Greet Wuyts

Addressing diagnostic challenges in primary immunodeficiencies: laboratory evaluation of Toll-like receptor- and NF-κB-mediated immune responses.

Abstract Toll-like receptors (TLRs) play an important role in immunity and mediate their actions via multiple signaling pathways, in particular, the nuclear factor of kappa light polypeptide gene enhancer in B-cells (NF-κB) pathway. Rare inherited defects of TLR- and NF-κB-dependent responses have recently been recognized. These primary immunodeficiencies predispose children to life-threatening infections and often remain undiagnosed. Establishing a sensitive, specific, cost-effective and simple method for diagnosis is therefore important. In this article, we review the known defects of TLR- and NF-κB-mediated pathways and the assays that can be used to screen for such defects.

PID in Disguise: Molecular Diagnosis of IRAK-4 Deficiency in an Adult Previously Misdiagnosed With Autosomal Dominant Hyper IgE Syndrome

Autosomal recessive IL-1R-associated kinase 4 (IRAK-4) deficiency is a rare cause of recurrent pyogenic infections with limited inflammatory responses. We describe an adult female patient with severe lung disease who was phenotypically diagnosed as suffering from autosomal dominant Hyper IgE syndrome (AD HIES) because of recurrent skin infections with Staphylococcus aureus, recurrent pneumonia and elevated serum IgE levels. In contrast to findings in AD HIES patients, no abnormalities were found in the Th17 and circulating follicular helper T cell subsets. A panel-based sequencing approach led to the identification of a homozygous IRAK4 stop mutation (c.877C > T, p.Gln293*).

Effect of previous vaccination with pneumococcal conjugate vaccine on pneumococcal polysaccharide vaccine antibody responses

During the past 10 years, pneumococcal conjugate vaccine (PCV) has become part of the standard childhood vaccination programme. This may impact upon the diagnosis of polysaccharide antibody deficiency by measurement of anti‐polysaccharide immunoglobulin (Ig)G after immunization with unconjugated pneumococcal polysaccharide vaccine (PPV). Indeed, contrary to PPV, PCV induces a T‐dependent, more pronounced memory response. The antibody response to PPV was studied retrospectively in patients referred for suspected humoral immunodeficiency. The study population was divided into four subgroups based on age (2–5 years versus ≥ 10 years) and time tested (1998–2005 versus 2010–12). Only 2–5‐year‐old children tested in 2010–12 had been vaccinated with PCV prior to PPV. The PCV primed group showed higher antibody responses for PCV–PPV shared serotypes 4 and 18C than the unprimed groups. To a lesser extent, this was also found for non‐PCV serotype 9N, but not for non‐PCV serotypes 19A and 8. Furthermore, PCV‐priming elicited a higher IgG2 response. In conclusion, previous PCV vaccination affects antibody response to PPV for shared serotypes, but can also influence antibody response to some non‐PCV serotypes (9N). With increasing number of serotypes included in PCV, the diagnostic assessment for polysaccharide antibody deficiency requires careful selection of serotypes that are not influenced by prior PCV (e.g. serotype 8). Further research is needed to identify more serotypes that are not influenced.

Distinct approaches to investigate the importance of the murine 4-1BB–4-1BBL interaction in the antibody response to Streptococcus pneumoniae

Protection against infection with Streptococcus pneumoniae is based mainly on the generation of antibodies to the pneumococcal capsular polysaccharides (caps‐PS). Although caps‐PS are considered thymus‐independent antigens, there is a growing body of evidence that T lymphocytes and costimulatory molecules are involved in the regulation of the antibody response to caps‐PS. We investigated whether the interaction between 4‐1BB and 4‐1BB ligand (4‐1BBL) is involved in the modulation of the antibody response to caps‐PS after immunization with Pneumovax® or with intact heat‐killed S. pneumoniae. Treatment with agonistic anti‐4‐1BB mAb, which mimics engagement of 4‐1BB by 4‐1BBL, had no effect on the IgG and IgM immune response to caps‐PS (Serotype 3) after immunization with Pneumovax or with S. pneumoniae Serotype 3. However, anti‐4‐1BB treatment strongly inhibited the IgG response to pneumococcal surface protein A (PspA). By contrast, the IgG anti‐caps‐PS (Serotype 3) antibody response was reduced strongly in 4‐1BBL−/− mice immunized with S. pneumoniae Serotype 3. The IgG anti‐PspA antibody response in the 4‐1BB−/− mice was comparable with the immune response in the wild‐type mice. We conclude that distinct pathways are involved in the humoral antibody response to pneumococcal antigens, depending on the nature of the antigen and the context in which the different antigens are presented. The 4‐1BB–4‐1BBL interaction is not involved in the antibody response to soluble caps‐PS. The influence of the 4‐1BB–4‐1BBL interaction in the immune reaction to S. pneumoniae Serotype 3 depends on the experimental system used.

Anti-Pneumococcal Capsular Polysaccharide Antibody Response and CD5 B Lymphocyte Subsets

ABSTRACT The role of CD19+ CD5+ and CD19+ CD5− B cell subpopulations in the antibody response to pneumococcal capsular polysaccharides (caps-PSs) is controversial. In the present study, we evaluated the role of human CD19+ CD5+ and CD19+ CD5− cell populations in the serotype-specific antibody response to caps-PS. After vaccination of 5 healthy human adults with Pneumovax (23-valent pneumococcal polysaccharide vaccine [PPV23]), IgG anti-caps-PS serotype 4 antibody-producing cells resided mainly in the CD19+ CD5− B cell subset, as assessed by enzyme-linked immunosorbent spot (ELISpot) analysis. Moreover, in a humanized SCID mouse model, CD19+ CD5− B cells were more effective than CD19+ CD5+ cells in producing IgG anti-cap-PS antibodies. Finally, an association was found between the level of IgG anti-caps-PS antibodies and the number of CD19+ CD5− B cells in 33 humans vaccinated with PPV23. Taken together, our data suggest that CD5 defines a functionally distinct population of B cells in humans in the anti-caps-PS immune response.

Identification of SP1683 as a pneumococcal protein that is protective against nasopharyngeal colonization

ABSTRACT Serotype-independent protein-based pneumococcal vaccines represent attractive alternatives to capsular polysaccharide-based vaccines. The aim of this study was to identify novel immunogenic proteins from Streptococcus pneumoniae that may be used in protein-based pneumococcal vaccine. An immunoproteomics approach and a humanized severe combined immunodeficient mouse model were used to identify S. pneumoniae proteins that are immunogenic for the human immune system. Among the several proteins identified, SP1683 was selected, recombinantly produced, and infection and colonization murine models were used to evaluate the capacity of SP1683 to elicit protective responses, in comparison to known pneumococcal immunogenic proteins (PhtD and detoxified pneumolysin, dPly). Immunisation with SP1683 elicited a weaker antibody response than immunisation with PhtD and did not provide protection in the model of invasive disease. However, similar to PhtD, it was able to significantly reduce colonization in the mouse model of nasopharyngeal carriage. Treatment with anti-IL17A and anti-IL17F antibodies abolished the protection against colonization elicited by SP1683 or PhtD + dPly, which indicated that the protection afforded in this model was Th17-dependent. In conclusion, intranasal immunization with the pneumococcal protein SP1683 conferred IL17-dependent protection against nasopharyngeal carriage in mice, but systemic immunization did not protect against invasive disease. These results do not support the use of SP1683 as an isolated pneumococcal vaccine antigen. Nevertheless, SP1683 could be used as a first line of defence in formulations combining several proteins.

Clinical characteristics of patients with low functional IL-6 production upon TLR/IL-1R stimulation.

Please cite this article as: Frans G, Van der Werff Ten Bosch J, Moens L, Wuyts G, Schaballie H, Tuerlinckx D, De Bie M, Vermeulen F, Schrijvers R, Meert W, Hestand MS, Delanghe J, Vermeesch Ir JR, Meyts I, Bossuyt X, Clinical characteristics of patients with low functional IL-6 production upon TLR/ IL-1R stimulation, Journal of Allergy and Clinical Immunology (2017), doi: 10.1016/j.jaci.2017.04.017.

Different Immunological Pathways Underlie the Immune Response to Pneumococcal Polysaccharides

Measurement of the antibody response to Pneumovax®/ Pneumo23®, a 23-valent pneumococcal polysaccharide vaccine, is an established method to evaluate B cell function. Such evaluation typically involves measurement of preand post-vaccination serotype-specific antibody levels. It is generally believed that the immune response to pneumococcal polysaccharides is MHC class II-independent. This is based on the observation that various polysaccharides (including pneumococcal polysaccharides) do not bind to MHC class II molecules (explaining the Tcell independence of polysaccharide antigens) and that MHC class II-deficient mice mount antibodies to the T cell-independent antigens dextran, phosphorylcholine, and levan [1, 2]. However, emerging evidence suggests that bacterial zwitterionic polysaccharides (which carry positively and negatively charges within one unit) are processed to low molecular weight carbohydrates and presented to T cells through the MHC class II endocytic pathway [3]. Examples of zwitterionic polysaccharides include polysaccharide A from Bacteroides fragilis and Streptococcus pneumoniae serotype 1 (most other pneumococcal polysaccharides are anionic) [3]. Although it is well recognized that the immune response to pneumococcal polysaccharides is complex and serotype dependent, little attention has been given to the difference between major histocompatibility complex class II (MHC class II)-dependent and MHC class II-independent pathways. In this letter, we illustrate that the immune response to the zwitterionic pneumococcal serotype 1 differs from the immune response to a non-zwitterionic (e.g., anionic) pneumococcal serotype (serotype 3 or serotype 4). In a first step, we immunized MHC class II knockout mice and control mice with Pneumo23® and measured the antibody response to serotype 1, serotype 3, and serotype 4. We found thatMHC class II knockout micemounted antibodies to serotype 3 but failed to do so (p < 0.0001) for serotype 1 (Fig. 1a), thereby confirming that the immune response to the zwitterionic serotype 1 is MHC class II-dependent. Neither wild type nor MHC class II knockout mice mounted detectable antibodies to serotype 4. Next, we evaluated the immune response to Pneumo23® in an 18-year-old MHC class II-deficient patient and in three controls. This patient has homozygous RFXANK deletion [4] and developed chronic bronchiectasis. He receives intravenous immunoglobulin substitution and prophylactic cotrimoxazole (P35 in [4]). Because the patient received immunoglobulin substitution, the immune response was evaluated by quantification of the antibody secreting cells (by ELISPOT) 7 days after vaccination. Antibodies to serotype 1 and serotype 4 were quantified (serotype 3 could not be reliably coated on the ELISPOT plates). In the MHC class II* Xavier Bossuyt xavier.bossuyt@uzleuven.be