Application of a novel affinity adsorbent for the capture and purification ofrecombinant Factor VIII compounds

摘要

Recombinant Factor VIII (FVIII) therapies have been created to provide treatment for Hemophilia A, aninherited bleeding disorder caused by mutation in the FVIII gene. A major challenge in the purificationof recombinant FVIII molecules is the development of an affinity chromatography step. Such a step mustbe highly specific and selective for the FVIII molecule, but also must be designed appropriately to ensurethe FVIII molecule can be effectively recovered without resorting to harsh elution conditions which maybe harmful to the product. Additionally, it is desirable to have affinity adsorbents designed to be reusableover a large number of column cycles while maintaining consistent purification performance. In thiswork, we describe the use of VIIISelect, a commercially available affinity adsorbent designed specificallyfor the purification of FVIII compounds. The VIIISelect adsorbent consists of a 13 kDa recombinant proteinligand attached to a cross-linked agarose base matrix. The structure of the recombinant ligand is basedupon Camelid-derived single domain antibody fragments. The VIIISelect adsorbent is produced using aprocess free of animal-derived raw materials, which is a highly desirable attribute for adsorbents used inthe purification processes of recombinant protein therapeutics. The VIIISelect adsorbent was used as theinitial capture column to purify a FVIII compound directly from clarified cell culture fluid prior to furtherdownstream purification. The purification performance of the VIIISelect was evaluated, which includedmeasurement of the static binding capacity, dynamic binding capacity, product recovery, impurity clear-ance, and adsorbent reuse. Following laboratory-scale process development, the VIIISelect adsorbent wasscaled up and used in the large scale manufacturing of a FVIII compound.