iR+ Seminar Series: Theory and Applications of Immune Repertoires
The iReceptor Plus Seminar Series highlights the Theory and Applications of immune repertoires. The overall goal of the iReceptor Plus Consortium is to facilitate sharing and analysis of Adaptive Immune Receptor Repertoire Data. These immune repertoires are rapidly increasing in size and importance to all aspects of immunotherapy, and the Consortium is dedicated to improving analysis and curation tools that will allow the entire immunology community to maximize the benefit of these data for biomedical research and patient care. In the seminar series we will invite one senior and one early career scientist each month to present their most exciting and impactful research in this area.
Zoom connection details will be sent to registrants two days before each seminar.
The seminar series talks will not be recorded.
iR+ Seminar Series - June 24, 2021
Keynote Speaker: Sachdev Sidhu, University of Toronto
Title: Rapid Development of Multivalent Anti-COVID-19 Antibodies
Abstract: We used protein engineering to develop tetravalent synthetic neutralizing antibodies for SARS-CoV-2. We show that these antibodies can be produced at large scale and possess stability and specificity comparable to approved antibody drugs. The best antibody targets the host receptor binding site of the virus spike protein, and thus, its tetravalent version can block virus infection with a potency that far exceeds that of the bivalent IgG.
Recombinant neutralizing antibodies (nAbs) derived from recovered patients hold promise as effective therapeutics against COVID-19. Here, we describe the use of advanced protein engineering and modular design principles to develop tetravalent synthetic nAbs that mimic the multi-valency exhibited by IgA molecules, which are especially effective natural inhibitors of viral disease.
At the same time, these nAbs display high affinity and modularity typical of IgG molecules, which are the preferred format for drugs. We show that highly specific tetravalent nAbs can be produced at large scale and possess stability and specificity comparable to approved antibody drugs. Moreover, structural studies reveal that the best nAb targets the host receptor binding site of the virus spike protein, and thus, its tetravalent version can block virus infection with a potency that exceeds that of the bivalent IgG by an order of magnitude.
Design principles defined here can be readily applied to any antibody drug, including IgGs that are showing efficacy in clinical trials. Thus, our results present a general framework to develop potent antiviral therapies against COVID-19, and the strategy can be readily deployed in response to future pathogenic threats.
Early Career Scientist: Ayelet Peres, Bar-Ilan University (BIU)
Title: Germline variability in adaptive immune repertoires
Abstract: Many recent studies utilize adaptive immune receptor repertoire sequencing (AIRR-seq) data to open new prospects in understanding the adaptive immune system. We and others use AIRR-seq data to gain insight on genomic variability within the T and B cell receptor encoding genomic loci.
To this end, we adapted and developed tools for genotype and haplotype inferences from AIRR-seq data. Applying our methods to several public data sets revealed a plethora of novel undocumented TRBV, IGHV, IGKV and IGLV alleles, structural variations, and variations in the untranslated regions including alternative splicing events.
In this talk I will cover our recent work and show how the discovered variations affect the expressed repertoires. I will also discuss challenges with these kinds of inferences, such as indistinguishable sequences coming from different genes and/or partial sequencing protocols, and our current thinking for how to overcome these challenges.
iR+ Seminar Series - May 27, 2021
Keynote Speaker: David Klatzmann, Sorbonne Universite
Title: Polyreactive CD8+ ab antiviral T cells: the immunologists’ 2nd “dirty little secret”
Abstract: How evolution shaped a process that produces TCRs that would effectively respond to diverse infectious agents is a central question of immunology. The paradigm is that a diverse enough repertoire of TCRs should always provide a proper, though rare, specificity for any given need. Expansion of such rare T cells would provide enough fighters for an efficacious immune response and enough antigen-experienced cells for memory.
We found that thymopoiesis releases a large population of CD8+ T cells harboring diverse α/βTCRs that are highly enriched for viral antigen recognition and have a fuzzy rather than tight specificity. In vitro, they bind to and are activated by multiple unrelated viral peptides. In vivo, T cells harboring such TCR respond to vaccination, infection, including COVID-19, and cancer.
Our results support an evolutionary selection of pleiospecific α/βTCRs for broad antiviral responses and heterologous immunity.
Early Career Scientist: Seth J. Zost, PhD, Postdoctoral Fellow, Crowe Laboratory, Vanderbilt Vaccine Center, Vanderbilt University Medical Center
Title: Leveraging single B cell sequencing to study human antibody responses to influenza and SARS-CoV-2
Abstract: Respiratory viruses such as influenza viruses and coronaviruses are major threats to public health, and both of these families of viruses have caused pandemics in the 21st century. In addition, circulation of seasonal influenza viruses results in significant morbidity and mortality.
In the case of both pathogens, antibodies play a major role in protection from infection, but there are important unanswered questions as to which epitopes are targeted in human immune responses, how antibody lineages evolve after repeated exposures, and the degree to which B cell clonotypes are shared between individuals.
Recent advances in single cell sequencing and gene synthesis have enabled paired sequencing of B cell populations and antibody discovery at previously unprecedented scales. Here, I will highlight several examples from our group where we have coupled repertoire sequencing and high-throughput antibody expression to both study the evolution of antibody lineages and to identify public clonotypes in the antibody responses to both influenza and SARS-CoV-2.
iR+ Seminar Series - April 22, 2021
Keynote Speaker: Hedda Wardemann, German Cancer Research Center (Deutsches Krebsforschungszentrum, DKFZ), Heidelberg, Germany
Title: Single-cell based antigen-receptor gene and function analyses of the human immune response to a complex pathogen
Abstract: We have developed a platform for the high-throughput analysis of Ig gene repertoires that preserves natural IgH and IgL gene associations and provides full-length Ig gene sequence information for direct gene cloning and the production of recombinant monoclonal antibodies. Recently, we have established a similar platform for the amplification and cloning of paired T cell receptor (TCR) alpha and beta chain genes to enable functional assessments of TCRs at single cell level. I will discuss how we use this platform to study the clonal evolution and functional quality of the human immune response to the malaria parasite Plasmodium falciparum with the long-term goal to develop improved vaccine design strategies.
Early Career Scientist: Jared Ostmeyer, UTSW
Title: Controlling for Confounding Factors when Diagnosing Cancer from T Cell Receptor Repertoires
Abstract: Our publications have shown that T cell receptor (TCR) repertoires sequenced from tissue biopsies can be used to distinguish tumor from control tissue with > 90% classification accuracies, suggesting that TCR repertoires can be used to diagnose cancer.
TCR repertoires sequenced from blood samples of cancer patients are now publicly available, and it is tempting to use these existing samples to develop blood-based assays for diagnosing cancer. Already, research groups including us have obtained promising results. However, these results are obtained by combining cancer and control samples from mismatching studies. Without controlling for age, sex, race, and even the reduced amount of blood volume collected from cancer patients relative to controls, these exciting results could merely be an artifact of confounding factors.
In this study, we control for possible confounding factors and observe that our ability to diagnose cancer from TCR repertoires sequenced from blood does not vanish. On patient-holdout cross-validations, we achieve diagnostic accuracies of 90% for colorectal cancer (controlling for depth coverage), 84% for breast cancer (controlling for depth coverage and age), and 83% for lung cancer (controlling for age, sex, race, and depth coverage).
While it is impossible to control for all potential confounding factors like variations in geographic locations, these results are encouraging. We propose future studies that (i) collect matching controls and cases and (ii) confirm TCR repertoire sequencing provides a distinct signal for diagnosing cancer independent of other blood-based assays and therefore can improve the diagnostic performance of these other blood-based assays.
iR+ Seminar Series - March 25, 2021
Keynote Speaker: Corey Watson, University of Louisville
Title: Germline polymorphisms in the heavy chain immunoglobulin locus contribute to variation in the antibody repertoire
Abstract: The immunoglobulin loci are among the most complex and polymorphic regions of mammalian genomes. The complexity of these loci has historically hindered our ability to fully characterize the extent of haplotype diversity that exists within and between species, and in turn, delineate the impact of germline variation on antibody-mediated responses.
A major focus of my lab is the development of novel genomic tools that leverage long read sequencing to comprehensively genotype the IG loci. In this talk, I will discuss our ongoing efforts to expand genomic resources for the human IG loci, and present data that demonstrate connections between IG polymorphism and inter-individual variation in the expressed antibody repertoire.
Early Career Scientist: Mamie Wang, (Interdepartmental Program in Computational Biology and Bioinformatics), Yale University
Title: High-throughput Single-cell Profiling of B cell Responses Following Inactivated Influenza Vaccination in Young and Older Adults
Abstract: Seasonal influenza contributes to a substantial disease burden annually, resulting in approximately 10 million hospital visits and 50 thousand deaths in a typical year in the US. 90% of the annual mortality from influenza occurs in people over the age of 65. While influenza vaccination is the best protection against the virus, it is less effective for the elderly. This may be due to differences in the quantity or type of B cells induced by vaccination in older individuals. To investigate this possibility, we leveraged recent development in single-cell technology that allows for simultaneous measurement of both gene expression profile and the B cell receptor (BCR) at single-cell resolution.
Pre- and post-vaccination peripheral blood B cells were sorted from three young and three older adults who responded to the inactivated influenza vaccine and were profiled using single-cell RNAseq with paired BCR sequencing. At pre-vaccination, we observed a higher somatic hypermutation frequency and a higher abundance of activated B cells in older adults than young adults. Following vaccination, young adults mounted a more clonal response than older adults. The response involved a mix of plasmablasts, activated B cells, and resting memory B cells in both age groups. The response in young adults was dominated by expansion in plasmablasts, while older adults’ response also involved activated B cells.
We observed a consistent change in gene expression in plasmablasts after vaccination between age groups but not in the activated B cells. These quantitative and qualitative differences in the B cell response may provide insights into the age-related change of influenza vaccination response.
iR+ Seminar Series - February 25, 2021
Keynote Speaker: Ludvig Sollid, University of Oslo
Title: Pathogenic T-cell/B-cell cross-talk in celiac disease: Studies of antigen receptors
Abstract: Adaptive immunity with involvement of antigen-specific T cells and B cells are likely essential in development of autoimmune diseases. Research on the involvement of autoimmune T and B cells is however hampered by the lack of knowledge of the disease driving antigens, particularly in humans. Celiac disease, a common disorder that has autoimmune features and this is caused by a maladapted immune response to cereal gluten proteins, is an exception in this regard. The disease is driven by activation of gluten-reactive CD4+ T cells and it is hallmarked by highly disease specific autoantibodies to the enzyme transglutaminase 2 (TG2) and to deamidated gluten peptides (DGP). I will discuss recent findings on the phenotype of gluten-reactive CD4+ T cells, the usage of disease-relevant T-cell receptors and B-cell receptors as well as the crosstalk between pathogenic T cells and B cells in this instructive human disorder.
Early Career Scientist: Jean-Philippe Buerckert, BISC Global
Title: A Deep Dive into the CDRH3 Landscape of SLE and RA Patients
Abstract: Systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) are two common polygenic autoimmune diseases characterized by elevated serum autoantibodies against a variety of tissue antigens and progressive damage in multiple organs and tissues. In both diseases, malfunctioning checkpoints result in the loss of self-reactivity control of the adaptive immune receptor repertoires (AIRR) manifesting in the production of autoreactive B-cells.
Here we show how AIRR-seq can be applied to dissect the CDRH3 repertoire from PBMC B-cell isolates. Sampling was performed from 10 SLE and 11 RA patients over two timepoints as well as those of 10 healthy controls yielding a total of about 20,000,000 sequences after filtering.
Globally, our data show increased non-functional BCR rearrangements in both diseases compared to healthy donors. We examined CDRH3 repertoire down to the single amino acid level and found that RA and to a lesser extent SLE patient’s repertoires are characterized by increased amounts of hydrophobic k-mers and amino acids compared to healthy controls.
Collectively, these observations shed new light on the underlying molecular basis of autoreactive B-cells in SLE and RA and indicate a complex cluster of B-cell development defects.
iR+ Seminar Series - January 28, 2021
Keynote Speaker: Steven H. Kleinstein, Yale School of Medicine, New Haven, CT, USA
Title: Analysis of B cell antibody repertoires from next-generation sequencing (in infection, vaccination and autoimmunity)
Abstract: Next-generation sequencing (NGS) technologies have revolutionized our ability to carry out large-scale adaptive immune receptor repertoire sequencing (AIRR-Seq) experiments. AIRR-Seq is increasingly being applied to profile B cell receptor (BCR) repertoires and gain insights into immune responses in healthy individuals and those with a range of diseases. As NGS technologies improve, these experiments are producing ever larger datasets, with tens- to hundreds-of-millions of BCR sequences. Although promising, repertoire-scale data present fundamental challenges for analysis requiring the development of new techniques and the rethinking of existing methods that are not scalable to the large number of sequences being generated .
To address these challenges, we have developed computational tools and methods that we currently make available to the wider scientific community through the Immcantation tool suite. This includes: raw read processing, novel V gene allele detection, subject-specific germline genotype identification, B cell clone assignment, lineage tree construction and analysis, somatic mutation profiling and selection analysis. Along with the underlying computational methodology, this presentation will discuss applications of BCR repertoire sequencing and lineage analysis to infection (Lyme disease, COVID-19 and West Nile Virus), vaccination (Influenza), autoimmunity (Multiple sclerosis, Myasthenia Gravis) and allergy/asthma.
- Yaari G, Kleinstein SH. Practical guidelines for B-cell receptor repertoire sequencing analysis. Genome Med. 2015 Nov 20;7:121. doi: 10.1186/s13073-015-0243-2.
Early Career Scientist: Milena Pavlovic, University of Oslo, Oslo, Norway
Title: immuneML: an open-source ecosystem for machine learning analysis of adaptive immune receptor repertoire data
Abstract: Adaptive immune receptor repertoires (AIRR) are key targets for immunological and pharmacological research as they provide a DNA-sequence record of all past and ongoing adaptive immune responses in health, disease, infection and vaccination. The capacity of machine learning (ML) to learn complex discriminative sequence patterns has led to its increasing use for AIRR-based diagnostics and therapeutics discovery. Previous developments have however been highly heterogeneous in terms of technical solutions, domain assumptions and user-interaction options, hampering transparent comparative evaluation and the ability to explore and select ML methodology most appropriate for a given study.
immuneML addresses these previous concerns by covering all major steps in AIRR ML within an open-source ecosystem and online user interface: AIRR sequence data read-in and encoding, training ML models of antigen specificity or immune state prediction as well as model assessment and interpretation. We demonstrate the broad applicability of immuneML for AIRR ML research by (i) replicating inside immuneML a published large-scale study on AIRR-based immune state prediction, (ii) applying a novel ML method for AIRR-based antigen specificity prediction and (iii) showcasing how immuneML may be used for AIRR ML method benchmarking. immuneML promotes reproducibility, customizability and shareability by (i) providing infrastructure for sharing complete ML workflows and intermediate steps, (ii) useful default parameters and workflows that shield beginners from common ML mistakes, (iii) and a user-friendly design based on the Galaxy framework.
iR+ Seminar Series - November 26, 2020
Keynote: Gunilla Karlsson Hedestam, Karolinska Institutet, Stockholm, Sweden
Title: Generation of a comprehensive database of rhesus and cynomolgus macaque IGH alleles
Abstract: We applied the immunoglobulin (IG) gene inference tool, IgDiscover, to define germline VDJ alleles in 45 macaques of different origins. Our analysis resulted in a comprehensive database comprising 1198 IGHV alleles, of which around 70% were not previously described. Haplotype analysis of the animals revealed a considerable level of structural variation in the IGH locus. This work will facilitate high-quality B cell studies in rhesus and cynomolgus macaques.
Early Career Scientist: Vanessa Mhanna, Sorbonne Universite, Paris, France
Title: Exhaustion of the regulatory T cell receptor repertoire instigates diabetes in NOD mice
Abstract: Non-obese diabetic (NOD) mice spontaneously develop autoimmune diabetes. We aimed to analyze their TCR repertoire to better understand NOD autoimmunity. We performed next-generation sequencing of TCRs from splenocytes of prediabetic NOD and normal B6 mice. We analyzed the repertoire of CD4+ effector T cells (Teffs), CD44low CD62Lhigh naïve regulatory T cells (nTregs) and CD44high CD62Llow activated/memory Tregs (amTregs). These latter are known to respond to self-antigens and to be involved in protection against autoimmune diseases.
NOD and B6 nTreg TCR β repertoires were very diverse and mostly composed of unexpanded clonotypes. In contrast, B6 amTregs contained frequent expanded clonotypes that were lost in NOD amTregs resulting in an increased diversity of their repertoire. This was also seen, albeit to a lesser extent, in NOD Teffs. These observations suggested that NOD mice had lost the amTreg clonotypes that could protect them from diabetes.
As IL-2 administration leads to Tregs expansion and activation, and correlatively to protection from diabetes occurrence, we investigated the effects of IL-2 on NOD TCR repertoires. IL-2 administration to NOD mice restored amTreg clonotype expansions and led to few and no clonotype expansions of nTreg and Teffs, respectively. Noteworthily, IL-2-expanded amTreg and nTreg clonotypes were markedly enriched for islet-antigen specific TCRs.
Altogether, our results establish a causal link between an IL-2-mediated impoverishment of Treg repertoires affecting self‑antigen-specific TCRs and the development of autoimmune disease.