis and ultimately the clinical phenotype. SUMMARY: Recent studies provide support for a unfolded protein response model of disease pathogenesis in cases of severe congenital neutropenia associated with ELA2 mutations and place severe congenital neutropenia in a growing list of human disease caused by misfolded proteins.     

Severe congenital neutropenia: new genes explain an old disease

PURPOSE OF REVIEW: This review summarizes the recent advances in the diagnosis and molecular characterization of isolated and syndromal forms of severe congenital neutropenia. RECENT FINDINGS: It has become evident that severe congenital neutropenia comprises several genetically distinct entities. In 1999, mutations were identified in the neutrophil elastase gene ELA2. ELA2 mutations have been found in cyclic, sporadic and autosomal dominant neutropenia. Recently, homozygous mutations in the antiapoptotic gene HAX1 were found in patients with autosomal recessive severe congenital neutropenia. Ongoing linkage studies suggest that more and, as yet unidentified, genes may be involved in the pathophysiology of severe congenital neutropenia. In other patients, congenital neutropenia is not an isolated finding but is associated with other abnormalities, in particular, lymphoid immunodeficiency and pigmentation defects such as Chédiak-Higashi syndrome, Griscelli syndrome type 2, Hermansky-Pudlak syndrome type 2, or deficiency of the endosomal adaptor p14. The molecular identification of these disorders originating from mutations in lysosome (related) proteins has advanced our knowledge of intracellular protein trafficking. SUMMARY: Recent insights into the molecular etiology of severe congenital neutropenia provide the opportunity for a definitive genetic classification system. Based on this knowledge, disease-related risks may be recognized and optimized therapeutic options may become available.     

Severe neutropenia consequent to sustained-release procainamide

A patient with reversible severe neutropenia and relative eosinophilia caused by sustained-release procainamide is described. Significant early and very late complications of therapy with procainamide preparations have been rarely described so far. The important pharmacokinetic aspects of sustained release procainamide are discussed, and the postulated mechanisms of induction of blood dyscrasias are reviewed.     

Severe neutropenia associated with sustained-release procainamide

Neutropenia is a rare complication of procainamide therapy. However, over a period of 20 months, 8 patients developed severe neutropenia while taking a sustained-release preparation of the drug. Seven patients presented with fever and constitutional symptoms and one patient was asymptomatic. Bone marrow examinations showed myeloid aplasia or maturation arrest in 5 patients and myeloid hyperplasia in 1. Neutropenia resolved within 30 days of drug withdrawal, and all patients survived. A case-control study showed a significant association between sustained-release procainamide therapy and severe neutropenia in 5 of 114 patients (4.4%) recovering from open-heart surgery (Mantel-Haenszel chi square = 13.84; p less than 0.001). Thus, life-threatening neutropenia may be common with sustained-release procainamide preparations.     

Prenatal diagnosis of congenital dysgranulopoietic neutropenia

Summary. Congenital dysgranulopoietic neutropenia is a severe life-threatening disease characterized by specific dysmorphic granulocytes. Prenatal diagnosis of congenital neutropenia was reported first in 1983. In 1989 we reported a family with congenital dysgranulopoietic neutropenia in two siblings (one male and one female): in 1992 the mother became pregnant and prenatal diagnosis was performed by cordocentesis. The results we obtained from the fetus at risk enabled us to suggest that it was not affected, and we advised the parents that the pregnancy could continue: a healthy female was born. The neutrophil count at 2 and 4 months was normal.     

Granulopoiesis in patients with congenital neutropenia

Granulopoiesis was investigated in five patients with congenital neutropenia (CN) (one Kostmann type, four benign forms). In semisolid agar culture, the marrow cells of all five patients produced normal numbers of CFU-c (colony-forming unit-culture). The size and classification of colonies were normal. In suspension culture in vitro with exogenous colony-stimulating factor (CSF) generated from omental-conditioned medium (OMCM), the myeloid precursors of all patients could proliferate and differentiate into normal polymorphonuclear neutrophils (PMNs). But in the absence of exogenous CSF, myeloid precursors of the patient with Kostmann-type CN did not proliferate or differentiate into PMNs at all. In the four patients with benign neutropenia, however, PMNs were found even without exogenous CSF similar to normal individuals. These results suggest that patients with CN may have normal granulopoietic stem cells with normal proliferative and differentiating capacity in response to exogenous CSF. When a small amount of normal human serum was added to normal marrow cultures stimulated by exogenous CSF, the colony growth increased in a superadditive manner. The enhancing activity of serum from neutropenic patients differed from that of normal serum. Especially, the addition of serum from the patient with Kostmann type CN to normal marrow cultures did not show this enhancement effect. The sera of patients with benign neutropenia had less enhancement effect than did normal control serum. These findings might be interpreted as showing an imbalance between CSF enhancer and inhibitors in the patients' serum.     

Neutrophil elastase mutations in congenital neutropenia

Severe congenital neutropenia (SCN) was originally described as an autosomal recessive disorder. Autosomal dominant and sporadic forms of the disease have subsequently been recognized. All forms of the disease are manifest by persistent severe neutropenia and recurrent bacterial infection. Cyclical neutropenia (CyN) is characterized by periodic neutropenia inter-spaced with (near) normal neutrophil counts. Recently, heterozygous mutations in the ELA2 gene encoding neutrophil elastase (NE) have been described in the majority of cases of CyN and sporadic and autosomal dominant SCN. A case of paternal mosaicism has provided genetic "proof" of the pathogenicity of such mutations, but the exact pathogenic mechanism remains elusive. This review will focus on the mosaic proof and examine possible pathogenic mechanisms. The lack of obvious associations and indeed overlap between the mutations that cause the two diseases will also be discussed. Clinically to date, the discovery of an elastase mutation has been of limited value to individual patients. However, it is hoped that further genotype/phenotype studies may improve assessment of patient prognosis.     

Granulopoiesis in severe congenital neutropenia.

The pathogenesis of the granulopoietic failure in three children with severe congenital neutropenia was studied. Mature neutrophils were absent from both peripheral blood and bone marrow. Assay of bone marrow granulocyte colony-forming cells (CFU-C) in a methylcellulose tissue culture system using colony-stimulating activity (CSA) from peripheral blood leukocytes demonstrated normal or increased concentrations of CFU-C compared to those from marrows of 60 age-matched controls. Colonies were of normal size and by light microscopy appeared to contain granulocytes in all stages of maturation including the mature polymorphonuclear neutrophil. CFU-C from peripheral blood of two patients were normal. Production and activity of CSA from the patients' peripheral blood leukocytes and urinary CSA excretion were normal. No serum inhibitors against CFU-C or CSA could be demonstrated using both control and autologous marrow. The defect did not appear to be due to a lack of granulocytic stem cells, a reduction of humoral stimulators of granulopoiesis, nor the presence of an inhibitor as measured by these techniques.     

Digenic mutations in severe congenital neutropenia

Severe congenital neutropenia a clinically and genetically heterogeneous disorder. Mutations in different genes have been described as causative for severe neutropenia, e.g. ELANE, HAX1 and G6PC3. Although congenital neutropenia is considered to be a group of monogenic disorders, the phenotypic heterogeneity even within the yet defined genetic subtypes points to additional genetic and/or epigenetic influences on the disease phenotype. We describe congenital neutropenia patients with mutations in two candidate genes each, including 6 novel mutations. Two of them had a heterozygous ELANE mutation combined with a homozygous mutation in G6PC3 or HAX1, respectively. The other 2 patients combined homozygous or compound heterozygous mutations in G6PC3 or HAX1 with a heterozygous mutation in the respective other gene. Our results suggest that digenicity may underlie this disorder of myelopoiesis at least in some congenital neutropenia patients.     

Kostmann syndrome and severe congenital neutropenia

Congenital neutropenia (CN) includes hematologic disorders characterized by severe neutropenia with an absolute neutrophil count (ANC) below 0.5 x 10(9)/L associated with severe systemic bacterial infections from early infancy. One subtype of CN, Kostmann syndrome, was originally described as an autosomal-recessive disorder, characterized by early-stage maturation arrest of myelopoiesis. Autosomal-dominant and sporadic cases have also been reported. Recent studies on the genetic bases of CN have detected different inherited or spontaneous point mutations in the neutrophil elastase gene. Development of additional genetic defects during the course of disease, such as granulocyte colony-stimulating factor (G-CSF)-receptor gene mutations and cytogenetic aberrations, indicates an underlying genetic instability. Data on more than 300 patients with CN collected by the Severe Chronic Neutropenia International Registry (SCNIR) since 1994 demonstrate that, independent of the CN subtype, more than 90% of patients respond to recombinant human (rHu)G-CSF with ANCs that can be maintained at approximately 1.0 x 10(9)/L. Adverse events include mild splenomegaly, moderate thrombocytopenia, osteoporosis, and malignant transformation into myelodysplasia (MDS)/leukemia. If and how rHuG-CSF treatment impacts on these adverse events remains unclear since there are no historical controls for comparison. Hematopoietic stem cell transplantation (HSCT) is still the only available treatment for patients refractory to rHuG-CSF treatment.