Abstract
Recombinant factor IX Fc (rFIXFc) fusion protein is the first of a new class of bioengineered long-acting factors approved for the treatment and prevention of bleeding episodes in haemophilia B. The aim of this work was to describe the manufacturing process for rFIXFc, to assess product quality and to evaluate the capacity of the process to remove impurities and viruses. This manufacturing process utilized a transferable and scalable platform approach established for therapeutic antibody manufacturing and adapted for production of the rFIXFc molecule. rFIXFc was produced using a process free of human- and animal-derived raw materials and a host cell line derived from human embryonic kidney (HEK) 293H cells. The process employed multi-step purification and viral clearance processing, including use of a protein A affinity capture chromatography step, which binds to the Fc portion of the rFIXFc molecule with high affinity and specificity, and a 15 nm pore size virus removal nanofilter. Process validation studies were performed to evaluate identity, purity, activity and safety. The manufacturing process produced rFIXFc with consistent product quality and high purity. Impurity clearance validation studies demonstrated robust and reproducible removal of process-related impurities and adventitious viruses. The rFIXFc manufacturing process produces a highly pure product, free of non-human glycan structures. Validation studies demonstrate that this product is produced with consistent quality and purity. In addition, the scalability and transferability of this process are key attributes to ensure consistent and continuous supply of rFIXFc.
Highlights
Current haemophilia B management involves replacement therapy with coagulation factor IX (FIX) concentrates, either given episodically to treat bleeding episodes or prophylactically to maintain minimum FIX activity and to prevent bleeding [1,2,3,4]
Recombinant FIX Fc Fusion Protein is the first of a new class of bioengineered longacting clotting factors. rFIXFc is composed of a single molecule of recombinant FIX (rFIX) covalently fused to the Fc domain of human immunoglobulin G1 (IgG1)
Clonal cell lines were derived and the optimal cell line selected was based on considerations of rFIXFc monomer productivity, rFIXFc activity [measured by a one-stage clotting assay based on activated partial thromboplastin time], propeptide processing, cell growth properties and stability
Summary
Current haemophilia B management involves replacement therapy with coagulation factor IX (FIX) concentrates, either given episodically (on demand) to treat bleeding episodes or prophylactically to maintain minimum FIX activity and to prevent bleeding [1,2,3,4]. FIX replacement therapy has been available since the 1970s, initially utilizing plasma-derived FIX concentrates [1,2]. Development of recombinant FIX (rFIX) in the 1990s circumvented viral safety concerns associated with plasma-derived concentrates and greatly reduced supply shortages [5] and continues to be important in the treatment of haemophilia B [6]. Due to viral contamination issues with previous plasma-derived FIX products, it is important to Accepted after revision 27 March 2014 characterize the manufacturing process of any new haemophilia therapy and to conduct validation studies that evaluate product quality and demonstrate the ability of the process to remove process-related impurities and viruses. Recombinant FIX Fc Fusion Protein (rFIXFc; Alprolix; Biogen Idec, Cambridge, MA, USA) is the first of a new class of bioengineered longacting clotting factors. The enhanced pharmacokinetic properties, prolonged halflife, safety and efficacy of rFIXFc have been described in animal models [11] and clinical studies [12,13]
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