Vojtech Thon

Clinical picture and treatment of 2212 patients with common variable immunodeficiency

BACKGROUND Common variable immunodeficiency (CVID) is an antibody deficiency with an equal sex distribution and a high variability in clinical presentation. The main features include respiratory tract infections and their associated complications, enteropathy, autoimmunity, and lymphoproliferative disorders. OBJECTIVE This study analyzes the clinical presentation, association between clinical features, and differences and effects of immunoglobulin treatment in Europe. METHODS Data on 2212 patients with CVID from 28 medical centers contributing to the European Society for Immunodeficiencies Database were analyzed retrospectively. RESULTS Early disease onset (<10 years) was very frequent in our cohort (33.7%), especially in male subjects (39.8%). Male subjects with early-onset CVID were more prone to pneumonia and less prone to other complications suggesting a distinct disease entity. The diagnostic delay of CVID ranges between 4 and 5 years in many countries and is particularly high in subjects with early-onset CVID. Enteropathy, autoimmunity, granulomas, and splenomegaly formed a set of interrelated features, whereas bronchiectasis was not associated with any other clinical feature. Patient survival in this cohort was associated with age at onset and age at diagnosis only. There were different treatment strategies in Europe, with considerable differences in immunoglobulin dosing, ranging from 130 up to 750 mg/kg/mo. Patients with very low trough levels of less than 4 g/L had poor clinical outcomes, whereas higher trough levels were associated with a reduced frequency of serious bacterial infections. CONCLUSION Patients with CVID are being managed differently throughout Europe, affecting various outcome measures. Clinically, CVID is a truly variable antibody deficiency syndrome.

Soluble BAFF Levels Inversely Correlate with Peripheral B Cell Numbers and the Expression of BAFF Receptors

The TNF family member protein BAFF/BLyS is essential for B cell survival and plays an important role in regulating class switch recombination as well as in the selection of autoreactive B cells. In humans, increased concentrations of soluble BAFF are found in different pathological conditions, which may be as diverse as autoimmune diseases, B cell malignancies, and primary Ab deficiencies (PAD). Because the mechanisms that regulate BAFF levels are not well understood, we newly developed a set of mAbs against human BAFF to study the parameters that determine the concentrations of soluble BAFF in circulation. Patients with PAD, including severe functional B cell defects such as BTK, BAFF-R, or TACI deficiency, were found to have higher BAFF levels than asplenic individuals, patients after anti-CD20 B cell depletion, chronic lymphocytic leukemia patients, or healthy donors. In a comparable manner, mice constitutively expressing human BAFF were found to have higher concentrations of BAFF in the absence than in the presence of B cells. Therefore, our data strongly suggest that BAFF steady-state concentrations mainly depend on the number of B cells as well as on the expression of BAFF-binding receptors. Because most patients with PAD have high levels of circulating BAFF, the increase in BAFF concentrations cannot compensate defects in B cell development and function.

Age dependency and mutual relations in T and B lymphocyte abnormalities in common variable immunodeficiency patients

Common variable immunodeficiency (CVID) is primary hypogammaglobulinaemia with an unknown aetiopathogenesis. Although various abnormalities of T and B cells have been described, their pathogenetic roles are unclear. We determined T and B lymphocyte subsets known to be abnormal in CVID in order to disclose possible relations between numerical abnormalities in those cells. Markers associated with B cell development (CD21, CD27, IgM, IgD) were determined on B lymphocytes (CD19+); T lymphocyte development (CD45RA, CD45RO, CD62L) and activation markers (CD25, CD27, CD28, CD29, CD38, CD57, HLA‐DR) were determined on CD4+ and CD8+ T lymphocytes in 42 CVID patients and in 33 healthy controls. Abnormalities in CD4+ T lymphocyte activation markers (increase in CD29, HLA‐DR, CD45RO, decrease in CD27, CD62L, CD45RA) were observed particularly in patients with a decreased number of memory (CD27+) and mature (CD21+) B cells (group Ia according to the Freiburg groups classification), while abnormalities observed in CD8+ cells (increase in CD27 and CD28 and decrease in HLA‐DR, CD57 and CD38) did not depend upon grouping patients together according to B lymphocyte developmental subpopulations. We observed correlations between immature B cells (IgM+ CD21–) and expression of CD27, CD62L, CD45RA, CD45RO and HLA‐DR on CD4+ T cells in CVID patients but not in the control group. The expression of CD27 and CD45RA on CD4+ T lymphocytes, such as the percentage of IgD+ CD27– and IgD+ CD27+ cells in B lymphocytes, showed age dependency to be more significant than in the control group. Our study demonstrates that T and B lymphocyte abnormalities in CVID are partially related to each other. Some of those abnormalities are not definite, but may evolve with age of the patient.

Double mutant and formaldehyde inactivated TSST-1 as vaccine candidates for TSST-1-induced toxic shock syndrome

Up to now there is no treatment for staphylococcal toxic shock syndrome, a disease mainly induced by toxic shock syndrome toxin-1(TSST-1). There is great demand in finding means to control the disease, one of them is the development of an effective and safe vaccine against TSST-1. In this study we constructed a series of vaccine candidates and investigated their biological activity, toxicity, and potential to invoke an immune response. TSST-1 was isolated from Stahylococcus aureus supernatants and recombinantly expressed as a N-terminal 6x histidine-tagged protein in Escherichia coli. In order to obtain molecules with minimal toxicity we constructed single mutants (G31R and H135A) and one double mutant (G31R/H135A) with both residues exchanged. We also detoxified native TSST-1 isolated from S. aureus, and recombinantly expressed TSST-1 by treatment with formaldehyde. Functional activity of native and recombinant TSST-1 and grade of inocuity of mutants and toxoids was determined by investigating mitogenity, T-cell activation, and cytokine release upon stimulation of human mononuclear cells with the vaccine candidates. All substances were tested in a rabbit immunization study. After primary immunization and three additional boosts all vaccinated animals developed antibody titers against TSST-1 and were protected against challenge with a lethal doses of superantigen potentiated with lipopolysaccharide.

Antibody forming cells and plasmablasts in peripheral blood in CVID patients after vaccination.

Common variable immunodeficiency (CVID), the most frequent primary antibody disorder, is characterized by hypogammaglobulinaemia and impaired antibody production. Poor vaccination response is essential for the diagnosis of CVID. Their under laying defects remain to be elucidated. Routine determination of antibody production in serum from CVID patients after vaccination and investigation of B cell function in vivo is complicated due to substitution therapy. Therefore we investigated antibody production on the B-cell level by ELISPOT and characterized changes in B-cell subpopulations in CVID patients, including plasmablasts, in peripheral blood by flow cytometry after vaccination for specification of the diagnosis. Thirty-seven CVID patients and eighty healthy volunteers were immunized with tetanus toxoid and pneumococcal polysaccharide vaccines. Specific antibody levels and B cell subpopulations were measured before vaccination and on day 7 after vaccination by ELISPOT assay and flow cytometry respectively. Of the thirty-seven well defined CVID patients studied, thirty lacked detectable spot forming cells producing specific IgG, IgA or IgM antibodies against employed vaccines and seven had only weak responses compared to controls. In the control group, an increase in circulating plasmablasts on day 7 post immunization corresponded with the appearance of antibody forming cells. In contrast, CVID patients failed to increase plasmablasts significantly in peripheral blood after antigen challenge. Our findings indicate that CVID patients have a block in terminal B-cell differentiation and that flow based assessment of plasmablasts in peripheral blood after vaccination serves as a surrogate diagnostic marker for assessing in vivo antibody responses in patients suspected to have CVID.

Interleukin-6 May Contribute to Mortality in Parkinson’s Disease Patients: A 4-Year Prospective Study

Objectives. The association between abnormal serum immunomarkers and mortality in 53 consecutive Parkinsons disease patients was studied. Materials and Methods. The plasma level of specific inflammatory cytokines was investigated: mannan-binding lectin (MBL), interleukin- (IL-) 6, and tumor necrosis factor-alpha (TNF-α). The baseline serum immunomarkers obtained from patients who died (n = 16) during a four-year follow-up period were compared with the data of patients who survived (n = 37). Results. The baseline level of IL-6 was significantly higher in the deceased patients than in the survivors. Elevated IL-6 levels and age were major independent contributors to disease mortality. Differences between other plasma cytokine level abnormalities were not significant. Conclusion. This study showed that IL-6 elevation may be a marker of increased mortality risk in Parkinsons disease patients. The inflammation may act in association with other factors and comorbidities in progressive neurodegenerative pathology.

Common variants at PVT1 , ATG13 – AMBRA1 , AHI1 and CLEC16A are associated with selective IgA deficiency

Selective immunoglobulin A deficiency (IgAD) is the most common primary immunodeficiency in Europeans. Our genome-wide association study (GWAS) meta-analysis of 1,635 patients with IgAD and 4,852 controls identified four new significant (P < 5 × 10−8) loci and association with a rare IFIH1 variant (p.Ile923Val). Peak new variants (PVT1, P = 4.3 × 10−11; ATG13–AMBRA1, P = 6.7 × 10−10; AHI1, P = 8.4 × 10−10; CLEC16A, P = 1.4 × 10−9) overlapped with autoimmune markers (3/4) and correlated with 21 putative regulatory variants, including expression quantitative trait loci (eQTLs) for AHI1 and DEXI and DNase hypersensitivity sites in FOXP3+ regulatory T cells. Pathway analysis of the meta-analysis results showed striking association with the KEGG pathway for IgA production (pathway P < 0.0001), with 22 of the 30 annotated pathway genes containing at least one variant with P ≤ 0.05 in the IgAD meta-analysis. These data suggest that a complex network of genetic effects, including genes known to influence the biology of IgA production, contributes to IgAD.

Detection of Impaired IgG Antibody Formation Facilitates the Decision on Early Immunoglobulin Replacement in Hypogammaglobulinemic Patients

Hypogammaglobulinemia (serum IgG lower than 2 SD below the age-matched mean) and clinical symptoms such as increased susceptibility to infection, autoimmune manifestations, granulomatous disease, and unexplained polyclonal lymphoproliferation are considered to be diagnostic hallmarks in patients with common variable immunodeficiency (CVID), the most frequent clinically severe primary immunodeficiency syndrome. In the present study, we investigated patients with hypogammaglobulinemia and no clinical or immunological signs of defective cell-mediated immunity and differentiated two groups on the basis of their IgG antibody formation capacity against a variety of different antigens (bacterial toxins, polysaccharide antigens, viral antigens). Patients with hypogammaglobulinemia and intact antibody production (HIAP) displayed no or only mild susceptibility to infections, while CVID patients showed marked susceptibility to bacterial infections that normalized following initiation of IVIG or subcutaneous immunoglobulin replacement therapy. There was a substantial overlap in IgG serum levels between the asymptomatic HIAP group and the CVID patients examined before immunoglobulin treatment. HIAP patients showed normal levels of switched B-memory cells (CD19+CD27+IgD−), while both decreased and normal levels of switched B-memory cells could be found in CVID patients. IgG antibody response to a primary antigen, tick-borne encephalitis virus (TBEV), was defective in CVID patients, thus confirming their substantial defect in IgG antibody production. Defective IgG antibody production against multiple antigens could also be demonstrated in an adult patient with recurrent infections but normal IgG levels. To facilitate early treatment before recurrent infections may lead to organ damage, the antibody formation capacity should be examined in hypogammaglobulinemic patients and the decision to treat should be based on the finding of impaired IgG antibody production.

Change in referral diagnoses and diagnostic delay in hypogammaglobulinaemic patients during 28 years in a single referral centre.

Background: The classical clinical manifestation of untreated immunoglobulin deficiency comprises predominantly recurrent and complicated respiratory tract infections. Before the 1980s, little was known about the clinical manifestation of immunodeficiency in the general medical population, and also the availability of serum immunoglobulin laboratory determination was not sufficient, leading to a significant diagnostic delay. Methods: We have analysed the diagnostic delay and referral diagnoses in patients in whom any form of primary hypogammaglobulinaemia had been diagnosed at our department, which was established in 1981. Results: Comparing the diagnostic delay in the 1980s (19 patients, median 5.5 years), the 1990s (37 patients, median 3.5 years) and the years 2001–2008 (33 patients, median 1 year), a significant decrease was observed (p < 0.05, Spearman’s correlation coefficient). Also, the median number of pneumonia episodes during the diagnostic delay decreased from 5 in the 1980s, to 1 in the 1990s and to 0 in the period of 2001–2008 (p < 0.05, Spearman’s correlation coefficient). While in the 1980s 17 of the 19 patients had pneumonia in their past history, in the period of 2001–2008 only 13 of the 33 patients were concerned. Conclusions: Our observation documents improved awareness of immunodeficiencies among physicians. It is supposed that earlier diagnosis will prevent complications, improve the quality of life and even survival of hypogammaglobulinaemic patients.

Anti-Gal IgM, IgA and IgG natural antibodies in childhood.

The target for the most abundant xenoreactive natural antibodies in humans is the α-Gal epitope. Anti-Gal could provide natural immune defense against pathogens that express the α-Gal epitope. Anti-Gal natural antibodies are usually studied in adult individuals. Data demonstrating the incidence and concentration of anti-Gal natural antibodies in childhood are in short supply and incomplete. In the present study we prospectively quantified anti-Gal IgM, IgA and IgG levels in different age groups of children from delivery to 24 months of age and compared these levels to the level of these antibodies in their respective mothers. Measurement of anti-Gal antibodies may broaden the spectrum of specific antibodies that are available for determination of specific antibody responses in physiological and pathological conditions in children. Plasma was collected from umbilical cord blood of full term newborn, from blood of infants at age 6, 12 and 24 months and from their respective mothers at time of delivery. Quantitative determination of anti-Gal antibodies IgM, IgA and IgG were made with the enzyme immunoassays Human Anti-Alpha Galactosyl IgM ELISA, IgG ELISA and IgA ELISA. Hemagglutination activity was titrated against rabbit erythrocytes. The kinetic processes for the formation of natural antibodies in the first two years of life, in general, compared with the kinetics for the formation of total immunoglobulins IgM, IgA and IgG. There were no detectable anti-Gal IgM and IgA in the cord blood, whereas anti-Gal IgG were found at similar levels in both neonate cord blood and peripheral blood of their respective mothers. When comparing the percentage of natural antibodies in the plasma of children, the level of natural antibodies in children at the age of two years was approximately 37% for IgM, 25% for IgG and 15% for IgA. The titration of antibodies required for agglutination of rabbit red blood cells over the 24 month period followed the same trend observed for the formation of natural antibodies. Our study demonstrates the kinetics of formation of anti-Gal IgM, IgA and IgG natural antibodies in the first two years of life. The relative lack of these antibodies in this period should be taken into account when assessing for humoral immunodeficiencies, particularly with regards to the potential for children to mount an anti-carbohydrate response.