Harry L. Malech

Chronic granulomatous disease. Report on a national registry of 368 patients.

A registry of United States residents with chronic granulomatous disease (CGD) was established in 1993 in order to estimate the minimum incidence of this uncommon primary immunodeficiency disease and characterize its epidemiologic and clinical features. To date, 368 patients have been registered; 259 have the X-linked recessive form of CGD, 81 have 1 of the autosomal recessive forms, and in 28 the mode of inheritance is unknown. The minimum estimate of birth rate is between 1/200,000 and 1/250,000 live births for the period 1980-1989. Pneumonia was the most prevalent infection (79% of patients; Aspergillus most prevalent cause), followed by suppurative adenitis (53% of patients; Staphylococcus most prevalent cause), subcutaneous abscess (42% of patients; Staphylococcus most prevalent cause), liver abscess (27% of patients; Staphylococcus most prevalent cause), osteomyelitis (25% of patients; Serratia most prevalent cause), and sepsis (18% of patients; Salmonella most prevalent cause). Fifteen percent of patients had gastric outlet obstruction, 10% urinary tract obstruction, and 17% colitis/enteritis. Ten percent of X-linked recessive kindreds and 3% of autosomal recessive kindreds had family members with lupus. Eighteen percent of patients either were deceased when registered or died after being registered. The most common causes of death were pneumonia and/or sepsis due to Aspergillus (23 patients) or Burkholderia cepacia (12 patients). Patients with the X-linked recessive form of the disease appear to have a more serious clinical phenotype than patients with the autosomal recessive forms of the disease, based on the fact that they are diagnosed significantly earlier (mean, 3.01 years of age versus 7.81 years of age, respectively), have a significantly higher prevalence of perirectal abscess (17% versus 7%), suppurative adenitis (59% versus 32%), bacteremia/fungemia (21% versus 10%), gastric obstruction (19% versus 5%), and urinary tract obstruction (11% versus 3%), and a higher mortality (21.2% versus 8.6%).

Genetic, biochemical, and clinical features of chronic granulomatous disease.

The reduced nicotinamide dinucleotide phosphate (NADPH) oxidase complex allows phagocytes to rapidly convert O2 to superoxide anion which then generates other antimicrobial reactive oxygen intermediates, such as H2O2, hydroxyl anion, and peroxynitrite anion. Chronic granulomatous disease (CGD) results from a defect in any of the 4 subunits of the NADPH oxidase and is characterized by recurrent life-threatening bacterial and fungal infections and abnormal tissue granuloma formation. Activation of the NADPH oxidase requires translocation of the cytosolic subunits p47phox (phagocyte oxidase), p67phox, and the low molecular weight GT-Pase Rac, to the membrane-bound flavocytochrome, a heterodimer composed of the heavy chain gp91phox and the light chain p22phox. This complex transfers electrons from NADPH on the cytoplasmic side to O2 on the vacuolar or extracellular side, thereby generating superoxide anion. Activation of the NADPH oxidase requires complex rearrangements between the protein subunits, which are in part mediated by noncovalent binding between src-homology 3 domains (SH3 domains) and proline-rich motifs. Outpatient management of CGD patients relies on the use of prophylactic antibiotics and interferon-gamma. When infection is suspected, aggressive effort to obtain culture material is required. Treatment of infections involves prolonged use of systemic antibiotics, surgical debridement when feasible, and, in severe infections, use of granulocyte transfusions. Mouse knockout models of CGD have been created in which to examine aspects of pathophysiology and therapy. Gene therapy and bone marrow transplantation trials in CGD patients are ongoing and show great promise.

Hyper-IgE Syndrome with Recurrent Infections — An Autosomal Dominant Multisystem Disorder

BACKGROUND The hyper-IgE syndrome with recurrent infections is a rare immunodeficiency characterized by recurrent skin and pulmonary abscesses and extremely elevated levels of IgE in serum. Associated facial and skeletal features have been recognized, but their frequency is unknown, and the genetic basis of the hyper-IgE syndrome is poorly understood. METHODS We studied 30 patients with the hyper-IgE syndrome and 70 of their relatives. We took histories, reviewed records, performed physical and dental examinations, took anthropometric measurements, and conducted laboratory studies. RESULTS Nonimmunologic features of the hyper-IgE syndrome were present in all patients older than eight years. Seventy-two percent had the previously unrecognized feature of failure or delay of shedding of the primary teeth owing to lack of root resorption. Common findings among patients were recurrent fractures (in 57 percent of patients), hyperextensible joints (in 68 percent), and scoliosis (in 76 percent of patients 16 years of age or older). The classic triad of abscesses, pneumonia, and an elevated IgE level was identified in 77 percent of all patients and in 85 percent of those older than eight. In 6 of 23 adults (26 percent), IgE levels declined over time and came closer to or fell within the normal range. Autosomal dominant transmission of the hyper-IgE syndrome was found, but with variable expressivity. Of the 27 relatives at risk for inheriting the hyper-IgE syndrome, 10 were fully affected, 11 were unaffected, and 6 had combinations of mild immunologic, dental, and skeletal features of the hyper-IgE syndrome. CONCLUSIONS The hyper-IgE syndrome is a multisystem disorder that affects the dentition, the skeleton, connective tissue, and the immune system. It is inherited as a single-locus autosomal dominant trait with variable expressivity.

Fungal infection in chronic granulomatous disease: The importance of the phagocyte in defense against fungi

Among 245 cases of chronic granulomatous disease which were evaluated, fungal infection occurred in 20.4 percent. Fungi encountered include Aspergillus, Torulopsis and Candida. In 18 percent of the patients with fungal infection, the disease was limited to soft tissue or bone; all did well. Most of the patients had fungal pneumonia and/or widely disseminated disease; diagnosis was usually confirmed by open lung biopsy. Patients with pneumonia or disseminated disease who received no therapy succumbed to infection, whereas more than half the patients who received antifungal therapy were cured. Modalities of treatment included antifungal chemotherapy, surgical removal of infected tissue and granulocyte transfusion. Although several patients showed dramatic improvement during granulocyte transfusions given in combination with antifungal chemotherapy, the improvement achieved was not statistically significant when compared with that achieved with chemotherapy alone. These results emphasized the importance of phagocytic cells in defense against fungi and the need for further evaluation of granulocyte transfusion therapy in compromised hosts in whom fungal infections develop.

Residual NADPH Oxidase and Survival in Chronic Granulomatous Disease

BACKGROUND Failure to generate phagocyte-derived superoxide and related reactive oxygen intermediates (ROIs) is the major defect in chronic granulomatous disease, causing recurrent infections and granulomatous complications. Chronic granulomatous disease is caused by missense, nonsense, frameshift, splice, or deletion mutations in the genes for p22(phox), p40(phox), p47(phox), p67(phox) (autosomal chronic granulomatous disease), or gp91(phox) (X-linked chronic granulomatous disease), which result in variable production of neutrophil-derived ROIs. We hypothesized that residual ROI production might be linked to survival in patients with chronic granulomatous disease. METHODS We assessed the risks of illness and death among 287 patients with chronic granulomatous disease from 244 kindreds. Residual ROI production was measured with the use of superoxide-dependent ferricytochrome c reduction and flow cytometry with dihydrorhodamine oxidation assays. Expression of NADPH oxidase component protein was detected by means of immunoblotting, and the affected genes were sequenced to identify causal mutations. RESULTS Survival of patients with chronic granulomatous disease was strongly associated with residual ROI production as a continuous variable, independently of the specific gene affected. Patients with mutations in p47(phox) and most missense mutations in gp91(phox) (with the exception of missense mutations in the nucleotide-binding and heme-binding domains) had more residual ROI production than patients with nonsense, frameshift, splice, or deletion mutations in gp91(phox). After adolescence, mortality curves diverged according to the extent of residual ROI production. CONCLUSIONS Patients with chronic granulomatous disease and modest residual production of ROI have significantly less severe illness and a greater likelihood of long-term survival than patients with little residual ROI production. The production of residual ROI is predicted by the specific NADPH oxidase mutation, regardless of the specific gene affected, and it is a predictor of survival in patients with chronic granulomatous disease. (Funded by the National Institutes of Health.).

Genetic Linkage of Hyper-IgE Syndrome to Chromosome 4

The hyper-IgE syndrome (HIES) is a rare primary immunodeficiency characterized by recurrent skin abscesses, pneumonia, and highly elevated levels of serum IgE. HIES is now recognized as a multisystem disorder, with nonimmunologic abnormalities of the dentition, bones, and connective tissue. HIES can be transmitted as an autosomal dominant trait with variable expressivity. Nineteen kindreds with multiple cases of HIES were scored for clinical and laboratory findings and were genotyped with polymorphic markers in a candidate region on human chromosome 4. Linkage analysis showed a maximum two-point LOD score of 3.61 at recombination fraction of 0 with marker D4S428. Multipoint analysis and simulation testing confirmed that the proximal 4q region contains a disease locus for HIES.

Neutrophil extracellular traps enriched in oxidized mitochondrial DNA are interferogenic and contribute to lupus-like disease

Neutrophil extracellular traps (NETs) are implicated in autoimmunity, but how they are generated and their roles in sterile inflammation remain unclear. Ribonucleoprotein immune complexes (RNP ICs), inducers of NETosis, require mitochondrial reactive oxygen species (ROS) for maximal NET stimulation. After RNP IC stimulation of neutrophils, mitochondria become hypopolarized and translocate to the cell surface. Extracellular release of oxidized mitochondrial DNA is proinflammatory in vitro, and when this DNA is injected into mice, it stimulates type I interferon (IFN) signaling through a pathway dependent on the DNA sensor STING. Mitochondrial ROS are also necessary for spontaneous NETosis of low-density granulocytes from individuals with systemic lupus erythematosus. This was also observed in individuals with chronic granulomatous disease, who lack NADPH oxidase activity but still develop autoimmunity and type I IFN signatures. Mitochondrial ROS inhibition in vivo reduces disease severity and type I IFN responses in a mouse model of lupus. Together, these findings highlight a role for mitochondria in the generation not only of NETs but also of pro-inflammatory oxidized mitochondrial DNA in autoimmune diseases.

ASPERGILLUS NIDULANS INFECTION IN CHRONIC GRANULOMATOUS DISEASE

Chronic granulomatous disease (CGD) is a rare inherited disorder of the NADPH oxidase complex in which phagocytes are defective in generating reactive oxidants. As a result, patients with CGD suffer from recurrent bacterial and fungal infections. The most common fungal infections are caused by Aspergillus species. Aspergillus nidulans is a rare pathogen in most patient populations with quantitative or qualitative neutrophil defects. We have reviewed all cases in which A. nidulans was isolated from patients at the National Institutes of Health (Bethesda, MD) between 1976 and 1997. A. nidulans infection occurred in 6 patients with CGD, but was not a pathogen in any other patient group. Aspergillus fumigatus was a more common pathogen in CGD compared with A. nidulans, but A. nidulans was more virulent. A. nidulans was significantly more likely to result in death compared with A. fumigatus, to involve adjacent bone, and to cause disseminated disease. Patients with A. nidulans received longer courses of amphotericin B therapy than patients with A. fumigatus, and were treated with surgery more often. In contrast to A. fumigatus, A. nidulans was generally refractory to intensive antifungal therapy, suggesting that early surgery may be important. These data show that A. nidulans is a distinct pathogen in CGD and its isolation carries more severe implications than that of A. fumigatus.

Hematologically important mutations: X-linked chronic granulomatous disease (third update)

Chronic granulomatous disease (CGD) is an immunodeficiency disorder affecting about 1 in 250,000 individuals. The disease is caused by a lack of superoxide production by the leukocyte enzyme NADPH oxidase. Superoxide is used to kill phagocytosed micro-organisms in neutrophils, eosinophils, monocytes and macrophages. The leukocyte NADPH oxidase is composed of five subunits, of which the enzymatic component is gp91-phox, also called Nox2. This protein is encoded by the CYBB gene on the X chromosome. Mutations in this gene are found in about 70% of all CGD patients. This article lists all mutations identified in CYBB in the X-linked form of CGD. Moreover, apparently benign polymorphisms in CYBB are also given, which should facilitate the recognition of future disease-causing mutations.

NAD(P)H Oxidase 1, a Product of Differentiated Colon Epithelial Cells, Can Partially Replace Glycoprotein 91phox in the Regulated Production of Superoxide by Phagocytes

Reactive oxygen species (ROS) serve several physiological functions; in some settings they act in host defense, while in others they function in cellular signaling or in biosynthetic reactions. We studied the expression and function of a recently described source of ROS, NAD(P)H oxidase 1 or Nox1, which has been associated with cell proliferation. In situ hybridization in mouse colon revealed high Nox1 expression within the lower two-thirds of colon crypts, where epithelial cells undergo proliferation and differentiation. Human multitumor tissue array analysis confirmed colon-specific Nox1 expression, predominantly in differentiated epithelial tumors. Differentiation of Caco2 and HT29 cells with 1α,25-dihydroxyvitamin D3 or IFN-γ enhances Nox1 expression and decreases cell proliferation, suggesting that Nox1 does not function as a mitogenic oxidase in colon epithelial cells. Transduction with retrovirus encoding Nox1 restored activation and differentiation-dependent superoxide production in gp91phox-deficient PLB-985 cells, indicating close functional similarities to the phagocyte oxidase (phox). Furthermore, coexpression of cytosolic components, p47phox and p67phox, augments Nox1 activity in reconstituted K562 cells. Finally, Nox1 partially restores superoxide production in neutrophils differentiating ex vivo from gp91phox-deficient CD34+ peripheral blood-derived stem cells derived from patients with X-linked chronic granulomatous disease. These studies demonstrate a significant functional homology (cofactor-dependent and activation-regulated superoxide production) between Nox1 and its closest homologue, gp91phox, suggesting that targeted up-regulation of Nox1 expression in phagocytic cells could provide a novel approach in the molecular treatment of chronic granulomatous disease.