Program: Oral and Poster Abstracts
Session: 322. Disorders of Coagulation or Fibrinolysis: Poster III
The manufacturing process for BAX 826 comprises several steps, starting with the bulk drug substance (BDS) of the ADVATE process which is subjected to polysialylation, i.e. covalent attachment of PSA of an average molecular weight of 20 kDa to rFVIII. Polysialylation is followed by a sequence of chromatographic purification steps and concentration of the conjugate by an ultra-/diafiltration step leading to the pre-formulated BDS. Final formulation of the BDS includes a filling and lyophilization step to obtain the final drug product. The process described is suited to manufacture BAX 826 in large scale and showed a good batch to batch consistency, ensuring an equivalent product quality for each batch.
BAX 826 was extensively structurally and functionally characterized. The methods used included reducing and non-reducing peptide mapping to determine amino acid sequence and post translational modifications, qualitative analysis of modification sites, SDS-PAGE and Western blot analysis, assessment of three dimensional structure similarities of BAX 826 and rFVIII, ADVATE, by Fourier-transformed infrared spectroscopy (FTIR), dynamic light scattering (DLS) and circular dichroism (CD). Functional characterization was performed by assessment of the kinetics of the assembly and activity of FIXa-FVIII (tenase) complex, determination of the rate of activation and inactivation of BAX 826 by thrombin, determination of overall hemostatic potency by a thrombin generation assay, measurement of the rate of inactivation of untreated or thrombin-activated BAX 826 by activated protein C, measurement of kinetics of the binding of BAX 826 to von Willebrand factor (VWF), to phospholipids, and to low-density lipoprotein (LDL)-receptor-related protein 1 (LRP1) by surface plasmon resonance spectroscopy.
While BAX 826 retained full hemostatic functionality of FVIII as a co-factor of the tenase complex, and can thus be considered a fully active FVIII molecule, it was found to have a reduced binding to VWF and to LRP1. Together with the conception that VWF as FVIII’s chaperone dictates the maximally achievable terminal half-life extension for FVIII reduced binding to VWF and FVIII’s major clearance receptor LRP1likely explains the prolonged pharmacokinetic (PK) properties when comparing BAX 826 with unmodified FL rFVIII in animal models. In summary, BAX 826, a polysialylated rFVIII derivative, can be manufactured reproducibly without relevant changes to the protein structure characteristic for a functional FVIII molecule with high specific activity.
Disclosures: Turecek: Baxalta Innovations GmbH: Employment . Siekmann: Baxalta Innovations GmbH: Employment . Mitterer: Baxalta Innovations GmH: Employment . Graninger: Baxalta Innovations GmbH: Employment . Schrenk: Baxalta Innovations GmbH: Employment . Matthiessen: Baxalta Innovations GmbH: Employment . Rottensteiner: Baxalta Innovations GmbH: Employment . Hoellriegl: Baxalta Innovations GmbH: Employment . Putz: Baxalta Innovations GmbH: Employment . Schwarz: Baxalta Innovations GmbH: Employment . Scheiflinger: Baxalta Innovations GmbH: Employment .
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