Selection of chromatographic methods for the purification of cell culture-derived Orf virus for its application as a vaccine or viral vector

Lothert, Keven; Pagallies, Felix; Feger, Thomas; Amann, Ralf; Wolff, Michael W.

In recent years, the Orf virus has become a promising tool for protective recombinant vaccines and oncolytic therapy. However, suitable methods for an Orf virus production, including up- and downstream, are very limited. The presented study focuses on downstream processing, describing the evaluation of different chromatographic unit operations. In this context, ion exchange-, pseudo-affinity- and steric exclusion chromatography were employed for the purification of the cell culture-derived Orf virus, aiming at a maximum in virus recovery and contaminant depletion. The most promising chromatographic methods for capturing the virus particles were the steric exclusion- or salt-tolerant anion exchange membrane chromatography, recovering 84% and 86% of the infectious virus. Combining the steric exclusion chromatography with a subsequent Capto(TM) Core 700 resin or hydrophobic interaction membrane chromatography as a secondary chromatographic step, overall virus recoveries of up to 76% were achieved. Furthermore, a complete cellular protein removal and a host cell DNA depletion of up to 82% was possible for the steric exclusion membranes and the Capto(TM) Core 700 combination. The study reveals a range of possible unit operations suited for the chromatographic purification of the cell culture-derived Orf virus, depending on the intended application, i.e. a human or veterinary use, and the required purity.