Cell Culture Process Scale-Up Challenges for Commercial-Scale Manufacturing of Allogeneic Pluripotent Stem Cell Products

Lee, Brian; Jung, Sunghoon; Hashimura, Yas; Lee, Maximilian; Borys, Breanna S.; Dang, Tiffany; Kallos, Michael S.; Rodrigues, Carlos Alberto Chirano; Silva, Teresa

doi:10.3390/bioengineering9030092

Cited by 16 publications

(10 citation statements)

References 38 publications

(45 reference statements)

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“…This method is not only time-consuming but also susceptible to personal bias. Relying solely on manual techniques for cell quality evaluation is not feasible for mass-producing therapeutic cells [ 112 ]. AI technology has consistently demonstrated its effectiveness in assisting the cultivation and maintenance of iPSC, particularly in identifying cell colonies and their functions.…”

Section: Discussionmentioning

confidence: 99%

The use of artificial intelligence in induced pluripotent stem cell-based technology over 10-year period: A systematic scoping review

Vo,

Saito,

Ida

et al. 2024

PLoS ONE

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Background Stem cell research, particularly in the domain of induced pluripotent stem cell (iPSC) technology, has shown significant progress. The integration of artificial intelligence (AI), especially machine learning (ML) and deep learning (DL), has played a pivotal role in refining iPSC classification, monitoring cell functionality, and conducting genetic analysis. These enhancements are broadening the applications of iPSC technology in disease modelling, drug screening, and regenerative medicine. This review aims to explore the role of AI in the advancement of iPSC research. Methods In December 2023, data were collected from three electronic databases (PubMed, Web of Science, and Science Direct) to investigate the application of AI technology in iPSC processing. Results This systematic scoping review encompassed 79 studies that met the inclusion criteria. The number of research studies in this area has increased over time, with the United States emerging as a leading contributor in this field. AI technologies have been diversely applied in iPSC technology, encompassing the classification of cell types, assessment of disease-specific phenotypes in iPSC-derived cells, and the facilitation of drug screening using iPSC. The precision of AI methodologies has improved significantly in recent years, creating a foundation for future advancements in iPSC-based technologies. Conclusions Our review offers insights into the role of AI in regenerative and personalized medicine, highlighting both challenges and opportunities. Although still in its early stages, AI technologies show significant promise in advancing our understanding of disease progression and development, paving the way for future clinical applications.

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Section: Discussionmentioning

confidence: 99%

The use of artificial intelligence in induced pluripotent stem cell-based technology over 10-year period: A systematic scoping review

Vo,

Saito,

Ida

et al. 2024

PLoS ONE

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“…Generating aggregates within the first 24 h at 110 rpm in PBS-Minis resulted in small aggregates, some of which may be unintentionally discarded in the supernatant during media changes without sufficient centrifugation or gravity settling time. However, considering that prolonged centrifugation (Archibald et al, 2016) and gravity settling (Lee et al, 2022) can negatively impact cell yield and viability, we chose to extend the aggregation period from 24 h to 48 h. This adjustment resulted in minimal viable cell loss in the supernatant. Finally, we observed that at a given agitation rate, higher aggregation yields could further impact cluster size resulting in larger clusters.…”

Section: Discussionmentioning

confidence: 99%

Analysis of the effects of bench-scale cell culture platforms and inoculum cell concentrations on PSC aggregate formation and culture

Iworima,

Baker,

Piret

et al. 2023

Front. Bioeng. Biotechnol.

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Introduction: Human pluripotent stem cells (hPSCs) provide many opportunities for application in regenerative medicine due to their ability to differentiate into cells from all three germ layers, proliferate indefinitely, and replace damaged or dysfunctional cells. However, such cell replacement therapies require the economical generation of clinically relevant cell numbers. Whereas culturing hPSCs as a two-dimensional monolayer is widely used and relatively simple to perform, their culture as suspended three-dimensional aggregates may enable more economical production in large-scale stirred tank bioreactors. To be more relevant to this biomanufacturing, bench-scale differentiation studies should be initiated from aggregated hPSC cultures.Methods: We compared five available bench-scale platforms for generating undifferentiated cell aggregates of human embryonic stem cells (hESCs) using AggreWell™ plates, low attachment plates on an orbital shaker, roller bottles, spinner flasks, and vertical-wheel bioreactors (PBS-Minis). Thereafter, we demonstrated the incorporation of an hPSC aggregation step prior to directed differentiation to pancreatic progenitors and endocrine cells.Results and discussion: The AggreWell™ system had the highest aggregation yield. The initial cell concentrations had an impact on the size of aggregates generated when using AggreWell™ plates as well as in roller bottles. However, aggregates made with low attachment plates, spinner flasks and PBS-Minis were similar regardless of the initial cell number. Aggregate morphology was compact and relatively homogenously distributed in all platforms except for the roller bottles. The size of aggregates formed in PBS-Minis was modulated by the agitation rate during the aggregation. In all cell culture platforms, the net growth rate of cells in 3D aggregates was lower (range: −0.01–0.022 h−1) than cells growing as a monolayer (range: 0.039–0.045 h−1). Overall, this study describes operating ranges that yield high-quality undifferentiated hESC aggregates using several of the most commonly used bench-scale cell culture platforms. In all of these systems, methods were identified to obtain PSC aggregates with greater than 70% viability, and mean diameters between 60 and 260 mm. Finally, we showed the capacity of hPSC aggregates formed with PBS-Minis to differentiate into viable pancreatic progenitors and endocrine cell types.

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“…It is widely accepted that static cultivation systems face challenges with regards to their scalability [9,32]. Hence, the compatibility of OSI in combination with a more scalable approach to hiPSC expansion under stirred conditions was evaluated, namely by using spherical Synthemax II coated MCs with a diameter of 125-212 µm and a rigid polystyrene Consequently, the significant differences in viable cell density per area (VCDA) observed over the 5 d cultivation period were primarily the result of lower initial cell attachment, with the higher doubling times playing a secondary role if VCD during freezing exceeded 50 × 10 6 cells mL −1 .…”

Section: One-step Inoculation Of Microcarrier-operated Stirred Biorea...mentioning

confidence: 99%

Section: One-step Inoculation Of Microcarrier-operated Stirred Biorea...mentioning

confidence: 99%

Process Intensification in Human Pluripotent Stem Cell Expansion with Microcarriers

Teale,

Schneider,

Eibl

et al. 2024

Processes

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Given the demands human induced pluripotent stem cell (hiPSC)-based therapeutics place on manufacturing, process intensification strategies which rapidly ensure the desired cell quality and quantity should be considered. Within the context of antibody and vaccine manufacturing, one-step inoculation has emerged as an effective strategy for intensifying the upstream process. This study therefore evaluated whether this approach could be applied to the expansion of hiPSCs in flasks under static and in microcarrier-operated stirred bioreactors under dynamic conditions. Our findings demonstrated that high density working cell banks containing hiPSCs at concentrations of up to 100 × 106 cells mL−1 in CryoStor® CS10 did not impair cell growth and quality upon thawing. Furthermore, while cell distribution, growth, and viability were comparable to routinely passaged hiPSCs, those subjected to one-step inoculation and expansion on microcarriers under stirred conditions were characterized by improved attachment efficiency (≈50%) following the first day of cultivation. Accordingly, the process development outlined in this study establishes the foundation for the implementation of this intensified approach at L-scale.

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Cell Culture Process Scale-Up Challenges for Commercial-Scale Manufacturing of Allogeneic Pluripotent Stem Cell Products

Cited by 16 publications

References 38 publications

The use of artificial intelligence in induced pluripotent stem cell-based technology over 10-year period: A systematic scoping review

The use of artificial intelligence in induced pluripotent stem cell-based technology over 10-year period: A systematic scoping review

Analysis of the effects of bench-scale cell culture platforms and inoculum cell concentrations on PSC aggregate formation and culture

Process Intensification in Human Pluripotent Stem Cell Expansion with Microcarriers

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