Testing the role of B cell receptor signalling in germinal centre responses

Lead Research Organisation: University of Birmingham
Department Name: Immunity and Infection

Abstract

In our highly mobile society the risks from infectious diseases are increasing due to increased travel, altered global migration and reduced security in the food-chain. Vaccination is a cost-effective approach that can protect against infection. However, vaccination does not always guarantee complete protection against infection and this is especially true in the very young and the elderly. As longevity is increasing, by 2034 it is estimated that 1 in 4 of the UK population will be aged over 65, this issue will continue to be at the forefront of preventive medicine. One of the main reasons for reduced vaccination responses is that the immune system is impaired at the extremes of age. Although poorly understood one cause is poor functioning of B cells, the immune cells that produce the antibodies that help identify and remove pathogens and thus prevent infection.

Production of long lived pathogen-specific B cells is the basis of most vaccinations. When a new pathogen or vaccine enters the body B cells respond by producing antibody and importantly they are conditioned through an optimization process called "affinity maturation" to continually try to improve the quality of the antibody they make. The process is akin to evolution in that B cells undergo repeated rounds of antibody gene mutation followed by selection of the "fittest" B cells to produce antibody that will best deal with the pathogen. How affinity maturation works has been under intense scrutiny over recent years and we know that it involves B cells receiving help from other immune cell, namely T cells. What we do not understand is how the direct interaction of B cells with the pathogen or vaccine shapes this process and having this missing piece of information should allow us to develop radically different approaches to vaccinations to improve the quantity and quality of antibody produced for example in older adults.

The ultimate aim of this project is to understand how this B cell and antibody selection process is regulated to allow us to produce better vaccines. This will be achieved through collaboration with the company Medimmune who will help to produce genetically modified mice where affinity maturation is enhanced. In addition to helping develop better vaccines, these mice will become valuable tools for industry in the quest to produce new monoclonal antibody therapeutics, as the process of generating new monoclonal antibodies itself involves vaccination.

The link with Medimmune will ensure that their capacity in animal model generation is integrated with Dr Toellner's knowledge of B cell and vaccination biology, to produce a project that will lead to a step change in the field and generate resources useful to researchers and industry alike.

Technical Summary

B cell activation leading to B cell differentiation is regulated by two major signals: Signals through B cell receptors (BCR) and signals from T helper cells. These signals regulate antibody responses during initial B cell activation after antigen entry, during the evolution to high affinity in germinal centres, and during the reactivation of a memory response. While signals from T cells are well studied, there is less information about the impact of differential BCR signals on B cell activation.
We will test the hypothesis that BCR signal strength is an important regulator of B cell differentiation. To test this, a range of mouse models with altered BCR signalling strength are developed. These involve hybrids of B cell receptors of naive B cells (IgM or IgD) and IgG1, leading to mice with hyperactive signalling in naïve B cells, and inducible mutations of kinases or phosphatases downstream of BCR signalling. These models will allow induction of differentiation stage dependent variation of BCR signalling strength in vivo, allowing us to comprehensively test the role of BCR signalling during differentiation stages in response to vaccination and B cell selection and differentiation. Importantly, all models proposed will lead to changes in BCR signalling activity when B cells enter the germinal centres. Therefore, we will be able to test whether and how BCR signalling strength within the germinal centre regulates B cell selection.
A spin-off from this project will be new mouse models that are more prone to produce high quality antibody responses to difficult targets, e.g. evolutionary conserved antigens or autoantigens. These may have commercial value in the production pipeline to new monoclonal antibody drugs, but will also be provided to the scientific community for non-commercial applications. The mice will also be useful to study the development of antibody mediated autoimmune diseases, and we started discussions with Rheumatologists at UoB about this.

Planned Impact

Society
The importance of this work extends significantly beyond supporting academic research. By understanding how antibody responses develop we are helping to understand how to improve vaccine development. This is vital. In our highly mobile society the risks from infectious diseases are increasing due to increased travel and altered global migration and extend also into the food-chain. This is because many of our infections are actively acquired from food (e.g. Salmonella) or livestock rearing acts as an incubator and enables pathogen diversity (e.g. influenza virus). In parallel, we have a decreasing efficacy of anti-microbial treatments due to resistance and only a modest number of anti-virals available. Vaccination is a cost-effective approach that can protect against infection at the extremes of age in those groups that are most susceptible. Furthermore, vaccination is an acceptable intervention to society at large, and as the media response to the recent measles outbreak demonstrates, is one that is diminishing in controversy. Indeed, the measles outbreak demonstrates the importance of vaccination programmes to protecting society and the consequences when there is insufficient vaccine coverage. Theoretical background how vaccines work, what signals regulate the emergence of high affinity B cells and antibody producing cells, and how immunological memory cells are regulated, is still in its infancy.
Industry
This collaboration between industry and UoB will not only increase our theoretical understanding of immune responses to vaccines, which may lead to better intelligent design of vaccines. It also will produce valuable information and tools (mouse strains) for the production pipeline towards new monoclonal antibody therapeutics. Generation of monoclonal antibody drugs to new targets involves vaccination of mice as a first step of raising specific antibodies. Understanding the immunological principles of vaccination will help designing and choosing antigens reliably inducing immune responses. Further, we expect the new mouse strains produced to produce antibody responses to difficult targets, e.g. autoantigens or conserved antigens that are of superior quality, i.e. broadness of the response to different antigenic epitopes and affinity of antibodies developing.

Publications

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Zhang Y (2016) Regulation of germinal center B-cell differentiation. in Immunological reviews

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Linterman MA (2017) T cells trump autoimmune antibody responses to limit sedition. in Nature immunology

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Barone F (2015) IL-22 regulates lymphoid chemokine production and assembly of tertiary lymphoid organs. in Proceedings of the National Academy of Sciences of the United States of America

 
Description We have developed a new mouse model for B cell anergy. B cells are the cells that make antibody. Antibodies protect us from infection after vaccination. Antibodies are used as drugs to target specific antigens causing disease, e.g. antigens on tumours. Antibodies can cause disease in antibody-mediated autoimmunity. Anergic B cells are cells that cross react with self-antigens, and therefor are driven into a suppressed (anergic) activation state during B cell development. These cells continue to circulate through the body, but cannot easily be activated. Our new mouse model of B cell anergy should be useful to study the induction of autoimmunity by studying how anergy can be broken. It should also help understanding how we can overcome anergy during monoclonal antibody drug development, where it is often necessary to activate anergic B cells.
Exploitation Route We plan to use the new mouse model to further study anergy, autoimmunity, monoclonal antibody drug design and the response of B cells to autoantigens in cancer.
Sectors Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology

 
Title GM mouse strains wth altered B cell receptor signalling in germinal centres 
Description GM mouse lines that should have altered B cell affinity maturation upon immunisation have been produced. The suitability of those to produce antibody of increased affinity is currently tested. 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Provided To Others? No  
Impact none yet 
 
Title IgMg1 mice 
Description B cell receptor signalling was altered by adding the IgG1 signalling chain onto IgM. 
Type Of Material Model of mechanisms or symptoms - mammalian in vivo 
Year Produced 2018 
Provided To Others? No  
Impact The new strain is a model of B cell anergy and will be useful to study autoimmunity and activation of B cells self-specific responses 
 
Description Gene expression in memory B cells 
Organisation Francis Crick Institute
Country United Kingdom 
Sector Academic/University 
PI Contribution Provision of mRNA from in vivo differentiated memory B cells
Collaborator Contribution Gene expression analysis from different in vivo differentiated memory B cells by RNAseq and bioinformatic analysis
Impact New data on gene expression in new memory B cell subsets.
Start Year 2015
 
Description In silico modelling of germinal centres 
Organisation Helmholtz Association of German Research Centres
Department Helmholtz Centre for Infection Research (HZI)
Country Germany 
Sector Academic/University 
PI Contribution Data provision for in silico models Work on the in silico model
Collaborator Contribution In silico modelling Predictions for optimal experimental in vivo setups
Impact The collaboration has led to several publications. It has produced a new in silico model to analyse the regulation of affinity maturation in germinal centres. This has lead to several further collaborations and publications for our collaborators.
Start Year 2006
 
Description Industrial partnership 
Organisation MedImmune
Department MedImmune Cambridge
Country United Kingdom 
Sector Private 
PI Contribution We are analysing GM mouse strains with altered B cell receptor signalling in germinal centres for their suitability as tools to produce new therapeutic monoclonal antibody drugs. The prediction is that the GM mice will easier produce specific high affinity antibodies to difficult target antigens. Further, we are analysing effects of these genetic modification on the regulation of high affinity antibody responses.
Collaborator Contribution The industrial partners design and produce the initial vectors for genetic manipulation of mice. They transfect these into embryonic stem cells and produce GM mice. After this step the mice are transferred to us. At a later stage they will test the mice for practical usability as vehicles to produce high affinity B cells to difficult targets for hybridoma fusions. They will use the mice and test whether they ca produce monoclonal antibodies using some of there identified human therapeutic targets.
Impact Three GM mouse strains have been produced and are currently analysed in our lab, publications in preparation. New iCASE PhD studentship to work on 4th strain.
Start Year 2014
 
Description Intravital imaging of memory B cell migration 
Organisation University of Cambridge
Country United Kingdom 
Sector Academic/University 
PI Contribution Provision of GMO mice,
Collaborator Contribution Immunisation of GMO mice and intravital imaging of migrating memory B cells. Training visits for UoB staff on intravital microscopy in preparation.
Impact Early results show new patterns of memory B cell migration and roles of chemokine receptors directing this.
Start Year 2016
 
Description Intravital imaging of plasma cell differentiation 
Organisation Leibniz Association
Department German Rheumatism Research Centre
Country Germany 
Sector Charity/Non Profit 
PI Contribution A postdoctoral research fellow spent twice 2 months at DRFZ to learn intravital imaging techniques and study plasma cell differentiation from germinal centres
Collaborator Contribution Hosting, training, and help with experiments. Provision of experimental animals
Impact Training of postdoctoral research fellow Preliminary data on plasma cell differentiation Submission of funding proposal to Wellcome Trust for an intravital microscope
Start Year 2011
 
Description Provision of GM mice 
Organisation RIKEN
Department RIKEN Center for Integrative Medical Sciences (IMS)
Country Japan 
Sector Hospitals 
PI Contribution Provision of mice that can be used to track germinal centre B cells
Collaborator Contribution Provision of mice that have an inducible expression of Cre recombinase under the control of the S1PR2 promotor
Impact We currently use these mice to generate new unique mouse models to study germinal centre B cell differentiation.
Start Year 2018
 
Description Testing the role of the cytoplasmic domain for B cell receptor signalling in vitro 
Organisation Albert Ludwig University of Freiburg
Country Germany 
Sector Academic/University 
PI Contribution We provide data on our experiments on in vivo activated B cells with genetic manipulation of B cell receptor signalling (IgMg1, IgMDg1, and IgDg1 mice).
Collaborator Contribution Gene manipulated mice with B cell specific Cre expression were provided. B cell lines are being developed with similar genetic manipulation of B cell receptor signalling, similar to our GM mouse lines. These will be suitable to test predictions and verify outcomes from our in vivo experiments in vitro.
Impact New cell lines and new genetically manipulated mouse lines
Start Year 2017