INsTRuCT

INsTRuCT ESR Projects

Host Institution: Trinity College Dublin, Ireland
Primary Supervisor: Dr. ZASLONA, Zbignew

Introduction: After emigrating from the blood to tissues, monocytes continue their maturation and undergo final differentiation into long-lived tissue-specific macrophages. Immune-metabolites, such as succinate and itaconate, markedly influence monocyte-to-macrophage transition and functional polarisation; however, we do not fully understand the mechanistic links between these metabolites and macrophage differentiation. A deeper appreciation of metabolic control of macrophage polarisation will lead to superior MRC-based therapies.

Aims: A1: We will assess how immunometabolites affect monocyte-to-macrophage differentiation using in vitro systems; A2: We will study the effects of immune-metabolites on monocyte recruitment and differentiation into macrophages in a mouse model of LPS-induced peritonitis. A3: In an OVA-induced murine asthma model, macrophages accumulate by local proliferation. We will explore how this process is affected by succinate and itaconate.

Expected Results: R1: The role of succinate and itaconate in regulation of monocyte differentiation and macrophage maturation will be eludicated. R2: We will define mechanisms by which these immunometabolites affect the process of monocyte recruitment and macrophage proliferation in vivo. Since succinate and itaconate production is altered by disease states, and because maturation status influences macrophage functions, our findings will have broad clinical implications.

Planned secondment: (1) An international 3-month secondment to Regensburg, Germany is planned as a networking, technical and scientific exchange. (2) An international, intersectoral 3-month secondment to GSK is planned, giving the ESR an opportunity to work with diseased human material.

Enrolment in Doctoral degree(s): Trinity College Dublin

Host Institution: Charité University Hospital, Berlin, Germany
Primary Supervisor: Prof. SAWITZKI, Birgit

Introduction: Recent evidence indicates that mitochondrial function controls macrophage and DC activity. By regulating substrates for histone acetylation and methylation, cellular metabolism controls transcription through epigenetic mechanisms. Mitochondrial dynamics control metabolic programming, hence immune cell differentiation. We hypothesise that mitochondrial fission underlies the shift towards glycolysis and inflammatory phenotype in DCs and macrophages, whereas enforced mitochondrial fusion maintains a stable regulatory phenotype. Our work suggests innovative approaches for developing more stable MRC that could be used as safer or more potent cell-based immunotherapies.

Aims: A1: To study mitochondrial dynamics, function and cellular metabolism in various MRC populations, including DC10, Tol-DC, ATDC and Mregs. A2: To explore the possibility of interfering with proteins regulating mitochondrial dynamics to generate more stable MRC.

Expected Results: R1: Characterising mitochondrial functions and cellular metabolism of different MRC subtypes will lead to a more complete scientific understanding of their interrelationships; R2: This work will determine whether a stable, maturation-resistant phenotype in MRC is dependent upon fusion-like morphology of mitochondria and their oxidative phosphorylation activity; R3: Small molecule inhibitors and genetic engineering will be used to reinforce mitochondrial fusion in MRC products; R4: As a pharmacodynamic indicator, immune monitoring methods will be implemented to analyse mitochondrial morphology and functions following in vivo transfer of MRC.

Planned secondment: An international, intersectoral 3-month secondment to Sanofi is planned to support preclinical development of gene therapy approaches to stabilising MRC function..

Enrolment in Doctoral degree(s): Charité University Hospital, Berlin

Host Institution: Ospedale San Raffaele (OSR)
Primary Supervisor: Dr. Silvia Gregori

Planned secondment: An international, intersectoral 3-month secondment to Sanofi is planned to support preclinical development of gene therapy approaches to stabilising MRC function.

Enrolment in Doctoral degree(s): Charité University Hospital, Berlin

Host Institution: Instituto de Salud, Carlos III, Madrid, Spain
Primary Supervisor: Prof. OCHANDO, Jordi

Introduction: The innate immune system displays long-term changes in its functional capacities through epigenetic programming of macrophages and monocytes, a phenomenon known as ‘trained immunity.’ Trained immunity is likely to be a critical issue in MRC-based therapy, affecting the quality of therapeutic cell products, their functional stability and their effects in patients. This project concerns an innovative treatment paradigm in which innate immune cells are specifically targeted and “untrained” using drug-loaded nanobiologics. A trained immunity-inhibiting nanobiologic library of high-density lipoprotein (HDL) derived nanomaterials with different immunomodulatory payloads will be created using advanced microfluidics technology. These highly innovation nanobiologics will complement future cell-based therapeutic approaches, both in terms of producing superior MRC-based products and co-treatment of patients receiving MRC-based therapies.

Aims: A1: To develop nanobiologic library technology to regulate trained immunity; A2: To evaluate the robustness of our trained immunity-inhibitory nanobiologics to prevent organ transplant rejection; A3: To test trained immunity-inhibitory nanobiologics as substances added to cell cultures during manufacture of MRC-based therapeutic products.

Expected Results: R1: Development of optimized nanoformulations that target different molecular pathways associated with trained immunity. This program’s successful completion will yield targeted immunotherapies not only to potentially treat transplant patients, but also autoimmune disorders and allergies.

Planned secondment: An international, intersectoral 6-month secondment to the non-academic group of Dr. Gregori (OSR, M22) is planned to support preclinical development of gene therapy approaches to stabilising MRC function.

Enrolment in Doctoral degree(s): Complutense University of Madrid

Host Institution: Newcastle University, United Kingdom
Supervisors: Dr. LORD, Phillip (primary) - Dr. HILKENS, Catharien

Introduction: Basic and translational research in immunoregulatory cell-based therapy has been limited by the intrinsically small-scale of clinical trials, which hampers comparison, standardization and reproducibility. Our group has spearheaded initiatives to define reporting guidelines for cell-based therapies, which serve as a foundation for further standardization efforts. This project aims to combine data from consortium partners with publically available knowledge two achieve two main goals: firstly, to unify descriptions of the pharmacological properties of tolerogenic cell productsin order to support standardisation of their quality-control; secondly, to predict the immune regulatory effects of tolerogenic cell products, which will inform the design of standardised immunomonitoring in clinical trials.

Aims: A1: To build a warehouse of publically available data on tolerogenic cell therapies and incorporating relevant background resources, including pathway, cell development and pharmacological data sets. A2: To embed good data management practice, enabling reproducibility and data sharing as the default behaviour within the consortium. A3: To enable semantic descriptions to cell products and immunomonitoring read-outs. A4: To predict optimal strategies for quality-control and immunomonitoring by using a combination of semantic similarity, rule-based learning and other computational learning techniques.

Expected Results: This research provides an innovative solution for predicting the immune regulatory effects of tolerogenic MRC-based products by creating an integrated, big data set scale through standardisation and data warehousing. R1: The proposed ‘data warehouse’ will represent a major scientific output of this project, which will be an extremely value resource for INsTRuCT partners and the wider field. R2: Development of recommendations about quality-control testing of MRC products; R3: Development of recommendations about the optimal conditions for ex vivo generation of MRC products.

Planned secondment: (1) 3-month secondment to SciBite (Cambridge, UK, M18) to learn new tools for the curation and management of biomedical data. (2) An international, 2-month secondment to Prof. Nathalie Cools’ lab to learn about new GMP-compliant technologies and regulations for manufacturing of MRC-based products.

Enrolment in Doctoral degree(s): Newcastle University

Host Institution: University of Nantes, France
Primary Supervisor: Dr. MOREAU, Aurélie

Introduction: Autologous Tolerogenic Dendritic Cells (ATDC) exhibit tolerogenic properties in vitro and in vivo. Human ATDCs are currently being investigated in a Phase I/II clinical trial in kidney transplantation. ATDC produce high levels of lactate that steer T cell responses towards tolerance. This project concerns further investigation of molecular mechanisms of ATDC action using cutting-edge techniques in order to develop of innovative methods for quantifying therapeutic effects of ATDC in solid organ transplant recipients.

Aims: A1: To examine the phenotypic and functional heterogeneity of ATDC by single cell RNASeq and epigenetic profiling studies. A2: Using LV coding for shRNA, we will delete molecules thought to be relevant to the regulatory function or stability of ATDCs. Gene-edited ATDCs will then be examined in vitro and in vivo using established assays and animal models. A3: The utility of functionally important molecules as biomarkers of ATDC effects in vivo will be explored using archived blood samples from patients treated with ATDCs or other immune regulatory cells.

Expected Results: R1: Understanding of the molecular mechanisms of ATDC action, allowing us to systematically compare ATDC function with other MRC products. R2: Novel molecules contributing to ATDC action will be mechanistically investigated. R3: Innovative tests for ATDC pharmacodynamic effects will be developed, standardised and clinically validated.

Planned secondment: (1) An intersectoral secondment of 2-months to Immunotech which produces CE/IVD-marked reagents and assays. (2) A 2-month intersectoral secondment to Silvia Gregori’s lab is critical to aim A1.

Enrolment in Doctoral degree(s): University of Nantes

Host Institution: University of Regensburg, Germany
Supervisors: Prof. REHLI, Michael - Prof. SPANG, Rainer

Introduction: The distinctive tolerogenic signatures of MRC (as well as the signals required to induce and maintain them) are poorly understood, as are the fate and phenotypic stability of adoptively transferred cells in vivo. This predominantly computational project is concerned with improving our understanding of differentiation routes, transcriptional and epigenetic features of MRCs in vitro, as well as their adaptation in vivo to optimize tolerogenic cell products.

Aims: A1: To devise computational methodology for the quantification and molecular characterization of MRC subsets in transcriptome or epigenome profiles from bulk samples such as blood or tissue. We will build on established deconvolution methods such as Cyber Sort and our in-house adaptive deconvolution algorithm based on loss-function learning. The method will be applied to public and consortium data. A2: Modelling myeloid differentiation both in vitro and in vivo from transcriptome and epigenetic data to unravel regulatory networks, co-expression modules and corresponding pathways important for tolerogenicity of myeloid cell products. SB considers the implementation risk of this project is low because all required data are public or already shared.

Expected Results: R1a: Novel deconvolution algorithms for an improved quantification of regulatory immune cells in bulk samples will be developed and implemented. The method will be open source and freely available to the scientific community. R1b: Annotatation of existing bulk profile data sets from transcriptomics and epigenomics resources such as TCGA with computationally estimated immune cell composition. These will be made freely available in web applications and and in portable containerized data warehouses that we will develop using the in-house LYRA system. R2a: Identification of latent factors that drive myeloid differentiation in cell therapy settings. These factors will inform the deconvolution models and guide their learning phase. R2b: Identification of signaling and regulatory pathways controlling tolerogenic features of cell products and computational prediction of candidate drugs that may improve the phenotypic stability or tolerogenic functions.

Planned secondment: The ESR will undertake a 2-month secondment at Miltenyi Biotec to expand their knowledge of management and analysis of immune cell-related NGS data.

Enrolment in Doctoral degree(s): University of Regensburg

Host Institution: Cimaas BV, The Netherlands
Primary Supervisor: Prof. GERMERAAD, Wilfred

Introduction: This proposal has the central question of how we can optimize a dendritic cell (DC) vaccine for cancer. To be successful as a clinical product, DCs should be strong activators of tumor specific cytotoxic T lymphocytes (CTLs). This can be accomplished by appropriate polarization of T helper (T_h) cells and activation of natural killer (NK) cells, which in turn positively stimulate T_h cells and CTLs. Furthermore, a GMP compliant culture system is necessary (and available), including good quality control. In this project we aim to optimize the DCs and measure the T cell responses induced in vitro.

Aims: A1a: To measure intracellular/extracellular vitamin C and extracellular vitamin D followed by studying the role of vitamin C and vitamin D (tolerizing DC) on DC-T cell and DC-NK cell interactions. A2: To reduce the IL-10 production in DCs to promote better T_h1 polarization and prevent immune suppression. A3: To characterize the MHC class II peptide presentation profile by mRNA electroporated DC and determine whether MHC class II targeting improves T_h responses to generate better CTLs. Finally, the ESR will be trained in a GMP environment. This project exemplifies how INsTRuCT can accelerate innovation in MRC-based therapies through commercial adoption of basic scientific discoveries. CiMaas already exchanged know-how with SQ, UA, UHREG and OSR. SB has minimized project implementation risks through detailed planning of collaborations and ensuring sufficient redundancy in scientific deliverables.

Expected Results: R1: Whether vitamin C enhances DC-T cell/NK cell interactions. R2: Optimized DCs capable of generating potent T cells, better T_h1 polarization and lower immunosuppression. R3: Profiling of peptides in MHCII molecules expressed by mRNA electroporated DC.

Planned secondment: The ESR will undertake a 2-month secondment at UHREG to learn about immune monitoring. The ESR will undertake a 2-month secondment at Sanquin to master DC-CD4 T cell co-cultures and optimized release assays (i.e. MHCII multimer analyses) to develop antigen-specific release assays for electroporated DC.

Enrolment in Doctoral degree(s): University of Maastricht

Host Institution: King’s College London, United Kingdom
Supervisors: Prof. SANCHEZ FUEYO, Alberto - Prof. LOMBARDI, Giovanna

Introduction: We propose to create a highly personalised approach based on surrogate immunological biomarkers to assess tolerance induction in order to direct treatment with MRC-based therapies according to patients’ individual needs.

Aims: A1: To perform sequential high-throughput phenotypic analysis of peripheral blood leucocytes from liver transplant recipients performing immunosuppression withdrawal to achieve tolerance by CyTOF, RNASeq and epigenetic landscaping (LIFT trial, >100 patients enrolled and centralized by KCL). A2: To evaluate antigen-specific responses in the same patients using in vitro functional assays, as well as profiling TCR repertoires of Tregs and effector T cells. A3: To correlate the distribution of antigen-specific effector and regulatory T cells based on TCR sequencing with peptide-specific MHC II tetramer staining in flow cytometry. A4: To develop bioinformatic tools to integrate phenotypic, transcriptional and epigenetic patterns in order to evaluate the function and stability of mixed leucocyte populations. A5: To apply the designed tolerogenic signatures and specific immune cell characterization methods to evaluate and compare the immunological effects of MRC-based and Treg-based therapies in liver (ThRIL, LITE studies at our center) and kidney transplant recipients (archived samples from UHREG, UN-CRTI).

Expected Results: R1: The immunomonitoring techniques established and validated on clinical trials will provide new tools to study immunological mechanism of tolerance, monitor disease progression, evaluate therapeutic effect, stratify candidates for MRC-based immunotherapy, and serve as prognostic markers of clinical outcome. R2: The implementation of bioinformatic platforms to improve the identification and characterization of regulatory and effector cells in complex multicellular samples, such as peripheral blood or biopsies, will identify novel biomarkers that can be developed as innovative clinical assays.

Planned secondment: An intersectoral secondment of 5-months to UCB Pharma is planned to provide the student with experience in market-driven biomarker research and to pursue aims A1 and A2.

Host Institution: Sanquin Blood Supply and University of Amsterdam, The Netherlands
Supervisors: Prof. ten BRINKE, Anja - Prof. van HAM, Marieke

Introduction: This project aims to develop versatile peptide-specific MHCII tetramers to increase the number of specific T cell responses that can be analysed in patients with autoimmune diseases. In addition, we will perform antigen-specific T cell receptor (TCR), B cell receptor (BCR) and single cell RNA sequencing. These innovations will allow more complete interpretation of the immunological and therapeutic impact of MRC-based therapies in clinical trials.

Aims: A1: To establish standardised and validated immunomonitoring assays of CD4+ T cell responses against disease-relevant antigens that will be useful for evaluating the efficacy and safety of tolerance-inducing MRC-based therapies. A2: To assess the effects of MRC-based therapies on auto-, allo- or recall- Ag-specific CD4+ T cell responses using new or archived samples from consortium partners; A3: To develop standardised and validated assays to measure Ag-specific memory B cell and Ag-specific antibody formation. Analysis of patient cohorts not treated with MRC-based therapies might be performed as an alternative proof-of-concept.

Expected Results: R1: Standardization of proliferation dye-based assay to monitor ag-specific CD4+ T cell responses; R2: Up-scaling production of patient and antigen-tailored in house developed peptide-exchangeable MHCII tetramers; R3: Establishment of an assay to measure antigen-specific antibody and memory B cell responses; R4: Analysis of Ag-specific T and B cell responses before, during and after tolerance inducing cell therapy to study safety and efficacy and to obtain insights in the mode-of-action of the applied immunotherapy.

Planned secondment: (1) An international 2-month secondment is planned to the lab of Dr. Nathalie Cools (Belgium) to provide the ESR with training in METC application and clinical trial immunomonitoring, as well as logistics of sample retrieval and clinical database registration during conduct of clinical trials. (2) An international, 2-month industrial secondment to Miltenyi (Germany) will support preclinical development of commercially viable antigen-specific immune monitoring assays.

Enrolment in Doctoral degree(s): University of Amsterdam, the Netherlands

Host Institution: University of Antwerp, Belgium
Primary Supervisor: Prof. COOLS, Nathalie

Introduction: Development of MRC-based products has been restricted by their cost and complexity. This is especially true when a unique batch is manufactured for each patient using labor-intensive, multiple-step processes. Successful future commercialization of cell-based, patient-specific products will require innovative solutions to unique manufacturing challenges and constraints; critically, any technological solutions will have to be affordable, reliable and not introduce new process-related risks. In this project, we aim to unlock new opportunities for the development of commercially viable and safe cell-based treatments by co-development of next generation technologies to facilitate lean, cost-effective manufacturing of MRC products, especially tol-DC. In collaboration with Antleron, an end-to-end closed and automated cell culture system will be developed that contains all features essential for safe production of cell therapies: precise control over temperature and fluid flow, mixing and cell seeding capability, internal refrigeration, light protection, various sterile access points to verify cell differentiation, biosensors to monitor cell performance and automated computer-controlled record keeping.

Aims: A1: To identify biological factors contributing to the optimal micro-environment for MRC generation that will allow redesign of MRC manufacturing processes to ensure product consistency and quality. A2: To engineer 4D environments for MRC generation that increase process reproducibility and robustness, and optimize roll-out of MRC products to the market.

Expected Results: R1: Novel biological insights into the differentiation process of tol-DC and other MRC types, such as the influence of environmenetal factors (eg. cell density, growth surface material, pH, dissolved oxygen and shear stress) affect quality-control measures and potency of MRC-based products. This information will be invaluable for the parallel development of a scalable and automated process. R2: Development of an R&D scale proof-of-concept bioreactor-driven tol-DC differentiation process with integrated up- and down-stream unit operations, and built-in quality control measures to monitor critical cell culture parameters. R3: Mapping of cost of goods in order to transition from R&D scale to an industrialized process.

Planned secondment: (1) An intersectoral secondment of 6 months is planned to provide the ESR with training in cell growth micro-environments and experience in designing scale-up and scale-out procedures in GMP conditions. Aim 2 is dependent upon this secondment. (2) An international, intersectoral secondment of 2 months to CiMaas BV will provide the ESR with a market-oriented perspective of manufacturing and roll-out of MRC-based products.

Enrolment in Doctoral degree(s): University of Antwerp

Host Institution: Miltenyi Biotec BV & Co. KG , Bergisch Gladbach, Germany
Primary Supervisor: Dr. KOLLET, Jutta

Introduction: It is critical to understand differences between myeloid cell populations in the context their surrounding tissue microenvironments. This project will identify and systematically characterize the MRC networks in tissues through development of computational methods to analyze multiparametric immunofluorescence image data (MACSima^TM technology). The composition and spatial distribution of MRC subsets in tissues will be classified using machine learning on a knowledge database.

Aims: A1: To apply model-based machine learning algorithms on multiparametric immunofluorescence image data of pancreatic cancer sections to identify and characterize the tolerogenic myeloid regulatory network. A2: To generate experimental image datasets from unrelated tissue sections to validate and optimize the initially generated classifier. A3: To optimize predictions and analysis tools by inclusion of self-learning components.

Expected Results: R1: The phenotype and spatial network of MRC in tissues (eg. tumour, transplant) will be characterized; thereby phenotypically distinct subpopulations of MRC will be identified. R2: Understanding of the correlation of transcriptome and proteome data and of the function of cell types will be obtained from collaborations with UHREG. This is particularly important, since public NGS datasets are frequently used for biomarker/ target discovery. R3: Data-driven marker collections (‘self-learning’ algorithms will automatically update a data base) will be used for identification and prediction of cell composition. R4: Optimization and commercialization of antibody panels for kits and diagnostics for use in identifying or isolating MRC subsets using the newly developed software tools. R5: Software for automated annotation and cell type classification in immunofluorescence images and novel visualization methods for multiparametric datasets will be established.

Planned secondment: A 2-month secondment to the labs of Professors Spang and Rehli is planned to train the ESR in analyzing different types of ‘Big Data’ in order to make correlations with imaging-based proteome data.

Enrolment in Doctoral degree(s): University of Regensburg

Host Institution: Institute of the Germans Trias i Pujol Foundation, Barcelona, Spain
Primary Supervisor: Prof. MARTINEZ CACERES, Eva

Introduction: Our research group has developed an autologous, antigen-specific cell-based therapeutic product for treatment of multiple sclerosis (MS) known as, “tolDC-VitD3.” We are now conducting a Phase-I dose-escalation clinical trial of tolDC-VitD3 therapy in patients with MS (EUDRA CT 2015-003541-26). 12 patients with active MS will receive tolDC-VitD3 by intranodal injection, followed by a 24-month observation and immune monitoring period. This project investigates pharmacodynamic effects of tolDC-VitD3 treatment in patients enrolled in this clinical trial.

Aims: A1: To perform silencing /downmodulation of candidate molecules on cells at different stages during the differentiation of monocytes into tol-DC-vitD3 or stimulatory DC with the aim of identifying transcriptional regulatory factors (TFs) governing the tolerogenic DC programme. A2: To analyse the contribution of TFs identified in A1 to overall tolerogenic phenotype human tolDC-vitD3 using established co-culture assays looking at T cell differentiation and effector functions. A3: To analyse the functional effect of tolDC VitD3 on peripheral blood T cells of multiple sclerosis patients compared to those of healthy donors. A4: To examine antigen-specific T and B cell responses of MS patients in the tolDC-VitD3 clinical trial (before, during and after cell therapy) as a low-risk project.

Expected Results: (1) New potential therapeutic targets involved in tolerance induction by tolDC-VitD3 will be identified through our in vitro experiments and analysis of patient samples. (2) Strategies will be developed to support tolDC-VitD3 treatment using drugs that interact with immunological networks affected by tolDC-VitD3 administration. (3) Novel clinical indications for tolDC-VitD3 will be investigated, especially other neuroinflammatory diseases and solid organ transplantation.

Planned secondment: (1) An international secondment of 2-months to Prof. van Ham is planned to undertake collaborative work on aim A4. (2) A 2-month intersectoral secondment to Aniling is planned. (3) An intersectoral secondment of 2-months will be spent in the academic department of Prof. Ochando where the student will learn about Transplant Immunology and solid organ transplant models.

Host Institution: University Hospital Regensburg, Germany
Primary Supervisor: Dr. HUTCHINSON, James Alexander

Introduction: Administration of human regulatory macrophages (Mreg) has shown early clinical promise as an adjunct immunosuppressive cell-based therapy in solid organ transplantation. This project proposal addresses the hypothesis that recipient T cell immunity is actively regulated through direct allorecognition of donor-derived Mregs and conversion of naive CD4+ T cells to FoxP3+ induced regulatory T cells. A sensitive and rapid functional screening assay has been developed to measure Mreg-mediated conversion of naïve CD4+ to FoxP3+ iTregs (miTregs). We propose to use this in vitro assay system to define novel mechanisms contributing to generation, stabilization or expansion of miTregs.

Aims: A1: To screen libraries of antibodies and small molecules for substances that affect miTreg generation using our established miTreg assay. A2: To investigate the contribution of Mreg-specific or miTreg-specific genes identified from previous transcriptional profiling studies to Mreg-induced Treg generation using gene-editing or –silencing techniques in our established assay system. A3: To investigate the pharmacological effects of substances identified by our screening assays in a human-into-mouse reconstitution model of human Mreg-driven allogeneic Treg conversion.

Expected Results: R1: Original basic scientific insights into the suppressor functions of Mregs, including identification of new suppressive mechanisms, will follow from the functional screening experiments; R2: agents that enhance or repress miTreg generation will be identified and their effects characterized; R3: New Mreg-specific mAbs and clinical tests will be developed.

Planned secondment: An international, intersectoral secondment of 3 months to AstraZeneca will provide the ESR with an industrial perspective of drug screening and a broader view of pharmaceutical development.

Enrolment in Doctoral degree(s):University of Regensburg

Host Institution: University of Newcastle, United Kingdom
Supervisors: Dr. HILKENS, Catharien (primary) - Prof. ISAACS, John - Dr. THELWALL, Pete

Introduction: We have developed a robust, GMP-compatible process to generate stable tolDC from patients with rheumatoid arthritis (RA). We recently completed a phase I trial showing that tolDC administration into an inflamed knee joint is safe. Diminished synovial inflammation was observed in the knee joint of patients treated with the highest tolDC dose; however no systemic changes were observed, either in overall disease activity or immune markers in blood. RA is a systemic disease, so tolDC may be more effective if they migrated to secondary lymphoid tissue. Therefore, we need to know (i) where tolDC migrate to after injection and (ii) whether they modulate T cell responses systemically. The aim of this clinically orientated project is to develop and validate tools required for addressing these questions in on-going clinical studies.

Aims: A1: To assess the distribution of tolDC loaded with relevant citrullinated peptides (citP) injected via different routes into patients by 19F-magnetic resonance imaging (MRI); A2: To assess whether tolDC treatment modulates citP-specific T cell responses assayed by the use of MHCII-citP tetramers in collaboration with SQ.

Expected Results: R1: Determination of the sensitivity of detecting 19F-labelled tolDC by MRI; R2: Confirmation that 19F-labelling of tolDC does not affect their migratory ability and tolerogenic functions; R3: Establishment of an immunmonitoring protocol to assess functional properties (e.g. cytokine production, suppressive capability) of citP-specific T cells from RA patients. For this work we will make use of relevant citP-MHCII tetramers provided by our partners of the RTCure consortium and collaborations with ESR10. R4: Application of these immunomonitoring protocols to samples obtained from RA patients before and after tolDC administration.

Enrolment in Doctoral degree(s):Newcastle University