Process data of allogeneic ex vivo-expanded ABCB5+ mesenchymal stromal cells for human use: Off-the-shelf GMP-manufactured donor-independent ATMP

© 2020, The Author(s). Background: Human dermal mesenchymal stromal cells (MSCs) expressing the ATP-binding cassette (ABC) efflux transporter ABCB5 represent an easily accessible MSC population that, based on preclinical and first-in-human data, holds significant promise to treat a broad spectrum of conditions associated not only with skin-related but also systemic inflammatory and/or degenerative processes. Methods: We have developed a validated Good Manufacturing Practice-compliant expansion and manufacturing process by which ABCB5+ MSCs derived from surgical discard skin tissues are processed to an advanced-therapy medicinal product (ATMP) for clinical use. Enrichment for ABCB5+ MSCs is achieved in a three-step process involving plastic adherence selection, expansion in a highly efficient MSC-selecting medium, and immunomagnetic isolation of the ABCB5+ cells from the mixed culture. Results: Product Quality Review data covering 324 cell expansions, 728 ABCB5+ MSC isolations, 66 ABCB5+ MSC batches, and 85 final drug products reveal high process robustness and reproducible, reliable quality of the manufactured cell therapy product. Conclusion: We have successfully established an expansion and manufacturing process that enables the generation of homogenous ABCB5+ MSC populations of proven biological activity manufactured as a standardized, donor-independent, highly pure, and highly functional off-the-shelf available ATMP, which is currently tested in multiple clinical trials

Comparative performance analysis of human iPSC-derived and primary neural progenitor cells (NPC) grown as neurospheres in vitro

Developmental neurotoxicity (DNT) testing performed in rats is resource-intensive (costs, time, animals) and bears the issue of species extrapolation. Thus, reliable alternative human-based approaches are needed for predicting neurodevelopmental toxicity. Human induced pluripotent stem cells (hiPSCs) represent a basis for an alternative method possibly being part of an alternative DNT testing strategy. Here, we compared two hiPSC neural induction protocols resulting in 3D neurospheres: one using noggin and one cultivating cells in neural induction medium (NIM protocol). Performance of Nestin+/SOX2+ hiPSC-derived neural progenitor cells (NPCs) was compared to primary human NPCs. Generally, primary hNPCs first differentiate into Nestin+ and/or GFAP+ radial glia-like cells, while the hiPSC-derived NPCs (hiPSC-NPC) first differentiate into βIII-Tubulin+ neurons suggesting an earlier developmental stage of hiPSC-NPC. In the ‘Neurosphere Assay’, NIM generated hiPSC-NPC produced neurons with higher performance than with the noggin protocol. After long-term differentiation, hiPSC-NPC form neuronal networks, which become electrically active on microelectrode arrays after 85 days. Finally, methylmercury chloride inhibits hiPSC-NPC and hNPC migration with similar potencies. hiPSC-NPCs-derived neurospheres seem to be useful for DNT evaluation representing early neural development in vitro. More system characterization by compound testing is needed to gain higher confidence in this method

Comparative performance analysis of human iPSC-derived and primary neural progenitor cells (NPC) grown as neurospheres in vitro

Developmental neurotoxicity (DNT) testing performed in rats is resource-intensive (costs, time, animals) and bears the issue of species extrapolation. Thus, reliable alternative human-based approaches are needed for predicting neurodevelopmental toxicity. Human induced pluripotent stem cells (hiPSCs) represent a basis for an alternative method possibly being part of an alternative DNT testing strategy. Here, we compared two hiPSC neural induction protocols resulting in 3D neurospheres: one using noggin and one cultivating cells in neural induction medium (NIM protocol). Performance of Nestin+/SOX2+ hiPSC-derived neural progenitor cells (NPCs) was compared to primary human NPCs. Generally, primary hNPCs first differentiate into Nestin+ and/or GFAP+ radial glia-like cells, while the hiPSC-derived NPCs (hiPSC-NPC) first differentiate into βIII-Tubulin+ neurons suggesting an earlier developmental stage of hiPSC-NPC. In the ‘Neurosphere Assay’, NIM generated hiPSC-NPC produced neurons with higher performance than with the noggin protocol. After long-term differentiation, hiPSC-NPC form neuronal networks, which become electrically active on microelectrode arrays after 85 days. Finally, methylmercury chloride inhibits hiPSC-NPC and hNPC migration with similar potencies. hiPSC-NPCs-derived neurospheres seem to be useful for DNT evaluation representing early neural development in vitro. More system characterization by compound testing is needed to gain higher confidence in this method