Downstream processing is another key area for bioProcessUK. “Downstream costs, including separation, are 80 percent of the final cost of goods, and separation is starting to become a serious bottleneck,” reported David Stuckey, Ph.D., of Imperial College London. “The cost of goods will only decrease if production processes are integrated and optimized. This is the norm in chemical engineering but not in biopharmaceuticals.”
The root of this disconnect is probably that, in production of a biologic, upstream processes like cell development are about biology, while the downstream processing is more about engineering. Dr. Stuckey believes that it is time to re-examine some of the older technologies, so the team at Imperial has been looking into solvent extraction. Traditionally, this has been nonselective and uses solvents that are hazardous and not biocompatible. What is needed is a more tunable system for extraction of biologic products.
The Imperial team is investigating reverse micelle (RM) extraction of proteins, where a surfactant traps a target protein thereby forming a micelle with a hydrophilic core. Forward and reverse extractions are carried out with a fermentation broth. Dr. Stuckey has tried this approach with lysozyme, ribonuclease, and cytochrome C to see if these could be selectively extracted from mixtures of the three. The experiments were also repeated with real fermentation broths. The back-extractions proved to be slow, but could be sped up by the addition of a counter-ion surfactant.
To scale up the procedure, a Graesser extractor is used to avoid the intractable emulsion that would otherwise inevitably form. “We have proved that this can work on a large scale,” said Dr. Stuckey. They are now looking at the RM approach for mAbs, working with Cambridge Antibody Technology, which is now a part of AstraZeneca (www.astrazeneca.com). The monoclonals, even though bigger molecules, are actually extracted faster than lysozyme.
While iso-octane has been used successfully as a solvent in this work, some companies don’t like it because it is flammable. However, good results can be obtained with vegetable oil, especially corn oil, extraction. Dr. Stuckey is now working on proving that extraction and back-extraction do not change the activity of the protein product.
At Sheffield University, Sheila MacNeil, Ph.D., cofounded CellTran™ (www.celltran.com), based upon work with skin cells. “The nonhealing ulcer costs 10–15 percent of the NHS budget and it is an increasing market because of the increase in diabetes,” she noted. “Although burns are not a big market, we did start out by looking at a better way of delivering cells to burn patients.”
Cultured epithelial autografts (CEA), which use the patient’s own cells, have been the standard approach for major burns and are clinically successful. An audit from Dr. MacNeil’s team found, however, that there are problems with the product, because more than 50% of the time, it is not used; the main problem being the timing of their detachment for application to the patient.
Simple detachment of these cultured skin cell grafts is difficult and, put simply, this is a product with an inflexible shelf life that demanded improvement.
Myskin™, the lead product of CellTran, is engineered to be a PostIt note-like autologous skin graft based on keratinocytes—easy to stick to the graft during transportation and easy to detach when needed by the surgeon.
“Myskin is a cell delivery surface for keratinocytes,” explained Dr. MacNeil. “The product is only part of a whole wound management system because it performs best if it goes onto a good wound bed. Myskin acts like a biological bandage and it has been used on a range of patients including those with diabetic foot ulcers and chronic venous ulcers.” Compared to CEA, Myskin is easier to transport from the lab to the patient, reported Dr. MacNeil. “We have really achieved flexibility in timing delivery.”
CellTran merged with Xcellentis in 2006, which gave it access to more allogeneic keratinocyte products. Other products in development include a contact lens system for treatment of corneal diseases and a keratinocyte/melanocyte treatment for the skin pigmentation disorder vitiligo.
“Tissue engineering in the U.K. is happening on a small scale at the moment,” commented Dr. MacNeil. “This will have to change as patient demand grows and there is more demand for scale-out.”
Other U.K. groups are looking at issues around the yield of biotech products. For instance, David Archer, Ph.D., of Nottingham Biopharm, a Centre of Excellence at the University of Nottingham, described the Centre’s work on yeast as a host, and the various factors that affect yield of heterologous proteins, commenting that the sequencing of over 60 yeast genomes is helping to drive the research.