We can help you understand and manage immune responses. Our unique solutions for immunology research can save you time, money and risk, whether your objective is basic research, preclinical or clinical development.
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MHC Pentamers used by team at GSK to study mRNA vaccines that can protect against several Flu strains
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ProImmune launches MutaMap™ Mutational Activity Map Service
How does MutaMap™ compare to molecular evolution technologies?
Molecular evolution techniques such as phage display and other phenotype-genotype
coupled randomization techniques are most commonly used in the affinity
maturation process for monoclonal antibodies and other binding
scaffolds. The advantage of these technologies is that they help explore
a very large sequence space of combined mutations.
There comes a point
however when final decisions have to be made on the implementation of a
protein sequence where individual point mutations may be considered in
an antibody or therapeutic protein to meet a variety of design
objectives. Randomized molecular evolution is not appropriate for this
step. Exploring individual point mutations is nothing new, but it has
been difficult to carry this step out in very high throughput way,
especially where the objective is to clone and express every mutant and
then measure its affinity/activity with reasonable accuracy. This
is what MutaMap™ can achieve.
Figure 5:Example work flow for pre-clinial protein engineering of therapeutic
monoclonal antibody; individual projects may differ.
How long does a MutaMap™ project take?
Our objective is to complete medium size projects of exploring 500-2000 mutations in
approximately 8-12 weeks from receiving the customer’s protein sequence.
Larger projects will take slightly longer, depending on complexity.
What is the stepwise process for carrying out MutaMap™?
we will discuss with you the sequence space you want to explore. This
may be the known paratope of a protein or the CDR and flanking regions
of a monoclonal antibody.
To save time and cost you may not want to explore substituting amino
acids into the sequence that are considered highly non-conservative or
prone to degradation when exposed. We will also discuss what you know
about the binding interaction of your protein with its target, whether
you have working immunoassays for this interaction and whether the wild
type protein and isolated ligand is available to work with in an
immunoassay system. Depending on the nature of the interaction and the
binding partners we will ensure that the base assay of binding wild type
protein to target works well.
details are agreed and the project is initiated we will run the
mutagenesis agreed for each position, express the protein at small scale
and carry out the equilibrium binding titration affinity measurement in
What will you get?
A final technical
report delivered via our secure webserver showing you the affinity or
activity determination for the wild type and each mutant with confidence
interval. This will be presented in various formats for ease of
interpretation, including a standard heatmap.
For customers that
want to carry out protein antigenicity studies in parallel, these can be
carried out in approximately the same timeframe as the MutaMap™. This
means that within a period of approximately 8-12 weeks we will have
determined experimentally both the putative T cell epitopes and the
MutaMap™ of permitted mutations in your protein sequence. This
information can allow you to proceed with much better informed decisions
on how to address immunogenicity related issues for your program while
addressing simultaneously other developability related design decisions
for your sequence.
Introducing: ProT2™ MHC Class II Tetramers
Oxford, UK, 14 January 2016
ProImmune, a leader in services for understanding and managing adaptive
immune responses, announced today the introduction of ProT2™,
a range of Human MHC Class II tetramer reagents for tetramer analyses in
the study of antigen-specific CD4+ T cell immune responses. ProImmune’s
offering represents the broadest choice of ready to order catalog and
custom items in this field and will enable research into CD4+ T cell
responses in a variety of disease areas including cancer, infectious
diseases, allergy, autoimmunity and transplantation. Another key area
for this product range is the detailed study of T cell help in unwanted
immune responses to biologics.
CD4+ T cell
responses are at the heart of steering the adaptive immune system
directing both cytotoxic T cell responses and antibody responses
mediated by B cells. ProT2™ MHC Class II Tetramers allow the direct
detection and separation of single antigen-specific CD4+T cells
accurately by flow cytometry.
Schwabe, CEO of ProImmune commented: “By introducing a broad and growing
ProT2™ Tetramer product range we intend to give MHC Class II tetramer
analysis the emphasis it deserves in the study of CD4+ T cell responses
which play such a pivotal role in adaptive immunity across so many
disease and treatment modalities.”
Class II MHC Monomer reagents are supported by ProImmune’s expert
customer service and application support, helping new users in
particular to establish and optimize robust protocols in their own
addition to ProImmune’s class II reagent offering further complements an
already extensive range of services for evaluating CD4+ T cell
responses, including ProImmune REVEAL® MHC peptide binding assays, CD4+
T cell and DC T cell proliferation assays and standard immune monitoring
services through ELISPOT and flow cytometry based assays.
information on ProT2™ reagents and their availability, or to place an
order, please visit:
, or call + 1 888 505 7765 in the United States or + 44 870 042 7279 in other countries.
Notes to Editors:
About ProImmune -
ProImmune’s mission is to
be a partner of choice for understanding and managing adaptive immune
responses. It does this by offering unique solutions for preclinical and
clinical immunology research, including a comprehensive and integrated
antigen characterization and immunogenicity testing platform, and
products and services for tracking antigen-specific immune responses
with state-of-the-art ELISpot and flow cytometry techniques. ProImmune
is committed to helping researcher’s achieve success through product
innovation, responsive service and focused application support, saving
time and money, and reducing risk.
For more information, please contact
Tel: +44 (0) 870 042 7279
Zyme Communications Ltd
Tel: +44 (0) 7771 730 919
ProImmune Introduces ProSentium™ a Unique Peptide Sequence Database to Investigate T Cell Immune System
Oxford,UK, 14 April 2015
– ProImmune, a leader in services for understanding and managing
adaptive immune responses, announced today the introduction of a new
antigen database service called ProSentium™, based on
assay results using sequencing mass spectrometry. ProSentium data
can show the precise peptide sequences from proteins of interest
visible to the body’s own T cell immune system. The data
collection covers many high value therapeutic targets and areas including the
majority of licensed replacement factor proteins, including clotting
factors, replacement enzymes, spanning development areas including
oncology, cardiovascular, CNS, immunology, gastroenterology and many
Under an exclusive or non-exclusive license, the bespoke database service
investigates and reports peptide hits against the ProSentium
database for a customer’s specific protein or protein family of
interest. Results are reported as overlapping nested peptide sets
and provide statistically relevant antigen sequence data that can be
used to further inform drug development decisions.
CEO of ProImmune commented: “ProSentium supports pivotal decision
making in drug design and development with the physical evidence
required for game changing approaches that target or interact with
the immune system. Through our service offering, we are delighted
that we can offer this ground-breaking resource to researchers so
they can better understand the body’s immune response and bring new
therapies to patients. ”
T cells play a
critical role in all immune responses, including in fighting
infections, cancer and in autoimmunity and in unwanted drug
reactions. To understand T cell responses, it is crucial to
understand the specific peptide antigens that the immune system
recognizes and are presented on the cell surface of both normal and
diseased cells. ProSentium data can be readily analyzed and
reported, and is fully compatible with ProImmune’s in vitro
assay services which can be combined to help investigators gain a
more complete understanding of relevant immune responses at a
For more information on ProSentium please visit:
, or call +1 888 505 7765 in the United States or +44 870 042 7279 inother countries.
More on ProSentium™
About ProImmune -
ProImmune’s mission is
to be a partner of choice for understanding and managing immune
responses. It does this by offering unique solutions for preclinical
and clinical immunology research, including a comprehensive and
integrated antigen characterization and immunogenicity testing
platform, and products and services for tracking specific immune
responses. ProImmune is committed to helping researcher’s achieve
success through product innovation, responsive service and focused
application support, saving time and money, and reducing risk.
14 Nov 2022 To 16 Nov 2022
PEGS Europe 2022, Barcelona
07 Sep 2022 To 10 Sep 2022
DGfI & OGAI Joint Meeting, Hanover, Germany
24 Mar 2022 To 24 Mar 2022
ProImmune SARS-CoV-2 Expert Symposium 2022
Dr. Tim Hickling discusses the importance in the application of immunogenicity prediction and mitigation in the production of biotherapeutics. He outlines some of the current methods for predicting immunogenicity and highlights the need for better accuracy in predictions.
Professor Tao Dong from the University of Oxford discusses her latest work in memory T cell responses in convalescent SARS-CoV-2 patients in the UK, highlighting the information from her publication in Nature Immunology and the use of ProImmune Pro5® Pentamers in her work.
Dr. Amy Rosenberg of CDER, FDA discusses the clinical context for the reappraisal of immunogenicity of therapeutic proteins that are being used to treat COVID-19. She highlights the importance of understanding the autoinflammatory or autoimmune response following infection SARS-CoV-2 and its potential to increase immunogenicity to self proteins leading to autoimmune and autoinflammatory disorders. This indicates the high probability that this could lead to higher immunogenicity in patients to therapeutics sued to treat SARS-CoV-2.
Dr. Nina Le Bert from Duke NUS, discusses her recent involvement in the study of SARS-CoV-2 specific T cell immunity, specifically looking at the induction and persistence of T cells using an overlapping peptide library of specific proteins in SARS-CoV-2 to test convalescent patients' IFN-gamma responses to these peptides. Using a cohort of individuals who were infected and recovered in 2003 with SARS, and a similar approach of overlapping peptides from SARS-CoV, she is able to show there are still persistent ex vivo T cell responses. Importantly, she also highlights the cross reactivity of NP peptides from SARS-CoV and SARS-CoV-2.
CAR-T cells are an engineered T cell therapeutic in which a chimeric antigen receptor (CAR) (a foreign protein) is expressed on the surface of autologous T cells. This foreign CAR protein has the potential to elicit immune responses from the host immune system.
The immune response could be (a) humoral, resulting in the formation of Anti-CAR antibodies and/or (b) cellular, resulting in the development of cytotoxic T cells that are specific to the CAR-T cell. A host vs CAR-T response may result in (a) the neutralization of the CAR, rendering the CAR-T therapeutic ineffective (b) or result in the CAR-T cells being killed by a cytotoxic T cell response akin to a host vs graft response. This may manifest as lack of persistence of CAR-Ts and as loss of efficacy. There may be other reasons for loss of efficacy, and hence understanding if immunogenicity is the reason for loss of efficacy will be key in determining whether a second dose may be meaningful, the timing of this second dose, the impact of lymphodepletion on immunogenicity etc.
Dr. Inderpal Singh is currently a Lead, Immunogenicity Prediction in Biologics, Assays & Technology at Teva Pharmaceuticals. In this talk he speaks to the value of deploying the ProScern DC-T cell proliferation assay to assess the impact of host-cell protein contamination.
Katerina describes the process of codon optimization and the role that it plays in a protein's rate of translation, expression and conformational properties. She describes the presence of synonymous mutations in various diseases and the immunogenicity implications relating to those mutations. She uses Factor IX as a model to describe the workflow to generate variants using CoCoPUTs and the evaluation of those variants for protein expression, confirmational differences, peptide presentation and many more attributes.
Beatriz discusses the use of cancer vaccines to transition ‘cold’ tumors in patients to ‘hot’ tumors. The UPenn group used Next Gen Sequencing to catalog tumor mutations and generated autologous dendritic cell vaccines to evaluate the immunogenicity of missense mutations and define the nature of patient T cells specific for melanoma neoantigens. They found that that immunological ignorance of clonal neoantigens is the basis for ineffective T cell immunity to melanoma and that therapeutic vaccination as an adjunct to checkpoint inhibitor treatment, should be considered to increase the breadth and diversity of neoantigen-specific CD8+ T cells.
Antigen processing and presentation by HLA class II is a complex process and current in silico predictions are not able to account for all the factors involved in immunodominance/immunogenicity as they focus only the binding component of the process. Generation of large training datasets from single HLA class II allele cell-based or cell-free MAPPS-like platforms is enabling development of algorithms for identification of naturally processed ligands. The combination of binding and processing is a more reliable approach to the identification of viable CD4+ T cell epitopes, and to rank molecules. In silico prediction still needs to be part of a more complex immunognicity risk asessment strategy.
The immune response plays an important role in fighting cancer; however, the tumor environment is immunosuppressive and limits effective anti-tumor immunity. A new and promising strategy of tumor immunotherapy blocks pathways used by tumors to inhibit anti-tumor immunity. This inhibitory strategy is called checkpoint blockade. One key immunoinhibitory pathway that inhibits tumor specific immunity is the PD-1 co-inhibitory pathway. This pathway consists of the PD-1 receptor and its ligands PD-L1 (B7-H1) and PD-L2 (B7-DC). The PD-1 pathway plays critical roles in maintaining immune control, and is a key mediator of T cell dysfunction (“exhaustion”) in cancer and chronic infections. This pathway is a promising therapeutic target in cancer. The remarkable effects of PD-1 pathway blockade in cancer demonstrate the key role of this pathway in inhibiting anti-tumor immunity. However, there are multiple co-inhibitory pathways that that limit T cell function, and these have become targets for cancer therapy. This talk will discuss the multifaceted immunoregulatory roles of PD-1 and its ligands in controlling T cell activation, tolerance and exhaustion. The role of the PD-1 pathway in cancer as well as therapeutic strategies that combine PD-1 blockade with other therapies also will be discussed.
The gastrointestinal (GI) tract is home to trillions of commensal bacteria that play an important role in nutrition, immune system development and host defence. In inflammatory bowel disease (IBD), a chronic debilitating disease of the gastrointestinal tract, there is a breakdown in the healthy dialogue between our body and our microbial residents resulting in chronic immune attack in the bowel. In this presentation I will review key host and microbial pathways that maintain intestinal homeostasis and discuss how understanding these pathways may provide new therapies for the treatment of chronic inflammatory diseases.
Studies over the past 20 years have clearly defined and extensively characterized Foxp3+ T regulatory cells (Tregs) as major players in the control of most aspects of immune responses as well as playing a potential role in non-immunologic sites (fat, muscle). We are now at a point where studies have been/or will be initiated in the clinic using cellular biotherapy with Tregs, augmentation of Treg numbers/function with IL-2, and modulation of Treg function with small molecules and antibodies. Yet, many factors involved in Treg biology remain poorly studied. I will focus this talk on three key issues: 1) control of Treg homeostasis by TCR and co-stimulatory signals; 2) the role of transcription factors in modifying certain Treg functions; and 3) a critical review of suppressor mechanisms used by polyclonal and antigen-specific Tregs.
Dr. David Wraith, founder of Apitope, describes how peptide immunotherapies can induce antigen-specific desensitization. He describes how the REVEAL binding assay was used to identify pan-allele binding epitopes, which can servce as potential targets for tolerance induction.
Dr. Federico Mingozzi details the AAV-2 gene therapy model in Phase I/II trial hemophelia patients. He showed that a loss of human FIX expression was associated with expansion of capsid-specific CD8 T cells and demonstrated a dose-dependent T cell activity against the vector.
The immune response to protein-therapeutics (immunogenicity) is an important safety and efficacy concern during drug development and regulation. Non-clinical assays that can be used in the early stages of clinical development and to identify at-risk individuals and sub-populations in the clinic are an important unmet need. The so-called MHC Associated Peptide Proteomic (MAPPs) assay directly identifies peptides derived from a protein of interest on a donor’s MHC-II proteins. Here we have applied this technique to address several questions related to the use Factor VIII (FVIII) replacement therapy, in the treatment of hemophilia A. Over a dozen FVIII products are marketed but most fall into 3 categories: (i) Purified from human plasma (PD-FVIII). (ii) Full-length FVIII manufactured using recombinant DNA technology (FL-rFVIII). (iii) A truncated, beta-domain deleted rFVIII (BDD-rFVIII). We investigated whether there are differences in the FVIII peptides found on the MHC-II proteins of the same individual when incubated with products from the 3 classes. Our principal findings are as follows:
The number of unique FVIII peptides isolated, average length of peptides and range of peptide length were comparable for MHC proteins immunoprecipitated from cells from hemophilia A patients and healthy donors.
When cells from the same donor was exposed to FL-rFVIII in the presence and absence of PD-VWF, fewer peptides were recovered in the absence of PD-VWF; FVIII peptides not recovered in the presence of VWF map regions of VWF-FVIII interaction.
When dendritic cells from the same donor were incubated with FL-rFVIII or PD-FVII (both in the presence of PD-VWF); fewer FVIII peptides were recovered from the PD-FVIII. The peptides not recovered in the PD-FVIII did not map to the locations of FVIII-VWF interactions.
For each donor, FVIII peptides identified by the MAPPs assay exhibited affinities for that donor’s MHC variants which was orders of magnitude higher than the affinities of all overlapping peptides from FVIII and VWF, or when compared to a million random peptides obtained from the human proteome.
Zuben Sauna is a Principal Investigator and a Reviewer at the US Food and Drug Administration. His research interests lie in understanding the pharmacogenetic basis of the immune response to proteins used in therapeutic interventions as these affect efficacy and safety. His laboratory exploits a combination of computational, in vitro and ex vivo approaches to understand why some individuals and/or sub-populations develop immune responses while others do not. His work has published extensively in high impact journals such as Nature Biotechnology, Nature Medicine, Science, Science Translational Medicine and Nature Reviews Genetics. He received his Ph.D. from Poona University, India with subsequent training at the National Cancer Institute, Bethesda, USA.
Dr. Herman Waldmann reviews the mechanisms of tolerance induction and discusses strategies to evoke tolerance to immunogenic antibodies.
Accumulated evidence gathered in the last three decades demonstrated that some members of the Parvoviridae family, belonging in particular to the species Rodent protoparvovirus, have natural antineoplastic activity in cell culture and animal models, and that human tumor cells can be targets for this activity. In an immunocompetent rat glioma model, the H-1 parvovirus (H-1PV) was able to efficiently cure gliomas while raising an antitumor memory immune response. This oncosuppressive effect was observed after virus administration through intratumoral, intravenous and intranasal routes, and appeared to rely on both the direct oncolytic activity of H-1PV, and its handover to the host immune system. The parvovirus was also capable of inducing the regression of human glioma xenografts in immunodeficient rats. These studies have laid the foundations for the launch of a first phase I/IIa clinical trial in which H-1PV is currently undergoing evaluation for its safety and first signs of efficacy in patients with recurrent resectable glioblastoma multiforme progressing in spite of conventional therapies (ParvOryx01 trial). While no clinical conclusions, besides safety, can be drawn at this stage from the trial, there are intriguing laboratory measurements in resected tumors and patients’ blood. Immunohistochemical and FISH analyses provided evidence for virus efficiency in crossing the blood-brain barrier, spreading throughout the tumor and getting expressed in tumor tissues. The possible impact of these direct viral effects on the tumor fate has to be balanced against evidence of immune infiltration and vascular disruption in infected tumors, and of systemic immune responses in treated patients.
Bernard Maillère is currently director of research and head of the laboratory of immunochemistry of the cellular immune response at Commissariat à l'Energie Atomique (CEA), a French Governmental Research organism. His works mainly deal with the prediction of immunogenicity and identification of T cell epitopes in humans. Bernard is currently workpackage leader of the European IMI project ABIRISK dedicated to immunogenicity of therapeutic proteins
Integrated platforms can be used to mitigate immunogenicity risk and characterize immune responses during the drug design and development stages. ProImmune offers mutational activity mapping for optimal protein design, DC-T/T cell proliferation assays for biologic lead selection/optimization, a Mass Spectrometry assay for characterization of antigen presentation; HLA-peptide binding assays to characterize individual epitopes & undiluted whole blood cytokine storm assays.
Dr. Butler is Assistant Professor at the Princess Margaret Cancer Centre Tumor Immunotherapy Programme where is he is the Clinical Head of the Immune Monitoring Team. Before joining he was an Instructor in Medicine at Harvard Medical School and Clinical Fellow in Medicine at the Dana Farber Cancer Institute. Dr. Butler’s expertise includes clinical trials in a variety of immune therapies including adoptive T cell therapy and checkpoint blockade.
Immunogenicity of biologics in the clinic has the potential to cause severer sequela and alter or neutralize efficacy resulting in exposing patients to risk and terminated development programs. Pre-clinical risk assessment utilizing orthogonal methods has made it possible to prioritize drug candidates with a decreased immunogenicity risk or utilize protein engineering o reduce immunogenicity risk. Here we will present the immunogenicity tool box utilized at Bristol Myers Squibb to pre-clinically minimize immunogenicity risk.
Jochem Gokemeijer has been working at Bristol Myers Squibb for 15 years in Immunogenicity and biologics drug discovery. Before that he was involved in two alternative scaffold biotech companies Phylos and Adnexus Therapeutics. Jochem did his undergraduate work in the Netherlands and graduate work at the Dana Farber Cancer Institute.
Dr. Paul Moss describes the CD8 and CD4 T cell immune responses to CMV infection. He describes the importance of Class II Tetramers to better understand the interplay of a chronic infection and immune control.
Dr. Emilee Knowlton of ProImmune describes the tools for epitope discovery and immune monitoring, providing case studies and considerations for both pre-clinical and clinical
Competition for limited, cell extrinsic survival factors is a general feature of the peripheral selection checkpoints involved in B lymphocyte maturation, activation and memory. Perhaps the best-characterized example involves BLyS family cytokines and receptors, which governs survival and differentiation within B cell subsets. Discovery of the BLyS cytokine and receptor family has proven a watershed event, significantly advancing our understanding of B lymphocyte selection and homeostasis. This family includes two ligands, BLyS (also termed BAFF) and APRIL; as well as three receptors, BR3 (also termed BAFFr), TACI, and BCMA. Members of this molecular family play critical roles in maintaining immunological tolerance, by impacting selection and survival in nearly all B cell subsets. Cells in the transitional and mature, pre-immune B lineage subsets rely on BLyS signals via the BR3 receptor for survival. In contrast to tolerogenic elimination at the BM immature B cell stage, the transitional checkpoint displays plasticity by integrating BCR-mediated selection with BLyS-mediated peripheral B cell homeostasis. Thus, when BLyS levels are elevated, transitional selection is “relaxed,” so that B cells that would normally be negatively selected instead survive to join mature naïve pools. Mounting, evidence suggests analogous competitive checkpoints for both germinal center B cells and plasma cells. In contrast to pre-immune pools, B cells in recently activated and antigen-experienced subsets shift their BLyS receptor profiles and hence their ligand reliance. For example, short-lived antibody-forming cells adopt a TACI dominated BLyS receptor signature, whereas germinal center (GC) B cells profoundly down-regulate TACI but retain BR3. The lack of TACI on GC B cells leads to a paucity of retained BLyS in the GC, such that the sole local source of BLyS in the GC is the TFH cell. Moreover, BLyS expression by GC TFH is crucial for appropriate GC evolution, since efficient affinity maturation fails if T cells are BLyS deficient. Finally, long-lived plasma cell pools express BCMA, shifting reliance to APRIL. Considered together, these observations suggest that deliberately altering BLyS levels might be a means for manipulating B cell repertoire selection in order to (1) restore self-tolerance in autoimmunity, (2) remodel the repertoire to accommodate neo-self antigens introduced through biologicals, transplantation and gene therapy, or (3) temporarily expand B cell repertoire diversity to reveal novel, therapeutically useful specificities.
The central role of T cell responses to clinical successes in immuno-oncology has been well appreciated, yet few technologies have been designed to exclusively generate effective and sustained T cell responses in vivo. The DPX platform is a unique oil based formulation that entraps its components and facilitates active uptake by antigen presenting cells at the site of injection. This unique presentation to the immune system results in robust and sustained T cell responses. The lead clinical product, DPX-Survivac, employs the DPX platform to induce T cells targeting the shared tumor associated antigen survivin, which is overexpressed in many human malignancies. DPX-Survivac has been demonstrated to induce high levels of circulating, polyfunctional T cell responses to survivin in clinical studies of advanced ovarian cancer patients, and some of these responses have been associated with an ability to mediate tumour regression. Preclinical and clinical data have supported combining this T cell targeting treatment with other immuno-oncology agents, such as checkpoint inhibitors.
Dr Lea Bartsch from Massachusetts General Hospital discusses her work with HBV, in trying to better the understanding of the immunological alterations in order to improve treatment strategies for HBV infection. She highlights the use of ProImmune’s custom ProT2 Class II Tetramers in tracking HBV antigen specific CD4 T cells to validate new HBV epitopes and analyze the different CD4 T cell responses in chronic vs. acute infections.
Dr. Kellie Smith from Johns Hopkins, discusses her group's global study of the associations of neoantigen specific T cell responses with clinical outcomes to checkpoint blockades in cancer. The talk outlines the use of their MANAFEST T cell receptor sequencing technology, and by sequencing the CDR3 region, they are able to track clones that recognise the same antigen of interest. Showing that with lower clonality there is an association with higher residual tumour burden in patients. Pro5 Pentamers are also shown being used as an accurate way to validate their findings, by using them to track the neoantigen specific CD8 T cells. She also highlights the use of single cell RNA-seq in determining the phenotype of the clones in responding and non-responding patients.
Professor David Withers from the University of Birmingham discusses his recent work in tracking dynamic changes of lymphocytes within tumours, to understand and characterise the phenotype of cells entering, leaving and being retained in the tumour through the use of the transgenic Kaede Photoconvertible mouse model. He also discusses the use of Single Cell RNA-seq to capture the heterogeneity within the tumour infiltrating lymphocyte population, and describing the use of ProImmune Pro5® Pentamers to track tumour neo-antigen specific CD8 T cells.
Hilario Ramos, Molecular Templates, discusses the use of Egineer Toxin Bodies (EBTs) – molecules with an antibody domain for targeting that is genetically fused to a de-immunized Shiga-like toxin A subunit which drives apoptotic cell death. These molecules can drive cell death by apoptosis, and Hilario discusses novel uses for EBTs of delivering antigenic peptides to tumours, in a novel way to make the tumour cells visible to the immune system. Hilario highlights the use of ProImmune's Pro5® MHC Class I Pentamers in tracking Antigen Specific CMV CD8 T cells in order to help build their CTL models.
Dr. Yeojun Yun discusses Synteka Bio's NEOscan™, a neoantigen prediction algorithm used to improve the selection and prioritization of neoantigens for inclusion in cancer therapies.
Christine Falk graduated in Biology at the Ludwig-Maximilians-University in Munich and performed her PhD thesis at the Institute for Immunology at LMU Munich (director G. Riethmüller) in the laboratory of D.J. Schendel on autoimmunity and the regulation of NK-like T cells and NK cells. As a postdoctoral fellow at the new Institute of Molecular Immunology (director D.J. Schendel) at the Helmholtz Center for Environment and Health (GMGU), she studied recognition of virus-infected cells and tumor cells by T and NK cells. As senior postdoctoral fellow and head of the research group “Immune Monitoring” at the German Cancer Research Center DKFZ in Heidelberg from 2006 until 2010, she was working on the microenviroment of solid tumors and its impact on tumor recognition by T and NK cells. In her current position since 2010, she is head of the Institute of Transplant Immunology, IFB-Tx, at Hannover Medical School MHH where she has established an immune monitoring laboratory investigating adaptive and innate immune responses in the context of stem cell as well as solid organ transplantation, in particular lung, heart, liver and kidney transplantation. The addition of transplant immunology to her former field of tumor immunology is reflected by focusing on common immunological mechanisms for rejection of both tumors and allografts that may open new perspectives for immunological interventions to achieve tumor rejection on one hand and on the other hand, immunological tolerance, the ultimate goal of solid organ transplantation
In cancer cells, the disruption of normal biochemical signaling results in aberrant post-translational modifications, including protein phosphorylation. These mis-phosphorylated proteins can be processed into antigenic peptides by the cellular machinery, resulting in presentation of this novel type of antigen on MHC molecules. PhosphoSynVax is Agenus’s third heat-shock protein-based vaccine platform. PhosphoSynVax is designed to induce immunity against this novel class of phospho-neo-epitopes specific to malignant cells of multiple donors in various cancer types. Based on mass-spectrometry and bio-informatic approaches, we have a library of proprietary phosphopeptide tumor target antigens from different cancers, including lung cancer, specific leukemias, ovarian cancer, colon cancer and others. This approach may enhance immune system recognition of phospho-neo-epitopes, leading to the destruction of cancer cells in various patients across cancer types.
The ability to clear many intracellular infections is likely to require a consorted immune response that includes CD8+ T cells. Although a large number of approaches have been taken to generating such responses in humans, achieving the magnitude and functionality of natural viral infections has been difficult. The most promising approach published to date is to employ a heterologous prime-boost, usually using two replicating or non-replicating viral vectors. The Jenner Institute has extensively used a replication-deficient adenovirus followed by a replication-deficient pox virus, and achieve ELISPOT levels of balanced CD4+ and CD8+ T cells in the many thousands of spot forming units per million PBMCs. Vaccitech has in-licensed programs based on the use of this strategy to control chronic HBV and chronic HPV. The progress of these programs will be discussed.
Jonathan Silk is the Head of Cell Research at Adaptimmune (www.adaptimmune.com), a biotechnology company located in Abingdon and Stevenage, UK, and Philadelphia, PA, USA. Adaptimmune is a world leader in the TCR T-cell therapy space, utilizing engineered Specific Enhanced Affinity T-cell receptor-expressing T-cells (SPEAR T-cells) to target both solid and hematologic cancer types. Jonathan and his team are investigating Next Generation SPEAR T-cells, to improve the function of T-cells for adoptive T-cell therapy, including overcoming immune-resistance mechanisms. Jonathan received his BSc in Biochemistry from the University of Kent at Canterbury (UK), with an industrial placement at Merck Sharp and Dohme (Harlow, UK). He obtained a PhD in Immunology studying at the MRC Clinical Sciences Centre (Imperial College London, UK). Subsequently he worked with Professor Vincenzo Cerundolo at the University of Oxford, UK, as a Post-doctoral Scientist, developing a number of cancer immunotherapy research programmes including the use of ligands for invariant NKT-cells as adjuvants and investigating mechanisms of tumour escape.
Rodd Polsky is an Investigator responsible for immunogenicity assay development within the Bioanalysis, Immunogenicity and Biomarkers organization at GlaxoSmithKline. Rodd received his Bachelor’s degree in Biology from Drexel University. Since that time, Rodd has spent the past 25 years providing preclinical and clinical immunogenicity support, including assay and study design, development, validation, and study support for the detection of anti-drug antibodies against therapeutic proteins.
Dr. Aaron Mansfield of Mayo Clinic Cancer Center discusses the current immunotherapy treatments for patients with mesothelioma, the relatively low tumour mutation burden in those patients and alternatives to non-synonymous point mutations as sources of neo-anitgens. The use of ProImmune REVEAL technology allowed him to compare the MHC binding scores of selected peptides from chromosomal rearrangements, of usually non-coding DNA to discover if any of these peptides were neo-antigenic.
Karen discusses how to measure antigenic diversity of B and T cell immune responses and how the structural components of those antigens drive immunogenicity. She describes technologies such as custom protein-peptide microarrays to detect antibodies; a mass spec based program called MHC TreASUre Hunt for immune peptidome discovery; and Molecular Dynamic Modeling to model peptide MHC interactions based on existing crystal structures to help develop computational systems to predict TCR.
Vibha discusses a proactive risk based approach to assessing immunogenicity in early development. She describes a suite of in silico risk assessment tools in combination with in vitro methods to evaluate immunogenicity or product attributes and how, when used in combination, those methods can reduce false positive identification. The stages of when to implement and report the data from the risk assessment tools is described as well as the current challenges the FDA reviewers face when evaluating the data package.
Rao describes Multiple Myeloma, a plasma cell cancer and the biomarkers that can be used to classify and monitor the immune response through the progression of the disease. They Harvard group looked at myeloid derived suppressor cells, plasmacytoid DC, T-regulatory cells, TH17 cells and B cell subpopulations in the interrogation of the cancer. There was a humoral immunity deficiency, increased PD1 expression on effector cells and increased levels of PDL1 expression detected in the multiple myeloma patients.
Paul discusses the challenges facing the identification and diagnosis of Lyme disease. He describes the life cycle stages of the carrier, Borrelia burgdorferi sensu lato species, as well as the incidence and transmission of Lyme disease in the United States. There are differential expressions of antigenic surface proteins that occur due to bacterial transcriptional changes and throughout the course of the disease. Highly reactive serum was screened for responses against 21 different proteins using 80 overlapping peptides in the ProArray Ultra linear B cell epitope mapping tool to improve the specificity and accuracy of current diagnostic methods.
Patrick describes the Children’s National cellular therapy program for treating viral infections following transplant surgeries. He discussed the generation of donor-derived virus specific T cells from cord using either an adenoviral vector or peptide mixes and the recognition of atypical epitopes by these cells. The team has treated over 80 patients with virus specific T cells with between 70-80% response rate.
Yanchun Peng is a senior Postdoctoral researcher working at the Human Immunology Unit, Weatherall Institute of Molecular Medicine, University of Oxford. After completing her undergraduate studies and then teaching in China, she moved the UK, first working as a research assistant, then completing her DPhil and remaining as a Postdoc in the research group led by Prof. Tao Dong since 2005. She has acquired extensive experience and skills in human T cell immunology with the major focus of her research being on evaluating anti-viral efficacy of virus-specific T cells and the interaction of host and virus in HIV infection, HIV/HCV co-infection and influenza infection. Her recent interest is to study the Viral OncoProtein(VOP) and tumor Specific Protein(TSP) specific T cell responses in virus associated cancer (ie HBV/HCC; EBV/NPC and HPV/CC) by combining cutting edge technologies (such as multi-colour Flow cytometry, Mass Cytometry (CyTOF), single cell RNASeq with SmartSeq 2 and 10X Chromium) with my expertise in isolating, expanding and characterising virus/tumor-specific T cells in vitro. Being equipped with these well-established platforms, skills and knowledge in human T cell immunology, she swiftly switched my work from cancer to COVID-19, as the pandemic started in 2020: exploring the role of SARS-CoV-2 -specific T cells in COVID-19 infection.
Dr Amy Flaxman is part of the Oxford Vaccine Group, Jenner Institute, University of Oxford. For her post-doctoral research she has worked on both pre-clinical studies developing vaccines for outbreak pathogens, such as Ebola, and clinical trials for outbreak pathogens, including MERS and Ebola. She is currently investigating immune responses to the Oxford ChAdOx1-nCoV vaccine for SARS-CoV-2 that was in Phase III clinical trials but is now approved. She leads the lab team carrying out antibody testing post-vaccination. Here, she is interested in the differences in antibody responses induced over time with different doses of vaccine, in different age groups and how this changes with administration of a booster vaccine.
Since the early days of the development of the CRISPR Cas9 gene editing technology there have been sporadic speculations that immune responses to Cas9 (which is of microbial origin) could have consequences for clinical applications. Since late last year, three studies have presented experimental evidence for T- and B-cell responses to Cas9. These results have prompted considerable commentary, in both the scientific and popular press, marking Cas9-immunogenicity as a major obstacle to bringing CRISPR genome-editing to the clinic. The studies on Cas9 immunogenicity have brought attention to a critical element in the eventual clinical application of Cas9-mediated gene editing. In this presentation I will provide results of recent studies using a broad array of assays including ELISAs to detect anti-Cas9 antibodies, T-cell proliferation assays to identify immune-hot spots on Cas9 and MHC Associated Peptide Proteomics. The experimental data demonstrate the need for fit-for-purpose assays, statistical methods and reagents that will be necessary for obtaining data that is sufficiently reliable to make decisions for drug development and licensure. The broad outline of my presentation is as follow:
• Unique challenges while assessing the immunogenicity risk to Cas9
• The need of validated reproducible assays that are fit-for-purpose
• The need for well characterized reagents and for controlling contaminants and impurities
• Model systems for evaluating the immunogenicity of Cas9
Sequence variations in CMV UL40-derived peptides control NK activation through HLA-E presentation.
Oesophageal cancer (OC) causes over 0.5 million deaths worldwide each year. Current therapeutic regimens focus on chemo-radiotherapy prior to surgery, however, only 20-30% of patients respond to treatment. Therefore, new treatments are badly needed. Natural killer T (NKT) cells are innate T cells that recognize lipid antigens presented by CD1d molecules. We enumerated circulating NKT cells in OC patients and control subjects by flow cytometry using monoclonal antibodies and CD1d tetramers loaded with glycolipid antigens.
This allowed for the detection of invariant NKT cells, which possess invariant T cell receptor (TCR) α-chains reactive with α-galactosylceramide and mediate anti-tumour immunity in mice and humans, as well as novel NKT cells that recognise other glycolipids. We found that invariant NKT cells were depleted and functionally-impaired in patients with OC. In contrast, NKT cells reactive against sulfatide and tetramyristoyl-cardiolipin (TCL) were present in significantly higher numbers in OC patients compared to controls. Most of these cells expressed γδ TCRs, specifically the Vδ1 subset. TCL induced the production of transforming growth factor-β by expanded Vδ1 T cells in vitro in a CD1d dependant manner. Supernatants of Vδ1 T cells activated with sulfatide or TCL inhibited degranulation by invariant NKT cells. These data suggest that invariant NKT cells protect against OC but other CD1d-restricted NKT cells may promote tumorigenesis in these patients. These findings have important implications for cellular therapies involving invariant NKT cells, which are currently being tested in clinical trials for multiple cancer types.
For forty years, I’ve been researching therapeutic antibodies with a particular interest in effector functions of the Fc region. With Herman Waldmann and Greg Winter, I took several of the early engineered antibodies into the clinic. We realised that activation of immune effector systems is not always desirable, but found that mutations designed to reduce binding to Fc receptors (e.g. ‘LALA’, aglycosyl) were not completely effective. At mAbsolve we have screened hundreds of new variants and identified some which are completely devoid of binding to Fc receptors.
Receptor binding is one thing, but cellular activity is more important. The modest binding levels of previous variants translated to quite substantial cellular activity, but our new variants remained completely negative. The risk of unwanted immunogenicity is always a concern when proteins are modified, but the variants gave no increased risk signals either in silico or in vitro.
The optimised Fc variant is comparable with wild type IgG for binding to FcRn, stability and manufacturability and is available for licensing.
Katie Ewer is Associate Professor and Senior Immunologist at the Jenner Institute, University of Oxford where she leads a program of exploratory immunology studying vaccine-induced immunity to malaria and outbreak pathogens, such as Ebola, MERS and now SARS-CoV-2. A key aim of her research is focused on characterizing T cell responses induced by vaccination with viral vectors. During the COVID-19 pandemic, she played a pivotal role in the clinical trials in an effort to find a vaccine, which has led to the successful roll-out of the first approved SARS-CoV-2 vaccine, ChAdOx1 in partnership with AstraZeneca in December 2020. In this talk she explains how the vaccine was developed in record-breaking time, the characterisation of the immune responses to the vaccine and the forthcoming challenges still faced including the question of booster doses, manufacturing and supply and how we respond to the next inevitable pandemic.
In this talk we learn how quantity and early induction of virus-sepcfici T cells is directly associated with mild disease; how higher functionality of SARS-CoV-2 specific T cells is found in assymptomatic cases of SARS-CoV-2 infection and how how early detection of SARS-CoV-2 specific T cells correlates with the the onset of vaccine efficacy.
Nina Le Bert is currently a Senior Research Fellow in the Emerging Infectious Diseases Programme of DUKE-NUS Medical School in Singapore. She earned her Master of Science degree at the Free University of Berlin in Germany, and her PhD from the University College London UK in 2011, working in the field of human innate and adaptive immunity. In 2014, she joined Professor Antonio Bertoletti’s lab, where she is investigating the virus-specific B and T cell responses during Hepatitis B Virus (HBV) infection to better understand their role in protection and/or liver pathology. Since the outbreak of the COVID19 pandemic, she is involved in studies of SARS-CoV-2 specific T cells.
In this talk we learn that SARS-CoV-2-specific PD-1+CD8+ T cells are not exhausted, but functional; that they are sustained for at least 8 months, as well as learning about the development of SARS-CoV-2-specific stem cell-like memory T cells (TSCM).
Eui-Cheol Shin received his M.D. (1996) and Ph.D. (2001) from Yonsei University, Seoul, Korea, and his postdoctoral training from National Institutes of Health, Bethesda, Maryland, USA. Then he joined Graduate School of Medical Science and Engineering, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Korea in 2007, where he is currently a professor. His laboratory, the Laboratory of Immunology and Infectious Diseases, focuses on the research of T cell responses in human viral disease and cancer.
In this presentation, Dr. Sette provides evidence that there are robust CD4+ and CD8+ T cell responses detected in uncomplicated SARS-CoV-2 convalescent cases; that reactivity is reproducibly detected in non-exposed subjects; specific CD4 and CD8 T cells targets in COVID-19 patients were identified; in acute and severe infection it appears that the speed of the adaptive response is key to protective immunity; that T cell responses are durable over at least 8 months and that there is negligible impact of SARS-CoV-2 variants on T cell responses.
Alessandro Sette has devoted more than 35 years in biotech and academia to understanding and measuring immune responses, and developing disease intervention strategies against cancer, autoimmunity, allergy, and infectious diseases. Dr. Sette’s laboratory is the world leader in the study of the specific structures, called epitopes, that the immune system recognizes. Dr. Sette has overseen the design and curation efforts of the national Immune Epitope Database (IEDB), a freely available, widely used bioinformatics resource. The IEDB catalogs all epitopes for humans and experimental animals for allergens, infectious diseases, autoantigens and transplants, and includes epitope prediction tools to accelerate immunology research around the world. Dr. Sette’s lab uses knowledge of epitopes to define the hallmarks of a beneficial immune response associated with effective vaccines, as opposed to immune responses that are ineffective or that cause harm. The laboratory’s infectious disease interests include SARS CoV2, dengue, Zika Chikungunya, herpesviruses, poxviruses, lassa fever, HIV and hepatitis viruses, and bacterial pathogens such as tuberculosis and bordetella pertussis. Our investigations outside infectious disease include allergic asthma and Parkinson’s disease.
Dr. Sette is a Doctor in Biological Sciences from the University of Rome and did postdoctoral work at the National Jewish Center for Immunology and Respiratory Medicine in Denver, Colorado. In 1988, Dr. Sette joined the newly founded company Cytel, in La Jolla, and was also appointed adjunct assistant professor at The Scripps Research Institute. He founded Epimmune in 1997, where he served both as Vice President of Research and Chief Scientific Officer until 2002, when he joined LJI as Head of the Division of Vaccine Discovery. He also heads the Center for Infectious Disease at LJI.
Questions addressed by the expert panelists include:
1. What is the potential effectiveness of booster vaccine doses to vunlerable populations, particularly in light of new VoCs.
2. Should we be using modified vaccines? Do we think that modified vaccines including mutations to cover VoCs are useful given the potency of T cell reponses that see against known VoCs. What about vaccine combinations?
3. Should we be vaccinating children when susceptible populations across the globe haven't yet got access to vaccines?
4. What is the future of the pandemic?
Our sincere thanks to the expert panel for their contribution:
Prof. Alessandro Sette (La Jolla Institute for Immunology, USA)
Prof. Antonio Bertoletti (Duke-NUS Medical School, Singapore)
Dr. Kate Broderick (Inovio, USA)
Questions addressed by the expert panelists include:
1. As viral immunologists you have all rapidly moved from your own area of expertise to characterising the immune response to SARS-CoV-2. What has most surprised and intrigued you in the study of the virus?
2. If SARS-CoV-2 variants totally evade vaccine-induced neutralising antibodies, can vaccine-induced memory T cells still contribute to host protection?
3. What insights do you have regarding the immune response in the context of re-infection of the same or different variants?
Our sincere thanks to the expert panel for their contribution:
Prof. Eui-Cheol Shin (KAIST, Korea)
Dr. Nina Le Bert (Duke-NUS Medical School, Singapore)
Prof. Tao Dong (University of Oxford, UK)
Dr. Yanchun Peng (University of Oxford, UK)
Dr Jeremy Fry provides a summary of ProImmune’s REVEAL and ProVE® service, the rapid epitope discovery system.
Dr Andrew Waller provides an overview of the Pro5 Pentamer technology and the benefits of using MHC pentamers to detect specific CD8 T Cell populations in your research.
Dr Heather Hensler introduces ProArray Ultra®, the high-throughput peptide and protein microarray assay service.
Dr Briana Betz explains the importance of understanding the HLA types of your study cohort, and how ProImmune can help you.
Dr Nikolai Schwabe introduces the ProPresent antigen presentation assay and some of its applications in the field of immunogenicity.
Managing immunogenicity risk should be a core focus in drug development. ProImmune’s CEO Nik Schwabe explains why.
thinkpeptides Peptide Ebook
Chulalongkorn University, Thailand
"I outsourced my tissue typing to ProImmune because they provide
me with the fastest service at the best price. Since the main
theme of my research is on T cell immunology, HLA-type is
inevitably needed. The customer service from ProImmune is of the
best quality. Their typing service allowed me to unambiguously
resolve the HLA type of many of my samples."
Duke Human Vaccine Institute, North Carolina, USA
"We have a programme of clinical studies and it is important for
our researchers to be able to correlate the immune responses
they observe with HLA type. We looked at a range of companies
offering tissue typing, and also at using an on-campus typing
facility, and we found that ProImmune were able to offer us the
best pricing and turnaround time. We now send them regular
batches of samples to type. They have been wonderfully easy to
work with, if we ask anything they get right back to us and
overall it has been a really good experience."
New York Medical College, New York, USA
Prof. Dattwyler used ProImmune's B cell Linear Epitope Mapping
Service to discover the linear epitopes present in two different
proteins. "I was really happy with the way that these guys took
the trouble to adjust my quote so that I had the best assay
design for the lowest price. They were really easy to work with,
always replied to my questions, and delivered the report on my
project on schedule as promised. It's great that I've been able
to go back to them and order my new epitopes as custom peptides
for follow on work, and I'm going to revisit my validated
epitopes with ProImmune's help and get some really fine mapping
of antibody binding to these sequences."
ProImmune REVEAL & ProVE® Epitope Discovery
HLA Tissue Typing
ProMap® T Cell Proliferation Assay
Pro5® MHC Class I Pentamers