Periodic Reporting for period 4 - iPSpine (Induced pluripotent stem cell-based therapy for spinal regeneration)
Période du rapport: 2023-07-01 au 2024-06-30
Low back pain (LBP) is a leading cause of disability worldwide. Degeneration of the spine’s intervertebral discs (IVDs) is a major contributor to LBP accounting for at least 40% (~200 million) of all LBP cases and an annual EU economic burden of ~€240 billion. To date, there are no treatments that can stop or reverse IVD degeneration. iPSpine develops for a subset of these patients estimated to be 40 million worldwide an advanced therapy medicinal product (ATMP), based on a game-changing therapeutic approach involving induced Pluripotent Stem Cells (iPSC) and smart biomaterials.
Main results achieved from month 54 to 66 on the three main objectives of iPSpine:
1. A strong evidence base for the effective and safe use of iPSC-based strategies for regenerative treatment of IDD, restoring the IVD to a functional unit.
The production method has been published and tested for >10 different human iPSC lines demonstrating robustness. Further molecular characterization of the iPSpine cells at single cell level (RNAseq) and at the epigenetic level at different stages of the differentiation protocol was conducted. Continuing effort was placed to show that the intermediate mesendoderm progenitors of iPS-notochordal-like-cells (iPS-NLCs) are relevant for further translational studies together with the two shortlisted biomaterials. The biomaterials supported viability and maturation of these progenitors after injection into the degenerate IVD in biomechanically-loaded tissue explants and in degenerate IVDs of sheep; at the biomechanical level, there was no improvement possibly due to the relative short follow up of 1 and 3 months, respectively. The xenogeneic approach did not elicit immunogenic responses in the sheep. The iPSpine aggrecan reporter iPSC line was used to demonstrate that the delivered progenitors have the capacity to obtain a healthy matrix-producing phenotype in degenerated IVDs of dogs, which is essential for long-term biomechanical and functional restoration of the IVD.
2. Enabling technologies for the development, characterisation, and validation of the iPSC-based strategy.
GMP-compliant sendai and mRNA reprogramming fosters broad applicability beyond the iPSpine therapy. Safety biodistribution/toxicity and tumorigenicity studies in mice shows that sorting of Bcln2-expressing iPSC, depletes the iPSpine therapy from undifferentiated iPSCs, enhances safety, and prevents tumor formation. Single-cell characterization of the transcriptomic and epigenetic landscapes of the cells that reside within the core of the healthy and degenerated dog IVDs, provides for novel insights in IVD (patho)physiology, as well as new strategies to rejuvenate degenerate human IVD cells. DigiWest technology enables targeted proteomic analysis of the iPSpine-cell phenotypic identity and studies of their anti-catabolic and regenerative effects on human IVD cells. CRISPRα targeting of multiple transcription factors, essential in NC physiology towards matrix producing cells, has identified a new combination of potent factors which could be transferred to an iTOP-based approach. The iTOP intracellular delivery technology was successfully applied on iPSC. Despite efforts, CRISPR-Cas9 proteins remains challenging to produce in an active form for the purpose of (epi)genetic reprogramming of iPSCs for lineage commitment using iTOP. The finalized in vitro and ex vivo models from cell-/tissue-/organ-scale together with culture media mimicking the healthy and degenerate IVD environment, the portable spine tester, and the newly developed iPSC reporter line for aggrecan, the most abundant protein of the the IVD tissue, have made a considerable contribution to objective 1. Furthermore, several published seminal research and review papers aid the scientific community in increased harmonization of methodology. Finally, the open digital platform prototype started accepting -OMICs data across the consortium. The smart iPSpine digital platform for ATMP development is available for download, including a video and user manual.
3. A recommendation on ethical and regulatory affairs to enhance and accelerate further development and implementation of iPSC-based therapeutic strategies.
The consortium analysed the complex ethical, policy-related, and regulatory aspects involved in ATMPs. An ethics workshop for stem cell researchers and feedback from patients fed into the framework for responsibilization. Policy studies explored the link between dynamic informed consent and governance, arguing for the need to shift focus from mere future use authorization to more interactive and robust participatory oversight of stem cell research and clinical translation.
Regulatory advice firmly guided the consortium through the regulatory field of ATMP. iPSpine prepared for and received their second national Scientific Advice from the Swedish Medicinal Product Agency. The authority considered the iPSpine therapy a first-in-class product paving the path for other ATMP products with a similar proposed manufacturing process or mode of action. The feedback was used to modify the planned nonclinical development and provided valuable input for characterisation and test specifications for future GMP production sothat studies and processes will more likely be acceptable to agencies when e.g. a clinical trial application or application for marketing authorization is filed. The efforts of the past 66 months have been captured in the Candidate Report, summarizing recommendations for the cell manufacturing and control, the nonclinical development of the product and the biomaterial development alone relevant for future product development and clinical translation.
Project management was conducted in different dimensions (scientific, financial, risk, ethical, IP aspects). The website (www.iPSpine.eu) remains up to date beyond iPSpine to maximize outreach. Whilst iPSpine continues to author numerous scientific publications, dissemination activities further focus on patient-targeted content. They culminated to a highlight: the patient-dedicated iPSpine symposium that realized direct interaction between researchers and patients, promoted dialogue and understanding of the patient needs, as well as the iPSpine outcomes.
1. A strong evidence base for the effective and safe use of iPSC-based strategies for regenerative treatment of IDD, restoring the IVD to a functional unit.
The production method has been published and tested for >10 different human iPSC lines demonstrating robustness. Further molecular characterization of the iPSpine cells at single cell level (RNAseq) and at the epigenetic level at different stages of the differentiation protocol was conducted. Continuing effort was placed to show that the intermediate mesendoderm progenitors of iPS-notochordal-like-cells (iPS-NLCs) are relevant for further translational studies together with the two shortlisted biomaterials. The biomaterials supported viability and maturation of these progenitors after injection into the degenerate IVD in biomechanically-loaded tissue explants and in degenerate IVDs of sheep; at the biomechanical level, there was no improvement possibly due to the relative short follow up of 1 and 3 months, respectively. The xenogeneic approach did not elicit immunogenic responses in the sheep. The iPSpine aggrecan reporter iPSC line was used to demonstrate that the delivered progenitors have the capacity to obtain a healthy matrix-producing phenotype in degenerated IVDs of dogs, which is essential for long-term biomechanical and functional restoration of the IVD.
2. Enabling technologies for the development, characterisation, and validation of the iPSC-based strategy.
GMP-compliant sendai and mRNA reprogramming fosters broad applicability beyond the iPSpine therapy. Safety biodistribution/toxicity and tumorigenicity studies in mice shows that sorting of Bcln2-expressing iPSC, depletes the iPSpine therapy from undifferentiated iPSCs, enhances safety, and prevents tumor formation. Single-cell characterization of the transcriptomic and epigenetic landscapes of the cells that reside within the core of the healthy and degenerated dog IVDs, provides for novel insights in IVD (patho)physiology, as well as new strategies to rejuvenate degenerate human IVD cells. DigiWest technology enables targeted proteomic analysis of the iPSpine-cell phenotypic identity and studies of their anti-catabolic and regenerative effects on human IVD cells. CRISPRα targeting of multiple transcription factors, essential in NC physiology towards matrix producing cells, has identified a new combination of potent factors which could be transferred to an iTOP-based approach. The iTOP intracellular delivery technology was successfully applied on iPSC. Despite efforts, CRISPR-Cas9 proteins remains challenging to produce in an active form for the purpose of (epi)genetic reprogramming of iPSCs for lineage commitment using iTOP. The finalized in vitro and ex vivo models from cell-/tissue-/organ-scale together with culture media mimicking the healthy and degenerate IVD environment, the portable spine tester, and the newly developed iPSC reporter line for aggrecan, the most abundant protein of the the IVD tissue, have made a considerable contribution to objective 1. Furthermore, several published seminal research and review papers aid the scientific community in increased harmonization of methodology. Finally, the open digital platform prototype started accepting -OMICs data across the consortium. The smart iPSpine digital platform for ATMP development is available for download, including a video and user manual.
3. A recommendation on ethical and regulatory affairs to enhance and accelerate further development and implementation of iPSC-based therapeutic strategies.
The consortium analysed the complex ethical, policy-related, and regulatory aspects involved in ATMPs. An ethics workshop for stem cell researchers and feedback from patients fed into the framework for responsibilization. Policy studies explored the link between dynamic informed consent and governance, arguing for the need to shift focus from mere future use authorization to more interactive and robust participatory oversight of stem cell research and clinical translation.
Regulatory advice firmly guided the consortium through the regulatory field of ATMP. iPSpine prepared for and received their second national Scientific Advice from the Swedish Medicinal Product Agency. The authority considered the iPSpine therapy a first-in-class product paving the path for other ATMP products with a similar proposed manufacturing process or mode of action. The feedback was used to modify the planned nonclinical development and provided valuable input for characterisation and test specifications for future GMP production sothat studies and processes will more likely be acceptable to agencies when e.g. a clinical trial application or application for marketing authorization is filed. The efforts of the past 66 months have been captured in the Candidate Report, summarizing recommendations for the cell manufacturing and control, the nonclinical development of the product and the biomaterial development alone relevant for future product development and clinical translation.
Project management was conducted in different dimensions (scientific, financial, risk, ethical, IP aspects). The website (www.iPSpine.eu) remains up to date beyond iPSpine to maximize outreach. Whilst iPSpine continues to author numerous scientific publications, dissemination activities further focus on patient-targeted content. They culminated to a highlight: the patient-dedicated iPSpine symposium that realized direct interaction between researchers and patients, promoted dialogue and understanding of the patient needs, as well as the iPSpine outcomes.
Upon successful implementation of the iPSpine therapy, we envision improved outcome and QoL for patients with IVD degeneration-related LBP, through effective regeneration of the degenerated tissue with restored biomechanical function of the disc, and long-lasting reduction of LBP. Altogether, this will result in reduced LBP-related premature retirement and improved socio-economic contribution.
The novel and extended knowledge, tools and technologies that will be the outcome of the iPSpine project, will provide unique platforms of knowledge, testing, and exploitation with broad applicability. These platforms will contribute to acceleration, innovation, and implementation of novel technologies and ATMPs for treatment of large patient groups. Hereby, iPSpine realizes its ambition to make a significant contribution by reducing translational bottlenecks through open innovation and taking European leadership in the development of ATMPs.
The novel and extended knowledge, tools and technologies that will be the outcome of the iPSpine project, will provide unique platforms of knowledge, testing, and exploitation with broad applicability. These platforms will contribute to acceleration, innovation, and implementation of novel technologies and ATMPs for treatment of large patient groups. Hereby, iPSpine realizes its ambition to make a significant contribution by reducing translational bottlenecks through open innovation and taking European leadership in the development of ATMPs.