Periodic Reporting for period 1 - NHPig (Reducing Non-Human Primates in Non-Clinical Safety Assessment: The European Initiative on Minipig and Micropig Models)
Reporting period: 2024-10-01 to 2025-09-30
NHPig (Reducing Non-Human Primates in Non-Clinical Safety Assessment: The European Initiative on Minipig and Micropig Models) aims to expand biological knowledge on mini- and micropig models and improve translational understanding between pigs, non-human primates (NHPs), and humans, with the goal of establishing pig models as viable alternatives to NHPs in nonclinical safety testing. This requires an evidence-based approach combining physiological and molecular characterisation with bioinformatics to identify shared and distinct biological pathways across species. The main objectives are: (1) replace NHPs in safety testing; (2) characterise and validate humanised and micropig models; (3) establish multi-organ biobanks and generate multi-omics data; (4) develop the iPig platform for digital monitoring; and (5) build a FAIR (Findable, Accessible, Interoperable and Reusable), publicly accessible database. In parallel, new approach methodologies (NAMs) based on pig models are being explored, including the porcine primary hepatocyte cultures. To achieve its ambitious goals, the NHPig consortium is structured into 5 experimental work packages (WPs), a WP focused on dissemination and outreach, exploitation and regulatory activities, and a WP covering the project management. To ensure strong ethical and regulatory oversight, NHPig has established both Regulatory and Ethics & Animal Welfare Advisory Boards to guide regulatory discussions and support the development of future standards for animal use in non-clinical safety testing. Alongside its scientific goals, the project prioritizes transparency and public engagement by promoting dialogue with the public, researchers, and regulators through activities such as Open Days and interactive workshops.
Project website: www.nhpig.eu
Project website: www.nhpig.eu
1) Biobanking and model development: Two full necropsy campaigns of 4.5-month-old wild-type Göttingen Minipigs (GMPs; 5 males, 5 females) and humanised (h) GMPs (5 males, 5 females) produced high-quality biobanks supporting immunological, transcriptomic, proteomic, and morphological studies. The molecular profiles to be generated will support the selection of the right model for safety studies. In addition, preparations for two disease-relevant models were completed: a type 1 diabetes model with established SOPs for glucose-homeostasis testing, and the humanised FcγR3B GMP model for testing human antibodies.
2) Digital monitoring & iPig platform: Biosensor and algorithm development (iPig) progressed from conceptual design to a functional intelligent housing system collecting synchronised behavioural, physiological, and acoustic data, supporting the development of non-invasive biomarkers of cardiovascular, respiratory, and neurobehavioural function. Achievements include an innovative wearable telemetry system based on a jacket with integrated 3D electrocardiography (ECG) electrodes and respiratory bands, and the prototype of a housing platform, enabling automated long-term monitoring with remote access, including a secure data-management infrastructure.
3) Immune system characterisation & immunosafety tools: For immune-system characterisation and immunosafety studies harmonised workflows for immune-cell isolation, storage, and flow cytometry across multiple tissues were established, along with refined whole-blood staining compatible with different cytometers. Specific achievements include validated reagents and advanced multi-colour panels for lymphoid and myeloid phenotyping, cytokine panels for ELISA and intracellular assays, and cytokine-release and T-dependent antibody-response (TDAR) assays. The first phase of a single-cell immune atlas began, including scRNA-seq of blood and bone marrow and optimisation of nuclei isolation from frozen tissue.
4) Safety studies & biomarker discovery: For safety studies and biomarker discovery eight compounds with known target-organ profiles were selected and dedicated study groups were formed to coordinate planning, ethics, analytical activities, and animal supply. The first study—a one-month toxicology study in GMPs with a siRNA—was completed, with standard toxicology, cytokine, complement, and miRNA analyses performed. Samples are now being prepared for transcriptomic and proteomic work. Ongoing studies include a 13-week monoclonal antibody study in WT and hGMPs, a pharmacokinetic study with tofacitinib to inform pivotal-study dosing, and a TDAR study with cyclophosphamide. In parallel, development of a microfluidic device for hepatocyte culture was initiated, supported by established hepatocyte-isolation workflows and optimisation for yield, viability, and cryopreservation.
5) Data integration, database infrastructure & FAIR implementation: For integrating multi-omics and toxicology data across preclinical species, the first operational version of the NHPig-DB, a robust, scalable, Standard for Exchange of Nonclinical Data (SEND)-compliant infrastructure, was delivered. Key achievements include a CDISC-compliant SEND schema, an SAP HANA backend extended from ProteomicsDB, integration of controlled vocabularies, and a web-based visualisation tool for interactive data exploration. A total of 37 SEND datasets from industry partners have been imported and validated. Early testing of AI-assisted querying showed the feasibility of natural-language interaction, a step toward self-service analytics. These developments provide the technical foundation for cross-species analyses and ensure alignment with FAIR (Findable, Accessible, Interoperable, Reusable) principles.
2) Digital monitoring & iPig platform: Biosensor and algorithm development (iPig) progressed from conceptual design to a functional intelligent housing system collecting synchronised behavioural, physiological, and acoustic data, supporting the development of non-invasive biomarkers of cardiovascular, respiratory, and neurobehavioural function. Achievements include an innovative wearable telemetry system based on a jacket with integrated 3D electrocardiography (ECG) electrodes and respiratory bands, and the prototype of a housing platform, enabling automated long-term monitoring with remote access, including a secure data-management infrastructure.
3) Immune system characterisation & immunosafety tools: For immune-system characterisation and immunosafety studies harmonised workflows for immune-cell isolation, storage, and flow cytometry across multiple tissues were established, along with refined whole-blood staining compatible with different cytometers. Specific achievements include validated reagents and advanced multi-colour panels for lymphoid and myeloid phenotyping, cytokine panels for ELISA and intracellular assays, and cytokine-release and T-dependent antibody-response (TDAR) assays. The first phase of a single-cell immune atlas began, including scRNA-seq of blood and bone marrow and optimisation of nuclei isolation from frozen tissue.
4) Safety studies & biomarker discovery: For safety studies and biomarker discovery eight compounds with known target-organ profiles were selected and dedicated study groups were formed to coordinate planning, ethics, analytical activities, and animal supply. The first study—a one-month toxicology study in GMPs with a siRNA—was completed, with standard toxicology, cytokine, complement, and miRNA analyses performed. Samples are now being prepared for transcriptomic and proteomic work. Ongoing studies include a 13-week monoclonal antibody study in WT and hGMPs, a pharmacokinetic study with tofacitinib to inform pivotal-study dosing, and a TDAR study with cyclophosphamide. In parallel, development of a microfluidic device for hepatocyte culture was initiated, supported by established hepatocyte-isolation workflows and optimisation for yield, viability, and cryopreservation.
5) Data integration, database infrastructure & FAIR implementation: For integrating multi-omics and toxicology data across preclinical species, the first operational version of the NHPig-DB, a robust, scalable, Standard for Exchange of Nonclinical Data (SEND)-compliant infrastructure, was delivered. Key achievements include a CDISC-compliant SEND schema, an SAP HANA backend extended from ProteomicsDB, integration of controlled vocabularies, and a web-based visualisation tool for interactive data exploration. A total of 37 SEND datasets from industry partners have been imported and validated. Early testing of AI-assisted querying showed the feasibility of natural-language interaction, a step toward self-service analytics. These developments provide the technical foundation for cross-species analyses and ensure alignment with FAIR (Findable, Accessible, Interoperable, Reusable) principles.
• GMP biobank: N = 5 male/5 female pigs; > 500 samples from 31 organs/tissues per pig; samples will be made publically available upon request via NHPig-DB pending approval by the NHPig steering committee.
• hGMP biobank: N = 5 male/5 female pigs; > 500 samples from 31 organs/tissues per pig; samples will be made publically available upon request via NHPig-DB pending approval by the NHPig steering committee.
• Innovative wearable telemetry system, including an ergonomic harness and jacket with integrated 3D electrocardiography (ECG) electrodes and respiratory bands, with excellent animal acceptance and robust ECG signal quality. The telemetry and frontend are currently accessible only to the consortium members. The corresponding
analyzed, structured data will be made public at a later stage via the NHPig-DB.
• Prototype of a housing platform, enabling automated long-term monitoring with remote access, including a secure data-management infrastructure. As the platform is still under development, it is currently accessible only within the consortium.
• Validated reagents and advanced multi-colour panels for lymphoid and myeloid phenotyping. Reagents and protocols will be published on the NHPig Webpage once the internal validation process is finished.
• Cytokine panels for ELISA and intracellular assays. Reagents and protocols will be published on the NHPig Webpage once the internal validation process is finished.
• Cytokine-release and T-dependent antibody-response (TDAR) assays. Reagents and protocols will be published on the NHPig Webpage once the internal validation process is finished.
• NHPig-DB, a robust, scalable, Standard for Exchange of Nonclinical Data (SEND)-compliant infrastructure. The NHPig-DB will be made available from Q3 2026 onward.
• Biobank samples and protocols will be made publicly available via the NHPig-DB from Q3 2026 onward; molecular profiles will be made publicly available immediately after acceptance of the primary publication.
Supporting links for some of the key outputs:
https://www.nhpig.eu/2025/03/26/eu-project-aims-to-boost-drug-testing-reliability-by-replacing-monkeys-with-pigs/(opens in new window)
https://insights.citeline.com/pink-sheet/drug-safety/eu-project-aims-to-boost-drug-testing-reliability-by-replacing-monkeys-with-pigs-S2GUYSKABNDRLMTFZ45GD65O3I/(opens in new window)
https://www.nhpig.eu/2025/03/26/new-article-on-the-nhpig-project-published-in-gottingen-minipigs-magazine/(opens in new window)
https://minipigs.dk/about-us/gottingen-minipigs-magazine(opens in new window)
https://www.nhpig.eu/2024/12/04/animal-welfare-reducing-lab-experiments-on-primates-interview-with-the-project-coordinator-eckhard-wolf-from-lmu/(opens in new window)
Eckstein, Y. et al. 2025. Protocol for in vivo assessment of glucose control and insulin secretion and sensitivity in the pig. STAR Protocols. 2025 Jun 20;6(2):103774. doi:10.1016/j.xpro.2025.103774
• hGMP biobank: N = 5 male/5 female pigs; > 500 samples from 31 organs/tissues per pig; samples will be made publically available upon request via NHPig-DB pending approval by the NHPig steering committee.
• Innovative wearable telemetry system, including an ergonomic harness and jacket with integrated 3D electrocardiography (ECG) electrodes and respiratory bands, with excellent animal acceptance and robust ECG signal quality. The telemetry and frontend are currently accessible only to the consortium members. The corresponding
analyzed, structured data will be made public at a later stage via the NHPig-DB.
• Prototype of a housing platform, enabling automated long-term monitoring with remote access, including a secure data-management infrastructure. As the platform is still under development, it is currently accessible only within the consortium.
• Validated reagents and advanced multi-colour panels for lymphoid and myeloid phenotyping. Reagents and protocols will be published on the NHPig Webpage once the internal validation process is finished.
• Cytokine panels for ELISA and intracellular assays. Reagents and protocols will be published on the NHPig Webpage once the internal validation process is finished.
• Cytokine-release and T-dependent antibody-response (TDAR) assays. Reagents and protocols will be published on the NHPig Webpage once the internal validation process is finished.
• NHPig-DB, a robust, scalable, Standard for Exchange of Nonclinical Data (SEND)-compliant infrastructure. The NHPig-DB will be made available from Q3 2026 onward.
• Biobank samples and protocols will be made publicly available via the NHPig-DB from Q3 2026 onward; molecular profiles will be made publicly available immediately after acceptance of the primary publication.
Supporting links for some of the key outputs:
https://www.nhpig.eu/2025/03/26/eu-project-aims-to-boost-drug-testing-reliability-by-replacing-monkeys-with-pigs/(opens in new window)
https://insights.citeline.com/pink-sheet/drug-safety/eu-project-aims-to-boost-drug-testing-reliability-by-replacing-monkeys-with-pigs-S2GUYSKABNDRLMTFZ45GD65O3I/(opens in new window)
https://www.nhpig.eu/2025/03/26/new-article-on-the-nhpig-project-published-in-gottingen-minipigs-magazine/(opens in new window)
https://minipigs.dk/about-us/gottingen-minipigs-magazine(opens in new window)
https://www.nhpig.eu/2024/12/04/animal-welfare-reducing-lab-experiments-on-primates-interview-with-the-project-coordinator-eckhard-wolf-from-lmu/(opens in new window)
Eckstein, Y. et al. 2025. Protocol for in vivo assessment of glucose control and insulin secretion and sensitivity in the pig. STAR Protocols. 2025 Jun 20;6(2):103774. doi:10.1016/j.xpro.2025.103774