February 15, 2016 (Vol. 36, No. 4)

Bioprocessors Are Applying Single-Use Technology to Accomplish What Was Previously Impossible

Single-use technology will continue to grow, but downstream processing remains the fly in the ointment for overall, or wider, adoption, says Surendra Balekai, senior global product manager-SUT, at Thermo Fisher Scientific.

Protein expression rates are in the 3–4 g/L range at large scale with 5–8 g/L achievable at laboratory volumes. “Eventually this level of productivity will reach large-scale production, maybe in as few as three years,” continues Balekai.

The near-doubling of protein titers that Balekai predicts will create an even stronger case for scaling out vs. scaling up, that is, running multiple 2,000 L disposable bioreactors rather than sourcing a single 5,000–10,000 L stainless steel bioreactor. “At high volumetric productivity, companies can better leverage their use of disposable bioreactors by scaling out,” adds Balekai, all the while benefiting from the versatility and flexibility of single use in multiproduct facilities.

He admits that wide adoption of upstream continuous or perfusion bioprocessing in disposable formats could change this picture, “but we won’t see that for at least five years.” Balekai is even less optimistic about downstream processing. “Companies are talking about continuous chromatography, with some implementations, but generally this technology has not made its way into industry,” he explains. “Downstream was, continues to be, and for at least three years will remain a bottleneck for overall adoption of single-use technology.”

An interesting growth area for single-use will be disposable sensors for nearly every process parameter one can think of. Sensors are not quite where they should be, however, in terms of cost, robustness, performance, and compatibility with disposable bioreactor materials and bioprocesses themselves. But Balekai predicts that “both sensor companies and biomanufacturers will continue working together to develop the sensors of tomorrow, and these products will begin to come online within one or two years for both upstream and downstream processing.”

A technician inserts a drive shaft into the Thermo Scientific HyPerforma Single-Use Mixer, a functionalized vessel that includes sterile contact surfaces for mixing, venting, and temperature sensing. The vessel’s ports also allow a choice of aseptic connections for filling, emptying, and sampling.

Inflection Point

The biopharmaceutical industry has recently moved through an inflection point with single-use technologies. For even the most conservative of manufacturers, the question is no longer whether single-use technologies are suitable, but how much efficiency and flexibility may be gained from their implementation.

“Process development and plant engineers are moving beyond simple replacement of traditional stainless-steel systems with single-use systems for operations like cell culture, media and buffer preparation, filtration, and chromatography,” notes Mike Felo, director, single-use product management at MilliporeSigma. “They are now exploring the application of single-use technologies to enable processing that was not previously possible.”

According to Felo, as the pharmaceutical industry evolves away from the blockbuster model, novel approaches to the entire process will be critical for driving greater productivity and process efficiency. “Already companies like Amgen, with their new commercial-scale, single-use facility in Singapore, and Genzyme, with multiple publications on clinical scale, continuous processes, are spearheading innovation in several fields, including process compression, continuous processing, and system integration. Single-use systems are not only suited for these developments in production technology but are enabling their implementation for clinical and commercial manufacturing.”

Additionally, Biogen, Merck & Co., and GlaxoSmithKline are developing single-use processes that enable closed processing within controlled, nonclassified (CNC) spaces. This greatly reduces or eliminates the need for intermediate aseptic filtration to control bioburden levels. This is particularly salient for vaccine processes, since some products can be retained on a sterilizing-grade membrane.

“Operation in CNC space reduces the need for environmental monitoring and testing and allows for more open, ballroom-style production rooms. This greatly reduces the costs of constructing and maintaining facilities,” Felo adds.

Standardization of single-use systems remains an unfulfilled desire of many manufacturers. There are two interesting areas to watch in this regard. Felo believes an agreement by end-users and suppliers on a common aseptic connector for noncritical applications should be attainable with some progress already made by Dave Wolton of the PM Group over the past two years.

BPOG’s (BioPhorum Operations Group) Standard Extractables Testing Protocol offers a second opportunity for standardization in how single-use components and systems are characterized and qualified by suppliers and end users. “While initial implementation will take time, eventually component interchangeability based on supplier’s model solvent extraction data could come about. And end-users’ risk analyses will offer a real opportunity for standardization, which does not currently exist,” notes Felo.

EMD Millipore Mobius® Bioreactors—scalable, stirred tank bioreactors providing flexibility by configuring software, hardware, and single-use assemblies.

Alphabet Soup

The past several years have seen much activity in the alphabet soup of industry organizations (standards, industry, and consensus bodies) regarding single-use systems. James Dean Vogel, director of The BioProcess Institute, says “these groups are responding to their members’ concerns, but are having difficulty reaching consensus on the top issues or even agreeing on what the top priority issues are.”

He notes that smaller companies seeking agility and flexibility employ single-use nearly exclusively, and that Amgen has done a “good job” with its Singapore facility, which primarily utilizes disposable equipment. “But there remains significant doubts regarding single use,” Vogel advises. “The technology is definitely a tool in the bioprocess toolbox and not going away, but will it be used every day by everybody? We haven’t yet reached the maturity level…yet.”

Hospitals, for example, employ single-use products for nearly every device or object that comes into contact with patients. “But people still argue for multi-use transfer lines in bioprocesses.”

The question of standards remains vexing. Vogel points out that companies should be rewarded for developing proprietary components, like sensors, connectors, or film materials that deliver superior function and high value. But if other industries are any guide, true growth and innovation occurs when products reach open-source status.

So why hasn’t the single-use marketplace delivered more standards? “Single-use process equipment is young, evolving, and not yet mature enough. I do foresee some type of landscape to allow more open-source references, perhaps utilizing standards or other collaboration methods evolving within the next decade, if only perhaps in one area,” continues Vogel. “Maybe even connectors? Or polymer materials of construction?”

Standards emerging through the efforts of the Bio-Process Systems Alliance (BPSA) and BPOG will lead to the increasing availability of high-quality information on single-use materials which, in turn, will provide greater technical insight to process performance.

“This will help to address one of the industry’s top concerns: that single-use materials do not negatively impact process performance or quality,” says Amit Dua, global market solutions leader for single-use bioprocessing, GE Healthcare’s Life Sciences.

“Longer-term, we will see higher levels of collaboration to drive process assurance and wider adoption of single-use in both clinical and commercial programs, as well as across the full bioprocessing workflow. We have already seen instances where collaborations between supplier and customer quickly remedy process challenges through new single-use product design using the latest information available.”

The lack of availability of chromatography resins in suitable formats for a wide range of clinical manufacturing programs has historically limited implementation of single-use during purification. Dua notes that industry is looking to access a wide range of ready-to-use, pre-packed, and validated chromatography columns to realize the efficiency benefits that this equipment may provide.

“The introduction of a wider range of formats for single-use chromatography columns, pre-packed with higher performance chromatography resins, will enable the economic implementation of single-use chromatography,” predicts Dua. “This is leading to increased implementation for clinical programs, and a significant number of biomanufacturing processes becoming 100% single-use.”

Dua believes that the full benefits of single-use will eventually be achieved through a “holistic approach.” Designing, engineering, and constructing a single-use GMP bioprocess facility, he says, is a risky, costly, and lengthy endeavor for biopharmaceutical companies. For example, successful unit operation integration is critical for GMP manufacturing as is smart automation and integration to ensure process control and data recording in accordance with GMP requirements. “Looking at the process as a whole today, single-use technology is implemented in various unit operations and teams in upstream, downstream, fill-finish, or support services are typically leading the implementation.

“Increasingly we will see companies appointing a ‘process owner’ who understands that optimizing a biologics workflow means looking at it in a truly holistic way, in order to make the best decisions and maximize the benefits single-use can provide. This means addressing unit operation optimization, automation and controls, and, finally, facility utilization.”

Ultimately, Dua points out, “we will see more efficient facilities, either greenfield or retrofit, and more multiproduct facilities, which give greater economic benefit by spreading facility fixed costs over multiple drug products.”

Reducing Facility Footprint

Single-use perfusion cell culture is becoming a significant strategy for upstream processing and that trend will continue, says Christel Fenge, Ph.D., vp of fermentation technology at Sartorius Stedim Biotech. Dr. Fenge, who began her career developing perfusion production processes for recombinant therapeutic proteins, notes that early excitement gave way to a lull in interest. “But now we’re seeing many companies, not just perfusion powerhouses, but even new players, examining continuous cell culture,” she says. “Customers are showing significant interest in small-scale perfusion models as well, particularly automated systems.”

Sartorius Stedim Biotech serves this market through its ambr® small-scale single-use bioreactors with which perfusion mimic approaches have been reported.

Reducing facility footprint is a primary benefit of perfusion cell culture. The compactness and productivity of continuous culture enables manufacturing at full scale in volumes as small as 500 L, notes Dr. Fenge. “This, and the ability to work with disposable equipment, is what drives this industry segment. A company can produce clinical trial material at their preferred scale and continue at that scale for producing commercial material. This could possibly save a scale-up step and take companies quicker to market.”

Dr. Fenge foresees improvement in materials of construction as well. Sartorius Stedim Biotech has been active in this area, through partnerships, particularly in films and tubing. “We have strong collaborations with companies that make these films, but also with resin providers. Our strategy has been to improve traceability and control of raw materials,” explains Dr. Fenge

One result of this effort has been Flexsafe® 3D bags for storage of liquids and intermediates and also Flexsafe RM and STR cell culture bioreactor bags.

Today, raw materials traceability and control over single-use product manufacturing help bioprocessors achieve reproducible profiles for leachables and extractables. Also, at the same time, it helps to control more fundamental process-related issues such as cell growth where it has been reported that a degradation product of a commonly added antioxidant has a cytotoxic effect.

“Comprehensive control of raw materials and manufacturing is a relatively new theme,” says Dr. Fenge. “For adoption of single-use processing to grow as much in the next ten years as in the previous decade, we must ensure control of these materials and through that assure consistent product quality, cell growth, and extractables and leachables profiles.”

From her role as vp for single-use technology at Pall and her participation in several single-use industry working groups, Hélène Pora has a unique take on the future of single-use bioprocessing. “Developing the capabilities of single-use technology fully will require not necessarily component standardization but more common approaches leading to standardization for key aspects like particulates, extractables/leachables, integrity testing, change control, and supply chain.”

What Pora calls “a more common approach and best practices” is expected to provide assurance that the technology is reliable and safe as it moves toward more critical applications. Progress already made is further a sign that “the industry is maturing.”

Achieving these goals will lead, over the next several years, to adoption of single-use technology in more critical applications at commercial scale. “And not only for media and buffer prep but in bioreactors, chromatography, tangential flow filtration, bulk and final filling, and others.”

A ten-year objective, according to Pora, will be wider adoption of continuous processing which, she says, “will not occur without single-use technology.” Continuous processing in the form of perfusion culture is a widely acknowledged trend upstream. GEN readers are similarly aware of the effect of rising titers. Together, these trends point to lower volumes for single-use upstream operations but present downstream purification with equally well-known issues.

“As upstream issues are solved downstream remains a bottleneck,” says Pore. “A possible but long-term solution will be continuous purification methodologies. Pall has been active in this area, with its acquisition of Tarpon Biosystems and its BioSMB purification system based on simulated moving bed chromatography,” Pora points out.

“Pall has also acquired an exclusive license from FloDesign Sonics for acoustic wave separation technology, which effects clarification for both batch and perfusion cultures. There is also hope for eventually implementing continuous concentration or diafiltration.”

Microscale modeling for a range of upstream processes is possible with ambr® 15, an automated bioreactor system from Sartorius Stedim Biotech. The system can manage multiple cell line development experiments (24 or 48) in parallel, and can be used to investigate single-use perfusion cell culture.

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