A Flexible, Intensified Seed Train to Boost Upstream Productivity

Yuliya Mikhed

September 28, 2022

4 Min Read

Biopharmaceutical companies continue to invest significantly in technologies that support process intensification (PI), including equipment for high-density and perfusion-mode cell cultures. During a July 2022 Ask the Expert webinar, Yuliya Mikhed (product manager for Biostat RM bioreactors at Sartorius) highlighted drivers for PI solutions, then described how an intensified modular seed train could help users to increase cell culture productivity and flexibility while reducing cost of goods (CoG).

Mikhed’s Presentation
Sartorius customers have identified several drug development “pain points.” Biologics developers are under pressure to shorten commercialization timelines for new therapies. However, mounting regulatory expectations are compelling companies to increase yields and generate enough material for testing. Companies such as small contract manufacturing organizations (CMOs) need flexible equipment to enable multiproduct manufacturing and to reduce turnover time between programs. And scale-up to commercial manufacturing remains error prone, time consuming, and resource intensive.

PI can help to increase yields and improve process economics, Mikhed explained. Sartorius customers that have implemented some form of PI have reported two- to threefold increases in overall productivity, footprint reductions of 50–70%, accelerated timelines for facility construction, and >30% reductions in CoG. Intensified processes based on single-use systems also enable flexible facility configurations and multiproduct manufacturing environments.

During upstream operations, Mikhed continued, an intensified modular seed train could provide significant value in product yield and operations. Thus, Sartorius has established a platform that leverages Biostat RM wave-mixing cell culture systems, third-generation Biostat STR bioreactor systems, and the Biobrain automation platform.

Conventional seed trains use three or four culture stages performed over 24–31 days to generate material for inoculating a production-scale bioreactor. Often, such a workflow involves multiple types of equipment and associated consumables. The intensified Sartorius process calls for a two-stage seed train in Biostat RM 50 and RM 200 bioreactor systems operating in perfusion mode over 21–23 days. Alternatively, users can apply a three-stage seed train, with N – 3 and N – 2 cultures running in batch mode and the N – 1 step performed in perfusion mode.

Those intensified workflows, Mikhed continued, are designed to minimize hardware requirements and long-term capital expenditure. Eliminating process steps can reduce equipment and consumables needs. Mikhed added that Biostat RM systems can operate in perfusion mode when paired with single-use bags with integrated membranes, eliminating need for an external cell retention device. Because the bioreactors can run in batch mode, users can choose to implement intensified culture protocols later in process development. Early adopters of the intensified Biostat RM workflow report that they can produce six more batches per year than what would be produced in traditional configurations.

Mikhed highlighted her company’s work with Intas Pharmaceuticals to intensify a five-transfer seed train beginning with shake-flask cultivation (N – 4) and culminating in inoculation of a 4,000-L bioreactor (N). The original process achieved viable cell counts (VCCs) of 7–8 million cells/mL of culture with 95% viability over 14–16 days of fed-batch culture. The intensified process needed to maintain an existing feeding strategy, operate in perfusion mode at the N – 1 stage, and generate 50 million cells/mL with >95% viability.

Leveraging Biostat RM technology and Biobrain automation, Sartorius established a three-transfer protocol. Cells were grown in shake flasks, transitioned to 25-L perfusion cultures in Biostat RM bioreactors, and used to inoculate a production-scale vessel. The new workflow generated VCCs of 50–150 million cells/mL with viability of 98–99% over 6–8 days of cultivation. Cultures based on the intensified process achieved growth rates and yields comparable with those from a 50-L control culture. Ultimately, the intensified workflow saved considerable time and capital while improving culture productivity 10-fold in a substantially reduced footprint.

Questions and Answers
Do Biostat RM bioreactors provide controls based on viable cell density (VCD) levels? Single-use probes are integrated into Biostat RM systems to measure relevant parameters. Users can program a Biobrain controller to automate adjustments to bioreactor parameters (e.g., cell bleed rates) based on incoming data.

What VCDs can the Biostat RM and STR systems achieve? Users of Biostat RM 50 and RM 200 systems report VCDs up to 170 million cells/mL and 100 million cells/mL, respectively. VCD can reach 190–200 million cells/mL in a Biostat STR system, although
150 million cells/mL is consistently achievable.

Find the full webinar online at www.bioprocessintl.com/category/webinars.

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