Introduction

Accelerate activity. Improve predictability. Drive higher process efficiency. Increase quality. Lower cost of goods sold (CoGS). Secure supply. In an era where the biomanufacturing wheels turn faster by the day, where the stakes are higher and the choices seemingly endless, it is easy to become overwhelmed. How can you make good biomanufacturing decisions and develop robust long-term strategies when the environment is constantly changing? Whether a political shift affects your product or market, a natural disaster disrupts your supply chain, or some other external influence suddenly alters your plan, you have needs that must be met and people counting on you to deliver product. In such situations, extensive bioprocessing knowledge and experience are invaluable tools that will make a difference. Good insights about technical advancements, process economy implications, and sustainability aspects can help improve your individual decisions and increase the likelihood of success.

Recently, four renowned subject-matter experts shared their insights and knowledge in four areas of biomanufacturing during the GE Bioprocess Insights webinar series with BioProcess International online. These webinars covered upstream through downstream bioprocessing, spotlighted strategies for implementing single-use technologies, and highlighted process economics of vaccine manufacturing. The topics were chosen with the objective to cover a broad range of topics related to biomanufacturing and provide insights into recent developments relevant for the overall bioprocess. The speakers discussed a variety of tools, trends, and technologies that can improve process efficiency, economy, and sustainability:

• Upstream process intensification and how clever process design can decrease your investments, decrease required floor space, and cut days from your overall bioprocess
• How downstream efficiencies can be achieved with a number of different options available in a virtual toolbox from which manufacturers can cherry-pick what is suitable for each specific process
• The environmental implications of implementing single-use technology compared with traditional reuseable options and how this is related to geography and process conditions
• Modern vaccine manufacturing and how a legacy process can be made more efficient with single-use technology and core-bead chromatography.

Each webinar was followed by a live question and answer (Q&A) session. In each case, questions from an engaged audience led to an enhanced discussion for the participants.

In this BioProcess International insert, you will find written summaries of the four webinars and some key questions that were discussed during the Q&A sessions. In addition, all webinars including those discussions are available on demand here.

Each topic and its respective insights and recommendations can be digested separately; however, the combined insights are what can really make a difference. Working in “silos” is a common pitfall for biomanufacturing companies and individuals. Gaining insights not just within one area of the bioprocess but to develop a good understanding of the overall picture will help eliminate the silos and improve the overall situation in a biopharmaceutical company.

Our industry is still evolving and blending novel ideas into existing processes. A certain amount of innovation and flexibility is required to stay ahead in this competitive environment. In addition, there are external factors we can never control. What we can influence, though, is our ability to be prepared with knowledge and insights that enable us to deal with those uncertainties in the best way possible. In doing so, we’ll be prepared to make not just good decisions, but those that will make a positive difference for our companies.

Brandon Pence
Global Marketing Leader, BioProcess GE Healthcare

GE, GE monogram, ÄKTA, Capto, Cellbag, Cytodex, HyClone, KUBio, ReadyToProcess, ReadyToProcess WAVE, WAVE Bioreactor, and Xcellerex are all trademarks of General Electric Company.

Intensified Seed-Train Strategies: Improve Productivity and Process Economy
with Andreas Castan

Andreas Castan, a staff scientist in research and development at GE Healthcare Sciences, discussed improving productivity and process economy through intensified seed-train strategies using perfusion. He outlined technical details and how such strategies affect capacity use, plant throughput, and costs of labor and batch production.

Recent decades have brought steadily increasing cell culture titers to several grams per liter, bringing down bioreactor volumes in turn. This has decreased cost of goods (CoG) to <$100/g product. But process bottlenecks are limiting how low CoG can go, and <$20/g is needed to make biopharmaceuticals available to the whole world.

To intensify an existing process, companies can either shorten the production bioreactor culture or focus on a perfusion-based seed train that feeds into it. The first option would apply perfusion in an N – 1 bioreactor to inoculate the production bioreactor with 20× more cells. But published studies indicate that approach could compromise product quality. So Castan used case studies to focus on intensifying the seed train with a high-density cell bank that is expanded in one step to generate enough cells to inoculate a 2,000-L bioreactor.

In the first case study, one 2-L GE perfusion Cellbag™ bioreactor was sufficient to produce a cell bank with 4.5-mL vials at a concentration of >50 million cells/mL. The whole procedure used one bioreactor without a centrifugation step. And the cell bank maintained good quality relative to a comparator. In the second case study, a 4.5-mL high–cell-density vial was used to inoculate a 20-L Cellbag bioreactor, with cells then expanded in one step to 10 L and >50 million cells/mL. That density was sufficient to directly inoculate a 2,000-L batch bioreactor.

Castan addressed process economy next, saying that a high-cell density cell bank makes traditional shake-flask steps redundant in a seed train and that perfusion can help to omit intermediate bioreactors. To quantify the influence on process economy in capacity use, plant throughput, and costs, GE created a production process-economic model. Results suggested that the high-density cell bank provides a time gain of five days for each manufacturing campaign, allowing for more batches to run each year. Fewer bioreactors fit in a smaller facility footprint. And hands-on time could be reduced by nearly half. So the strategy reduces upstream production costs per batch by 12–20% compared with a traditional approach.

Questions and Answers
Does this affect product quality attributes? We have investigated process performance in terms of in-process controls and quality attributes and haven’t seen an impact.

Does this approach work with bacterial expression systems? Because bacteria double faster, the benefits might not be as clear. Reaching very high cell densities would require a lot of oxygen that might be difficult to transfer. So I would say that this application is more suitable for mammalian culture.

Does media composition or perfusion rate need to change for the higher cell density? Of course, you need to supply more nutrients when increasing cell density. We have solved that by keeping cell-specific perfusion rates constant at >100 pL/cell/d so as not to push the system — without developing a specific medium for this application — and maintained a very high viability.

Are there any problems with oxygen supply during perfusion? No. Dissolved oxygen was regulated in a cascade with rocking speeds, and O₂ supplementation was sufficient to support high cell densities without reaching the limits of the system.

What is the biggest challenge in establishing this method? There were no technical challenges. It was more a matter of convincing quality assurance personnel to explore the technology.

Are you studying cell-bank stability? We have done limited stability studies on our high-density cell bank over a couple of weeks and have seen very stable revival in those tests. For a commercial cell bank, you would do more, also looking into cell properties at the end of a production run and so on. We have focused on performance in a production bioreactor.

An Evaluation of Process Economy: Vaccine Manufacturing with Single-Use Technology
with Dr. Mats Lundgren

Mats Lundgren, a custom applications director at GE Healthcare’s life sciences business in Sweden, discussed the need for updating vaccine processes and provided case studies involving single-use process technologies. He also described the economic implications of introducing such technologies upstream and using modern downstream technology.

Manufacturing costs can represent >50% of the overall costs of vaccines, and companies face severe cost pressures. Yields from labor-intensive processes can be low. Dedicated facilities built for decades-old processes are based on stainless steel reusable equipment. Technology transfer requires significant expertise, and scale-up is difficult with technologies such as roller bottles and centrifugation. And regulatory quality requirements are increasing, especially regarding cross-contamination risks, batch variability, and animal-derived components. Disposables address many of those concerns.

GE has compiled five years of data covering the process economies of single-use technologies, and Lundgren showed a table listing results from eight companies — large and small, sponsors and CMOs. They reported increased consumables use but lowered facility costs, footprints, and labor. Facility build times were reduced, as were batch turnover times and consumption of water and energy. One company reported a 30% increase in operating capacity. Overall, the cost of goods (CoG) is reduced significantly.

Next, Lundgren presented some internal results that included data from customers. After a detailed scheduling of unit operations, GE calculated capital investments in terms of hardware and facility as well as costs of labor, disposables, raw materials, utilities, and waste disposal.

From an upstream case study, Lundgren reported “increased profit opportunity” based on higher capacity and lower costs with microcarriers in single-use bioreactors. From a downstream case study, he reported that core-bead chromatography could enable single-use operations and facilitate scale-up to industrial scales. It also lowered downstream operational costs and increased productivity compared with size-exclusion chromatography. Thus, single-use technology helps increase vaccine profits significantly by lowering production costs. It can also help companies increase their plant production capacities.

Question and Answers
Are presterilized microcarriers commercially available and free of animal-derived components? We are launching two versions soon: animal-component–free Cytodex™ 1 for human vaccine support (presented here) and gelatin-coated Cytodex™ 3 for the veterinary industry.

How are the microcarriers presterilized and delivered? What is their estimated shelf-life? They are gamma irradiated and come in bottles that can be connected to bioreactors. We expect the shelf life to be similar to that of standard microcarriers.

How are they loaded into a single-use bioreactor? You can do that in different ways. For example, you can weld tubing to connect the bottle and then use air to move carriers into the bioreactor.

Can core-beads be sterilized? Yes. They are based on a robust resin you can run in aseptic mode. You need a clean-in-place procedure. Sterilizing whole plastic columns is tricky with steam-in-place, which is better with stainless steel.

How many times can core beads be reused? That depends on the seed material and cleaning procedures. We can supply cleaning protocols. Because of the strong ligand, you need harsh cleaning conditions, but you can reuse the beads a number of times.

Are there applications for the core-bead chromatography other than virus purification? You can use it for other things: if you want to separate something small from something that is large. You can even use it for cell purification. You can also use it for other types of large molecules depending on the pore cut-off size. For example, Capto™ Core 700 beads can purify very large protein molecules, polysaccharides, and DNA as well as viruses. Monoclonal antibodies are a bit too small to purify using this current technology because they actually enter the beads.

Are you aware of single-use technologies being used for BSL-3 or higher classified manufacturing? That is being investigated quite a lot these days. You need to ensure containment and design the facilities and surrounding equipment very carefully.

Are the cost savings different for different vaccines? This is something you would have to look for with every vaccine you’re interested in. If the manufacturing technology and cell line are similar, you could expect similar results.

Unlocking the Potential for Efficiency in Downstream Bioprocesses
with Madhu Raghunathan

Madhu Raghunathan, product strategy leader for downstream bioprocessing equipment at GE Healthcare, introduced the concept of downstream process intensification. He focused on chromatography unit operations, on creating functionally closed systems to increase process efficiency, on single-use technologies, and on continuous/connected processing. Automation ties them all together. These solutions can be applied independently or jointly to keep up with changing process and market dynamics toward the goal of process intensification.

Pack-in-place columns require several manual preparation steps. Automated columns simplify the process through axial-compression technologies and pretested/preverified methods, with significantly reduced bioburden risk and simplified maintenance. Raghunathan described a case study in which an automated approach provided ~50% reduction in time spent packing/unpacking and maintaining columns compared with a manual pack-in-place method.

Prepacked columns for campaign use can eliminate seven of 11 common steps associated with column packing. Before choosing them, users must consider production volume, scale, manufacturing frequency, and flexibility. Companies need internal know-how and must consider capital/operating expenses and replicability. A knowledgeable vendor can help them decide between cleanable and reusable and prepacked formats.

Raghunathan also discussed the benefits of single-use technologies. To successfully and properly implement them, companies need to ensure that their vendors have a robust supply chain. Users thus won’t have to worry about issues such as extractables/leachables, bag integrity, and lot qualification to reap the full benefits.

Next, Raghunathan introduced the concept of straight-through processing in continuous chromatography using periodic-countercurrent chromatography. Key benefits include equipment size reduction and significant gains in resin capacity use. Fully continuous bioprocessing (from a perfusion bioreactor through to product concentration) is especially useful for stability-challenged biologics.

Straight-through processing isn’t limited to continuous chromatography. Elution from one process step can be directly loaded to the next without hold-up time or intermediate processing. That reduces total equipment footprint and overall cycle times in downstream processing. A related topic, inline conditioning, involves formulating buffers on demand from basic stock solutions. Intelligent algorithms accommodate varying input parameters to generate desired outputs.

Finally, Raghunathan described the degrees of automation. Disconnected “islands” can use the same or different platforms, with their data output later integrated for further analysis. Ultimately an integrated user interface can run all workflows together, with unit operations combined under a single “recipe.” The end goal would be a facility run from a command center with all data acquisition and analytics integrated into a single “back-end” while all processes run dynamically based on quality by design (QbD) and process analytical technology (PAT). That provides for real-time product release and dynamic process adaptation to varying inputs.

Questions and Answers
What is the biggest challenge in implementing continuous processing downstream? The complexity of continuous processing is a challenge. People are concerned about contamination and facility fit. We need robust sensors for product quality and technology for ensuring process sterility.

Do you know of a commercial process using these methods? About 18–20 commercial processes are using perfusion in upstream processing. We haven’t found any commercially approved processes with continuous downstream processing.

How can “intelligent” columns handle significantly different resins? They use axial compression methodology and leverage preverified packing methods for a broader “operating range” and deliver robust outcomes across that range.

What is the role of PAT here? When running a continuous process over an extended period, a process should adjust to minor variations in input parameters. So PAT has a significantly greater role to play here than in batch processing.

What is the minimum process size? Continuous processing is especially suited to smaller scales given that large-scale processes run over longer periods and already may be optimized to maximize efficiency and lower cost of goods (CoG).

Single-Use Technology and Sustainability: Quantifying Environmental Impact
with Bill Flanagan

Bill Flanagan leads GE’s Ecoassessment Center of Excellence and chairs the American Center for Life Cycle Assessment’s board of directors. He discussed the environmental impact and sustainability of single-use technologies, beginning with bioprocess industry trends and challenges: targeted therapies and novel biologics; cost pressures; globalization and localized manufacturing. Biomanufacturers need to do more within smaller footprints with higher productivities and facility use rates. Over the past decade, adoption of single-use technologies has enabled them to do so. But Flanagan emphasized the importance of environmental lifecycle assessments (LCAs).

LCAs provide a holistic perspective of the entire value chain from cradle to grave and “a wide variety of environmental-impact categories, not just carbon footprint.” Insights gained from comparing single-use and traditional technologies in this way can reveal environmental benefits and impacts related to bioprocess operations. Flanagan described an LCA study GE performed on its WAVE Bioreactor™ and ReadyToProcess™ systems. Many people in the industry expected disposables to be less environmentally sound than traditional technologies. But results showed the opposite. He pointed to significant reductions in energy, water-for-injection (WFI) and process water, clean steam and cleaning chemicals as major drivers in favor of single-use technologies.

Along with Quantis International and Biopharm Services, GE is performing a new LCA study to incorporate aspects of geography, a wider range of process scales, and equipment use (100% conventional, 100% single-use, and hybrid scenarios). This was partly in response to customer interest and market trends. The original study focused on monoclonal antibody (MAb) manufacturing; the new study will expand on that to include vaccines and hybrid facilities; Xcellerex™ bioreactors, HyClone™ growth media and buffers, ÄKTA™ ready chromatography systems and ReadyToProcess chromatography columns; and a broader range of end-of-life disposal options.

After describing the study methodology, Flanagan presented some preliminary results comparing a traditional stainless steel facility with single-use technology in a retrofit traditional facility as well as an optimized floor-plan design such as a modular KUBio™ facility. The study initially considers MAb production at 2 × 2,000-L scale with 6 g/L titers in a 10-batch campaign.

“What we’ve seen so far very much correlates with our previous LCA results,” Flanagan said. “For the scenarios we’ve looked at so far, the single-use process technology exhibits lower life cycle environmental impact than traditional technology. End-of-life contributions are pretty small relative to the overall life cycle. As we start looking at different installation geographies, we see that single-use technology exhibits varying levels of environmental-impact benefits.”

Next the study will example a broader range of MAb process scales and hybrid configurations, then vaccines at smaller scales.

Question and Answers
Did the LCA include water consumed by the disposables manufacturer? Yes, we looked very closely at carbon, energy, and fresh-water consumption.

As the batch number rises, does the single-use savings decrease? A 10-batch campaign well represents most campaigns, as for more batches, everything expands fairly linearly. So the savings would be proportionate.

In the LCA, is the single-use equipment landfilled or recycled? Our default assumptions would be incineration without heat recovery. We also think about landfill for some materials. There is some recycling in the model, and we will be looking at a wider variety of end-of-life options (e.g., incineration with energy recovery). We have actually solicited input from some manufacturers about their disposal mechanisms — which also varies by geography.

What processes for recycling did you look at? So far we are looking at fairly straightforward recycling of materials. As we get further into this, there could be a broader range of recycling involved. And it’s going to be different for the packaging. We’ll be looking much more specifically at that as we get further into the study because many end users are interested in these programs. We recognize the importance of end-of-life management, and we want to make sure that we have the LCA modeling to help people make decisions about some of those ongoing programs to deal with solid wastes.