student Osaka Medical and Pharmaceutical University Takatsuki, Osaka, Japan
Abstract: Human induced pluripotent stem cells were first produced over 15 years ago. Although this breakthrough in stem cell research has provided a novel platform for the development of individualized cell therapies, iPSC-based clinical products are yet to achieve wide global adoption. The biggest obstacle is the cost of cell manufacturing and quality control tests required to ensure safety of the cell therapies. Substantial reduction of the cost, while assuring safety of the cell therapies, requires major changes to the existing protocols of reprogramming and differentiation. We believe that development of the process in 3D system will simplify and streamline the workflows. The end goal is to create a customized proprietary system, but due to the complexity of the process consisting of reprogramming somatic cells, cloning, engineering and expansion of PSCs, and final differentiation, we are working with commercially available instruments to evaluate and optimize all steps of the process. We have also learned that we still need to clarify certain aspects of stem cell biology, since established characteristics of the cells often seem to be dependent on specific parts of the culture system. Here we report our progress in development of an automated 3D platform for generation, expansion, cloning and differentiation of pluripotent stem cells for clinical applications. We have established a reprogramming protocol of hematopoietic progenitors in the Elplasia ultra-low attachment microplates (Corning). To manipulate cell aggregates we have utilized CellCelector platform (ALS/Sartorius). For single-cell cloning we have used either CellCelector or UP.SIGHT (Cellink/Cytena) with Elplasia microplates or SIEVEWELL slides (Tokyo Ohka Kogyo). Scaling up of 3D PSC culture was done on Elplasia system ranging from 96-well microplates up to 12K microcavities flask as a stationary culture, followed by 100- and 500 mL MAG vessels driven by PBS-MINI MagDrive Bioreactor (STEMCELL Technologies). We are currently working on the differentiation protocols development starting from cell aggregates in Elplasia plates, as well as utilizing microcarriers. Regarding cell quality control, the biggest issue is genetic stability. To address this problem in the most cost-effective manner, we are working on the multi-omics automated approach.
Funding Source: Japan Agency for Medical Research and Development (AMED), grant JP24bm1323001h0102
