A high-throughput media design approach for high performance mammalian fed-batch cultures

Abstract

An innovative high-throughput medium development method based on media blending was successfully used to improve the performance of a Chinese hamster ovary fed-batch medium in shaking 96-deepwell plates. Starting from a proprietary chemically-defined medium, 16 formulations testing 43 of 47 components at 3 different levels were designed. Media blending was performed following a custom-made mixture design of experiments considering binary blends, resulting in 376 different blends that were tested during both cell expansion and fed-batch production phases in one single experiment. Three approaches were chosen to provide the best output of the large amount of data obtained. A simple ranking of conditions was first used as a quick approach to select new formulations with promising features. Then, prediction of the best mixes was done to maximize both growth and titer using the Design Expert software. Finally, a multivariate analysis enabled identification of individual potential critical components for further optimization. Applying this high-throughput method on a fed-batch, rather than on a simple batch, process opens new perspectives for medium and feed development that enables identification of an optimized process in a short time frame.

Tailoring recombinant protein quality by rational media design

“…Cell culture media design has a great potential to modulate the key quality attributes of the molecule 25 , and in particular by means of high-throughput design of experiment (DoE) approaches 26,27 , due to its central role in the upstream manufacturing 19 . The environment the cell is cultured in closely influences its metabolism to a great extent [20][21][22] .…”

“…The adjustment of the levels of a choice of medium components and the addition of novel compounds either in the medium or the in bolus feeds, influence the enzymatic reactions in the endoplasmic reticulum and in the Golgi apparatus, the substrate generation as well as their transport into the two organelles 23,24 . Further enhancements and technology advancements of high-throughput cell culture systems, including 96-deepwell plates and TubeSpin bioreactor tubes, allow to test a plethora of different supplements at various concentrations simultaneously [25][26][27] . Moreover, the observed trends in these systems may be reproduced in controlled bioreactor systems, including lab-scale bioreactors 28 , which highlights the great value of small-scale high-throughput compound screening for the identification of new levers that tailor recombinant protein quality attributes.…”

“…Supplementation was assessed in high-throughput systems including 96-deepwell plates and shake tubes. 96-deepwell plates have proven to be a reliable system to screen raw materials impacting product quality attributes of recombinant proteins 25,27,28 . Confirmation of the findings was conducted in lab-scale 3.5-liter bioreactors.…”

“…This chapter presents potential compounds and mechanisms influencing the production of low-molecular-species. High-throughput media blending approaches 26,27,251 and mechanistic studies were called upon based on the information provided by the literature and hypotheses resulting from the experimental results. Due to the relatively few number of scientific reports that address LMW formation in mammalian cell culture, it was decided to perform a general reshuffling of the platform medium components to identify potential levers for LMW content reduction.…”

“…High-throughput cell culture systems opened the door for testing many conditions simultaneously [266][267][268] . The technology developed for media optimization in 96-deepwell plates (96-DWP) fed-batch cultures 26,27,251 can also be applied for product quality modulation 25,64 . Good alignment of the glycan profiles between 96-DWP and shake tubes has been shown, and in the same study trends observed in high-throughput systems have been confirmed in lab-scale bioreactors 28 .…”