Novo Nordisk A/s

Chiron Corporation

Other

Zymogenetics, Inc.

Novartis Vaccines & Diagnostics, Gmbh

Nordisk Insulinlaboratorium

Nordisk Gentofte A/s

Nordisk Gentofte

Mirella Ezban Rasmussen

Rae Lyn Burke

George Kuo

Pablo D Valenzuela

Frank R Masiarz

Martha Truett

Ole Juul Nordfang

Barbara S Chapman

Jan Moller Mikkelson

Jennifer M Favaloro

Charles E Hart

Lars Christian Petersen

Carol A Pachl

Ulla Hedner

Daniel Caput

Frank W Rasmussen

Jorgen F Hansen

Helle Worsaae

Jorgen V Jensen

Methods and compositions are provided for recombinant DNA production of Factor VIIIC and truncated derivatives thereof. Based on amino acid sequences, probes are developed for isolating messenger RNA, cDNA and/or chromosomal DNA encoding for Factor VIIIC. The Factor VIIIC gene in its entirety, a fragment thereof, or a cDNA is then used for expression of Factor VIIIC in a host. The bacteriophage &#x3bb;FVIII23D containing the 14.43 kb EcoRI fragment was deposited at the A.T.C.C. on Jan. 4, 1984 and given Accession No. 40094. Also, the vector pSVF8-200 was deposited at the A.T.C.C. on Jul. 17, 1985 and given Accession No. 40190.

Factor VIII:C nucleic acid molecules

Recombinant protein complexes having human Factor VIII:C activity are expressed in a eukaryotic host cell by transforming the host cell with first and second expression cassettes encoding a first polypeptide substantially homologous to human Factor VIII:C A domain and a second polypeptide substantially homologous to human Factor VIII:C C domain, respectively. In the present invention, the first polypeptide may be extended having at its C-terminal a human Factor VIII:C B domain N-terminal peptide, a polypeptide spacer of 3-40 amino acids, and a human Factor VIII:C B domain C-terminal peptide. Expression of the second polypeptide is improved by employing an &agr;,-antitrypsin signal sequence.

Protein complexes having factor VIII:C activity and production thereof

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. The invention relates to novel methods of treatment and uses of modified Factor VII for preventing or treating myocardial injury associated with post-ischemic reperfusion, for improving regional myocardial blood flow during reperfusion, and maintaining or improving vascular patency in a patient, as well as topical application of modified Factor VII at vascular sites susceptible to thrombus formation.

Modified factor VII

Recombinant protein complexes having human Factor VIII:C activity are expressed in a eukaryotic host cell by transforming the host cell with first and second expression cassettes encoding a first polypeptide substantially homologous to human Factor VIII:C A domain and a second polypeptide substantially homologous to human Factor VIII:C C domain, respectively. In the present invention, the first polypeptide may be extended having at its C-terminal a human Factor VIII:C B domain N-terminal peptide, a polypeptide spacer of 3-40 amino acids, and a human Factor VIII:C B domain C-terminal peptide. Expression of the second polypeptide is improved by employing an .alpha..sub.1 -antitrypsin signal sequence.

Protein complexes having Factor VIII:C activity and production thereof

Methods and compositions are provided for recombinant DNA production of Factor VIIIC and truncated derivatives thereof. Based on amino acid sequences, probes are developed for isolating messenger RNA and/or chromosomal DNA encoding for Factor VIIIC. The Factor VIIIC gene in its entirety or encoding for a fragment thereof is then used for expression of Factor VIIIC in a host. The bacteriophage .lambda.FVIII23D containing the 14.43 kb EcoRI fragment was deposited at the A.T.C.C. on Jan. 4, 1984 and given Accession No. 40094.

Method and composition for preparation of factor VIIIC

The present invention relates to the use of compounds of formula I ##STR1## wherein R.sup.1 is independently hydrogen, hydroxy, alkyl with 1 to 6 carbon atoms, acyloxy groups with 1 to 6 carbon atoms, alkyloxy with 1 to 6 carbon atoms or from 1 to 5 sugar moieties; and R.sup.2 is independently hydrogen, or alkyl with 1 to 6 carbon atoms, in the reduction of coagulation time in mammals.

Use of a low molecular weight metabolite from fungus for reducing

A coagulation active complex of Factor VIII fragments is produced by causing coagulation inactive FVIII heavy chain to react with coagulation inactive FVIII light chain in the presence of a complex forming agent. Thus, FVIII-HC and FVIII-LC are converted to coagulation active FVIII complex in the presence of divalent metal ions, such as Mn.sup.2+, Ca.sup.2+ or C.sup.2+, or a component of the pro-thrombin complex.

Process for producing a coagulation active complex of factor VIII

A coagulation active complex of Factor VIII fragments is produced by causing a coagulation inactive FVIII heavy chain to react with a coagulation inactive FVIII light chain in the presence of a complex forming agent. Thus, FVIII-HC and FVIII-LC are converted to coagulation active FVIII complex in the presence of metal ions, such as Mn.sup.2+, Ca.sup.2+, or Co.sup.2+ or a component of the prothrombin complex or a substance having reactivity to compounds containing the group --SH and/or --S--S.

Methods and compositions are provided for recombinant DNA production of Factor VIIIC and truncated derivatives thereof. Based on amino acid sequences, probes are developed for isolating messenger RNA, cDNA and/or chromosomal DNA encoding for Factor VIIIC. The Factor VIIIC gene in its entirety, a fragment thereof, or a cDNA is then used for expression of Factor VIIIC in a host. The bacteriophage .lambda.FVIII23D containing the 14.43kb EcoRI fragment was deposited at the A.T.C.C. on Jan. 4, 1984 and given Accession No. 40094. Also, the vector pSVF8-200 was deposited at the A.T.C.C. on July 17, 1985 and given Accession No. 40190.

Factor VIII:C cDNA cloning and expression

Hybridomas producing monoclonal antibodies specific for polypeptide fragments derived from human Factor VIIIC are provided. Class I hybridomas produce monoclonal antibodies reactive with a 80/77 kd doublet fragment or with both the 80/77 kd doublet and a 240 kd polypeptide. Class III hybridomas produce monoclonal antibodies reactive with the 240 kd polypeptide as well as a 92.5 kd fragment and its precursors. Class III antibodies show additional reactivity with a 40 kd thrombin digestion product. The monoclonal antibodies are useful for the separation of Factor VIIIC and its constituent polypeptides, as well as for the immunoassay of Factor VIIIC in biological samples.

Monoclonal antibodies to factor VIIIC

Concentrate of the antihemophilic factor VIII and a process for

An antihemophilic factor preparation (AHF) of high solubility and long half-life is produced in high yields by complete or partial thawing of deep-frozen human blood plasma by irradiation with electromagnetic waves of a frequency of about 10.sup.8 -10.sup.15 for a period of time and with an energy penetration such that the temperature in the thawed plasma does not exceed 10.degree. C. at any point, and processing in a manner known per se by centrifuging the thawed product to form a cryoprecipitate, redissolving the cryoprecipitate in a buffer and isolating a concentrated solution which, if desired, is freeze-dried. Irradiation is preferably effected by microwaves of a frequency of 10.sup.8 -3.times.10.sup.11 Hz, in particular 2.times.10.sup.9 -3.times.10.sup.10 Hz, or by infrared light, said frozen plasma being preheated if desired. The preparation is suitable for injection performed by the patient himself without medical assistance.

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Copenhagen, Denmark

Mirella Ezban Rasmussen