A Look into Biologic Scale-Up Strategies

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Optimizing cell-culture processes for monoclonal antibody (mAb) production is a key factor in scaling up manufacturing to commercial levels. To maximize cell-culture output, strategy is required that takes into consideration factors such as cell culture media, environmental monitoring, and process controls. Yet, despite decades of process optimization, there remain stubborn challenges—or new challenges that have arisen–during the scale up of cell culture processes.

Persistent challenges

Previous process optimization in mAb production meant that mAb titer and cell density were once low, and a successful scale up was determined through quantity, that is, achieving the same mAb titer at larger scales, notes Masa Nakamura, PhD, global bioprocess science specialist at 3M Separation and Purification Sciences Division. However, he continues, now that the biopharma industry is in an era of higher titer and cell density, the focus has shifted from “quantity” to “quality.”

“Engineered modern mAb products require higher degrees of process control to produce high-quality products along the scale up, which demands a holistic approach in cell-culture scale up,” Nakamura says.

Not only do biomanufacturers need to control cell densities and titers, but they must also manage post-translational modifications such as glycosylation profiles, says Susan Riley, leader, Cell Culture business, Cytiva. As titers have increased, the balance of the bioproduction workflow must also be well managed, she states.

“It does little good to increase the expression of protein to 10 g/L [for example] if significant yield will get lost in the downstream filtration, purification, or even final filling steps,” Riley cautions. In terms of upstream hardware—with the possibility of failure, even in late-stage clinical trials—Riley notes that biomanufacturers want to minimize risk by delaying critical capacity decisions until as close to final approval as possible.

Andy Topping, PhD, chief scientific officer at FUJIFILM Diosynth Biotechnologies, points out that, over time, as expression systems have become more efficient, the viable bioreactor scale has decreased, with many commercial products able to be produced in few 2000-L single-use bioreactor runs per year and the industry has responded by developing both scale-up and now scale-out strategies. This trend has placed enormous pressure on solution management and downstream processing in legacy assets, and novel fluid management systems, including in-line dilution and blending, are now maturing. “As a result, scale and capability of single-use process systems with full process closure are continuously expanding,” says Topping.

Meanwhile, processes that are expected to require manufacture at very large scale will often need to be transitioned from single-use bioreactors to stainless-steel bioreactors, Topping emphasizes. Although recent innovations have made the performance of these single-use systems closer to stainless steel, many processes will still have been developed and operated in bioreactors with fundamentally different engineering, mixing, and gas transfer capabilities, where harvesting will be conducted by depth filtration rather than centrifugation. Scale up of these processes therefore also means translating the process correctly to the new systems, according to Topping.

“Moving forward, the industry finds itself in a secular trend of increased diversity as more capable systems and fundamental understanding become available at an accelerated pace,” says Thomas Page, PhD, vice-president of Engineering and Development, FUJIFILM Diosynth Biotechnologies.

Scale-up innovations

Multiple innovations have arisen for the sake of making cell culture scale-up more efficient. According to Topping, the most fundamental innovations have been improved cell lines and media, which enable the higher cell densities and higher titers. That increase, in turn, allows many processes to use a scale-out approach at or near to the scale used for clinical supply, rather than a scale-up approach.

Single-use manufacturing systems are also an innovation that by design enables an optimum solution for closed processing and better aseptic techniques as well as ease of use. Single-use platforms offer both standard and flexible workflows. Furthermore, the impact of having a closed process reduces contamination risk while enhancing efficiency and predictability, benefits that cannot be overstated, says Page.

“For example, use of full isolators for flask work significantly decreases flask contamination rates and resultant schedule and manufacturing cadence impacts. Integration of standard and custom sensors with the single-use consumables further reduces risks of contamination,” Page states.

“Full understanding through detailed risk assessment of process closure allows facilities to operate with multi-product cell culture on a predictable high cadence rate. This significantly reduces overhead and significantly increases availability of capacity to the market,” Page adds.

Meanwhile, innovations for bioreactors include new fluid mixing formats that create more efficient oxygen transfer with lower shear; this helps to support higher oxygen demand for both intensified and perfusion cultures, says Nakamura. Outside of the bioreactor and when looking at the harvest and clarification of cell cultures, single-use centrifuge and chromatographic clarification are new additions to the bioprocess toolbox for high density cell culture scale up, he adds.

“However, for cell retention in perfusion cultures, the cell retention devices would need to evolve further to include more precise pore membrane structures that achieve high precision separation with minimal clogging,” Nakamura states.

Page makes the observation that growth in the cell therapy space, which has resulted in the concomitant development of robust cold storage containers for mammalian cells, has opened the door for the use of high-density cell banks. “High-density cell banks offer the potential to massively decrease labor, risk, and time in plant. It is a critical capability for maturing clinical phase perfusion offerings,” Page states.

Other developments, such as novel sensors (e.g., Raman soft sensors), have allowed greater real-time insight into process performance at all scales, Topping also states. These novel sensors are being applied in the continuous monitoring of metabolites, such as glutamine and lactate, as well as mAb titers in the bioreactor, Topping explains.

“Other emerging technologies benefiting large-scale biotherapeutic manufacturing are artificial intelligence data science for integrated processing modeling, robotics, automation, and the related transition to paperless program management and data management systems,” says Page. Digital twins, for instance, offer the potential for enhancing both speed-to-market and post-approval optimization. In-silico modeling offers the potential to select optimal cell lines and modified sequences that can optimize across a broad arc—from potency to selectivity to manufacturability. All these benefits offer the promise of increased patient access and sustainability, adds Page.

Innovation compatibility with single-use technology

Vaccines were traditionally manufactured in chicken eggs or in stainless-steel systems, but the expansion into single-use technologies, and particularly the growing adoption of single-use bioreactors, has helped transform the global capacity for vaccines, both in terms of efficiency and flexibility, Riley points out.

Specifically, Cytiva has expanded the single-use technology approach to include viral vector-based therapeutics, including oncolytic vaccines, in the design of its prefabricated, modular facility (KUBio, Cytiva); recent expansion of its modular facility design is aimed at accelerating the manufacture of and reducing the risks of viral vector-based therapeutics.

Nakamura, meanwhile, notes that innovations in the harvest and clarification of cell cultures involve single-use technologies, specifically single-use centrifuge and chromatographic clarification processes. “These technologies are compatible with current single-use bioprocess workflows and would often provide process economic advantages for mAb production in high density cell cultures,” he states.

Page emphasizes that it is now possible to engineer large, capable single-use assets that are fully electric and devoid of any steam production. This innovation, coupled with high density production and process closure means that energy usage could be decreased up to an order of magnitude; however, such an energy decrease will need to be verified as next-generation plants come online, Page explains.

It is important to keep in mind that many innovations taking place are agnostic to the production platform, but some are specific to either a single-use or stainless-steel system. Improved bioreactor performance, increases in scale, development of more single-use harvesting options, and novel cell retention devices for perfusion have all pushed bioproduction capabilities forward, according to Page.

“In general, it is important for any company working in this space to have a good risk-based model to understand the type, scale, and pathway for hazards and to have a clear, explicit control strategy. Aligning this understanding end-to-end in an organization is critical,” says Page.

For example, to optimally move a development project from clinical single-use manufacturing through to large-scale stainless-steel manufacturing, one needs to understand the impacts of product and unit operations. This understanding must then be mapped out to differentiate between product-specific requirements and facility and equipment controls. This mapping must be done in such a way that the chemistry, manufacturing, and controls sections may be developed to fit the target assets used for the product lifecycle, explains Page.

“With the target lifecycle map or potential pathways mapped, process knowledge may be subjected to directed evolution such that costly remediations, such as equipment changes, validation extensions, or redevelopment may be avoided,” says Page.

Another area for mapping back and forth between single-use and stainless-steel systems requires the development of unit operations that were previously unavailable in a single-use system. These include continuous centrifugation and homogenization, for example, says Page. However, these topics go beyond mAb processing and will have profound impact on advanced therapies, microbial products, and vaccines.

“The key is for industry to have a mappable process that allows easy transition into and then later back out of large-scale stainless-steel production and product lifecycle. Ideally each product could be produced in its most efficient embodiment over this lifecycle. As these capabilities mature, there will clearly be opportunities for hybrid deployment,” Page states.

Strategy effectiveness

To have successful scale-up, it is imperative that product consistency, in both quality and quantity, be addressed. Having product consistency requires a holistic approach in the upstream stage, the clarification step, and downstream purification, notes Nakamura. Focusing solely on mAb titer would not be enough to provide successful scale up and consistent manufacturing.

“An example of an effective strategy in scaling up mAb cell culture to commercial scale includes optimizing cell culture from both upstream and downstream point of views,” Nakamura intones. “Cell culture is not only the source of products, but also the source of all soluble and insoluble contaminants (cells, debris, host cell proteins, DNA, product variants), which need to be removed in downstream. Harvest criteria and timing can be intentionally selected to balance the high titer and low contaminants to achieve the most overall benefits.”

In Cytiva’s experience, Riley notes, the most effective strategies for successful cell culture scale-up involve a deep understanding of how the entire bioproduction workflow must work together in a balanced way. “Paying attention to the vortex speed or air sparging in the bioreactor, the crossflow and pressures across TFF [tangential flow filtration] membranes, and the aspect ratios associated with resins in purification columns all contribute to the ultimate yield of the final product. And at the end of the process, if the therapeutic is of absolute premium value, such as in gene therapy, the use of robotic aseptic filling [is important] so that not a drop will be lost,” she explains.

Riley further notes that automation and digital strategies are solutions that can be deployed to streamline the cell-culture process, enable the continuous manufacturing concept, reduce process variability, and lower production costs. “It should be noted that collaboration between tool/solution providers and therapeutic developing organizations is greatly beneficial for accelerating the technology advancement,” she adds.

As molecules diversify and precision medicine continues to advance, says Riley, there will be smaller production runs that require maximum flexibility, and a variety of single-use components will become increasingly important. “And, frankly, some of the newer modalities use little cell-culture media at all but require a deep understanding of the role that other fluid components can play in processing,” she states.

Meanwhile, Page explains that traditional strategies, including mapping exercises from scale-to-scale, allow small-scale design of experiments, which are representative of production-scale parameter space and therefore useful. In the past, stainless-steel bioreactors were designed within a relatively narrow range of capability, but today, the advent of single-use bioreactors has required the industry to develop ways of migrating from stainless-steel to single-use systems as well as across different vendor-supplied single-use bioreactors.

“Today, many programs may migrate across different bioreactors over their lifecycle. The design-of-experiment approach will continue and will likely be enhanced as next-generation single-use bioreactors with enormous operation space become available,” says Page.

The key issue for contract development and manufacturing organization (CDMO)-specific operations today, as Page sees it, is for CDMOs to select equipment that have the broadest operating space and to nest such equipment in a flexible facility. Facilities must increasingly have the broad capability to efficiently support different modes of operation, Page concludes.

About the author

Feliza Mirasol is the science editor for BioPharm International.

Article Details

BioPharm International
Volume 36, No. 8
August 2023
Pages: 15–18

Citation

When referring to this article, please cite it as Mirasol, F. A Look into Biologic Scale-Up Strategies. BioPharm International 2023, 36 (8), 15–18.