Replacing the first epidermal growth factor-like domain of factor IX with that of factor VII enhances activity in vitro and in canine hemophilia B.
Using the techniques of molecular biology, we made a chimeric Factor IX by replacing the first epidermal growth factor-like domain with that of Factor VII. The resulting recombinant chimeric molecule, Factor IXVIIEGF1, had at least a twofold increase in functional activity in the one-stage clotting assay when compared to recombinant wild-type Factor IX. The increased activity was not due to contamination with activated Factor IX, nor was it due to an increased rate of activation by Factor VIIa-tissue factor or by Factor XIa. Rather, the increased activity was due to a higher affinity of Factor IXVIIEGF1 for Factor VIIIa with a Kd for Factor VIIIa about one order of magnitude lower than that of recombinant wild-type Factor IXa. In addition, results from animal studies show that this chimeric Factor IX, when infused into a dog with hemophilia B, exhibits a greater than threefold increase in clotting activity, and has a biological half-life equivalent to recombinant wild-type Factor IX.
Chang
J Y
JY
Center for Thrombosis and Hemostasis, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-7035, USA. jychang@med.unc.edu
Monroe
D M
DM
Stafford
D W
DW
Brinkhous
K M
KM
Roberts
H R
HR
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D004274:DNA, Recombinant / Q000378:metabolism; D003619:Dansyl Compounds / Q000494:pharmacology; D004285:Dogs; D004815:Epidermal Growth Factor / Q000235:genetics / Q000378:metabolism*; D005164:Factor IX / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D015949:Factor IXa / Q000378:metabolism; D015944:Factor VIIIa / Q000378:metabolism; D015942:Factor VIIa / Q000494:pharmacology; D015945:Factor XIa / Q000494:pharmacology; D015951:Factor Xa / Q000378:metabolism; D065427:Factor Xa Inhibitors; D002836:Hemophilia B / Q000378:metabolism*; D006801:Humans; D010314:Partial Thromboplastin Time; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism*
D000590:Amino Acid Chloromethyl Ketones; D004274:DNA, Recombinant; D003619:Dansyl Compounds; D065427:Factor Xa Inhibitors; D011993:Recombinant Fusion Proteins; D004815:Epidermal Growth Factor; C030824:dansylglutamyl-glycyl-arginine chloromethyl ketone; D015944:Factor VIIIa; D005164:Factor IX; D015942:Factor VIIa; D015949:Factor IXa; D015945:Factor XIa; D015951:Factor Xa