Recombinant activated factor VII is reabsorbed in renal proximal tubules and is a ligand to megalin and cubilin.
OBJECTIVE
Recombinant activated factor VIIa (rFVIIa) is used for treatment of haemophilia patients with inhibitors. Tissue distribution studies in rats have shown that injected (125)I-rFVIIa accumulates in organs such as the liver and the kidneys. In this study, we explored which mechanism could be involved in renal clearance of rFVIIa.


METHODS
Immunohistochemistry was used for examination of the renal distribution in detail after injection of rFVIIa to mice and rats. Surface plasmon resonance evaluated specific binding of rFVIIa to megalin and cubilin. The biological function of megalin and cubilin in rFVIIa endocytosis was explored in opossum kidney (OK) cells.


RESULTS
Staining of rFVIIa was observed only in endosomes and lysosomes within proximal convoluted tubules from renal cortex of mice and rats. Specific binding of rFVIIa to megalin and cubilin was in the presence of receptor-associated protein (RAP) obliterated and reduced by approximately 50%, respectively. Immunofluorescence microscopy and a quantitative cellular endocytosis showed uptake in OK cells of either rFVIIa or (125)I-rFVIIa, and this uptake was significantly decreased in the presence of RAP.


CONCLUSIONS
We suggest that the renal cortex plays a significant role in clearance of injected rFVIIa and that endocytosis and degradation of rFVIIa in proximal tubule cells is mediated via binding to megalin and cubilin.
Seested
Torben
T
Exploratory ADME, Biopharmaceuticals, Novo Nordisk A/S, Måløv, Denmark. tojo@novonordisk.com
Appa
Rupa Shree
RS
Jacobsen
Christian
C
Christensen
Erik Ilso
EI
D000818:Animals; D002478:Cells, Cultured; D004705:Endocytosis; D015942:Factor VIIa / Q000378:metabolism*; D007668:Kidney / Q000378:metabolism; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D026561:Low Density Lipoprotein Receptor-Related Protein-2 / Q000378:metabolism*; D008297:Male; D051379:Mice; D009893:Opossums; D051381:Rats; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D020349:Surface Plasmon Resonance; D014018:Tissue Distribution
D026561:Low Density Lipoprotein Receptor-Related Protein-2; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; C029486:intrinsic factor-cobalamin receptor; C103587:recombinant FVIIa; D015942:Factor VIIa