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Improving Continuous Process Efficiencies Through Development of a High-Performing Perfusion Media
The biopharmaceutical industry has a growing interest in continuous processing as a way to improve manufacturing efficiency and facility utilization. One reason is that continuous processing has proved a very successful manufacturing model in many other industries. Furthermore, certain continuous technologies have already been incorporated into existing biopharmaceutical manufacturing with many benefits. One continuous process technology that is currently being utilized in some commercial biopharmaceutical manufacturing is perfusion culture.
In several applications, perfusion culture provides many benefits over traditional fed-batch processing and has been used successfully for many years to manufacture unstable proteins. These benefits include improving product quality and stability, better scalability, increased facility utilization and cost savings. Please see our blog “Perfusion Bioreactors -With so much to offer they deserve a closer look,” for more details on how perfusion culture operates.
In a recent webinar titled “Development Strategies for High-Performing Perfusion Media,” Andreas Castan, Ph.D., Staff Scientist, GE Healthcare Life Sciences, did an excellent job discussing the benefits of continuous processing and describing enabling technologies including development of a high performing perfusion media.
Future of Biopharmaceutical Manufacturing
Dr. Castan, began his talk by discussing his vision of the future of biopharmaceutical manufacturing and where the industry is heading over the next 25 years. One area highlighted was continuous manufacturing and the associated benefits. These benefits include, providing a cleaner, flexible, more efficient manufacturing platform. Another benefit stated was that the FDA and other regulatory institutions have encouraged the incorporation of a continuous process in biopharmaceutical manufacturing.
Drivers Toward A Continuous Process
Dr. Castan continued the talk by describing the drivers that have encouraged the biopharmaceutical industry to consider incorporating continuous processing. One of these drivers is the success and proven track record of perfusion culture in manufacturing unstable proteins including coagulation factors and enzymes.
Additional drivers include:
- High volumetric productivity
- Closed system operation and elimination of intermediate hold steps
- Integrated operations vs. several unit operations
- Less manual operations and a high level of automation
These drivers address challenges currently found in traditional fed-batch processing and help meet future goals for the industry. For example, in continuous process, the operations are integrated with one flowing into the other vs. in batch where there are several unit operations with storage steps between operations. These storage steps necessitate monitoring for in process controls after each step and add to the workload for QA/QC groups. In addition these hold steps requires large tanks for storage. Thus adding time and capital to the manufacturing process. Furthermore, continuous processing by nature requires a high level of automation. Automation reduces the amount of manual operations, thus freeing operator time for other tasks and reducing the risk of operator error.
More specifically, a continuous process can address several challenges with respect to the key areas of product quality and manufacturing flexibility. Those challenges include:
- High and consistent product quality through operation at optimal steady state conditions
- Lower level of impurities due to high cell viability
- Improved product quality through decrease of product hold time
- Minimizes bioburden risk through closed system operation
- Less intermediate testing through elimination of hold steps
- Compatibility with QbD and PAT methodologies
- Flexibility due to smaller equipment size
- Compatible with disposable products
- Rapid capacity increase/decrease
- Short cycle times and simplified logistics
- Simplified transfer to new facility through modularity, standardization and small equipment foot print
The idea of incorporating a continuous process throughout the entire biopharmaceutical manufacturing process is relatively new and as such, enabling technologies needed to be developed in order to achieve success. Technologies like improved process control, single-use products and cell retention systems made the opportunity of continuous processing in upstream possible.
Enabling technologies include:
- Process control
- Single-use products
- Facility design
- Continuous downstream processing
- Cell retention systems
- Cell culture media design
In order to have a fully continuous process from upstream through downstream, downstream continuous processes also had to be developed. This included creation of multi-column chromatography, inline conditioning systems and straight through processing. For more information about continuous downstream processing, please see our blog “Continuous Downstream Processing – A Tool to Address Key Manufacturing Challenges.”
Perfusion Cell Culture
In upstream, perfusion culture is a key component of continuous manufacturing and in any culture system whether it be fed-batch or perfusion, media is a cornerstone for cell health, viability and productivity. Cell culture media design and optimization is key to achieving the highest productivity in manufacturing.
Dr. Castan describes in his talk how many perfusion processes are based on maintaining a constant cell specific perfusion rate (CSPR). He goes on to describe two scenarios, one involves high CSPR, which means that the medium is not adapted to cell metabolism and many medium components leave the bioreactor unmetabolized. High CSPR also requires high volumetric perfusion and the operation is ultimately not cost-efficient. In contrast a low CSPR means that the cell culture medium is meeting the cell line’s nutritional needs. The goal is to use high performing media, which allows you to run your process at low CSPR.
Highlights of Case Study to Develop High Performing Media for Perfusion
In an effort to optimize perfusion culture productivity, GE Healthcare Life Sciences began a study to develop a high performing perfusion media. They began by using their ActiCHO™ platform production media and two ActiCHO feeds. The study was conducted using the ReadyToProcess WAVE™ 25 and 2 liter perfusion Cellbag™ bioreactor chambers with floating filters. For the host they used a monoclonal antibody producing cell line licensed from Cellca GmbH. Dr. Castan, went on to describe the method for the case study and the results, which I have summarized below.
Two Approaches to Media Design
GE Healthcare Life Sciences utilized two different approaches to media design and then compared the two approaches – batch approach vs. steady state approach.
They began the batch approach design by conducting a screening design of experiment (DOE) study and an optimization DOE study. Lastly they verified the media by using it in a perfusion culture run. They found that the medium performance could be enhanced with the addition of feed solutions and ultimately identified an optimum blend of the platform media (ActiCHO) with ActiCHO Feed A and ActiCHO Feed B. By using this media they were able to achieve 1 gram/liter titer at a volumetric perfusion rate of 1 reactor volume per day with a cell growth rate that was low.
Steady State Approach
Using the steady state approach the media was developed using perfusion culture and spent media analysis was conducted. This analysis informed a new medium design that was then verified in perfusion.
During the process, Dr. Castan explained that the team was able to objectively measure specific productivity and the amino acid consumption rate as the perfusion rate was decreased step wise from 100 to 25 pL/c/d. They also examined the impact of CSPR on cell specific productivity and on amino acid consumption and created a heat map for 7 limiting amino acids at different CSPR. By using this data they were able to design a high performing perfusion media optimized for their process.
The team then verified that the medium developed performed well in perfusion and the result was 1.5 gram per liter titer at a volumetric perfusion rate of 1 reactor volume per day with low cell growth rate.
Conclusions From the Case Study
Dr. Castan summarized that both approaches were successful in developing a high performing perfusion media and were methodologies that could be applied to other types of cell culture media. They both offered a quick route to an efficient upstream perfusion process with the batch fed design taking 2 months and the steady state approach taking 4 months.
Other Points to Consider:
- The steady state approach showed improved performance, but required twice the development time compared with the batch process.
- Use of the steady state developed media resulted in a final process with more than a 75% decrease in cell specific perfusion rate (CSPR) compared with starting process conditions.
- Using this information it was calculated that medium consumption per cell could be decreased 4.5 times to a CSPR of 20 pL/c/d.
Dr. Castan stated that during a 3 week time period, using a 500 L process run with 50 million cells/mL, the optimized media compared to the original process would save more than 35,000 liters of media. Thus saving more than $550,000 in USD assuming a cost of 15 USD/L media. This example demonstrates the tremendous upside in developing a high performance perfusion medium.