August 1, 2016 (Vol. 36, No. 14)

Entertain Hybrid Approaches and Comprehensive Views of Process Analytics, Automation, and Quality Assurance

Whoever said there’s nothing new under the sun was not, evidently, referring to bioreactors. Yes, we may feel as though we have heard everything about the relative benefits and capabilities of both stainless-steel equipment and single-use systems.

But familiar distinctions are becoming less relevant, and new developments are changing the stainless/plastic calculus. With single-use systems gaining ground every day, it becomes impossible to ignore plastic in any discussion of bioreactors, even discussions purportedly focused on steel.

With the adoption of single-use bioprocess equipment reaching a turning point in terms of acceptance, industry has taken a more neutral—perhaps philosophical is a better term—stand regarding the relative merits of stainless steel and plastic. The arguments and rivalries faded long ago. “There are compelling business reasons for either one,” notes Brady Cole, vice president, commercial operations, ABEC. “We take the same custom approach to single-use manufacturing equipment as we have for stainless for over 40 years.”

Bioprocessors must still perform due diligence when specifying new bioreactor capacity. Production scale and economics remain pertinent issues. Yet if anything, adoption of single-use equipment has had a positive effect on how the industry views stainless-steel equipment. Paraphrasing Mark Twain, Cole notes that “reports of the death of stainless-steel equipment have been greatly exaggerated.” Companies are still using large stainless-steel bioreactor tanks in new facilities, and despite rises in titers for many products, tank sizes have remained in the 10 kL to 20 kL working volume range.

Larger volumes plus higher titers have generated bottlenecks here and there for downstream operations, but these problems are solvable through appropriate engineering approaches. At the same time, a renewed focus on product quality has provided parallel quality incentives for manufacturers of stainless-steel equipment. “We need to align our products with customers’ quality systems,” Cole adds.

To that end, he sees a rededication to lifecycle documentation, and “a lot more” qualification at the bioreactor production facility rather than at user sites. “That makes a huge impact on quality as well as a scheduling advantage,” Cole insists. “If you go through installation and operation qualification (IQ/OQ) at ABEC, you will decrease your schedule risk and increase your chance of achieving high levels of quality during operation.”

Realizing Quality

Sensors and probes are relatively straightforward modifications that make bioreactors more suitable for process analytic technology (PAT) and quality by design (QbD). According to Surendra Balekai, senior global product manager at Thermo Fisher Scientific, the lack of standardization in single-use sensors has hampered PAT and QbD efforts in the world of stainless-steel bioreactors and fermenters.

Companies adopt single-use sensors for the usual benefits, which overall translate to lower costs. Sensors for stainless-steel bioreactors are multi-use, so their deployment involves steam sterilization and cleaning that disposable probes cannot tolerate. This, says Balekai, is what makes reusable sensors so expensive.

Theoretically, single-use probes could be used in fixed-tank systems, but their individualized, unique dimensions prevent this for lack of suitable connectivity. “There’s a need for our industry to consider connectivity standards to reduce development costs,” Balekai says. Such standards would open up fixed-tank processing to the diversity and ease of use of sensors developed for single-use bioreactors, which in turn will improve PAT-ability.

Harmonizing connectivity between plastic and steel will require development of two components. The first is an SIP (steam in place)-tolerant plastic port on the bioreactor, either built-in or aseptically retrofitted. This part must be constructed of sterilization-friendly material such as polyethersulfone. If this idea is to become attractive for fixed-tank systems, the second connection, on the sensor, must be available in a ready-to-use sterile format.

“Unfortunately, not much work has taken place to implement this idea,” Balekai tells GEN. Such hybrid structures have lagged stainless- and plastic-only connectivity in terms of actualization. “Yet large-scale manufacturers who have already invested in fixed-tank equipment are eager to use the same analytics in steel and plastic, which would make production methods more easily transferable. This would open up a new world of sensing for stainless steel, while lowering costs.”

Standardization would also provide new options for other unit operations requiring sensors or controls (operations such as mixing, virus inactivation, and final formulation, which have unique sensing requirements), and improve the ability to integrate these process steps, both within the realm of stainless steel and for hybrid systems. Since installed stainless-steel infrastructure will stay for at least another 10+ years, there is a need to harmonize analytical capabilities between stainless-steel and single-use technology for easy technology transfer in hybrid infrastructure.

A technician inserts a single-use probe into a 1,000 L Thermo Scientific HyPerforma single-use bioreactor. Theoretically, single-use probes could be used in stainless-steel bioreactors irrespective of their manufacturers. Practically, such interoperability is unlikely until standards efforts resolve connectivity issues such as probe/port dimensions and sterilization-friendly formats.

Importance of Hybrid Systems

Hybrid systems raise questions of economics that supersede the usual arguments for single-use systems. Connectors deployed during a process must be sterile, and such a process usually requires that critical components be assembled in highly classified space with the usual issues of cost, time, and inconvenience. Several experts interviewed for this article cited the benefits of Steam-Thru® Connectors from CPC, which was happy to provide additional information.

The Steam-Thru Connector is not new, but its relevance has grown with the idea of marrying stainless-steel and plastic components. The device’s three-port design allows sterile connectivity in low-classification space, thus relieving bioprocessors of having to weld or perform other ad hoc operations in a laminar flow hood prior to assembly.

Simply perform an SIP cycle through the connector’s lower two ports, then move the connector’s valve to create a sterile flow path between the bioreactor and single-use system. This increases process flexibility while maintaining sterility for feed, harvest, or seed train transfer applications. For operations where sterile disconnect is also desired, the Steam-Thru II Connector would allow a second SIP cycle to perform a kill cycle prior to disconnecting the plastic system from the bioreactor.

“Customers use these as replacements for stainless-steel valves,” says John Boehm, the bioprocessing business unit manager at CPC. Previously, the preferred way to connect systems in sterile fashion was the “block and bleed” method consisting of isolation valves coming off the side of the vessel.

This type of connector technology can enable the retrofit or construction of facility designs that drive down production and operating costs. Engineering design firms are working with manufacturers on new facility designs based on closed systems that support production in lower classification clean room.

“Maintaining process flexibility and product sterility are critical aspects of the continuous improvement required to meet the cost challenge facing the industry,” Boehm notes. “Steam-Thru Connectors have become an enabling technology.”

This bioreactor seed-train, which makes use of CPC’s Steam-Thru Connectors, illustrates the advantages of stainless-plastic hybrid systems.

Role of Automation

Deployment possibilities for single-use and stainless-steel technologies are in constant flux. Some of the possibilities involve replacement; others, integration. “Where possible, customers are trying to identify areas in production where single-use fits,” says Ken Clapp, senior manager of applications, technology, and integration at Xcellerex-GE Healthcare Life Sciences.

Bioprocessing has always been driven by automation and integration. Today, however, bioprocessing is being driven even more strongly by these factors now that there is a need to integrate SIP and single-use technology.

“Since stainless-steel bioreactors came first and are well-established, single-use bioreactors are often considered differently, but it all boils down to scale/size and materials,” Clapp says. “Performance between the two should be equivalent, if not in practice then certainly as a goal.”

Process analytics need to be appropriate, practical, reliable, accurate, and cost-effective. Sensors exist for most current process variables. But Clapp notes that “one perennial issue is the capability of sensors to withstand either autoclaving or SIP.” Many sensors in bioprocessing are adapted from devices that have been used in the chemical industry for decades.

“End-product and intermediate measurements, I believe, are still the holy grail in bioprocess sensing,” Clapp says, “and the ability to know, say, product quality or product quantity by direct in-process measurement would be priceless.”

According to Clapp, stainless-steel equipment usage is growing outside of North America and Europe, particularly in India, Korea, and China. Far Eastern companies have been entering large-molecule markets with a vengeance. These companies have the ability to adopt single-use technology, but they may also feel constrained. “They’re also tied to the product they’re pursuing, which could be biosimilars or biobetters,” explains Clapp. “These companies will, whenever possible, attempt to replicate the original manufacturing process, even if that happens to be stainless steel.”

Automation has for decades been a constant for stainless-steel bioreactors. In the early phase of modern bioprocessing, the controls for agitation, temperature, and pH  were often manual. Today’s systems feature near-hands-off operation for SIP, cultivation, feeding, and cleaning protocols. Analytical measurements occur online through primary and redundant sensors for every critical process parameter.

The benefits of automation, says Michael Smith, vice president of operations at Techniserv, include more robust process control, a higher level of process optimization, PAT techniques that improve statistical process control, higher titers at lower costs, and more rapid processing: “These translate to more product per volume per unit time, with a lower likelihood of unforeseen problems causing product loss.”

In this sense, the promises of automation have pretty much been realized. Smith refers to 10–15 years ago when industry seers predicted a never-realized capacity crunch (and everybody else listened). Bioprocessors bought into this idea by planning more stainless capacity at higher volumes. “It turns out that by improving and optimizing processes through automation, bioprocessor increased titers at least two- to threefold during the subsequent 15 years,” Smith says. As a result, the industry experienced an excess of idle fixed-tank capacity in the mid-2000s.

There’s no disputing, Smith says, that changes in processes through automation made a big difference in capacity utilization. However, even this trend is beginning to reverse. “Everybody,” as Smith puts it, “is trying to sink Ireland with stainless steel,” a reference to the $8 billion in new bioprocess facilities that have appeared in that country over the last decade. And further capacity is being built as you read this.

Automated systems do cost more, which would cause startups and smaller companies to think hard about turning control over to machines. Smith maintains that is a near-sighted view.

“Yes automation is more expensive, but in the long term, automation adopters will get a substantial payback in reliability with fewer operator-generated issues or lost batches,” he maintains. “The most common manufacturing glitches occur because multiple operators do not follow standard operating procedures in a consistent manner. Automation minimizes that issue.”

As single-use bioreactors become more prevalent, biomanufacturers show more interest in replacement and integration strategies, which rely on single-use/stainless-steel equivalency, an instance of which is illustrated by the Xcellerex XDR bioreactor. The bioreactor, from GE Healthcare Life Sciences, has a probe belt that is equivalent between single-use and stainless-steel bioreactors. It provides an access point for conventional as well as advanced sensors.

Effective Scale-Up

At the recent Sartorius European Upstream and Downstream Technology Forum, held in Göttingen, Germany, the point was made that to accelerate clinical candidate molecules through the developmental process, it is necessary to have effective scale-up technology. The challenge to speed the move to the clinic was considered by Jincal Li, Ph.D., executive director of biologics process development at Wuxi AppTec Biopharmaceuticals.

Dr. Li’s team has extensive experience with Sartorius Stedim Biotech’s ambr® high-throughput single-use bioreactor system for parallel fermentation or cell culture using different size units controlled by an automated liquid-handling platform. The system provides a scale-down model to explore a range of conditions with scalability to larger bioreactors, replicating classical laboratory-scale bioreactors. 

According to Dr. Li, because the ambr technology allows control of pH and osmolarity, he was able to obtain consistent results, even in cell lines which generate large amounts of lactate. Based on the accuracy of these observations, he said his team was able to scale up from 15 mL to liter levels in their cultures. He added that the viability and growth overall matched quite well as he increased the scale of the cultures all the way to 2,000 L.

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