Process analysis of pluripotent stem cell differentiation to megakaryocytes to make platelets applying European GMP

Abstract

Quality, traceability and reproducibility are crucial factors in the reliable manufacture of cellular therapeutics, as part of the overall framework of Good Manufacturing Practice (GMP). As more and more cellular therapeutics progress towards the clinic and research protocols are adapted to comply with GMP standards, guidelines for safe and efficient adaptation have become increasingly relevant. In this paper, we describe the process analysis of megakaryocyte manufacture from induced pluripotent stem cells with a view to manufacturing in vitro platelets to European GMP for transfusion. This process analysis has allowed us an overview of the entire manufacturing process, enabling us to pinpoint the cause and severity of critical risks. Risk mitigations were then proposed for each risk, designed to be GMP compliant. These mitigations will be key in advancing this iPS-derived therapy towards the clinic and have broad applicability to other iPS-derived cellular therapeutics, many of which are currently advancing towards GMP-compliance. Taking these factors into account during protocol design could potentially save time and money, expediting the advent of safe, novel therapeutics from stem cells.

Mapping the biogenesis of forward programmed megakaryocytes from induced pluripotent stem cells

“…Optimizing cell culture conditions to increase cell output and diminish the cost of goods remains one of the major challenges in translating iPSC-derived cellular products to the clinic ( 21 , 53 ). Using the set of MKP markers identified here, we found that adjusting oxygen and cytokine conditions can increase MKP generation, maximizing mature MK generation.…”

Improving Human Induced Pluripotent Stem Cell-Derived Megakaryocyte Differentiation and Platelet Production

“…Platelet numbers in the range of >10 8 , which are currently obtained by many researchers in laboratory scale, will be sufficient to perform well-designed mechanistic studies towards better understanding regenerative platelet functions. Further improvements of hiPSC-derived platelet production technologies will consider GMP requirements [12,25,50] and might focus on using particularly histocompatible hiPSCs [12,14,25,51,52]. This will enable providing clinical platelet units, particularly for patients suffering from alloimmunization against frequent antigens, otherwise lacking suitable donors, in the near future.…”

Manufacturing with pluripotent stem cells (PSConf 2021): Key issues for future research and development

“… 7 He also emphasized that understanding the culture and differentiation process through process analytical tools, with a focus on those stages that were most challenging to develop into a GMP environment, had been key to progress towards a final manufacturing process. 8 Dr Ghevaert outlined the significant work involved in the development of new potency assays including an in vitro thrombus formation and a laser‐induced injury model in NSG mice. 6 , 8 A major issue that the whole hPSC manufacturing field is facing today, is the selection of appropriate manufacturing cell lines.…”

“… 8 Dr Ghevaert outlined the significant work involved in the development of new potency assays including an in vitro thrombus formation and a laser‐induced injury model in NSG mice. 6 , 8 A major issue that the whole hPSC manufacturing field is facing today, is the selection of appropriate manufacturing cell lines. Dr Ghevaert explained the significant diversity that the UKRMP programme had discovered in the differentiation characteristics of different hESC lines.…”

“…He explained the significant benefits in the optimisation of platelet yield that this process had delivered 7 . He also emphasized that understanding the culture and differentiation process through process analytical tools, with a focus on those stages that were most challenging to develop into a GMP environment, had been key to progress towards a final manufacturing process 8 . Dr Ghevaert outlined the significant work involved in the development of new potency assays including an in vitro thrombus formation and a laser‐induced injury model in NSG mice 6,8 .…”