Development, Validation, and Application of a Novel Ligand-Binding Assay to Selectively Measure PEGylated Recombinant Human Coagulation Factor VIII (BAX 855).
BAX 855 is a PEGylated recombinant factor VIII preparation that showed prolonged circulatory half-life in nonclinical and clinical studies. This paper describes the development, validation, and application of a novel ligand-binding assay (LBA) to selectively measure BAX 855 in plasma. The LBA is based on PEG-specific capture of BAX 855, followed by immunological factor VIII (FVIII)-specific detection of the antibody-bound BAX 855. This assay principle enabled sensitive measurement of BAX 855 down to the low nanomolar range without interference from non-PEGylated FVIII as demonstrated by validation data for plasma from animals typically used for nonclinical characterization of FVIII. The selectivity of an in-house-developed anti-PEG and a commercially available preparation, shown by competition studies to primarily target the terminating methoxy group of PEG, also allowed assessment of the intactness of the attached PEG chains. Altogether, this new LBA adds to the group of methods to selectively, accurately, and precisely measure a PEGylated drug in complex biological matrices. The feasibility and convenience of using this method was demonstrated during extensive nonclinical characterization of BAX 855.
Baxalta Innovations GmbH , 1220 Vienna, Austria.
Alfred
A
Weber
Baxalta Innovations GmbH , 1220 Vienna, Austria.
Andrea
A
Engelmaier
Baxalta Innovations GmbH , 1220 Vienna, Austria.
Sandra
S
Hainzelmayer
Baxalta Innovations GmbH , 1220 Vienna, Austria.
Eva
E
Minibeck
Baxalta Innovations GmbH , 1220 Vienna, Austria.
Heinz
H
Anderle
Baxalta Innovations GmbH , 1220 Vienna, Austria.
Hans Peter
HP
Schwarz
Baxalta Innovations GmbH , 1220 Vienna, Austria.
Peter L
PL
Turecek
D000818:Animals; D000906:Antibodies / Q000737:chemistry; D001681:Biological Assay / Q000379:methods*; D005169:Factor VIII / Q000032:analysis* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D006207:Half-Life; D006801:Humans; D007074:Immunoglobulin G; D008024:Ligands; D008251:Macaca; D051379:Mice; D011092:Polyethylene Glycols / Q000737:chemistry; D011817:Rabbits; D051381:Rats; D011994:Recombinant Proteins / Q000032:analysis* / Q000097:blood / Q000235:genetics / Q000378:metabolism; D012680:Sensitivity and Specificity
D000906:Antibodies; D007074:Immunoglobulin G; D008024:Ligands; D011994:Recombinant Proteins; D011092:Polyethylene Glycols; C000609799:BAX 855; D005169:Factor VIII