Periodic Reporting for period 1 - ABCardionostics (Human Antibody-enabled Cardiovascular Personalized Theranosis)
Periodo di rendicontazione: 2024-04-01 al 2025-03-31
Cardiovascular diseases (CVDs) are the leading cause of mortality worldwide, with profound health and socioeconomic impacts. In Europe, they caused 38% of deaths in 2019 and are projected to contribute to 23.6 million deaths annually by 2030. Atherosclerotic vascular disease (ASVD) is a major contributor, yet accurate diagnosis and effective treatment remain challenging. Limitations of current tools underscore the urgent need for innovative approaches.
ABCardionostics aims to transform ASVD management by combining advanced biotechnology, imaging systems, and personalized therapies. The project addresses critical gaps in patient stratification, diagnosis, and treatment of those at high risk of severe cardiovascular events such as heart attacks and strokes.
The project will develop multi-marker PET imaging combined with high-resolution MRI, using specialized human antibodies for detailed plaque characterization. It will also create bispecific antibodies for targeted immunotherapy, designed to reprogram harmful monocyte/macrophage populations toward protective states. By integrating biotechnologies, in silico analyses, and innovative engineering, ABCardionostics seeks to improve ASVD diagnosis and treatment, enabling earlier intervention.
A unique library of human antibodies (HuAbs) was extracted from injured endothelium and lesional tissue in a rabbit model of atherosclerosis. Using this set, ABCardionostics will:
- Develop 68Ga-labelled PET tracers and translational multi-marker PET/MR imaging protocols.
- Create bispecific antibodies (bsAbs) for targeted immunotherapy and plaque regression.
- Characterize plaque vulnerability by identifying culprit and protective monocyte/macrophage subsets.
- Identify HuAb molecular targets and evaluate their potential as soluble ASVD biomarkers.
ABCardionostics has potential for transformative societal and economic impact by improving diagnosis and treatment of ASVD. Early detection and personalized care can reduce severe cardiovascular events, healthcare costs, and improve patient quality of life. The project supports healthier aging, mitigates the long-term socioeconomic burden of CVDs, and strengthens Europe’s leadership in cutting-edge CVD management, with benefits extending globally.
ABCardionostics aims to transform ASVD management by combining advanced biotechnology, imaging systems, and personalized therapies. The project addresses critical gaps in patient stratification, diagnosis, and treatment of those at high risk of severe cardiovascular events such as heart attacks and strokes.
The project will develop multi-marker PET imaging combined with high-resolution MRI, using specialized human antibodies for detailed plaque characterization. It will also create bispecific antibodies for targeted immunotherapy, designed to reprogram harmful monocyte/macrophage populations toward protective states. By integrating biotechnologies, in silico analyses, and innovative engineering, ABCardionostics seeks to improve ASVD diagnosis and treatment, enabling earlier intervention.
A unique library of human antibodies (HuAbs) was extracted from injured endothelium and lesional tissue in a rabbit model of atherosclerosis. Using this set, ABCardionostics will:
- Develop 68Ga-labelled PET tracers and translational multi-marker PET/MR imaging protocols.
- Create bispecific antibodies (bsAbs) for targeted immunotherapy and plaque regression.
- Characterize plaque vulnerability by identifying culprit and protective monocyte/macrophage subsets.
- Identify HuAb molecular targets and evaluate their potential as soluble ASVD biomarkers.
ABCardionostics has potential for transformative societal and economic impact by improving diagnosis and treatment of ASVD. Early detection and personalized care can reduce severe cardiovascular events, healthcare costs, and improve patient quality of life. The project supports healthier aging, mitigates the long-term socioeconomic burden of CVDs, and strengthens Europe’s leadership in cutting-edge CVD management, with benefits extending globally.
In WP1, novel methods mapped macrophage and monocyte subsets in human carotid plaques. Advanced multispectral and high-plex imaging identified eight distinct populations, including pro- and anti-inflammatory and lipid-associated macrophages. A refined antibody panel was established, integrating new markers to characterize immune cells in relation to plaque features and patient traits. Disease-relevant in vitro macrophage models, including inflammatory lipid-associated macrophages (ILAMs), are under development to capture key aspects of human plaque biology.
In WP2, strategies were developed to identify HuAbs binding macrophage subsets in vulnerable plaques. A next-generation sequencing (NGS) pipeline was implemented for phage display outputs using PacBio long-read sequencing, enabling full-length coverage of scFv antibody sequences, accurate annotation, and clonotype identification. Collaboration with IMGT ensured precise filtering, annotation, and clustering of sequences, complemented by Sanger-sequenced clones to validate recurrent clones and affinity-impacting mutations. This enabled selection of the top 100 enriched scFv sequences, now undergoing validation through flow cytometry, immunohistochemistry, and in vitro assays.
Within WP3, we focused on optimizing production and purification of selected HuAb fragments. Novel vector systems and tag-based purification strategies improved yields and stability, enabling efficient tracer grafting and milligram-scale production for biological validation.
In WP4, proteomic studies of aortic samples identified proteins differentially expressed in diseased versus healthy vessels. Computational analyses confirmed that two of the most promising antibodies recognise Galectin-3, a protein implicated in plaque inflammation. Biochips have been designed to further assess antibody-target interactions and specificity across macrophage subsets.
In WP5, two lead antibodies (P3, C10) targeting Galectin-3 were radiolabelled with zirconium-89 and tested in animals. Both showed high stability and suitable pharmacokinetics. PET imaging revealed uptake in plaque-prone regions, with P3 showing promise for detecting early atherosclerotic lesions. Therapeutic studies in advanced mouse models are planned.
In WP2, strategies were developed to identify HuAbs binding macrophage subsets in vulnerable plaques. A next-generation sequencing (NGS) pipeline was implemented for phage display outputs using PacBio long-read sequencing, enabling full-length coverage of scFv antibody sequences, accurate annotation, and clonotype identification. Collaboration with IMGT ensured precise filtering, annotation, and clustering of sequences, complemented by Sanger-sequenced clones to validate recurrent clones and affinity-impacting mutations. This enabled selection of the top 100 enriched scFv sequences, now undergoing validation through flow cytometry, immunohistochemistry, and in vitro assays.
Within WP3, we focused on optimizing production and purification of selected HuAb fragments. Novel vector systems and tag-based purification strategies improved yields and stability, enabling efficient tracer grafting and milligram-scale production for biological validation.
In WP4, proteomic studies of aortic samples identified proteins differentially expressed in diseased versus healthy vessels. Computational analyses confirmed that two of the most promising antibodies recognise Galectin-3, a protein implicated in plaque inflammation. Biochips have been designed to further assess antibody-target interactions and specificity across macrophage subsets.
In WP5, two lead antibodies (P3, C10) targeting Galectin-3 were radiolabelled with zirconium-89 and tested in animals. Both showed high stability and suitable pharmacokinetics. PET imaging revealed uptake in plaque-prone regions, with P3 showing promise for detecting early atherosclerotic lesions. Therapeutic studies in advanced mouse models are planned.
During the first year, ABCardionostics achieved results beyond current cardiovascular imaging and immunotherapy standards. Novel human antibody fragments specifically recognize immune cells driving atherosclerosis. Radiolabelled candidates demonstrated vascular uptake in animal models, marking the first antibody-based radiotracers for vessel wall imaging. This enables multimodal imaging combining PET and 4D-flow MRI for early plaque detection, offering in vivo “deep phenotyping”.
Proteomic analyses identified candidate plaque biomarkers, with Galectin-3 validated as relevant. Selected antibodies are being reformatted into larger therapeutic formats, laying the groundwork for plaque-targeted interventions beyond systemic anti-inflammatory treatments, potentially achieving higher efficacy with fewer side effects.
Key needs for translation include:
- Further preclinical validation in larger cohorts and advanced models
- Optimization of antibody affinity, stability, and safety
- Expansion of PET/MRI hemodynamic analyses
- Standardization of biomarker assays and plasma correlations
- Industrial-scale production and early regulatory engagement
Proteomic analyses identified candidate plaque biomarkers, with Galectin-3 validated as relevant. Selected antibodies are being reformatted into larger therapeutic formats, laying the groundwork for plaque-targeted interventions beyond systemic anti-inflammatory treatments, potentially achieving higher efficacy with fewer side effects.
Key needs for translation include:
- Further preclinical validation in larger cohorts and advanced models
- Optimization of antibody affinity, stability, and safety
- Expansion of PET/MRI hemodynamic analyses
- Standardization of biomarker assays and plasma correlations
- Industrial-scale production and early regulatory engagement