Zymogenetics, Inc.

Novo Nordisk A/s

Other

Zymogenetrics, Inc.

Kathleen L Berkner

Charles E Hart

Lars Christian Petersen

Ulla Hedner

Claus Bregengaard

Mark J Murray

Donald C Foster

Frederick S Hagen

Sharon J Busby

Margaret Y Insley

Charles L Gray

Richard G Woodbury

Methods are disclosed for producing proteins having biological activity for blood coagulation mediated by Factor VIIa or Factor IX. The proteins are produced by mammalian host cells which have been stably transfected with a DNA construct containing a nucleotide sequence which codes at least partially for either Factor VII or Factor IX. The nucleotide sequence comprises a first nucleotide sequence encoding a calcium binding domain, joined to a second nucleotide sequence positioned downstream of the first sequence. The second sequence encodes a catalytic domain for the serine protease activity of either Factor VIIa or Factor IX. The joined sequences code for proteins having substantially the same biological activity for blood coagulation as either Factor VIIa or Factor IX.

Expression of factor VII and IX activities in mammalian cells

The catalytic active site of Factor VII is modified to produce a compound which effectively interrupts the blood coagulation cascade. The modifications render Factor VIIa substantially unable to activate plasma Factors X or IX. Pharmaceutical compositions of the modified Factor VII are used to treat a variety of coagulation-related disorders, including platelet deposition, vascular thrombosis, ischemic reperfusion, acute closure of a coronary artery, vascular restenosis secondary to balloon angioplasty, endarterectomy, reductive atherectomy, stent placement, laser therapy or rotablation.

Modified factor VII

The catalytic active site of Factor VII is modified to produce a compound which effectively interrupts the blood coagulation cascade. The modifications render Factor VIIa substantially unable to activate plasma Factors X or IX. Pharmaceutical compositions of the modified Factor VII are used to treat a variety of coagulation-related disorders.

Modified Factor VII

Factor VII of the coagulation cascade is modified to act as an anticoagulant. Amino acid modifications are employed to produce a modified Factor VII having a substantially reduced susceptibility to activation by enzymes which typically activate wild-type Factor VII. The modified Factor VII is able to compete with wild-type Factor VII and/or VIIa for binding tissue factor, inhibiting clotting activity. As the modified Factor VII acts specifically to interrupt the coagulation cascade, pharmaceutical compositions of modified Factor VII may be administered in place of, or in conjunction with lower doses of, conventional anticoagulant therapies.

Modified factor VII anticoagulant proteins

Methods are disclosed for producing a protein which has substantially the same biological activity as human protein C or human activated protein C. The protein is produced by mammalian host cells transfected with a plasmid capable of integration in mammalian host cell DNA. The plasmid includes a promoter followed downstream by a nucleotide sequence which encodes a protein having substantially the same structure and/or activity as human protein C or human activated protein C the nucleotide sequence being followed downstream by a polyadenylation signal.

Production of activated protein C

DNA sequence encoding Factor VII with an amino acid substitution at

Genomic and cDNA sequences coding for a protein having substantially the same biological activity as human protein C and recombinant transfer vectors comprising these sequences are disclosed. Methods are disclosed for producing a protein which has substantially the same biological activity as human protein C. The protein, which may be in the form of activated protein C, is produced by mammalian host cells transfected with a plasmid capable of integration in mammalian host cell DNA. The plasmid includes a promoter followed downstream by a nucleotide sequence which encodes a protein having substantially the same structure and/or activity as human protein C, the nucleotide sequence being followed downstream by a polyadenylation signal.

Expression of protein C

Methods are disclosed for producing proteins having biological activity for blood coagulation mediated by Factor VIIa. The proteins are produced by mammalian host cells which have been stably transfected with a DNA construct containing a nucleotide sequence which codes at least partially for either Factor VII. The nucleotide sequence comprises a first nucleotide sequence encoding a calcium binding domain, joined to a second nucleotide sequence positioned downstream of the first sequence. In particular, the first nucleotide sequence may be derived from a genomic clone or cDNA clone of Factor VII. The second sequence encodes a catalytic domain for the serine protease activity of Factor VIIA. The joined sequences code for proteins having substantially the same biological activity for blood coagulation as Factor VIIa.

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Seattle, WA, United States of America

Kathleen L Berkner