IntroductionIn 2020, the first Dutch West Nile virus (WNV) infected birds were detected through risk-targeted surveillance of songbirds. Retrospective testing of patients with unexplained... Show moreIntroductionIn 2020, the first Dutch West Nile virus (WNV) infected birds were detected through risk-targeted surveillance of songbirds. Retrospective testing of patients with unexplained neurological disease revealed human WNV infections in July and August 2020. Bird ringers are highly exposed to mosquito bites and possibly avian excrements during ringing activities. This study therefore investigates whether bird ringers are at higher risk of exposure to WNV and Usutu virus (USUV).MethodsDutch bird ringers were asked to provide a single serum sample (May – September 2021) and to fill out a survey. Sera were screened by protein microarray for presence of specific IgG against WNV and USUV non-structural protein 1 (NS1), followed by focus reduction virus neutralization tests (FRNT). Healthcare workers (2009–2010), the national immunity cohort (2016–2017) and blood donors (2021) were used as control groups without this occupational exposure.ResultsThe majority of the 157 participating bird ringers was male (132/157, 84%) and the median age was 62 years. Thirty-seven participants (37/157, 23.6%) showed WNV and USUV IgG microarray signals above background, compared to 6.4% (6/94) in the community cohort and 2.1% (2/96) in blood donors (p < 0.01). Two seroreactive bird ringers were confirmed WNV or USUV positive by FRNT. The majority of seroreactive bird ringers travelled to EU countries with reported WNV human cases (30/37, 81%) (p = 0.07). No difference was observed between bird ringers with and without previous yellow fever vaccination.DiscussionThe higher frequency of WNV and/or USUV IgG reactive bird ringers indicates increased flavivirus exposure compared to the general population, suggesting that individuals with high-exposure professions may be considered to complement existing surveillance systems. However, the complexity of serological interpretation in relation to location-specific exposure (including travel), and antibody cross-reactivity, remain a challenge when performing surveillance of emerging flaviviruses in low-prevalence settings. Show less
In this thesis, mathematical modeling and simulation was applied as a tool to inform quantitative decision making in oncology drug discovery and development. Modeling based approaches were shown to... Show moreIn this thesis, mathematical modeling and simulation was applied as a tool to inform quantitative decision making in oncology drug discovery and development. Modeling based approaches were shown to be useful to understand the mechanism of action and deconvolve the complexities of novel biotherapeutic modalities being used to treat cancer, including monospecific and bispecific monoclonal antibodies and antibody drug conjugates. Several key observations and learnings were made. For example, modeling was shown to be a useful method to reduce animal experimentation, by enabling in vitro to in vivo correlations or use of simulation to replace experimental methodologies. Mechanism based modeling and simulation was found to be a useful means to translate from preclinical studies to the clinic to ensure progression of the best drug to clinical trials. These models could then be used to optimize design of clinical studies from selection of starting doses to recommended efficacious doses for pivotal trials. Modeling was shown to be beneficial to understand variability in the clinic and to identify factors impacting drug response in individual patients, paving the way for precision medicine strategies, informing clinical diagnostics, biomarkers, and doses for different oncology indications. Show less
Therapeutic proteins have become very successful in the treatment of various chronic and life-threatening diseases. However, besides their benefits, therapeutic proteins seem to have a common... Show moreTherapeutic proteins have become very successful in the treatment of various chronic and life-threatening diseases. However, besides their benefits, therapeutic proteins seem to have a common problem - the response of a patient’s immune system against the protein. This means that the immune system of the patient actively removes the drug from the body, thereby potentially decreasing or reversing the effect of the therapy. By now there is strong consensus that damaged and aggregated proteins are important risk factors. Protein aggregates are, due to their heterogeneity and often low quantity, challanging to characterize. Further, there is a large academic interest in understanding the mechanisms of aggregation and the role of non-proteinaceous particles in the process of protein aggregation and unwanted immunogenicity in order to design more effective and safe protein-based medicines. This PhD thesis supported that research effort by developing and improving analytical methodologies to detect the size, quantity and other properties of protein aggregates and particles, especially in the relevant nano- and micrometer size range. These techniques were then applied to study a so far unknown nanoparticulate impurity in pharmaceutical-grade sugars. Further, the results shown in this thesis revealed that these nanoparticulate impurities pose a threat to protein stability. Show less
The work presented in this thesis aimed at increasing our understanding of the effect of helminths on Plasmodium spp. immune response in co-infected individuals living in endemic countries. It... Show moreThe work presented in this thesis aimed at increasing our understanding of the effect of helminths on Plasmodium spp. immune response in co-infected individuals living in endemic countries. It presents data from studies conducted in rural and semi urban areas of Lambaréné (Gabon) where the burden of malaria and helminths is particularly important. Although scarce previous studies have indicated an effect of helminths on malaria outcomes and immune response to Plasmodium spp. parasite in co-infected subjects. However it is still debated how consistent is this effect across study sites and teams and what its immunological basis is. Show less
Immunoglobulin G (IgG) represents the most abundant antibody class in the human circulation. IgG consists of two heavy chains and two light chains. Parts of the heavy chains, together with the... Show moreImmunoglobulin G (IgG) represents the most abundant antibody class in the human circulation. IgG consists of two heavy chains and two light chains. Parts of the heavy chains, together with the light chains, form two fragment antigen binding (Fab) moieties, whilst the remainders of the two heavy chains form the fragment crystallizable (Fc) moiety. Human IgGs are glycosylated at the highly conserved N-glycosylation site asparagine 297 in the CH2 domain of each heavy polypeptide chain of the Fc part. Fully galactosylated N-glycans are positioned between the Fc polypeptide chains, resulting in an open Fc conformation which is required for high affinity binding to Fc_ receptors. Small changes in the Fc glycosylation can already have a profound influence on the interaction of the Fc portion with receptors modulating the anti and pro-inflammatory properties of IgG. Mass spectrometry provides great opportunities for deta iled structural characterization of protein glycosylation including protein identification, determination of site-specific glycosylation profiles, and structural characterization of glycans at the level of released glycans and glycopeptides. In this thesis novel approaches for fast, miniaturized and high-throughput analysis of IgG Fc N-glycosylation are presented, and the utility of these methods has been demonstrated for clinically relevant research questions. Show less
A vaccine against Plasmodium falciparum is needed to augment currently available malaria control tools. A handful of parasite antigens are at various stages of pre-clinical and clinical development... Show moreA vaccine against Plasmodium falciparum is needed to augment currently available malaria control tools. A handful of parasite antigens are at various stages of pre-clinical and clinical development. Amongst these is P. falciparum apical membrane antigen 1 (PfAMA1), an antigen expressed by asexual stage parasites and believed to be important for the parasite__s invasion of both red blood cells and liver cells. The immune response to PfAMA1 is mediated mainly by antibodies that prevent parasite invasion of host cells. PfAMA1 however shows allelic polymorphism, with anti-PfAMA1 antibody responses exhibiting strain-specificity. This thesis investigated multi-allele vaccine formulation strategies that would overcome the strain-specificity of antibody responses to PfAMA1. The main findings of this thesis are that i) different PfAMA1 alleles share epitopes to which functional cross-strain antibodies can be induced, ii) a three-allele PfAMA1 formulation yields the greatest proportion of functional cross-strain antibodies, and iii) three PfAMA1 alleles, irrespective of the adjuvant used for formulation and whether they are administrated as a multi-allele formulation or sequentially, induce similar proportions of cross-strain antibodies. Overall, a multi-allele formulation with three in silico-designed PfAMA1 candidates yields antibodies that inhibit several parasites in vitro and warrant their development as a human blood stage vaccine. Show less
In this research heavy chain antibody fragments (VHHs) are developed as potential tools for non-invasive in vivo imaging of Alzheimer__s disease (AD) and Cerebral Amyloid Angiopathy (CAA). First... Show moreIn this research heavy chain antibody fragments (VHHs) are developed as potential tools for non-invasive in vivo imaging of Alzheimer__s disease (AD) and Cerebral Amyloid Angiopathy (CAA). First the generation of antibody fragments, directed against A_ is described. VHHs that were selected from different libraries showed differential affinity for different A_ epitopes when used for immunohistochemistry. These observations indicate that the VHHs are the first immunologic probes with the capacity to differentiate between parenchymal and vascular beta amyloid aggregates. Next, in chapter three the VHH are assessed on their ability to cross the blood-brain barrier using an established in vitro blood brain barrier co-culture system. VHH ni3A showed the highest transmigration efficiency of all tested VHHs. This transport is, in part, facilitated by a 3 amino acid substitution in its N-terminal domain. Additional studies indicated that the mechanism of transcellular passage of ni3A is by active transport. The data described in chapter four provides preliminary proof that these VHHs have the capacity to target A_ depositions in vivo. Consequently, the VHHs are promising tools for further development as imaging agents for the differential diagnosis of A_-related neurodegenerative diseases like CAA and AD. Show less
Major advancements in the understanding of the immune system have provided us with the opportunity for rational design of therapeutic immunological interventions. The notion that dendritic cells ... Show moreMajor advancements in the understanding of the immune system have provided us with the opportunity for rational design of therapeutic immunological interventions. The notion that dendritic cells (DC) play a crucial role in the activation of T lymphocytes has made DC biology of central importance for vaccine development. Accordingly, efficient delivery of antigen to DCs is one of main objectives in vaccine development. In this thesis, antibody-mediated antigen targeting is evaluated as a potential antigen delivery strategy for therapeutic vaccination. Complexes of protein antigen and antigen-specific antibodies are natural formulations that bind to Fc__ receptors. Fc__R ligation on DCs leads to efficient uptake, DC maturation and presentation of the antigen to T lymphocytes. Interaction of Ag-Ab complexes with Fc__Rs on DCs provides a link between the humoral and cellular arms of the immune response. This thesis contains an extensive evaluation of Fc__R-mediated antigen delivery to dendritic cells in the context of T lymphocyte-mediated immunotherapy. In addition, it contains a detailed analysis of Fc__R function on DCs and addresses the kinetics of cross-presentation of antigen after Fc__R-mediated uptake. Show less
Worldwide, more than a billion people are infected with helminths. These worm infections are chronic in nature and can lead to considerable morbidity. Immunologically these infections are... Show moreWorldwide, more than a billion people are infected with helminths. These worm infections are chronic in nature and can lead to considerable morbidity. Immunologically these infections are interesting; chronic helminth infections are characterized by skewing towards a T helper 2 type response as well as regulatory responses. The latter is thought to prevent strong immune responses against parasitic worms, allowing their long-term survival and restricting pathology. This regulatory network is thought to also temper responses to non-helminth antigens, like allergens or self antigens, possibly leading to lower prevalence of allergies and autoimmune diseases in subjects that are chronically infected with helminths. Thus, helminths may bear molecules that have potential therapeutic action against allergies and possibly other inflammatory diseases. However, on the other side of the coin, this would predict that helminth infected subjects might not respond strongly to third party antigens like vaccines. Helminth derived molecules have been identified that induce T helper 2 and regulatory responses via modulation of dendritic cells and some appear to do so via TLR signaling. New insights into these pathways could be useful to antagonize suppression and hence boost vaccine efficacy or to optimize suppression induced by helminth derived molecules and control inflammatory diseases. Show less
Huidig beleid schrijft voor dat voor de bloedtransfusie plaatsvindt, het serum van de patient getest wordt op irreguliere anti-stoffen van rode bloedcellen. In dit boek onderzoekt Henk Schonewille... Show moreHuidig beleid schrijft voor dat voor de bloedtransfusie plaatsvindt, het serum van de patient getest wordt op irreguliere anti-stoffen van rode bloedcellen. In dit boek onderzoekt Henk Schonewille of dit beleid moet veranderen om bloedtransfusie veiliger te maken. Current pre-transfusion policy requires the patient's serum to be tested for the presence of irregular red blood cell antibodies. In case of an alloantibody, red blood cells lacking the corresponding antigen are transfused after an antoglobulin crossmatch. The aim of the study in this thesis is primarily to investigate whether this policy should change to improve transfusion safety Show less
