Interdisciplinary research

A collaborative effort between The Biomolecular Interaction Facility and the Nanobody Facility

This project aims to identify broadly-reactive/protective nanobodies that specifically bind to rare conserved epitopes on influenza A viruses or coronaviruses. Broadly protective nanobodies developed against these viruses can be used for diagnosis and therapy, but also for the design for novel broadly-protective vaccine antigens. This is important as influenza A viruses and coronaviruses are antigenically diverse, while there is a constant threat of the emergence of novel viruses from animal reservoirs as was dramatically demonstrated by SARS-coronavirus-2.

From June 2020, this project is continuing with collaborative efforts between the labs of  Sabrina Oliviera (Cell Biology Group, Dept. of Biology) and UMI Hub members Paul van Bergen en Henegouwen (Nanobody Facility) en Xander de Haan (The Biomolecular Interaction Facility) for the selection of nanobodies against different influenza A viruses and coronaviruses.

A collaborative effort between  The Crystal and Structural Chemistry Facility  and the Nanobody Facility

We study the molecular interactions that are the root of the complement system that plays a role in the defence against microbes and clearance of apoptotic and necrotic cells. To prevent unwanted complement activation host cells protect themselves with a range of regulators of complement activation (RCA’s), such as MCP/CD46, DAF/CD55 and CD59 that promote cleavage (MCP) or accelerated decay (DAF) of the C3 convertases that opsonise the targeted surfaces or prevent the assembly (CD59) of the terminal MAC complex. Complement receptors 1 and 2 (CR1/CD35 and CR2/CD21) are members of the RCA family that are expressed on nucleated blood cells and erythrocytes (CR1). CR1 is the main receptor for the processing and clearance of opsonised particles and also acts as an inhibitor of complement. CR2 is a receptor for the C3b cleavage-products iC3b, C3d and C3dg and forms a complex with CD19 and CD81; the B-cell co-receptor complex. CR2 plays a role in B-cell activation and maturation and enhances B-cell response to antigens.
Complement plays a dual role in cancer biology; Limited complement activation has been suggested to promote cell proliferation and neovascularization and may have a tumor-promoting effect, whereas unbridled induction of complement results in tumorcell-lysis. Anti-tumor immune therapies benefit from complement activation both directly, through initiation of complement dependent cytotoxicity (CDC), and indirectly, by enhancement of antibody-dependent cellular toxicity  via CR2. Most cancers over-express several complement components, including RCA’s and RCA expression is correlated with CDC resistance and poor prognosis.
It has been shown that complement activation in combination with either immune- or chemotherapy has a positive effect on therapy outcome and within this project we hope to obtain a selection of nanobodies that target RCA’s to used 1. study the role of RCA’s in tumor-growth and anti-tumor therapy, and 2. stimulate CDC by inhibiting RCA activity.

June 2020 - project is ongoing

A collaborative effort between the Tumor Immunology Facility and the Mass Spectrometry Department 

Identifying ligands recognized by γδT cells is still highly relevant. In the past year several papers shed new light on the recognition mechanism of ligands by γδTCRs, most of these studies imply that the TCR can interact with multiple ligands. Depending on the Vγ-gene usage γδTCRs can bind to butyrophilin-family members, however this interaction doesn’t lead to a strong activation signal. Binding to a second ligand using regions in the Vδ-domain and CDR3γ loop they can recognize a second ligand which is essential for potent T cell activation(1-4). For many γδTCRs one or both of these ligands have not yet been identified, so additional efforts are needed to identify these ligands. 

This project is ongoing.

A collaborative effort between The Faculty of Science and the Mass Spectrometry Department 

A collaborative effort between the University Medical Centre Utrecht and the Nanobody Facility

Staphylococcus aureus causes a range of clinical infections that are becoming increasingly difficult to treat due to antibiotic resistance (e.g. MRSA). New treatment strategies are needed to curtail this alarming development. Opsonic antibodies are regarded indispensable for effective immunity against S. aureus, since they enhance phagocytosis and killing by neutrophils. Therefore, there is increasing interest to apply monoclonal antibodies (mAbs), which can either be used prophylactically to temporary eradicate S. aureus in individuals at risk for infection or for treatment of infections. Thus far however, all passive immunization strategies in humans have failed to show efficacy against S. aureus infections. Strain diversity, the large repertoire of (human-specific) virulence factors, the difficulty of translating animal models to humans, and lack of good vaccine targets are all contributing to the challenge.
My research focuses on the abundant S. aureus cell wall glycopolymers called wall teichoic acids (WTAs), which have attracted major attention as targets for new antibiotics and possibly vaccines. WTAs display limited structural variation through glycosylation, which occurs through the activity of three distinct enzymes: TarM, TarS and TarP. In the natural antibody repertoire, the TarS-coat represents an immunodominant epitope on S. aureus for antibody recognition, while the TarP coat blocks antibody binding and support immune evasion. Similarly, antibodies recognizing the TarM-coat are very low abundant. Hence, we hypothesize that boosting immune detection of the TarM- and TarP-coat interferes with bacterial immune evasion and enhances bacterial clearance.

This project is ongoing.

A collaborative effort between the Infection and Immunity Department and the Monoclonal Antibody Facility

Epithelial transmembrane (TM) mucins are key players in host-microbe interactions and carcinogenesis, but they are poorly studied. Due to their complex glycosylation and size, the generation of tools to study TM mucins is challenging. The family of TM mucins consists of at least 8 members with various levels of expression at mucosal surfaces such as the lung and intestine. TM mucins have large highly glycosylated extracellular domains with barrier or receptor functions for pathogenic bacteria and viruses. The cytoplasmic tails connect to signaling pathways that regulate inflammatory responses, cell proliferation and apoptosis. The TM mucin MUC1 is protective during infection with Campylobacter, Helicobacter or Influenza A virus. However, we demonstrated that the MUC1 serves as a receptor for Salmonella invasion into epithelial cells. We conclude that mucins can have highly specific functions depending on the pathogen in question. The roles of different TM mucins in Klebsiella pneumoniae or corona virus infections in the respiratory tract have not been established. However, interactions of pathogens with mucin proteins might greatly impact therapeutic efficiency. In my group, we have a good toolbox available to study MUC1. However, for other mucins we lack the necessary tools starting with specific antibodies.

The TM mucin MUC13 is highly expressed in the intestinal tract and moderately expressed in the lungs. Adenocarcinomas such as colorectal carcinoma and pancreatic carcinoma overexpress MUC13 and overexpression is associated with metastasis and poor prognosis. We generated CRISPR/Cas9 knockout cell lines and demonstrated that MUC13 regulates cell proliferation by interaction with the EGFR and coordinates cellular migration during wound healing. Next, we need to investigate the role of MUC13 during mucosal infection with bacteria and viruses. Novel MUC13 antibodies are instrumental to study the role of MUC13 during inflammation and could serve as tumor biomarkers and targeting of MUC13-overexpressing adenocarcinomas.

This project was approved in May 2020.

A collaborative effort between the UMCU and the UMab Facility

In the past decade T cells have gained interest for their potential as immunotherapeutic agents for malignancies of diverse origin. They are known to be important for recognition of foreign pathogens, stress signatures of infected cells but also cancer cells. In vitro, T lymphocytes display very potent and broad tumor recognition; they can target and lyse cancer cells of both hematological and solid origin. However, the adoptive transfer of ex vivo expanded polyclonal T cells associates so far with little clinical response, most likely because of an heavily underestimated diversity and mechanisms of tolerance(1). Therefore there is a need to develop novel treatment strategies utilizing the tumor reactive potential of  TCRs while overcoming the limitations of the current therapies.
We developed a TCR based T cell engager by linking soluble tumor-reactive  TCRs  to a CD3 binding moiety, creating TCR Anti-CD3 bispecific molecules (GABs). These GABs can bind to tumor cells with the TCR and then recruit and activate T lymphocytes by binding to CD3 on the lymphocyte. The CD3 binding moiety of the GAB consists of a single chain antibody fragment (scFv) derived from a CD3 antibody, currently clone OKT3 which is used in many preclinical bispecific T cell engagers. However, many recent publications have shown that the (pre)clinical success of these T cell engagers depends on the binding properties of both the tumor targeting part as well as the T cell (CD3) binding part of the bispecific molecule(2).  In order to study the role of the CD3 binding part in the context of GABs, we propose to utilize the excellent immunization pipeline of the UMAB facility to generate novel anti CD3 monoclonal antibodies with different binding properties.

This project was approved in May 2020.

A collaborative effort between the Faculty of Veterinary Medicine and the Nanobody Facility

The picornaviridae form a large family of viruses that comprise many human and animal pathogens, including causative agents for hepatitis, aseptic meningitis, and foot-and-mouth disease. The life cycle of these naked viruses typically culminates in the induction of cell lysis and consequent release of viral progeny. To counteract virus spread, the host mounts a neutralizing antibody response directed against the viral capsid. Importantly, recent discoveries indicate that already in the pre-lytic phase of infection virus particles can escape infected cells via packaging and release within extracellular vesicles (EV). Inside these naturally occurring nanoparticles, viruses are cloaked by a completely host-derived membrane and are therefore inaccessible to neutralizing antibodies. EV-enclosed viruses can be abundantly present in the circulation of viremic patients (Feng 2013). Enclosure in EV not only protects viruses against humoral immunity, but also affects their uptake by cells. EV have been shown to enhance infection efficiency and allow delivery of virus to otherwise inaccessible tissues including the brain (Hudry 2016). Inhibition of EV-enclosed virus release is therefore an interesting target for antiviral therapies. However, the molecular mechanisms underlying this newly discovered means of viral dissemination are largely unknown. My research group at the division CMC has a long-standing expertise in EV research and recently discovered that EV-enclosed viruses are released during infection with EMCV (Grein 2019) and CVB3, two excellent models representing a range of pathogenic picornaviridae. However, further research is hampered by the lack of antibodies against these widely studied viruses. We propose to produce nanobodies to EMCV and CVB3, which will be used in advanced microscopic imaging and to develop screening methods for the identification of molecular players in EV-enclosed virus formation. We expect that these nanobodies will greatly enhance our knowledge of host-virus interactions and will help to identify therapeutic targets aimed at EV-mediated virus spread.

This project was approved in May 2020.

A collaborative effort between the Nanobody Facility and the research group of Joost Snijder,  Science Faculty, UU.

This project involves the investif\gation of the role of Myelin Associated Glycoprotein and other neural cell adhesion molecules as receptors in herpes virus infection.

This project was approved in July 2021.

A collaborative effort between the Nanobody Facility and the research group of Piet Gros, Science Faculty, UU.

This project aims to develop nanobodies that can specifically target a large variety of intracellular cancer biomarkers by exploiting nature’s design of presenting peptides derived from these proteins at the cell surface through MHC-1 molecules.

This project was approved in July 2021

A collaborative effort between Celia Berkers (metabolomics) and the Biomeolecular Mass Spectrometry group  of  Albert Heck

The host kinases pathways that regulate metabolism after picornavirus infection is largely unknown. Targeted phosphoproteomics of specific phosphorylations in kinase activating loops is used to efficiently identify the activity of many major kinases. We aim to use this technique to study the role of the security proteins L and 2Apro in regulating host kinases to better understand how L and 2Apro suppress innate immune responses and metabolism.

This project was approved in July 2021

A collaborative effort between the group of Frank van Kuppeveld, Faculty of Veterinary Medicine and Celia Berkers (metabolomics). 

This project aims to identify (druggable) metabolic pathways that are commonly affected upon infection with coronaviruses using a fluxomics approach. In this project, we bring together ‘viral infections’, one of the main biological drivers within the UMI Hub, with mass spectrometry, one of its key technologies. The proposed work contributes to the UMI aim of developing novel ways of immune interventions, particularly to combat RNA viruses.

This project was approved in July 2021

A collaborative effort between the group of Berend Jan Bosch, Faculty of Veterinary Medicine and the Tumour Immunology Facility at the UMCU.

This project aims to test the potency of a novel vaccine candidate. Because of the high mutation rate of the coronavirus genome, rapid evolution of new variants is expected and current vaccines could easily become obsolete or would need periodical updates in order to build immunity against new virus variants. 

This project was approved in July 2021