Biomedical Engineering and Bioengineering Commons^™

Incidence And Potential Implications Of Methylglyoxal In Industrial Cell Culture Revisited, Frank Chaplen

Cell Culture Engineering XV

Methylglyoxal is a toxic by-product of glycolysis and amino acid metabolism in mammalian systems. The major route for methylglyoxal detoxification is the glyoxalase pathway, which consists of the enzymes glyoxalase I (GLO1) and glyoxalase II (GLO2). A required co-factor for the glyoxalase pathway is reduced glutathione. Evidence suggests that GLOI and methylglyoxal have important roles to play in the signal pathways associated with oxidative stress and necrotic cell death¹. Previous work has demonstrated that growth conditions found in industrial cell culture have marked effects on endogenous methylglyoxal levels in Chinese hamster ovary (CHO)². Furthermore, decreased levels …

Targeting Product Quality: Where Systems Biotechnology And Process Design Meet, Markus Michael Muller, Jan Bechmann, Ingo Gorr, Harald Bradl, Jan Visser

Cell Culture Engineering XV

Product quality is a result of the entire production process including protein sequence, host cell, media and process parameters. Many of the desired product properties are defined by posttranslational modifications with impact on biological activity, immunogenicity, half-life or stability. In‑depth understanding of the host cells capabilities as well as of the process interactions enables the targeted modulation of product quality attributes by rational selection of host cells and design of bioprocesses. This is valuable for new biological molecules in order to improve efficacy, reduce side effects, access new patient populations. For biosimilars this allows developing into defined quality attribute profiles. …

Development Of A Qualtiy Driven Integrated Continuous Biomanufacturing Process, Daniel Karst, Fabian Steinenbach, Thomas Villiger, Miroslav Soos, Massimo Morbidelli

Cell Culture Engineering XV

The recent improvements of continuous up- and downstream processes in the production of therapeutic proteins suggest their final integration to a single process stream. Besides general benefits of continuous manufacturing, such as reduced equipment size, enhanced cost efficiency and high volumetric productivity, the steady state operation favors constant or even improved product quality. However, N-linked glycosylation, charge variants, aggregates and other critical quality attributes of therapeutic proteins depend on operating conditions in up- and downstream processing. The combination of long process durations and the accompanied lack of experimental high-throughput capabilities of continuous bioprocesses demand for model based optimization regarding product …

Integrated Continuous Bioprocessing - A Gold Mine For Cell Culture Process Understanding?, Mats Akesson, Martin Heitmann, Ditte Skibstrup, Peter Tiainen

Cell Culture Engineering XV

We have developed an integrated continuous processing framework for end-to-end production of complex fragile proteins based on perfusion cultivation and automated multi-step purification [1,2]. Upstream, the integrated system consists of a stirred tank bioreactor with an ATF cell retention system. The clarified harvest directly enters an off-the-shelf ÄKTA chromatography system converted into a continuous purification unit. Two alternating capture columns precede a multi-step purification train with full flexibility and control of individual columns.

The integrated set-up enables compact automated bench-top factories converting cell culture media to purified protein in an efficient manner without intermediate storage. It provides monitoring of the …

Optimization Of Glycosylation And Charge Distribution Through Culture Parameters And Supplements, Sigma Mostafa, Ventaka Tayi, Shahid Rameez, Nathan Oien, Jaspreet Notey, Brian Baker, Jimmy Smedley, Abhinav Shukla

Cell Culture Engineering XV

Culture parameters are known to have significant impacts on product quality, although these effects are sometimes cell-line dependent and the directionality of the effect has to be determined empirically. We will present data from three case studies where glycosylation or charge distribution was modified to match the reference molecule or to reduce variability. In the first case study, glycan optimization for a biosimilar will be described. Galactose, fucose, and mannose levels were optimized through screening of raw materials and process parameters. A range of media, feed, alternate sugars, metals and other supplements as well as temperature set points were tested. …

Challenges Of Scale Down Model For Disposable Bioreactors: Case Studies On Growth & Product Quality Impacts, Jincai Li, Guangming Zhang, Huilin Zhu, Weichang Zhou

Cell Culture Engineering XV

Despite wide-spread use of disposable bioreactors, there is a lack of well-established scale-down model for larger scale SUBs. Here we report a case of NS0 cell culture process transfer from 2000L stainless steel bioreactor (SST) to 2000L disposable bioreactor (SUB). Initial attempts in trying to grow the NS0 cells in the small scale 2D bags yielded non-satisfactory results, as growth was impacted by bag material type as well as by suppliers of the same bag material type. However, 3D bags of 50L and above proved to be supportive of the NS0 cell line growth.

Even for cell lines that do …

Strategies For Optimizing A Cell Culture Platform To Achieve High Recombinant Protein Titer Without Impacting Product Quality, Natarajan Vijayasankaran, Robin Gruver, Steven Meier

Cell Culture Engineering XV

Recent advances in cell line development and in the design of culture media have significantly increased both the volumetric and specific productivity of fed batch processes, which is our standard platform technology for synthesis of therapeutic proteins. Cell culture processes are generally simplified when needed as a robust platform technology to meet the imperatives of speedy timelines for early process development and ease of manufacturing. However, both culture media and feeding strategies need to be customized for specific cell lines when there is a need to achieve higher titer due to higher product demand. This presentation will focus on the …

Efforts To Reduce Impact Of Media Variability On Product Quality For A Commercial Perfusion Process, Nirel Rillera, Kevin Yeh, Benjamin Youn, Jerome Acuna, John Henstrand, Sean Forestell

Cell Culture Engineering XV

Strict control of raw materials used in bioprocesses is necessary to ensure consistent product quality. However, use of poorly characterized complex components makes achieving such strict control difficult. A strong correlation between product quality variability among bulk drug substance lots and changes in a complex medium component was observed in a commercial perfusion process. This correlation was subsequently confirmed experimentally at both large and small scales. A multifaceted strategy was applied to investigate and mitigate the impact of the media variability on product quality, which included (1) studies in small scale perfusion bioreactors; (2) development of a simple cell based …

Overcoming Process Intensification Challenges To Deliver A Manufacturable And Competitive Integrated Continuous Biomanufacturing Platform, Jason Walther, Neha Shah, Myles Hollenbach, Jonathan Wang, Marcella Yu, Jiuyi Lu, Yang Yang, Konstantin Konstantinov, Chris Hwang

Cell Culture Engineering XV

Groups in both industry and academia have achieved high densities and productivities in perfusion cell culture processes. At Sanofi, we have demonstrated perfusion densities greater than 100 million cells/mL (with associated high productivities) at a cell-specific perfusion rate of only 20 pL/cell/day. This process intensification reduces the footprint of upstream unit operations as well as capital and operating expenses of manufacturing facilities. The continuous nature of perfusion cell culture also creates opportunities for integration of continuous downstream operations, leading to further process intensifications and volume reductions.

In this presentation, we will discuss our work on several upstream challenges that must …