Efficacy, safety, tolerability and pharmacokinetics of a novel human immune globulin subcutaneous, 20%: a Phase 2/3 study in Europe in patients with primary immunodeficiencies.

A highly concentrated (20%) immunoglobulin (Ig)G preparation for subcutaneous administration (IGSC 20%), would offer a new option for antibody replacement therapy in patients with primary immunodeficiency diseases (PIDD). The efficacy, safety, tolerability and pharmacokinetics of IGSC 20% were evaluated in a prospective trial in Europe in 49 patients with PIDD aged 2-67 years. Over a median of 358 days, patients received 2349 IGSC 20% infusions at monthly doses equivalent to those administered for previous intravenous or subcutaneous IgG treatment. The rate of validated acute bacterial infections (VASBIs) was significantly lower than 1 per year (0·022/patient-year, P < 0·0001); the rate of all infections was 4·38/patient-year. Median trough IgG concentrations were ≥ 8 g/l. There was no serious adverse event (AE) deemed related to IGSC 20% treatment; related non-serious AEs occurred at a rate of 0·101 event/infusion. The incidence of local related AEs was 0·069 event/infusion (0·036 event/infusion, when excluding a 13-year-old patient who reported 79 of 162 total related local AEs). The incidence of related systemic AEs was 0·032 event/infusion. Most related AEs were mild, none were severe. For 64·6% of patients and in 94·8% of IGSC 20% infusions, no local related AE occurred. The median infusion duration was 0·95 (range = 0·3-4·1) h using mainly one to two administration sites [median = 2 sites (range = 1-5)]. Almost all infusions (99·8%) were administered without interruption/stopping or rate reduction. These results demonstrate that IGSC 20% provides an effective and well-tolerated therapy for patients previously on intravenous or subcutaneous treatment, without the need for dose adjustment.

T and B cell deficiency associated with yellow nail syndrome.

Yellow nail syndrome (YNS) is a rare disorder of unknown aetiology that is characterized by yellow nails associated with lymphoedema and chronic respiratory manifestations. There are no detailed immunological studies in YNS. In this study, we present first extensive immunological analysis of both adaptive and innate immunity in two patients with YNS. One patient has common variable immunodeficiency, whereas second patient has specific antibody deficiency syndrome. Severe lymphopaenia, a striking deficiency of naïve CD4+ and CD8+ T cells and total B cells, and increased transitional B cells were observed. T cell proliferative response to mitogens and antigens was significantly reduced in both patients. Both patients failed to make specific antibody response to pneumococci. Complement, natural killer cell activity and neutrophil oxidative burst were normal. Immunoglobulin administration resulted in decreased frequency and severity of infections, and an impressive effect was observed on lymphoedema and on the recurrence of pleural effusion. Our data show that YNS is associated with both T and B cell defects. Furthermore, Immunoglobulin may be beneficial in clinical manifestations of lymphoedema.

H2AX gene does not have a modifier effect on ataxia-telangiectasia phenotype.

Ataxia-telangiectasia (AT) is a complex disorder characterized by progressive neurodegeneration, immunodeficiency, hypersensitivity to DNA damaging agents and cancer predisposition. Clinical heterogeneity is observed even among the affected siblings with AT. Mutations of the ataxia-telangiectasia mutated (ATM) gene are responsible for AT. H2AX, an essential histone protein, is phosphorylated by ATM in response to double-strand breaks, and H2AX-deficient mice share some clinical and laboratory findings with AT. Therefore, we sought a possible modifier effect of H2AX gene on various clinical features in a group of patients with AT and healthy controls. We performed sequence analysis of H2AX gene in 81 patients with AT, and in 51 of them, we analysed methylation. We examined H2AX gene expression in 25 patients. We investigated 48 healthy individuals as a control group. We did not detect any mutation or sequence variation in the H2AX gene, or any altered methylation pattern in any of the patients. Although H2AX gene expression was markedly increased (2.5- to 11.8-fold) in five of 25 patients, and slightly increased (1.5- to 2.4-fold) in four patients, the correlations between H2AX gene expression and the evaluated clinical features of the patients were not significant. Other potential modifier genes that might be scrutinized in AT patients include p53, 53BP1 and TIP60, as well as the genes that effect mitochondrial function and the oxidative response.

Differential sensitivity of naïve and subsets of memory CD4+ and CD8+ T cells to hydrogen peroxide-induced apoptosis.

CD4+ and CD8+ memory T cells are identified into central and effector memory subsets, which are characterized by distinct homing patterns and functions. In this investigation, we show that naïve and central memory CD4+ and CD8+ T cells are sensitive to hydrogen peroxide (H2O2)-induced apoptosis, whereas effector memory CD4+ and CD8+ T cells are relatively resistant to H2O2-induced apoptosis. Apoptosis in naïve and central memory CD4+ and CD8+ is associated with the release of cytochrome c and activation of caspase-9 and caspase-3, upregulation of Bax and voltage-dependent anion channel (VDAC) expression, and decreased intracellular glutathione (GSH). In vitro GSH and a superoxide dismutase mimetic Mn(III) tetrakis (1-methyl-4-pyridyl) porphyrin inhibited H2O2-induced apoptosis in both naïve and central memory CD4+ and CD8+ T cells. Furthermore, VDAC inhibitor 4,4'-diisothiocynostilbene-2,2'-disulfonic acid blocked H2O2-induced apoptosis. These data demonstrate that H2O2 induces apoptosis preferentially in human naïve and central memory CD4+ and CD8+ T cells via the mitochondrial pathway by regulating intracellular GSH and the expression of Bax and VDAC.

Role of NF-kappaB signaling pathway in increased tumor necrosis factor-alpha-induced apoptosis of lymphocytes in aged humans.

In human aging, lymphocytes display increased sensitivity to tumor necrosis factor-alpha (TNF-alpha)-induced apoptosis. TNF-alpha induces both survival and apoptotic signals. The survival signal is mediated by the activation of NF-kappaB. Although a role of certain proapoptotic molecules in aging has been reported, a role of altered NF-kappaB signaling pathway has not been explored in detail. In this study, we have compared TNF-alpha-induced activation of NF-kappaB, phosphorylation of IkappaBalpha, and the expression of IKKbeta between lymphocytes from young and aged humans. Furthermore, we have explored a role of IKKbeta in increased susceptibility of lymphocytes from aged humans to TNF-alpha-induced apoptosis. Lymphocytes from aged humans displayed decreased activation of NF-kappaB, reduced phosphorylation of IkappaBalpha, and decreased expression of IKKbeta. In addition, overexpression of IKKbeta in lymphocytes from aged humans normalized TNF-alpha-induced apoptosis to the level of young subjects. These data suggest a deficiency of NF-kappaB signaling pathway and a role of IKKbeta, at least in part, for increased sensitivity of lymphocytes from aged humans to TNF-alpha-induced apoptosis.

Biochemical and molecular basis of thimerosal-induced apoptosis in T cells: a major role of mitochondrial pathway.

The major source of thimerosal (ethyl mercury thiosalicylate) exposure is childhood vaccines. It is believed that the children are exposed to significant accumulative dosage of thimerosal during the first 2 years of life via immunization. Because of health-related concerns for exposure to mercury, we examined the effects of thimerosal on the biochemical and molecular steps of mitochondrial pathway of apoptosis in Jurkat T cells. Thimerosal and not thiosalcylic acid (non-mercury component of thimerosal), in a concentration-dependent manner, induced apoptosis in T cells as determined by TUNEL and propidium iodide assays, suggesting a role of mercury in T cell apoptosis. Apoptosis was associated with depolarization of mitochondrial membrane, release of cytochrome c and apoptosis inducing factor (AIF) from the mitochondria, and activation of caspase-9 and caspase-3, but not of caspase-8. In addition, thimerosal in a concentration-dependent manner inhibited the expression of XIAP, cIAP-1 but did not influence cIAP-2 expression. Furthermore, thimerosal enhanced intracellular reactive oxygen species and reduced intracellular glutathione (GSH). Finally, exogenous glutathione protected T cells from thimerosal-induced apoptosis by upregulation of XIAP and cIAP1 and by inhibiting activation of both caspase-9 and caspase-3. These data suggest that thimerosal induces apoptosis in T cells via mitochondrial pathway by inducing oxidative stress and depletion of GSH.

T-ALL with monoclonal gammopathy and hairy cell features.

The presented case is a boy with T-cell acute lymphoblastic leukemia (ALL) with hairy cell (HC) features and monoclonal gammopathy. The disease process had an acute onset and followed a rapid, progressive course. The patient had minimal splenomegaly and bicytopenia, but the bone marrow displayed increased numbers of reticulin fibers. The blasts were positive for tartrate-resistant acid phosphatase (TRAP) and CD11c. Molecular analysis revealed rearrangement of immunoglobulin heavy chain genes and a rearranged T-cell receptor (TcRbeta) beta gene. The patient responded to conventional ALL therapy. Acute T-cell ALL with HC features in childhood has not been reported previously, either alone or in association with monoclonal gammopathy. We propose "T-ALL with hairy cell features" to describe this case.

Isolated cutaneous response to granulocyte-monocyte colony stimulating factor in fatal idiopathic disseminated Bacillus-Calmette-Guerin infection.

UNLABELLED: Severe disseminated Bacillus-Calmette-Guerin (BCG) infection is very rare and has been regarded as idiopathic when no immunodeficiency is present. This entity seems to be due to several new types of inherited abnormalities in the pathways important in defence against Mycobacteria. Although improvement with interferon-gamma (IFN-gamma) has been reported in some patients, to our knowledge there are no reports on the effect of other cytokines in the treatment of these patients. We report here the clinical response to IFN-gamma and granulocyte-monocyte colony stimulating factor (GM-CSF) treatment in a patient with idiopathic disseminated BCG infection who failed to respond to multiple antimycobacterial agents. The patient showed partial and transitory response to IFN-gamma, however, GM-CSF treatment led to rapid improvement of skin lesions within 2 weeks without any effect on the progression of the disease in the other organ systems. CONCLUSION: The response of idiopathic disseminated Bacillus-Calmette-Guerin infection to granulocyte-monocyte colony stimulating factor treatment was limited to cutaneous lesions. Granulocyte-monocyte colony stimulating factor may have acted to promote wound healing or the levels of this factor achieved in other affected organs may have been inadequate.

TNF-alpha-induced apoptosis in neonatal lymphocytes: TNFRp55 expression and downstream pathways of apoptosis.

Previously we have shown decreased Fas-mediated apoptosis in cord blood lymphocyte subsets. In this study, we compared tumor necrosis factor (TNF)-alpha-induced apoptosis in T lymphocytes and their subsets between cord blood and peripheral blood from healthy young controls. The expression of TNF receptor I (TNFR-I) was assessed by flow cytometry and quantitative RT-PCR. The expression of adapter molecules TNF receptor-associated death domain (TRADD), Fas-associated death domain (FADD) and TNF-associated factor-2 (TRAF-2) and caspase 3 was analyzed by Western blotting. The activity of caspase 3 and caspase 8 was measured by colorimetric assay. The susceptibility of CD4+ and CD8+ T cells to TNF-alpha-induced apoptosis was measured by terminal deoxytidyl transferase-mediated dUTP nick end labelling (TUNEL) assay. Both CD4+ and CD8+ T cell subsets from cord blood demonstrated decreased susceptibility to TNF-alpha-induced apoptosis that was associated with decreased activation of both caspase 8 and caspase 3 as compared to T cell subsets in peripheral blood. Furthermore, expression of TNFR-I, TRADD and caspase 3 was decreased in cord blood lymphocytes as compared to peripheral blood lymphocytes. The significance of these observations is discussed.

Impaired IgG antibody production to pneumococcal polysaccharides in patients with ataxia-telangiectasia.

Various factors seem to be etiologic in the susceptibility to sinopulmonary infections in ataxia-telangiectasia (A-T) patients, i.e., low serum and salivary IgA, low serum IgG2, and even aspiration of saliva. S. pneumoniae is a common pathogen responsible from pulmonary infections and impaired antibody response to polysaccharide antigens is seen in patients with IgG2 and IgA deficiency as well as patients with CVID and WAS. We studied IgG-type antibody production to six pneumococcal serotypes in 29 A-T patients by ELISA before and 3-4 weeks after pneumococcal vaccine. The response was considered positive when the antibody titer was >10 U/ml but weak when the titer was 10-20 U/ml. Twenty-two of 29 (76%) patients did not respond to any of the serotypes, 5 (17%) showed a positive response to one serotype, 1 (3.4%) to two serotypes, and 1 (3.4%) to four serotypes. With conversion to gravimetric units (ng IgG/ml) and >1800 ng/ml (300 ng Ab N/ml) considered a positive response, 5 of 29 (17.2%) patients showed a positive response (300 ng ab N/ml) to two or fewer serotypes. All patients tested produced IgG antibody to tetanus toxoid. Sixteen of 27 (59.3%) patients had low IgG2 and four (14.8%) had low IgG3 levels, while 18 (62.1%) of 29 patients had low serum IgA. No correlation was found either between serum Ig isotype levels and antipolysaccharide antibody response or between susceptibility to infection and antibody production. The mechanism responsible for disturbed antipolysaccharide (TI-2 antigen) antibody production in patients with A-T needs to be investigated. It may provide additional information on the function of the ATM gene product and be helpful in clarifying the role of B cells and contribution of T cells in TI-2 responses.

Cartilage-hair hypoplasia syndrome: increased apoptosis of T lymphocytes is associated with altered expression of Fas (CD95), FasL (CD95L), IAP, Bax, and Bcl2.

Cartilage-hair hypoplasia (CHH) is a rare autosomal recessive short-limbed dwarfism associated with thin and sparse hair and cell mediated or combined immunodeficiency. However, the basis of immune deficiency in CHH is unclear. In this study, we investigated a role of apoptosis in immunodeficiency in a patient with CHH. An increased apoptosis of both CD4+ and CD8+ T cells, as determined by TUNEL assay, was observed in CHH compared to an age-matched healthy dwarf control. Increased apoptosis in CHH was associated with increased expression of Fas (CD95), CD95L, and Bax and decreased expression of Bcl-2 and inhibitor of apoptosis protein (IAP) compared to the control. These data suggest that lymphopenia and immunodeficiency in CHH may be, at least in part, due to increased apoptosis of T cells, possibly through the Fas/ FasL signaling pathway.

Bronchiectasis due to ciliary aplasia in Turner's syndrome.

A seven-year-old girl with Turner's syndrome, who suffered from recurrent respiratory system infections since birth, was investigated to determine the etiology of bronchiectasis. Electron microscopy of recurrent nasal biopsy specimens revealed ciliary aplasia. Ciliary aplasia in Turner's syndrome, has not previously been reported.

A case of chronic severe neutropenia: oral findings and consequences of short-term granulocyte colony-stimulating factor treatment.

Neutropenia is an absolute decrease in the number of circulating neutrophils in the blood which results in susceptibility to severe pyogenic infections. Various oral findings such as periodontitis, alveolar bone loss and ulceration may be seen in neutropenic patients. A case is presented of a 6 year old girl with chronic, probably congenital, severe neutropenia with frequent respiratory tract infections, recurrent oral ulcerations and significant periodontal break-down resembling prepubertal periodontitis. She was given granulocyte-colony stimulating factor (G-CSF) treatment which resulted in an increase in granulocyte count within two weeks and resolution of the neutropenic ulceration. It is suggested that G-CSF together with dental care regimens is a promising treatment model in chronic severe neutropenia cases presenting with oral manifestations.

Defective serum opsonization activity in children aged 6-48 months having acute purulent otitis media.

Serum opsonization of yeast (Saccharomyces) was investigated in 51 patients whose ages were between six and 48 months (median 15 months) with acute purulent otitis media and in an age-matched control group (median 13 months). Opsonization was assessed by measuring yeast particle uptake in an assay based on an electronic count of the unphagocytosed particles in serum by polymorphonuclear leukocytes. Despite normal levels of CH50 and serum immunoglobulins, a defective opsonization was determined in 13.7 percent of the patients (7 in 51). The corresponding figure was 2.9 percent in 103 healthy controls (p < 0.001). On the other hand, 218 percent (5 in 23) of the children having a history of recurrent purulent otitis media showed defective opsonization (p < 0.001). Previously, the presence of an opsonization defect has been linked to low levels of mannan binding lectin (MBL), a calcium dependent serum lectin that acts as an opsonin. Therefore, our findings indirectly support the idea that MBL has an important role as host defense, particularly in the earlier period of life when the antibody repertoire is restricted.

Mutations in the mu heavy-chain gene in patients with agammaglobulinemia.

BACKGROUND: Most patients with congenital hypogammaglobulinemia and absent B cells are males with X-linked agammaglobulinemia, which is caused by mutations in the gene for Bruton's tyrosine kinase (Btk); however, there are females with a similar disorder who do not have mutations in this gene. We studied two families with autosomal recessive defects in B-cell development and patients with presumed X-linked agammaglobulinemia who did not have mutations in Btk. METHODS: A series of candidate genes that encode proteins involved in B-cell signal-transduction pathways were analyzed by linkage studies and mutation screening. RESULTS: Four different mutations were identified in the mu heavy-chain gene on chromosome 14. In one family, there was a homozygous 75-to-100-kb deletion that included D-region genes, J-region genes, and the mu constant-region gene. In a second family, there was a homozygous base-pair substitution in the alternative splice site of the mu heavy-chain gene. This mutation would inhibit production of the membrane form of the mu chain and produce an amino acid substitution in the secreted form. In addition, a patient previously thought to have X-linked agammaglobulinemia was found to have an amino acid substitution on one chromosome at an invariant cysteine that is required for the intrachain disulfide bond and, on the other chromosome, a large deletion that included the immunoglobulin locus. CONCLUSIONS: Defects in the mu heavy-chain gene are a cause of agammaglobulinemia in humans. This implies that an intact membrane-bound mu chain is essential for B-cell development.

Dyskeratosis congenita: unusual onset with isolated neutropenia at an early age.

A 3.5 year old male patient with dyskeratosis congenita (DC) presented at the age of 13 months with isolated neutropenia preceding characteristic skin findings. The average absolute neutrophil count of 500/mm3 persisted without the presence of anemia or thrombocytopenia during the follow up. Neutropenia responded to granulocyte-colony stimulating factor (G-CSF) at a dose of 10 micrograms/kg per day. Immunologic findings were normal as was the chromosomal stability and sister chromatid exchange.

Homozygous C2 deficiency: association with defective alternative pathway function and immunoglobulin deficiency.

Deficiency in the second component of complement (C2) is the most common homozygous complement deficiency. While approximately half of the affected individuals are apparently healthy, C2 deficiency may be associated with autoimmune diseases and rarely increased susceptibility to infection. We report 5 patients who had homozygous type I C2 deficiency in two families. Three of them suffered from frequent infections. These symptomatic patients had additional risk factors; the index cases in the first and the second family had IgG2 deficiency and IgA deficiency, respectively, and alternative complement pathway hemolytic activity was also low in both of them and in the sibling of the first index case. These results emphasize the probable role of other immunologic defects in the clinical presentation of C2 deficiency.

Low expression of T-cell receptor-CD3 complex: a case with a clinical presentation resembling humoral immunodeficiency.

Low expression of T-cell receptor-CD3 (TCR-CD3) complex, a rare cause of combined immunodeficiency, has only recently begun to be recognized. Here we report a four-year-old boy who has defective TCR-CD3 complex expression presenting with recurrent chest infections and pulmonary symptoms implying bronchial asthma. In our opinion, this entity should be borne in mind as the possible underlying defect in children with combined immunodeficiency having signs and symptoms of humoral immunodeficiency.

Selective IgA deficiency with unusual features: development of common variable immunodeficiency, Sjögren's syndrome, autoimmune hemolytic anemia and immune thrombocytopenic purpura.

We report on a girl with selective IgA deficiency and persistently low complement component 4 (C4) levels compatible with heterozygous C4 deficiency. Deterioration of her serum immunoglobulin levels and transition to common variable immunodeficiency were observed within a 5 year follow-up. She also developed Sjögren's syndrome, autoimmune hemolytic anemia and immune thrombocytopenic purpura. While these abnormalities have been described before in various combinations, to our knowledge, they have not been reported in a single individual.