Periodic Reporting for period 3 - Immune-Image (Immune-Image: Specific Imaging of Immune Cell Dynamics Using Novel Tracer Strategies) Reporting period: 2021-10-01 to 2022-09-30 Summary of the context and overall objectives of the project Immune-Image aims to develop new imaging technologies to study the effects of modern immune therapies in cancer and inflammation patients (Fig. A), which are very successful but unfortunately only in a subset of patients. Our research should lead to new technologies where we can use Positron Emission Tomography (PET), Magnetic Resonance Imaging (MRI) and Optical Imaging (OI) to assess if an immune therapy is successful for an individual patient and, even better, to predict if an immune therapy will be effective. If we understand why patients do not respond to immune therapy, we could also design new drugs that will benefit these patients. Imaging is optimally suited for this purpose because a highly sensitive, whole body signal will be measured and the results will be complementary to cellular biology measurements on biopsies. 22 partner institutions work closely together to achieve this overall objective. The work program is divided in 5 scientific and 2 supportive work packages (Fig. B). Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far WP1 Communication activities towards public and patients were continued. By promoting sustainable thinking within the consortium through organizing workshops and discussions the consortium is being guided towards sustainability either of individual achievements or the organizational and technological structure.A dedicated Task Force defined a transparent procedure to prioritise newly developed immunotracers for development towards clinical application.WP2 offered centrally organised solutions to harmonize data collection, processing, sharing, and analysis and in addition collected information regarding multi-centre data management to design a data management plan. Solutions were developed and implemented to enable future clinical data capture, exchange of images, and findability of biosample meta-data. The accreditation programs for clinical PET image acquisition were implemented in several imaging centres. Harmonization of pre-clinical PET imaging progressed through implementation of phantom experiments. A legal report highlighting applicable regulations and high-level templates for SOPs was shared within the consortium. A prototype of analytical PET/CT methods for semi-automatic quantification of tracer intensity/volume was developed.WP3 The in vitro and first in vivo characterizations of CD8β-, CD69- and CD163-targeting nanobodies are progressing as expected. Generation of the bispecific CD8β/CD69-targeting nanobody has resulted in some compounds showing promising results. S100A9-targeting small molecules and nanobodies are being optimized and characterized. A workflow to generate PD-1 and PD-L1-targeting peptides was established and resulted in the first promising hits. Protocols to label immune cells for longitudinal cell tracking are being fine-tuned. A GMP-grade fluorescent labelled PD-L1 antibody was transferred to clinical use.WP4 partners established precise imaging protocols for some already available immunotracers. In addition, models for in vivo cross validation were further improved. Consequently, the already available immunotracers were successfully transferred in WP5 for longitudinal assessment of immune cell tracking as well as monitoring of different treatment approaches in IMID and tumor models. Moreover, WP4 focused on the establishment of an experimental vasculitis model and humanized mouse models.In WP5 the impact of immune therapeutic interventions in various disease models in a longitudinal manner was investigated with existing immune tracers and multi-modal imaging technologies. The results indicate the high value of immunotracers to assess the dynamics of immune cells in various cancer and IMIDs disease model systems in vivo and underline the value of the proposed imaging platform.WP6 drafted or already submitted the clinical trial protocols and worked extensively on making the sites ready for execution of the clinical studies across sites. Moreover, a trial in rheumatoid arthritis patients using [18F]fluoro-PEG-folate PET as measurement outcome of αTNF treatment has currently included 8 patients.WP7 continued to assess the relevant ethics requirement for research activities at all partners. Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far) It is too early within the project to report about actual results of the project that have societal / economic impact. The overall progress beyond the state of art will be:Impact on advancing the field of immune cell imaging1. Immune-Image will deliver a systematic immunotracer generation platform and ensure that the platform endures beyond Immune-Image.2. Ready-to-use set of validated PET and OI tracers will be produced. Additionally, MRI approaches to assess immunotherapy will be developed.3. Immune-Image will produce regulatory accepted standardised protocols with validated immune-imaging approaches.Advancing clinical and healthcare practice, improving European citizens' health and wellbeing, and making these sustainableImmune-Image will provide insights into the immunological status of individual patients, which will lead to prediction of response to therapeutic interventions and provide insights into individual therapy responses. The patient will benefit with optimised therapies, increased safety and prevention of unnecessary side effects.Boost & optimise drug discoveryThe immunotracers resulting from Immune-Image will find immediate application in clinical trials, especially phase I and II. There, the immunotracers will be used to visualise and quantify the impact of immunotherapy on specific targets and immunological pathways, thereby reducing ambiguity in the evaluation of immunotherapy efficacy. This will lead to a reduction in the duration and costs of drug development and attract clinical research of pharmaceutical companies with new immune therapies in Europe.Impact on basic & translational researchWe envision Immune-Image will generate a number of academic demonstration studies, which will lead to opportunities for the development of novel therapeutic interventions. The results of Immune-Image will enable and facilitate R&D activities relevant to other diseases that have an immunological component. Finally, within Immune-Image new scientists in the field will be trained, who will drive future research.Strengthening competitiveness and industrial leadership and addressing specific societal challenges1. The immunotracer development platform will be used for addressing additional immune system-driven healthcare challenges. This will result in the creation of new jobs and attract additional private investments.2. Immune-Image will facilitate trans-disciplinary and trans-sectoral collaboration and will boost the development of appropriate business models and generate the desired economic and employment impact at a global level.3. Support European industrial leadership to successfully compete in the immunotherapy market worldwide .4. We will actively reach out to European SMEs that focus on immunotherapy drug development to involve them within Immune-Image.5. Support economic growth of industry by reducing unnecessary costs. For example, visualisation and quantification of the impact of therapy on specific sites and pathways is expected to decrease ambiguity in the evaluation of treatment efficacy in early stage clinical trials.6. Increase awareness of the benefits that molecular imaging with immunotracers can create for patients and for all EU citizens. Figure A: Scheme of the sustainable and flexible Immune-Image platform.