November 1, 2010 (Vol. 30, No. 19)

Single-Use Systems Slowly but Surely Gaining Acceptance in Critial Unit Operations

Disposable bioreactors have become the vanguard of single-use processing, a litmus test of sorts for the acceptance of single-use equipment in biomanufacturing. Adoption has been slower than expected, but progress has been encouraging nonetheless. The steady uptake of disposables in critical unit operations suggests that, eventually, single-use bioreactors will be more the rule than the exception, particularly at small- to mid-scale.

Jeffery Craig, global director for business development at ATMI, calls the evolution of disposable bioreactors “part of a natural maturation process” for single-use.

Until recently, ATMI maintained a low profile in the life sciences. The Belgium-based company, which manufactures its own animal-derived component-free plastic films, has for years supplied microelectronics firms with ultraclean, single-use bags. It subsequently introduced its own disposable mixing technology and, in 2008, acquired LevTech, which specialized in disposable, magnetically levitated impeller mixing driven by a superconducting magnet.

This year ATMI extended its commercial alliance with Sartorius Stedim Biotech by adding new technologies to the marketing and distribution partnership, specifically ATMI’s Magnetic Mixer. Sartorius Stedim Biotech retains distribution and marketing rights to ATMI’s LevMixer® mixing system, as it has since early 2008. The agreement also opens up ATMI’s bioreactor-related products to Sartorius Stedim Biotech.

Integration, regulatory compliance, and the emergence of standards have been among the predominant trends in single-use bioreactors, and these have been driven by compliance and quality concerns. Users are more interested in complete, plug-and-play processes rather than standalone unit operations, says Craig, despite the persistence of a “custom mentality”—a leftover from disposables’ early days.

“The role of standards in the acceptance of single-use bioreactors is analogous to the adoption of plastics in the microelectronics industry. Standards, particularly for leachables and extractables, benefit everyone.”

Standardization does not imply a lack of innovation. ATMI’s Integrity™ disposable bioreactors, which range in size up to 1,000 L, are cube-shaped vs. the more common cylinders or flat, oblong bags. The reactors are designed to “move a lot of fluid,” provide good mixing and oxygenation with low shear, and are scalable from bench to manufacturing scale.

When Xcellerex  broke the 1,000 L ceiling in 2008 with its XDR-2000 disposable bioreactor (with a fully inflated volume of 2,500 L), observers believed the upper size limit for bio-bags had been reached. Supporting massive, fluid-filled bags presents engineering issues, disposal of used bags raises environmental concerns, and bioreactions above 1,000 L capacity were said to be the exclusive domain of stainless steel.

Despite these objections, Xcellerex is working on even larger disposable bioreactors, with a 5,000 L unit under development.

Parrish Galliher, CTO, explains that such large systems benefit from economies of scale. “It takes one operator to run a 1,000 L reactor and one operator to run a 5,000 L reactor. Quality control and release-related costs are also lower because you’re producing fewer batches. And while a 5,000 L bag will be more expensive than a 2,000 L bioreactor, it won’t be two-and-a-half times as expensive.”

The standard rule of thumb for costing stainless steel reactors states that cost goes up by a factor of 20.4 power (a factor of about 1.32) when size doubles. This rule applies to disposable bioreactors as well.

Galliher also suggests that the value of disposables relative to stainless reactors improves at larger scale. More steam, cleaning fluids, time, and personnel are required to sanitize a stainless steel reactor, while outlays for the disposable’s cleaning and cleaning validation remain at zero.

In September, Project GreenVax announced that it would use XDR GMP single-use bioreactors from Xcellerex in a new facility under construction in Bryan, TX. Project GreenVax is an offshoot of the Texas Plant-Expressed Vaccine Consortium consisting of G-Con  and Texas A&M University.

Located at the Texas A&M Health Science Center, the 21-acre facility will house 145,000 square feet of biomanufacturing space that will be built and managed by G-Con.

Project GreenVax, which uses tobacco plants rather than the current egg-based or cell-based vaccine technology, could shorten vaccine production to a fraction of the current time, allowing rapid response to newly emerging viruses. Eventually the consortium hopes to produce 100 million vaccine doses per month.

Earlier, Xcellerex entered an agreement with SK Chemicals one of South Korea’s largest conglomerates, to commercialize SK-developed vaccines with Xcellerex’ FlexFactory® biomanufacturing platform. At the time, SK’s CEO Chang Geun Kim noted that the flexibility provided by disposable bioreactors would accelerate development and commercialization of his company’s cell culture-derived influenza and pneumococcal conjugate vaccines.

FlexFactory is based on the combination of single-use technologies, controlled-environmental modules, and process automation including electronic batch records. The company’s TransPlant services facilitate installation, validation, and operation of FlexFactory-based manufacturing plants.

G-Con president Barry Holz said that the XDR system was chosen because of its “competitive capital costs” and lower operating costs compared with stainless steel bioreactors.

The XDR single-use bioreactor line from Xcellerex ranges from 50 L (shown) to 2,000 L. All systems are GMP ready and directly scalable.

Smaller Bags Still the Sweet Spot

While some suppliers have demonstrated the feasibility of 1,000 and even 2,000 L bioreactor bags, smaller units remain the largest group in terms of sales. In fact rather than being held back by disposables’ real or imaginary size limitations, suppliers are embracing the technology’s unquestioned benefits, particularly for development groups. A good deal of innovation is occurring within the 500 L and smaller category as well.

GE Healthcare’s 2007 acquisition of Wave Biotech provided what Gerard Gach, director of marketing for RTP (Ready to Process) at GE, calls “a great foundation” in the disposable bioreactor marketplace. While GE remains committed to the Wave product line and its 500 L working volume limit, the company is “exploring other platforms.”

Gach admits that while demand exists for disposable bioreactors of 1,000 and 2,000 L working volumes, the sweet spot for 500 L bioreactor bags continues to grow.

Rising volumetric productivity has greatly expanded markets for the small-by-comparison Wave bioreactors. Five hundred liters is not such a small process volume for many vaccines and diagnostics, and serves clinical programs for many therapeutic proteins as well.

Gach notes that the decline of the blockbuster model and the rise of targeted and personalized treatments means that batch sizes will continue trending downward to serve shrinking patient populations. And where 500 L is too small, several Wave reactors may be daisy-chained.

Not unnoticed by GE in the discussion of cell culture sizes and throughput is the impact of upstream processing on downstream operations, particularly with respect to process bottlenecks. Running smaller batches helps processors synchronize upstream and downstream events.

“Many people who saw the 500 liter limit as a shortcoming are reassessing that opinion,” Gach reports.

The recent flurry of activity around small, benchtop-capable reactor bags suggests that this market is far from exhausted.

GE Healthcare reports that its Wave Cellbag bioreactors and ReadyCircuit assemblies are quickly configurable with ReadyToProcess filters, including both cross flow and nominal flow, for sterile bioprocessing operations.

Sensing and Control

Michael Cunningham, Ph.D., senior research scientist at Millipore, observes that early single-use bioreactors suffered from less-than-optimal cell densities, product titers, and oxygen delivery limits (e.g., as measured by kLa or volumetric oxygen transfer coefficient). Improvements were often accompanied by shear damage to cells.

Some blamed those engineering limitations on bag design and agitation methods, but “the problem was much more complex. There were probably other things going on.”

Today, Dr. Cunningham says, disposable systems—even very small ones—more closely mimic conventional bioreactors.

Indeed the gap is closing rapidly. During three conferences this past year, contract manufacturer Avid Bioservices presented data on the comparability between Thermo Fisher Scientific’s Single Use Bioreactor (SUB) and conventional stainless steel bioreactors.

In August, CMC Biologics installed a disposables-based manufacturing facility at its Seattle location. The multipurpose plant, undertaken in collaboration with Thermo Fisher Scientific, will produce early-stage clinical batches of monoclonal antibodies and other cell culture-derived therapeutic proteins.

Central to the upstream production capabilities are Hyclone/Thermo 100 L and 200 L SUBs and disposable mixers.

One way plastic is catching up to steel is through advanced sensing and control. Many suppliers are capitalizing on this user-driven trend, even in their smaller reactor bags. For example, Xcellerex expects to launch a line of single-use sensors for its disposable bioreactors and mixers by the end of 2010.

Earlier this year Finesse Solutions introduced a line of single-use TruFluor sensors for headspace pressure and dissolved oxygen or pH plus process temperature. The sensors are available in Thermo Scientific (HyClone) BioProcess Containers and SUBs as turnkey bioreactor components, in volumes from 25 L to 2,000 L. TruFluor sensors consist of a disposable sheath, optical reader, cable, and transmitter, and operate through real-time phase fluorimetry.

Also last summer, New Brunswick Scientific (NBS) and Pall  announced a joint venture to produce single-use bioreactors with control capabilities. The collaboration is based on the combination of Pall’s Allegro™ single-use biocontainer with NBS’s CelliGen® bioprocess controller.

NBS also offers a CelliGen® disposable benchtop bioreactor that combines single-use and stirred-tank technologies. The pre-sterilized 5 or 14 L vessels feature a non-invasive sensor technology that significantly reduces turnaround time for reactors of this size from ten hours to about one hour, according to product manager Rich Mirro.

Millipore has entered the disposable bioreactor marketplace with a 3 L product, CellReady, which uses probes for controlling temperature, dissolved oxygen, and pH.

“Three liters is the workhorse in product development,” says Dr. Cunningham. The company recently made a development-stage 250 L single-use reactor available to select customers.

While built-in sensors and controls are becoming the norm, some off-line measurements still require old-fashioned sampling of a bioreactor’s contents. Sampling has always been problematic since not all bioreactors—whether steel or disposable—allow ready access to process fluids in real or near-real time.

Groton Biosystems  recently introduced a sterilizable, disposable filter-sampling probe for automated and manual extraction of cell-free samples. The product simplifies analysis by eliminating filtration or centrifugation while reducing cross-contamination risk, according to CEO Bill Dinardo.

Single-use bioreactors have improved dramatically since the earliest implementations. And, while disposables’ benefits are now well known, other issues have arisen.

GE has begun investigating how supply issues affect customers because, says Gach, “some customers struggle with inventory management, particularly for bioreactors.” This is one reason GE expects to expand beyond rocking-type bag systems and eventually to cover the biomanufacturing space more comprehensively.

The company is also undertaking an analysis of the environmental impact and benefits of single-use bioreactors. This study goes beyond a comparative analysis of acquisition and operating costs for stainless steel and plastic to include carbon footprint, energy, and water consumption. “We made it one of our missions to understand these issues, and translate them to customers as they validate their decision on disposables.”

One might assume that single use is on everyone’s agenda, but that is not so. The worldwide capacity glut still puts off firms with large investments in stainless steel despite constant volumetric productivity improvements. Dr. Cunningham reports that at a recent meeting bioprocessors indicated a preference for running hard-piped equipment at less than full capacity, rather than switching to plastic.

Whether this trend comes to dominate large biopharma’s mindset toward disposable reactors, at least over the next five to ten years, is anyone’s guess.

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