The Scripps Research Institute

Jerry L Ware

Zaverio M Ruggeri

A polypeptide which is capable of inhibiting binding of von Willebrand factor (vWF) to platelets and comprising an amino acid sequence that corresponds to the amino acid sequence of that fragment of mature von Willebrand factor subunit having its amino terminus at about Cys.sup.509 and its carboxy terminus at about Cys.sup.695, said polypeptide comprising optionally a second and/or a third domain, the second domain corresponding to the amino acid sequence of that fragment of mature vWF subunit having its amino terminus at about Thr.sup.450 and its carboxy terminus at about Tyr.sup.508, or a subfragment or combination of subfragments thereof, and a third domain corresponding to the amino acid sequence of that fragment of mature vWF subunit having its amino terminus at about Asp.sup.696 and its carboxy terminus at about Gly.sup.727, or a subfragment or combination of subfragments thereof; and also a process for producing said polypeptides from encoding DNA sequences, and also a method of inhibiting or treating thrombosis in a patient which comprising administering to such patient an effective amount of a therapeutic composition comprising one or more polypeptides of the invention.

Therapeutic domains of van Willebrand factor

A method of inhibiting platelet transfusion refractoriness comprising administering third-party platelets to a patient whose platelet glycoprotein Ib.alpha. polypeptides expressed from the patient's DNA have been determined to have either threonine or methionine residues, or both, at amino acid residue position 145 thereof, the third-party platelets having expressed at residue position 145 of the glycoprotein Ib.alpha. polypeptide thereof amino acids that are the same as those expressed on the glycoprotein Ib.alpha. polypeptides of the patient's platelets. Also, an antibody, or a fragment thereof, in substantially pure form having as its target epitope a domain of glycoprotein Ib.alpha., the affinity of the antibody, or fragment thereof, for said epitope being dependent on the identity of the amino acid residue at position 145 of the glycoprotein. In addition, in a process for storing a supply of platelets for therapeutic use in a patient, the improvement comprising segregating the platelets into three stocks dependent upon whether residue 145 of glycoprotein Ib.alpha. polypeptides of the platelets includes: (A) threonine; (B) methionine; or (C) both threonine and methionine. Also, a method of immunoassay comprising contacting a sample of blood, or a composition derived therefrom, with alloantigenic Thr or Met.sup.145 form of glycoprotein Ib.alpha., or a fragment thereof, containing the Thr/Met.sup.145 locus, and determining whether there is present a complex of antibody and a form of the glycoprotein or a fragment thereof.

Antibodies and immunoassays for detecting platelet transfusion

A method of inhibiting platelet transfusion refractoriness comprising administering third-party platelets to a patient whose platelet glycoprotein Ib.alpha. polypeptides expressed from the patient's DNA have been determined to have either threonine or methionine residues, or both, at amino acid residue position 145 thereof, said third-party platelets having expressed at residue position 145 of the glycoprotein Ib.alpha. polypeptide thereof amino acids that are the same as those expressed on the glycoprotein Ib.alpha. polypeptides of the patient's platelets. Also, an antibody, or a fragment thereof, in substantially pure form having as its target epitope a domain of glycoprotein Ib.alpha., the affinity of said antibody, or fragment thereof, for said epitope being dependent on the identity of the amino acid residue at position 145 of said glycoprotein. In addition, in a process for storing a supply of platelets for therapeutic use in a patient, the improvement comprising segregating said platelets into three stocks dependent upon whether residue 145 of glycoprotein Ib.alpha. polypeptides of said platelets includes: (A) threonine; (B) methionine; or (C) both threonine and methionine. Also, a method of immunoassay comprising contacting a sample of blood, or a composition derived therefrom, with alloantigenic Thr or Met.sup.145 form of glycoprotein Ib.alpha., or a fragment thereof, containing said Thr/Met.sup.145 locus, and determining whether there is present a complex of antibody and a form of said glycoprotein or a fragment thereof.

Expression vectors encoding alloantigens of glycoprotein Ib.alpha.

Recombinant DNA expression vectors encoding a peptide which inhibits binding of von Willebrand factor to platelet membrane glycoprotein Ib, said vector including a nucleotide sequence encoding the amino acid sequence from HIS.sup.1 to LEU.sup.610, inclusive, of the amino terminal region of platelet membrane glycoprotein Ib.alpha., or any sequential subset thereof; mammalian host cells transformed by said vectors; and a process for producing a peptide having the identifying characteristics of the 45 kDa tryptic fragment of glycocalicin comprising the steps of (A) providing a stable, extrachromosomally replicable vector capable of directing in mammalian cells the expression of a nucleotide sequence encoding an amino acid sequence which includes said fragment, said nucleotide sequence further encoding as part of said amino acid sequence amino acids which are not native to said fragment, and which are oriented at the carboxy terminus of said fragment, (B) transforming said mammalian cells with said vector, and (C) maintaining said transformed mammalian cells under conditions permitting the expression of said peptide.

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Encinitas, CA, United States of America

Jerry L Ware