Economics is driving bioseparations that minimize waste and maximize output, and moving continuous chromatography into some small-molecule niches.
By Angelo De Palma, Ph.D., Contributing Editor
For manufacturing therapeutic proteins, chromatography is nothing less than the “Holy Grail of bioseparations,” notes Howard Levine, Ph.D., President of BioProcess Technology Consultants (Acton, Mass.). As competition heats up and biopharma awaits the emergence of biogenerics, cost-consciousness is driving new bioseparation technologies that minimize waste and boost output. Higher flow velocity and binding capacity mark today’s chromatography systems, while disposable and membrane chromatography systems (see Disposables and Membrane Adsorbers
, below) are helping to reduce cost and waste even further.
At the same time, economics is taking chromatography — at least, in its continuous form — where it has rarely gone before, into more small-molecule process niches (see Small-Molecule Chromatography: A Hard Sell
Overall, whether in large or small molecule production, chromatography technology is driven by the need to minimize process waste. “Biotech can no longer accept 50% losses per step,” says John Jenco, Ph.D., senior staff scientist at Pall Life Sciences (East Hills, N.Y.).
Downstream bioprocessing is also racing to keep up with tremendous gains achieved upstream. “We’ve seen huge improvements both upstream and downstream, but generally, downstream purification has not kept pace with upstream production,” noted Duncan Low, scientific director at Amgen (Thousand Oaks, Calif.) at September’s IBC BioProcess International conference in Boston.
Chromatography possesses poor economics of scale compared to fermentation, says Low. “You can often scale fermentations without changing the bioreactor footprint, but that just isn’t possible with chromatography columns,” he says.
According to Low, protein titers have increased thousandfold, from 5-50 mg/L in 1980 to 5g/L and higher today, with a target of 10-20 g/L for the next three to five years. Higher titers generate lower volumes per quantity of protein, which aids downstream operations, as well as more biomass, which does not.
Purification experts have responded with numerous improvements, including:
- process integration and intensification;
- the use of compound filters during clarification;
- more efficient resin utilization (including reuse) and higher product recovery;
- alternative downstream strategies that employ membrane adsorbers, liquid-liquid separations, and of course, disposables.
High process volumes and protein titers stress the capabilities of downstream operations in general, and chromatography in particular, notes Eric Grund, director of Fast Trak Biopharma at GE Healthcare (Piscataway, N.J.). So far, vendors have responded with resins and other chromatography products that offer improved flow velocity and binding capacity.Wyeth prepares for ton-scale MAb production
Fermentation has been moving to larger and larger vessels, in good part due to the success of high-dose monoclonal antibody (MAb) products. “Clearly, the industry is planning for protein manufacture at the ton scale,” says Brian Kelley, director of purification process development at Wyeth Biopharma (Andover, Mass.). Wyeth, with three MAb products in late-stage development and a total of twelve in the pipeline, is countering the downstream challenge with process intensification and platform chromatography strategies that are easily transferred both internally and to contract manufacturers.
Wyeth development scientists have reduced downstream MAb chromatography steps to just two — protein A resin for initial capture followed by anion exchange polishing — versus the more common three columns. Wyeth eliminates an intermediate column, typically ion exchange, hydrophobic interaction or hydroxyapatite, by carefully removing cellular debris before the capture to optimize that step, and tweaking the capture column and anion exchanger for maximum efficiency and throughput.
|A technician operates a Capto MMC column cation exchanger, which allows rapid protein purification from undiluted feedstock. Photo courtesy of GE Healthcare.|
The company uses resin-impregnated microwells that serve as “microcolumns” for testing adsorption/desorption at varying pH, ionic strength and elution conditions. Optimizing the ion exchanger to do the work of two columns takes about one day and consumes only about one gram of product. “We’re finding the sweet spot for the anion exchange step,” says Jeff Deetz, senior director for drug substance development.
Other companies and vendors are catching on to micromethods for optimizing very large columns. Through its process proteomics collaboration with Ciphergen (Fremont, Calif.), Pall now offers high-throughput chromatography screening services for rapid process-scale chromatography methods development. Based on SELDI mass spectrometry, the service rapidly screens binding and elution conditions on the Ciphergen ProteinChip array coated with various adsorptive chromatography chemistries. “This technique allows you to do in a few hours what previously took days or weeks with small columns,” says Jerold Martin, Pall’s senior VP for scientific affairs.
Since the workload of downstream steps depends on upstream productivity, bioprocessors must decide rather early how to deal with low-capacity stationary phases, and whether to use one column (with split batches) or multiple columns. Splitting batches to accommodate shortfalls in chromatography capacity introduces uncertainty into a process in that, simply, more things can go wrong.