055 Manufactured Cells: High-throughput development of cryopreservation design space

Autor: Ryan Shorr, Greg Russotti, Dan DeWitt, Brian Murphy
Rok vydání: 2013
Předmět:
Zdroj: Cryobiology. 67:413
ISSN: 0011-2240
DOI: 10.1016/j.cryobiol.2013.09.061
Popis: Cryopreservation of cellular therapeutics requires detailed development of the formulation and cryopreservation processes to optimize cell health and mitigate freeze/thaw damage. In keeping with pharmaceutical best practices and Quality-by-Design (QbD) initiatives there is an increased focus on understanding how inputs affect cryopreservation outputs, and in moving from traditional “recipe” validation towards multivariate characterization and understanding of the design space. Cryopreservation outcomes are determined by a large number of parameters including: cellular properties, complex formulation mixtures, multi-step freezing and thawing processes, final container geometry, and interactions with the upstream and downstream processes. Thorough characterization of this multivariate space is difficult and costly, and finite developmental resources should be allocated using a modern science and risk-based approach. We have developed a 96-well method for investigating interactions between different formulation and process variables in cryopreservation. This method uses a 96-well freeze/thaw process with a 96-well post-thaw regrowth assay (alamarBlue). Use of plate-based methods increases experimental capacity about 10-fold over vial-based methods, which enables the screening of more parameters, with better resolution and higher statistical power. A sample experiment used 5 Me 2 SO concentrations, 4 salt concentrations, and 3 different freeze rates with 4 replicates per condition. In this and other multivariate experiments we found several interactions that would not be apparent from univariate experiments. Additionally, the increased spatial resolution allowed better assessment of non-linear responses such as optima and plateaus. These experiments accelerate the characterization of formulation design space, and enable better process understanding. Consistent with QbD principles, this increased understanding guides development and delivers a more robust and reliable process, as well as enabling process changes with lower risk of unanticipated interactions. The plate-based method can be generally applied to study formulation components, freezing parameters, pre-freeze parameters or post-thaw stability. One major limitation of this method is that the results are often scale-dependent. While it is useful for understanding cellular fundamentals and constraints under tightly controlled conditions, the scale-up to product containers requires additional experiments. Source of funding: None declared. Conflict of interest: None declared. bmurphy@celgene.com
Databáze: OpenAIRE
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