Nature vs. Nurture
Despite strides in productivity, biomanufacturers remain committed to even higher titers. “I don’t think we’ve reached the ceiling yet,” says Hitto Kaufmann, Ph.D., vp of process science at Boehringer-Ingelheim.
Before process development even begins, companies need to gain a better understanding of how molecular attributes affect expression. Boehringer-Ingelheim is developing sophisticated analytics to screen for “expressability” within a bioreactor. Miniaturized (scale-down) systems are the key, Dr. Kaufmann says.
The question of nature or nurture—innate cellular capabilities versus growth conditions—always arises in these discussions.
Many cell culture experts believe that media and feed strategies have been most responsible for the run-up in volumetric productivity, and Dr. Kaufmann believes that significant, further improvements are coming. Improvement depends on techniques for rapidly screening these conditions and tailoring them to specific cell lines.
Melville believes the relative contributions of nature and nurture are equal. “You can’t separate cell line from process; they are part of the same continuum. You design the process to fit the cell line, but you also select the cell line to fit the process.”
On the “nature” side, targeted integration of genes into the CHO genome has become the new “City of Gold.” The driver here is not volumetric productivity, but faster development. “CHO cells are prone to genetic instability,” Dr. Kaufmann says. “To have designed loci within the genome that ensure stable, high expression will be a key area of focus for biomanufacturers. If successful, this approach may allow developers to skip screening for clones and media.”
Melville disagrees. “I don’t think I’ve seen any site-directed integration that has yielded higher expressions than a good cell-line screening program.” He describes a “good” screen as a reliable model that assures what occurs during clone selection will be relevant throughout process development and beyond.
“Stable integration sites and site integration have come a long way in providing a reliable, minimal expression level, but they’ve never been able to achieve ten grams per liter or higher, which we’re seeing in other systems nowadays.”
Process conditions can affect a protein product’s quality and help fine-tune its in vivo behavior—characteristics that do not translate directly into “grams per liter,” but are just as significant.
Post-translational modifications (PTMs) can affect any number of properties. Developers of therapeutic proteins have only begun to leverage knowledge of the more than 100 PTMs, which in some cases are fine-tunable by merely altering process conditions like pH or temperature.
We know that fucosylation affects the activity of antitumor monoclonal antibodies—lower levels of this sugar increase therapeutic efficacy. Similarly, higher sialylation results in a longer circulating half-life.
“The more we understand relationships between process conditions, glycosylation, and physiologic function, the more companies will exploit these strategies,” Dr. Kaufmann explains. “A few of these functional relationships are known, but many more remain mysterious, at least to the degree that we can be certain of in vivo relevance. That will change.”
In the future, companies may develop several CHO cell lines, each producing unique glycosylation patterns that tune in desired characteristics. Manufacturers of biosimilars may be able to exploit PTMs to differentiate their products as well.
The “food chain” for deploying such technology is typical for biotech. Early work and proof of concept in cells typically occurs at universities. Smaller biotechnology firms will license and optimize the relevant technologies.
If successful, they will be acquired by a much larger firm. For example GlyCart was formed in 2000 as a spinoff of the Swiss Federal Institute of Technology, Zurich. Roche acquired the company in 2005 for, among other things, GlycoMab®, a technique for directing and optimizing glycosylation in antibodies.