Protein A: Extending this Platform Process Paradigm
The use of affinity resins for recombinant protein purification has the potential to revolutionize downstream manufacturing processes. Such platform processes, based around affinity chromatography, will accomplish what Protein-A (PA) resin has done for monoclonal antibody therapeutics (MAb’s), a $85 billion market. For non-MAb recombinant proteins the use of affinity approaches will substantially reduce process development time and bring new products to market more quickly. Although a unique affinity resin will likely be needed for each protein of interest, the benefits of affinity resin development will outweigh the development requirements of time and cost.
Protein A works because IgG monoclonal antibodies have similar Fc regions, which bind to a specific Protein A domain. Non-antibody therapeutic proteins do not share common binding regions. The challenge becomes identifying a specific affinity ligand around which to develop a capture resin. Such a unique affinity ligand/resin system, one that binds the target with nM to pM affinity, can often yield a highly pure target protein with high recovery. The ligand should specifically bind the target in culture broth or other crude post-upstream streams and release the target under conditions that maintain the target protein’s biological activity.
What would the process look like? An additional two to three columns would likely be used to polish the target material post the affinity capture column. The second column, in a bind elute mode, would be specific for removing the major impurity(s) in the product, such as aggregates, fragments, or improperly glycosylated or phosphate forms, etc. The third column, in a flow through mode, might be an anion membrane used to reduce residual materials such as DNA, endotoxin (if from E. coli culture) and host cell proteins.
Affinity ligands include affimers, aptamers, peptides, dyes and other small molecules. The discovery and initial development of any of these ligands can take several months and producing a custom commercial resin will take additional time. An approach, to reduce the development and implementation time, would involve applying rational design, and/or high throughput screening, during the research phase when the therapeutic protein is being characterized. The ligand identification would be completed as efficacy is determined in the appropriate animal model. The purpose would be to obtain an appropriate ligand (in-house or external) for Phase I downstream development. Small batches of resin from the resin manufacturer would be used to develop the process. At the completion of process development sufficient affinity resin would be available from the resin vendor for early clinical studies. The ability to produce commercial lots would be established during Phase II. A well established, large scale, resin manufacturer would be charged with developing a robust reproducible manufacturing process at the appropriate scale for the client’s needs and to keep sufficient inventory at hand.
The use of affinity chromatography simplifies development and condenses the development timeline for commercial implementation. Combining affinity capture with scaling- out (adding a second production line when additional commercial capacity is needed) should simplify facility design as well.
Blog article by: Frank Riske, Ph.D.