Large-scale single-use bioreactors prove successful as limits are tested.
Commercial-scale success
The 2000-L-scale SUB has been the mainstay of large-scale SUBs since its introduction. Yasser Kehail, Biomanufacturing Business Development leader at Cytiva explains that 2000 L is the sweet spot for large-scale manufacturing. Among the reasons for this are, first, that recent technological advancements in cell-culture media and cell-line development have increased product titer and yield significantly. What was once produced at 15,000-L-scale in stainless-steel bioreactors 10–15 years ago can now be produced at smaller sizes, such as 2000 L, states Kehail. Second, current biological entities are designed for specific patient populations, which are smaller and based on specific biomarkers, such as a gene or specific protein. “Hence the clinical material needed for studies and clinical trials are lower in terms of scale,” Kehail says.
According to Nephi Jones, senior manager, Research and Development, BioProduction, Thermo Fisher Scientific, most of the established SUB manufacturers have had a 2000 L or larger product offering for many years. “Clearly most users see the benefit of single-use bioreactors and have been requesting more capacity (displacing 10,000–15,000 L stainless-steel bioreactors), but not at the cost of ease of use, reliability, or performance,” Jones states.
However, as the biopharmaceutical market continues to diversify and grow, future demand in biologics is expected to go far beyond current capabilities of established technologies, Alex Chatel, product manager at Univercells Technologies, cautions. Chatel notes that manufacturers are continuously searching for solutions to reach higher production capacities but must remain vigilant of large footprints and consequent costs. “Currently high-demand applications require scaling-out of production with several bioreactors (i.e., placing multiple bioreactors side by side) which can be sub-optimal,” he adds.
Chatel also points out that there is a demand for significantly high-volume SUBs, as demonstrated by the launch of larger bioreactors; however, there must be a balance between operational complexity, footprint, and batch failure risk and cost with the scaling-out option.
Market pressures
Market pressure pushing the drive toward further adoption of large-scale SUBs primarily centers around demand, according to Adrian Mazzone, director, Process Development, Pharma Services, Thermo Fisher Scientific. These bioreactors allow for more traditional scale-up (single-process stream) and, by utilizing high turn-down ratios, add flexibility. Importantly, large-scale SUBs allow contract development and manufacturing organizations (CDMOs) to retain existing clients with a “start here, stay here” strategy, Mazzone says. This strategy is meant to ensure that clients have an uninterrupted pathway from development through commercialization and don’t feel the need to transition to a CDMO with stainless-steel capacity, he asserts.
In addition, large-scale SUBs enable manufacturers to increase their production throughput at
limited capital expenditure (CapEx) compared to stainless-steel facilities, says Chatel. “The growth of the cell and gene therapy market as well as the phenomenal needs in cell-based vaccines that will be sustained in the years to come means that large scale-bioreactors will be in demand,” he states.
Chatel adds that manufacturers must always keep two important parameters in mind when assessing biomanufacturing needs: scalability of the technologies they use and the cost of goods related to the final product. In his opinion, current capabilities offered by established technologies will end up constraining the availability of affordable biologics. Although the biopharma industry is addressing this issue through solutions such as process intensification to reduce equipment and facility footprint while increasing throughput at considerably lowered operational costs and cost of goods. Maintaining quality and safety is also a requirement.
Another key driver in the growing use of large-scale SUBs is the growth of gene therapies that rely on adeno-associated virus (AAV) vectors to deliver therapeutic transgenes, specifies Laura Juckem, PhD, vice-president of Scientific Operations at Mirus Bio. The commercialization of gene therapies requires high-titer AAV vector preparations, but the large-scale manufacture of AAV in adherent human embryonic kidney (HEK) 293 cells is both costly and inefficient, she points out. “We can therefore expect to see further additions of large-scale single-use bioreactors as manufacturers transition to suspension HEK 293 platforms,” she states.
Juckem points out that one of the performance limitations of SUBs in gene therapy manufacturing is the inefficiency of polyethylenimine-based transient transfection methods. Novel transfection modalities, however, can lead to multi-fold increases in viral titers, which result in significant gains in process efficiency and a corresponding reduction in manufacturing cost, she states. Examples of some novel transfection modalities include reagents and enhancers found in Mirus Bio’s good manufacturing practice (GMP) platform (VirusGEN).
“Optimization of the transient transfection process will lead to higher success rates in single-use bioreactors as vector production is scaled from research to industrial-scale manufacturing,” Juckem emphasizes.
Kevin Mullen, director of Product Management, BioProduction, at Thermo Fisher Scientific states that SUT has been adopted in abundance to meet early- and late-stage clinical processes. Furthermore, as the molecules in development progress from clinical to commercial (larger) scale, Mullen notes that there is market preference to stay within single use rather than shifting to stainless-steel bioreactors. The market has therefore asked for large-scale SUBs that scale well from small-scale process design to large-scale commercial manufacturing, he states.
“Not only has the market been asking for large-scale SUBs; they have also been asking for large-scale SUBs with significantly improved mass transfer and mixing capabilities to support high-cell density cultures,” Mullen adds.
“The single use bioprocess industry has crossed the chasm where this technology can be used beyond its original uses of evaluation, early adoption, and bench marking against stainless-steel processes,” states Kehail, who points out that the biopharma industry is now seeing GMP implementation of SUT in approved clinical drugs, where the technology provides robust data, comparable product quality, and positive business outcomes. Kehail says that these results demonstrate that the industry has matured in regard to SUT utilization. “The current [market] pressure comes from having more and more clients transitioning from evaluation to GMP, which places further pressure on the supply chain,” he says.
However, Kevin Jose, senior group leader of Upstream Process Development at Catalent Biologics, Madison, takes a different view and remains unconvinced that demand would grow for large-scale SUBs in the long run. He observes that there already exists a variety of options for companies looking for large-scale single-use manufacturing. One approach involves running multiple small-scale bioreactors in tandem (for example, 2 x 2000 L) with a large-scale downstream set up. Another approach is to leverage end-stage perfusion (i.e., continuous manufacturing), which would allow for the production of larger amounts of drug substance in a smaller single-use vessel.
“[As] there are options to address large-scale manufacturing with smaller bioreactors, it remains to be seen whether there will be high demand for large-scale SUBs themselves. Additionally, as the market moves towards smaller patient population sizes (such as rare/orphan diseases and personalized therapies), and as technologies improve to increase cell/process productivity, there may be less demand for large-scale manufacturing in the future,” Jose states.
Fixtures in the industry
As large-scale SUBs establish a firmer foothold in commercial manufacturing, their continued performance will be further scrutinized. For instance, according to Kehail, it has not yet been determined how well single-use bags will withstand high pressure, temperature, and aeration at volumes greater than 4000 L. He stresses that the robustness of the bioreactor bag is essential. “Any leak, pin hole, or puncture could result in significant financial and product loss,” he states, adding, “we have only demonstrated and proven performance of single-use bags at 2000-L scale.”
Meanwhile, Jose notes that the introduction of high turndown ratio (i.e., ratio of total volume to working volume) SUBs has addressed logistical and technical/engineering issues with respect to seed train and production culture flexibility for large-scale single-use manufacturing. “The technological advances made in SUBs have led to the development of contact layer films with ‘low’ to ‘no’ extractables and leachables that could affect cell culture performance. Also, the shift from cylindrical to cuboidal design for SUBs has improved the efficiency of gas transfer and mixing to ensure optimum scalability up to 5000 L,” Jones says.
Moreover, the use of real-time technology, in conjunction with SUT such as Raman spectroscopy, can improve processes even further to ensure optimum cell culture performance, Jones states.
“From a SUB performance standpoint, ease of use (set-up, installation, execution, tear-down) and reliability are key to the success of any large-scale SUB offering,” adds Mazzone.
Meanwhile, Mullen notes that SUT has become the standard over the past 15 years, in which the technology has proved its reliability throughout an everchanging world of new technology, supply chain challenges, and a global pandemic. “The market demand is to have improved power input per volume (PIV) and mass transfer (kLa) to support high cell densities and support high product quality. The performance in the large-scale reactors needs to be similar to the performance in PD [process development] small-scale reactors so that processes can be optimized at PD small scale and scale up to the larger-scaled reactors,” he says.
While SUBs are mainly stirred-tank vessels, fixed-bed bioreactors are another family of large-scale bioreactors, says Chatel. These fixed-bed bioreactors are much smaller than an equivalent stirred tank vessel for an equivalent throughput. Both SUBs and fixed-beds are fitted with technology to control the basic needs of cell culture (pH, dissolved oxygen, temperature), but beyond the basic bioreactor features, process analytical technology (PAT) is increasingly developed and used to provide automated measurement of advanced production parameters, such as product titer and metabolites concentration, Chatel explains.
In all cases, Chatel emphasizes, the operational environment allowing cell cultures at high densities requires sufficient gas supply and, as mentioned earlier, kLa, which must be enabled by either sparging or gas overlays. Meanwhile, control over foam formation and carbon dioxide stripping must be ensured to prevent cytotoxic effects. “Good mixing that ensures homogeneous availability of nutrients and gases is essential to successful [bioreactor] performance, and this must be ensured by a carefully studied mixing environment,” says Chatel.
Finally, manufacturers must ensure that future SUBs are designed as part of a family that enables smooth scale-up and scale-down. This holistic approach will enable end-users to successfully translate from development to commercial production, Chatel concludes.