From protocol to product: ventral midbrain dopaminergic neuron differentiation for the treatment of Parkinson's disease

Current cell therapy product limitations include the need for in-depth product understanding to ensure product potency, safety and purity. New technologies require development and validation to address issues of production scale-up to meet clinical need; assays are required for process control, validation and release. Prior to clinical realization, an understanding of production processes is required to implement process changes that are essential for process control. Identification of key parameters forms the basis of process tolerances, allowing for validated, adaptive manufacturing processes. This enables greater process control and yield while withstanding regulatory scrutiny. This report summaries key milestones in specifically for ventral midbrain dopaminergic neuroprogenitor differentiation and key translational considerations and recommendations to enable successful, robust and reproducible current cell therapy product-manufacturing

The productivity limit of manufacturing blood cell therapy in scalable stirred bioreactors

Manufacture of red blood cells (RBCs) from progenitors has been proposed as a method to reduce reliance on donors. Such a process would need to be extremely efficient for economic viability given a relatively low value product and high 2E12 cell dose. Therefore, the aim of these studies was to define the productivity of an industry standard stirred-tank bioreactor and determine engineering limitations of commercial RBC production. Cord blood derived CD34+ cells were cultured under erythroid differentiation conditions in a stirred micro-bioreactor (ambr™). Enucleated cells of 80% purity could be created under optimal physical conditions: pH 7.5, 50% oxygen, without gas-sparging (which damaged cells) and with mechanical agitation (which directly increased enucleation). O2 consumption was low (~5x10(-8) µg/cell.hr) theoretically enabling erythroblast densities in excess of 5x10(8) /ml in commercial bioreactors and sub-10 L/unit production volumes. The bioreactor process achieved a 24% and 42% reduction in media volume and culture time respectively relative to unoptimized flask processing. However, media exchange limited productivity to 1 unit of erythroblasts per 500 L of media. Systematic replacement of media constituents, as well as screening for inhibitory levels of ammonia, lactate and key cytokines did not identify a reason for this limitation. We conclude that the properties of erythroblasts are such that the conventional constraints on cell manufacturing efficiency, such as mass transfer and metabolic demand, should not prevent high intensity production; furthermore this could be achieved in industry standard equipment. However, identification and removal of an inhibitory mediator is required to enable these economies to be realized

Qualification of academic facilities for small-scale automated manufacture of autologous cell-based products

Academic centres, hospitals and small companies, as typical development settings for UK regenerative medicine assets, are significant contributors to the development of autologous cell-based therapies. Often lacking the appropriate funding, quality assurance heritage or specialist regulatory expertise, qualifying aseptic cell processing facilities for Good Manufacturing Practice (GMP) compliance is a significant challenge. The qualification of a new Cell Therapy Manufacturing Facility (CTMF) with automated processing capability, the first of its kind in a UK academic setting, provides a unique demonstrator for the qualification of small-scale, automated facilities for GMP compliant manufacture of autologous cell-based products in these settings. This paper shares our experiences in qualifying the CTMF, focussing on our approach to streamlining the qualification effort, the challenges, project delays and inefficiencies we encountered and the subsequent lessons learned

Quality approaches to allow multi-site equivalence in pluripotent stem cell based product manufacturing [Abstract]

Quality approaches to allow multi-site equivalence in pluripotent stem cell based product manufacturing [Abstract

Distributed automated manufacturing of pluripotent stem cell products

Establishing how to effectively manufacture cell therapies is an industry-level problem. Decentralised manufacturing is of increasing importance, and its challenges are recognised by healthcare regulators with deviations and comparability issues receiving specific attention from them. This paper is the first to report the deviations and other risks encountered when implementing the expansion of human pluripotent stem cells (hPSCs) in an automated three international site–decentralised manufacturing setting. An experimental demonstrator project expanded a human embryonal carcinoma cell line (2102Ep) at three development sites in France, Germany and the UK using the CompacT SelecT (Sartorius Stedim, Royston, UK) automated cell culture platform. Anticipated variations between sites spanned material input, features of the process itself and production system details including different quality management systems and personnel. Where possible, these were pre-addressed by implementing strategies including standardisation, cell bank mycoplasma testing and specific engineering and process improvements. However, despite such measures, unexpected deviations occurred between sites including software incompatibility and machine/process errors together with uncharacteristic contaminations. Many only became apparent during process proving or during the process run. Further, parameters including growth rate and viability discrepancies could only be determined post-run, preventing ‘live’ corrective measures. The work confirms the critical nature of approaches usually taken in Good Manufacturing Practice (GMP) manufacturing settings and especially emphasises the requirement for monitoring steps to be included within the production system. Real-time process monitoring coupled with carefully structured quality systems is essential for multiple site working including clarity of decision-making roles. Additionally, an over-reliance upon post-process visual microscopic comparisons has major limitations; it is difficult for non-experts to detect deleterious culture changes and such detection is slow

Pluripotent stem cell based medicinal products: A case study of process transfer related technical and manufacturing issues [Abstract]

Pluripotent stem cell based medicinal products: A case study of process transfer related technical and manufacturing issues [Abstract

Supplementary Information Files for 'Distributed automated manufacturing of pluripotent stem cell products'

Supplementary Information Files for 'Distributed automated manufacturing of pluripotent stem cell products'Abstract:Establishing how to effectively manufacture cell therapies is an industry-level problem. Decentralised manufacturing is of increasing importance, and its challenges are recognised by healthcare regulators with deviations and comparability issues receiving specific attention from them. This paper is the first to report the deviations and other risks encountered when implementing the expansion of human pluripotent stem cells (hPSCs) in an automated three international site–decentralised manufacturing setting. An experimental demonstrator project expanded a human embryonal carcinoma cell line (2102Ep) at three development sites in France, Germany and the UK using the CompacT SelecT (Sartorius Stedim, Royston, UK) automated cell culture platform. Anticipated variations between sites spanned material input, features of the process itself and production system details including different quality management systems and personnel. Where possible, these were pre-addressed by implementing strategies including standardisation, cell bank mycoplasma testing and specific engineering and process improvements. However, despite such measures, unexpected deviations occurred between sites including software incompatibility and machine/process errors together with uncharacteristic contaminations. Many only became apparent during process proving or during the process run. Further, parameters including growth rate and viability discrepancies could only be determined post-run, preventing ‘live’ corrective measures. The work confirms the critical nature of approaches usually taken in Good Manufacturing Practice (GMP) manufacturing settings and especially emphasises the requirement for monitoring steps to be included within the production system. Real-time process monitoring coupled with carefully structured quality systems is essential for multiple site working including clarity of decision-making roles. Additionally, an over-reliance upon post-process visual microscopic comparisons has major limitations; it is difficult for non-experts to detect deleterious culture changes and such detection is slow.</div