Uwe Kölsch

Hypomorphic homozygous mutations in phosphoglucomutase 3 (PGM3) impair immunity and increase serum IgE levels

BACKGROUND Recurrent bacterial and fungal infections, eczema, and increased serum IgE levels characterize patients with the hyper-IgE syndrome (HIES). Known genetic causes for HIES are mutations in signal transducer and activator of transcription 3 (STAT3) and dedicator of cytokinesis 8 (DOCK8), which are involved in signal transduction pathways. However, glycosylation defects have not been described in patients with HIES. One crucial enzyme in the glycosylation pathway is phosphoglucomutase 3 (PGM3), which catalyzes a key step in the synthesis of uridine diphosphate N-acetylglucosamine, which is required for the biosynthesis of N-glycans. OBJECTIVE We sought to elucidate the genetic cause in patients with HIES who do not carry mutations in STAT3 or DOCK8. METHODS After establishing a linkage interval by means of SNPchip genotyping and homozygosity mapping in 2 families with HIES from Tunisia, mutational analysis was performed with selector-based, high-throughput sequencing. Protein expression was analyzed by means of Western blotting, and glycosylation was profiled by using mass spectrometry. RESULTS Mutational analysis of candidate genes in an 11.9-Mb linkage region on chromosome 6 shared by 2 multiplex families identified 2 homozygous mutations in PGM3 that segregated with disease status and followed recessive inheritance. The mutations predict amino acid changes in PGM3 (p.Glu340del and p.Leu83Ser). A third homozygous mutation (p.Asp502Tyr) and the p.Leu83Ser variant were identified in 2 other affected families, respectively. These hypomorphic mutations have an effect on the biosynthetic reactions involving uridine diphosphate N-acetylglucosamine. Glycomic analysis revealed an aberrant glycosylation pattern in leukocytes demonstrated by a reduced level of tri-antennary and tetra-antennary N-glycans. T-cell proliferation and differentiation were impaired in patients. Most patients had developmental delay, and many had psychomotor retardation. CONCLUSION Impairment of PGM3 function leads to a novel primary (inborn) error of development and immunity because biallelic hypomorphic mutations are associated with impaired glycosylation and a hyper-IgE-like phenotype.

Combined immunodeficiency develops with age in Immunodeficiency-centromeric instability-facial anomalies syndrome 2 (ICF2)

The autosomal recessive immunodeficiency-centromeric instability-facial anomalies syndrome (ICF) is characterized by immunodeficiency, developmental delay, and facial anomalies. ICF2, caused by biallelic ZBTB24 gene mutations, is acknowledged primarily as an isolated B-cell defect. Here, we extend the phenotype spectrum by describing, in particular, for the first time the development of a combined immune defect throughout the disease course as well as putative autoimmune phenomena such as granulomatous hepatitis and nephritis. We also demonstrate impaired cell-proliferation and increased cell death of immune and non-immune cells as well as data suggesting a chromosome separation defect in addition to the known chromosome condensation defect.

Even in pneumococcal sepsis CD62L shedding on granulocytes proves to be a reliable functional test for the diagnosis of interleukin-1 receptor-associated kinase-4 deficiency.

A 9-month-old infant presented with fatal pneumococcal sepsis and attenuated inflammation indices. Even in septic conditions, flow cytometry–based CD62L shedding test on granulocytes proved to be a fast and reliable diagnostic tool for the detection of a defect in the innate immunity. Confirmatory immunologic and genetic assays identified an autosomal-recessive interleukin-1 receptor–associated kinase-4 deficiency due to compound heterozygous mutations.

Liver abscess complicated by diaphragm perforation and pleural empyema leads to the discovery of interleukin-1 receptor-associated kinase 4 deficiency.

Interleukin-1 receptor-associated kinase 4 (IRAK-4) deficiency predisposes to severe invasive bacterial infections in infancy and early childhood, often with a fatal course caused by a defect in Toll-like receptor and interleukin-1 receptor signaling. Despite severe invasive infections, acute phase responses are often diminished. We report the successful treatment of a child with multiple liver abscesses, diaphragm perforation and pleural empyema, accompanied by strong acute phase responses as a unique presentation of IRAK-4 deficiency.

Septic arthritis or juvenile idiopathic arthritis ‐ the case of a 2 year old boy

Peanut epitopes for IgE and IgG4 in peanutsensitized children in relation to severity of peanut allergy. J Allergy Clin Immunol 2008: 121 : 737.e10–43.e10. 3. Hong X, Caruso D, Kumar R, et al. IgE, but not IgG4, antibodies to Ara h 2 distinguish peanut allergy from asymptomatic peanut sensitization. Allergy 2012: 67: 1538–46. 4. Stapel SO, Asero R, Ballmer-Weber BK, et al. Testing for IgG4 against foods is not recommended as a diagnostic tool: EAACI Task Force Report. Allergy 2008: 63: 793–6. 5. Sverremark-Ekstrom E, Hultgren EH, Borres MP, Nilsson C. Peanut sensitization during the first 5 yr of life is associated with elevated levels of peanut-specific IgG. Pediatr Allergy Immunol 2012: 23: 224–9. 6. Caubet JC, Bencharitiwong R,Moshier E, Godbold JH, Sampson HA, Nowak-Wegrzyn A. Significance of ovomucoidand ovalbuminspecific IgE/IgG(4) ratios in egg allergy. J Allergy Clin Immunol 2012: 129: 739–47. 7. Astier C, Morisset M, Roitel O, et al. Predictive value of skin prick tests using recombinant allergens for diagnosis of peanut allergy. J Allergy Clin Immunol 2006: 118: 250–6. 8. Tay SS, Clark AT, Deighton J, King Y, Ewan PW. Patterns of immunoglobulin G responses to egg and peanut allergens are distinct: ovalbumin-specific immunoglobulin responses are ubiquitous, but peanut-specific immunoglobulin responses are up-regulated in peanut allergy. Clin Exp Allergy 2007: 37: 1512–8. 9. Uermosi C, Beerli RR, Bauer M, et al. Mechanisms of allergen-specific desensitization. J Allergy Clin Immunol 2010: 126: 375–83. 10. Aalberse RC, Stapel SO, Schuurman J, Rispens T. Immunoglobulin G4: an odd antibody. Clin Exp Allergy 2009: 39: 469–77.

DC generation from peripheral blood mononuclear cells in patients with chronic myeloid leukemia: Influence of interferons on DC yield and functional properties.

ABSTRACT In Chronic Myeloid Leukemia (CML), standard treatment consists of modern tyrosine-kinase inhibitors (TKI). Nevertheless, there is evidence that immune responses against leukemia-associated antigens (LAA) may play an important role in disease control. Dendritic cell (DC)- based immunotherapy is able to induce T cell responses against LAA and might therefore pose an interesting therapeutic option in CML, especially in the setting of minimal residual disease (MRD). GMP production of DC for clinical vaccination remains a time- and cost- intensive procedure and standardized DC generation is warranted. We asked whether maturation-induction with IFN-γ and IFN-α has an influence on functional properties of DC derived from peripheral blood mononuclear cells (PBMC) in CML patients. Monocyte-derived DC from healthy donors and from patients with CML were analyzed after maturation-induction with our TNF-α-containing standard cytokine cocktail with or without addition of IFN-α and/or IFN-γ. Our results confirm that the addition of IFN-γ leads to enhanced IL-12 secretion in healthy donors. In contrast, in CML patients, IFN-γ was not able to increase IL-12 secretion, possibly due to a higher degree of cell adherence and lower cell yield during the cell culture. Our data suggest, that- in contrast to healthy donors-, additional interferons are not beneficial for maturation induction during large-scale DC production in patients with CML.

Severe macrophage activation syndrome. Is there a causative role for a homozygous A91V mutation in the perforin gene

A 16 year old lebanese girl with consanguinous parents presented with a severe “abdominal” sepsis supposedly resulting from an infected vaginal tampon (ESBL E.coli). She had been healthy before. She developed severe hepatic functional disorder, infarction of the spleen, cardiovascular and renal insufficiency, as well as anemia and thrombocytopenia. Macrophage activation syndrome was diagnosed subsequently and systemic glucocorticoid treatment initiated. The girl recovered clinically. Of note, she developed severe cushingoid syndrome. Due to limited compliance she discontinued all anti-inflammatory medication (nsaids, gc and ciclosporin) 4 weeks later. The following 18 months inflammatory parameters were persistantly elevated (ESR>100mm/h). Familial mediteranean fever was excluded. Genetic analysis revealed a homozygous perforin I gene mutation 91 (GCG) -> Valin (GTG)/.pAla91Val-/A91V in exon 2. Familial hemophagocytic lymphohistiocytosis type II was discussed as a potential diagnosis. Perforin expression was diminshed to about 50% in NK-cells, however functional NK-cell cytotoxicity was in the lower normal range, considered not impaired. On the basis of these findings, we want to discuss the role of the homozygous A91V perforin mutation for the initiation or perpetuation of a life-threatening macrophage activation syndrome, in addition to the further management of the patient.