Validation—The Critical Link
Thorough validation of scale-up strategies can save major headaches as the process is built up and out. The point is to demonstrate that products manufactured at large scale for market distribution are chemically and biologically equivalent to those produced at small scale for preclinical and early-stage safety testing.
In biomanufacturing the process is the product, as Genzyme learned with Myozyme/ Lumizyme treatment for Pompe disease. U.S. regulators ruled that the product produced at 2,000 L scale was glycosylated significantly differently than the material manufactured in 160 L batches. “Scale-up is a risk,” observes Harald Dinter, Ph.D., vp of global biological development at Bayer HealthCare. “Whenever you change the process, even if only minimally, there is the risk of changing the product.”
Bayer’s Berkeley, CA, facility manufactures clotting factors for hemophilia, while its Wuppertal, Germany, site produces monoclonal antibodies and other biologicals.
Normally a bioprocessor will validate a large-scale process through scale-down studies, where the bioreactors, clarification, chromatography columns, and filtration systems are precise scaled-down versions of the production-scale processes. Not every process enjoys that luxury. For example, Bayer could not produce a reliable scale-down model for producing Factor 8, a large, complex, highly glycosylated molecule in perfusion reactors, so it performed development work at full commercial scale. “It was obviously costly to do,” notes Dr. Dinter, “but we felt we needed to do that to assure the safety of the product.” Purification studies were performed in scale-down mode, however.
Factor 8 is perhaps an extreme example compared with monoclonal antibodies, since it is larger and carries more glycosylation sites. The potential variations are, therefore, greater than with antibodies and warrant a two-track development model for expression and purification.
Validation is somewhat easier when platform manufacturing technologies are available, which is common with mAbs. Companies that specialize in antibodies often boast of cell culture, harvest, and purification that follows an almost identical path from product to product.
Disposable bioprocess equipment is likely to present novel validation problems as processes progress from 500 and 1,000 L bags to 2,000 L stainless steel vessels and beyond. Plastic and steel appear to produce equivalent products, as studies at Bayer and other firms suggest, but the number of molecules compared in this fashion is still relatively small. The plastic-steel issue will probably not last long, however. As Dr. Dinter points out, cell culture titers are rising rapidly enough that some products in development today could eventually be manufactured in plastic vessels of 1,000–3,000 L.
Biomanufacturers have numerous options for scale-up validation support. Charles River Laboratories provides analytical support for scaling from master cell bank through full production. While processing occurs at the sponsor’s site or at a CMO, Charles River becomes the eyes and ears for scale-up validation, says Barry Rosenblatt, Ph.D., director of biopharmaceutical technology development. The company examines product attributes such as identity, safety, potency, chemical characterization, and virologic profile at every scale-up stage, looking for biological or physiochemical changes that might affect safety.
The most common discrepancy between scales, according to Dr. Rosenblatt, are in post-translational modifications. For mAbs this includes, but is not limited to, glycosylation, a factor critical in some antibodies for biodistribution, potency, and immunogenicity. Post-translational changes such as glycosylation and deamidation, which affects a protein’s charge profile, commonly occur as a result of changes in the cells’ housekeeping functions. Other common differences include changes in impurity aggregation behavior and impurity profile.
“The easiest mistake to make is to assume that if it works at the bench, then scale-up involves nothing more than multiplying the sizes of vessels and equipment,” says Dr. Rosenblatt. Maintaining identical conditions for mixing and column chromatography, for example, require a good deal of engineering, he observes. Scale-down models, when available, are useful for predicting how larger processes will proceed, but to validate a scale-up requires “thinking in both directions, scale-up and scale-down,” he says.