January 15, 2008 (Vol. 28, No. 2)

Customized Cell Culture Media Predicted to Increase Productivity while Decreasing Costs

Cell culture media have become central to biomanufacturers’ efforts to lower cost of goods and improve productivity. Bioprocessors continue to look for better-defined media in convenient formats to help streamline scale-up and manufacturing. Media vendors continue to deliver on all fronts.

Now a part of Thermo Fisher Scientific (www.thermofisher.com), HyClone has upgraded its cell culture media analytic capabilities during media development and manufacturing, according to Brandon Pence, bioproduction market manager. It utilizes Thermo mass spectrometers and near-infrared spectroscopy, with the former working in realtime alongside single-use bioreactors.

“This emerging real-time analytics approach greatly expands our ability to optimize and control both media and the processes they serve,” comments Pence. HyClone’s media analytic initiative dovetails nicely with process analytic technology.

Pence is one of several experts who note the increased use of fed-batch cultures, and with that, the demand for customized feed supplements. “Fed-batch cultures can boost productivity significantly without a large increase in raw material costs,” he says. Among HyClone’s fed-batch supplement products is the CellBoost™ line of six media supplements for boosting culture productivity in middle- to late-growth stages. Supplementation can substantially extend the productive life of a mammalian cell culture well beyond the 7–10 day norm for conventional batch cultures.

Culture formats often come down to a cost and value proposition, specifically the value of enhanced protein production versus the cost of achieving it. Pence views fed-batch strategies as a kind of compromise between perfusion cultures and straight batch culture. Perfusion strategies produce prodigious yields of protein over an extended period, but they consume large volumes of costly media and raw materials, while demanding significant maintenance.

Fed-batch supplementation provides high protein-expression levels more economically, albeit over a shorter time period compared with perfusion cultures. Fed-batch cultures have the added benefits of ease of use, scalability, and manufacturing flexibility. “That is why fed-batch cultures are most popular at larger bioproduction scales,” says Pence.

Demand for Fed-batch Products

Bob Burrier, Ph.D., vp of R&D for Invitrogen’s (www.invitrogen.com) cell culture systems division, also notes building momentum for fed-batch cell culture strategies. Invitrogen’s latest product offerings in this area include the CHO CD EfficientFeed™ Kit, which contains CHO CD EfficientFeed A and CHO CD EfficientFeed B supplements. These supplements are protein-free, chemically defined, concentrated liquids that provide an extra metabolic boost to cell cultures.

Fed-batch cultures are eclipsing both traditional batch and continuous processes, according to Dr. Burrier. “Perfusion cultures are still important when products are unstable or toxic to the cells,” he says, “but fed-batch culture to extend the useful productivity time of the culture has become the norm.”

Since fed-batch cultures remain viable longer, equipment turnaround is lower and protein titers are higher. To counteract the effects of foam that occurs in high-density cultures, Invitrogen has introduced a new antifoaming agent, FoamAway™ Irradiated, a gamma-irradiated emulsion containing 3% Simethicone in disposable packaging.

Invitrogen has also been improving and expanding products based on its granulated AGT™ media. Originally developed for basal media, AGT has now been applied to feed supplements and is used to make the CHO CD EfficientFeed products. “The AGT-based feed supplements are ideal for those who wish to develop a fed-batch process but don’t have the time or resources to do extensive media analysis or development,” Dr. Burrier notes. Further feed strategy optimization is possible through Invitrogen’s PD-Direct™ services or independently, he adds.

AGT provides the benefits of a powdered media but with less dust generation. The granulation process provides complete formulations without the need to worry about osmolality or pH adjustments, according to Dr. Burrier. Granulated media wets much more quickly, translating to reduced preparation time—a significant consideration for processors who reconstitute tens of thousands of liters of media and feed additives per year. Custom AGT-generated media may also contain growth factors and trace components.

Media vs. Cell-line Engineering

Experts still dispute whether cell line engineering or culture medium are predominantly responsible for the dramatic improvement in protein titers. One can find experts on both sides of the issue. Dr. Burrier believes both factors contribute and that coupling media technologies with cell line development provides the best chance of arriving at a super-productive culture. He stresses the importance of a high-titer clone as a starting point from which the benefits of process optimization, including media work, increase productivity.

Dr. Burrier agrees that media becomes the productivity opportunity but only if cells are already programmed to produce protein in the 1–2 g/L range. At this point two- to fivefold productivity gains become a media and process optimization exercise.

“If you start with a cell line producing only 100 milligrams per liter, there is no way you’re getting up to one to five grams per liter simply by optimizing the media and process. Companies first need to utilize technologies for cell line improvement, then concentrate on the media,” Dr. Burrier explains.

Bruce Lehr, director of marketing at SAFC Biosciences (www.safcbiosciences.com), similarly believes that media, while important, pales next to cell-line engineering as a productivity exercise. “There certainly is interplay between cell engineering and media, and if you can match those activities earlier, there is a real potential to obtain productivity gains more easily.”

To Lehr, however, the gradual replacement of animal-derived components (ADCs) in cell cultures has been the most significant trend over the past year.

Biomanufacturers continue to demand not just ADC-free media, particularly for new processes, but are specifying that such media be manufactured in an ADC-free production environment. HyClone, for example, maintains manufacturing suites for liquid and powder products that have never seen animal components, in addition to conventional media formulation suites for formulations that require animal-derived ingredients.

Moreover bioprocessors are beginning to avoid, whenever possible, all nondefined or undefined components like soy or wheat hydrosylates and yeastolates (yeast hydrolysates).

Many established processes still employ serum-based media, however. “Biomanufacturers are not going back and redoing old processes with defined media,” Lehr notes. “Most work on defined media takes place during process development and early clinical trials.”

Best-in-class biomanufacturers are also increasingly relying on media platforms to standardize production in preferred cell lines, while boosting productivity during individual cell cultures through feed additives. Top companies increasingly look to media suppliers for supply chain management, raw material characterization, sourcing, risk management, and compliance.

“Companies are increasingly willing to outsource these noncore activities,” Lehr comments. The trend toward outsourcing creates opportunities for media suppliers, “and not just for supplying media.”

SAFC Biosciences provides the full spectrum of media products and services, from front-end cell line cloning, medium and feed development, optimization, process scale-up, manufacturability, and manufacturing-scale media and supplement supply, according to Lehr. The company has also entered the downstream processing marketplace through its Bioease™ line of biodisposables, and with liquid and powder buffers for purification processes. “With increases in upstream productivity, downstream opportunities continue to arise,” says Lehr.

Although about 90% of SAFC’s media sales are of the customized variety, developers often begin with off-the-shelf media and work with the company on modifications, eventually arriving at a production-worthy medium.

Powdered media has become the norm for large processes. “Generally people switch from liquid to powder as processes grow,” says Tom Fletcher, director of technology at Irvine Scientific (www.irvinesci.com). Irvine sells animal-derived-component-free media, primarily for CHO cell cultures. The company’s business nearly doubled last year, with much of that growth coming from European and Asian markets.

Like many media suppliers, Irvine is not just a media vendor, but a provider of custom media and feeding strategy services. The company sells mostly custom media, although many customers still demand off-the-shelf basal or classical media.

In development-services mode, Irvine works with customers’ cells at its facility or may collaborate remotely by sending test batches of media to customers and tweaking the formulation based on experimental results. “We’re very flexible in that regard,” says Fletcher.

The company recently expanded powder manufacturing to keep up with demand for custom media, which according to Fletcher has nearly doubled during the past year. The upgrade includes continuous milling technology, acquired more than a year ago but just recently validated. Irvine reports that it can now supply powdered media in quantities from 1 kg to 5,000 kg. It has also opened a cell culture, media-development, and prototype-production facility in Tokyo.

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