Format

Send to

Choose Destination
Protein Expr Purif. 2015 Nov;115:165-75. doi: 10.1016/j.pep.2015.08.023. Epub 2015 Aug 28.

Development, upscaling and validation of the purification process for human-cl rhFVIII (Nuwiq®), a new generation recombinant factor VIII produced in a human cell-line.

Author information

1
Octapharma AB, Elersvägen 40, SE-112 75 Stockholm, Sweden. Electronic address: stefan.winge@octapharma.se.
2
Octapharma AB, Elersvägen 40, SE-112 75 Stockholm, Sweden.
3
Octapharma Molecular Biochemistry, Walther-Nernst-Str. 3, 12489 Berlin, Germany.
4
BAC BV - Thermo Fisher Scientific, 28 Huizerstraatweg, 1411 GP Naarden, The Netherlands.
5
Octapharma GmbH, Altenhöferallee 3, D-60438 Frankfurt, Germany.
6
Octapharma GmbH, Im Neuenheimer Feld 590, 69120 Heidelberg, Germany.
7
Octapharma AG, Seidenstrasse 2, 8853 Lachen, Switzerland.

Abstract

INTRODUCTION:

Human-cl rhFVIII (Nuwiq®), a new generation recombinant factor VIII (rFVIII), is the first rFVIII produced in a human cell-line approved by the European Medicines Agency.

AIMS:

To describe the development, upscaling and process validation for industrial-scale human-cl rhFVIII purification.

METHODS AND RESULTS:

The purification process involves one centrifugation, two filtration, five chromatography columns and two dedicated pathogen clearance steps (solvent/detergent treatment and 20 nm nanofiltration). The key purification step uses an affinity resin (VIIISelect) with high specificity for FVIII, removing essentially all host-cell proteins with >80% product recovery. The production-scale multi-step purification process efficiently removes process- and product-related impurities and results in a high-purity rhFVIII product, with an overall yield of ∼50%. Specific activity of the final product was >9000 IU/mg, and the ratio between active FVIII and total FVIII protein present was >0.9. The entire production process is free of animal-derived products. Leaching of potential harmful compounds from chromatography resins and all pathogens tested were below the limit of quantification in the final product.

CONCLUSIONS:

Human-cl rhFVIII can be produced at 500 L bioreactor scale, maintaining high purity and recoveries. The innovative purification process ensures a high-purity and high-quality human-cl rhFVIII product with a high pathogen safety margin.

KEYWORDS:

Haemophilia A; Human-cl rhFVIII; Nuwiq®; Pathogen safety; Purification process; Recombinant factor VIII

PMID:
26318235
DOI:
10.1016/j.pep.2015.08.023
[Indexed for MEDLINE]
Free full text

Supplemental Content

Full text links

Icon for Elsevier Science
Loading ...
Support Center