Accumulation of the recombinant factor VIIa in rat bone: importance of the Gla-domain and relevance to factor IX, another vitamin K-dependent clotting factor.
The vitamin K-dependent clotting factors II, VII, IX, and X are proteins which undergo gamma-carboxylation of specific glutamic acid residues prior to secretion from the liver. These unique Ca2+ binding amino acids allow the interaction of the proteins with cell surface phospholipids, a function that is crucial for expression of full procoagulant activity of the proteins. The N-terminal region of the molecule contains the gamma-carboxylation sites and is termed the Gla-domain. A preliminary observation in rats suggested that mineralized bone accumulated activated recombinant FVII (rFVIIa: NovoSeven) as well as the non-activated, single chain rFVII. The present study investigated the role of the Gla-domain in the accumulation of rFVII in bone, as well as the influence of the activation state of FVII on this phenomenon. Rats were treated with 125I-labelled rFVII, rFVIIa, Gla-domainless rFVIIa, factor IX, iodide, or recombinant human growth hormone (rhGH). Following sacrifice, radioactivity was measured in mineralized bone, among other tissues. Following administration of 125I-radiolabelled rFVII, rFVIIa and factor IX, but not Gla-domainless rFVIIa, iodide or rhGH, extensive sequestration occurred in endochondrally as well as intramenbranously ossified bones. The results indicate that the proteins containing a Gla-domain, and only these, are sequestered in bone. Additionally, the normally occurring form of FVII in the circulation, the single-chain FVII, exhibited similar kinetics in rat bone and plasma, as the two-chain rFVIIa. The half-life of rFVII/rFVIIa in mineralized bone was between 3 and 4 days, implying that significant bone accumulation of the factor will take place at steady state.(ABSTRACT TRUNCATED AT 250 WORDS)
Thomsen
M K
MK
Biopharmaceuticals Division, Novo Nordisk A/S, Gentofte, Denmark.
Wildgoose
P
P
Nilsson
P
P
Hedner
U
U
D000818:Animals; D001842:Bone and Bones / Q000378:metabolism*; D002113:Calcification, Physiologic; D006224:Cricetinae; D005164:Factor IX / Q000737:chemistry / Q000378:metabolism*; D015942:Factor VIIa / Q000737:chemistry / Q000378:metabolism*; D005260:Female; D005971:Glutamates / Q000737:chemistry / Q000378:metabolism*; D018698:Glutamic Acid; D013006:Growth Hormone / Q000378:metabolism; D007457:Iodine Radioisotopes; D007700:Kinetics; D008297:Male; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013329:Structure-Activity Relationship; D014812:Vitamin K / Q000378:metabolism*
D005971:Glutamates; D007457:Iodine Radioisotopes; D011994:Recombinant Proteins; D014812:Vitamin K; D018698:Glutamic Acid; D005164:Factor IX; D013006:Growth Hormone; D015942:Factor VIIa