Due to the complexity of therapeutic protein purification, downstream bioprocess bottlenecks can arise from almost anywhere, from capacity or facility mismatches to scheduling. Although their sources and bottom-line impact may be debated, bottlenecks are real and their alleviation has become a significant activity, to the point where companies are seriously considering alternatives to traditional unit operations.
As product titers rise, negative chromatography is becoming a viable alternative to a dedicated capture step. Negative chromatography captures impurities while allowing products to flow through. BioToolomics is working with several biopharmaceutical clients on negative chromatography for mAbs based on high-capacity, low-cost, single-use media in disposable columns.
Biotoolomics first employs high-throughput methods to characterize impurities and screen its library of mixed-mode ligand resins. The goal, says Chad Zhang, Ph.D., managing director, is a one- or two-column process that clears all impurities to desired levels.
Protein A capture no longer makes sense, Dr. Zhang argues, because of its high cost and failure to concentrate product. “When titers were one gram per liter and the resin had a capacity of 15 grams per liter it was possible, through capture and elution alone, to reduce feedstock volumes 10-fold. But with today’s titers in the 15 to 20 gram per liter range and resins holding 40 grams or so of protein per liter, you need 3 or 4 column volumes to elute the product. Instead of concentrating you wind up with the same or higher volumes than before capture.”
Host cell proteins (HCPs) are a challenge for negative chromatography, but no more so than with traditional capture chromatography. According to Dr. Zhang, HCPs linger as four to five percent of total protein after traditional capture, which is one reason why bioprocessors employ more than one column. The same is true for negative chromatography. Dr. Zhang is confident that negative chromatography can eventually remove HCPs, as well as viruses and other contaminants, to the 10 ppm range.
Biotoolomics hopes that negative chromatography will eventually become a technology toolbox that biomanufacturers turn to for purifying therapeutic proteins. But the technique is by no means one-size-fits-all. “Deploying it successfully requires close examination of a customer’s antibody product, cell culture process, and impurities,” says Dr. Zhang.