WO1999046299A1 - Improved methods for producing factor viii proteins - Google Patents
Improved methods for producing factor viii proteins Download PDFInfo
- Publication number
- WO1999046299A1 WO1999046299A1 PCT/US1999/005329 US9905329W WO9946299A1 WO 1999046299 A1 WO1999046299 A1 WO 1999046299A1 US 9905329 W US9905329 W US 9905329W WO 9946299 A1 WO9946299 A1 WO 9946299A1
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- solution
- polypeptide
- factor
- fold
- dilution
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/745—Blood coagulation or fibrinolysis factors
- C07K14/755—Factors VIII, e.g. factor VIII C (AHF), factor VIII Ag (VWF)
Definitions
- the present invention relates to improved methods for the purification of procoagulant proteins, particularly recombinant production of Factor Vi ⁇ and related proteins.
- Hemophilia is an inherited disease which has been known for centuries, but it is only within the last few decades that it has been possible to differentiate among the various forms; hemophilia A and hemophilia B. Hemophilia A is caused by strongly decreased level or absence of biologically active coagulation factor VTH, which is a protein normally present in plasma. Until recently, therapeutic factor VQI concentrates have been prepared by fractionation of plasma.
- variants and analogs of FNHI protein, and D ⁇ A sequences encoding them have also been reported [see e.g. Toole, United States Patent 4,868,112; EP 0786474; WO 86/06101 and WO 87/07144].
- variants and analogs are modified such that part or all of the B domain are missing and or specific amino acid positions are modified, for example, such that normally protease-labile sites are resistant to proteolysis, e.g. by thrombin or activated Protein C.
- Other analogs include modification at one or more lysine and/or tyrosine residues.
- the B domain is dispensable for the procoagulant activity of FVm, and that active procoagulant protein can be expressed and secreted by expression of a FVHI-encoding D ⁇ A in which the nucleotide region encoding part or all of the B domain is lacking.
- active protein of these variants produced and secreted, it accumulates in the media at higher levels than when expressed by the full- length DNA.
- the reduced level of active procoagulant FVHI protein in the media has been attributed, at least in part to several factors [see e.g. WO 87/04187, WO 88/08035 and WO 88/03558].
- the present invention provides improved methods for the purification of procoagulant proteins, including both FVm and variants thereof, which may be produced by recombinant techniques in higher yield and/or in more homogeneous form.
- the present invention provides improved methods of purification of FVILI protein from cell cultures, preferably from recombinant cell cultures.
- the methods provide for obtaining FVIH protein of a higher purity than methods currently in use.
- the methods of the present invention comprise diluting the eluate from the immunoaffinity column with a solution of higher ionic strength than the eluate solution.
- the methods of the present invention comprise diluting the eluate from an immunoaffinity column with a solution containing lower amounts of ethylene glycol than contained in the eluate solution.
- the present invention provides improved methods for purification of a Factor VDI polypeptide comprising: a) adding a mixture containing Factor VDI polypeptide to be purified to an immunoaffinity matrix which binds by hydrophobic attraction to the FVm polypeptide; b) eluting the Factor VDI polypeptide from the immunoaffinity matrix with a desorbing solution which causes desorption of the Factor VTLI polypeptide, which is released in an elution solution; c) diluting the elution solution with a solution comprising higher ionic strength than that of the elution solution, resulting in a diluted Factor VIH solution; d) passing the diluted Factor VHI solution through an ion exchange column capable of binding to the Factor VHI polypeptide, thereby binding the Factor VDI polypeptide while allowing contaminants to pass through the ion exchange column; and e) eluting the purified Factor
- the desorbing solution of step (b) may contain no salt, or very low salt.
- the dilution of step (c) is preferably performed using a solution comprising from about 5 to about 20 mM NaCl, most preferably about 5 to about 15 mM NaCl.
- the eluting solution is preferably diluted with salt-containing solution from about 3-fold to about 5-fold , most preferably about 3-fold.
- the present invention comprises improved methods for purification of a Factor VIE polypeptide comprising: a) adding a mixture containing Factor VHI polypeptide to be purified to an immunoaffinity matrix which binds by hydrophobic attraction to the FVIH polypeptide; b) eluting the Factor VDI polypeptide from the immunoaffinity matrix with a desorbing solution which causes desorption of the Factor VIII polypeptide, which is released in an elution solution, wherein the desorbing solution comprises a non-polar agent; c) diluting the elution solution with a solution comprising lower concentration of the non-polar agent than that of the desorbing solution, resulting in a diluted Factor VIQ solution; d) passing the diluted Factor VDI solution through an ion exchange column capable of binding to the Factor VIE polypeptide, thereby binding the Factor VIE polypeptide while allowing contaminants to pass through the ion exchange column; and e) eluting the purified
- the desorbing solution of step (b) contains ethylene glycol, more preferably about 50% (v/v) ethylene glycol, and the dilution of step (c) is performed using a solution comprising less than about 50% (v/v) ethylene glycol, such that the final concentration of ethylene glycol is from about 17% to about 33% (v/v).
- the desorbing solution of step (b) contains 50% (v/v) ethylene glycol, and the dilution of step (c) is performed using a solution comprising no ethylene glycol, such that the final concentration of ethylene glycol is from about 17 to about 33% (v/v), most preferably about 33% (v/v) ethylene glycol.
- the elution solution is diluted from about 1.5-fold to about 3-fold, most preferably about 1.5 fold, or 2:3.
- the Factor VIE polypeptide of the present invention is generally produced recombinantly, but may also be purified from plasma.
- the recombinant Factor VIE polypeptide may be natural full length Factor VHI polypeptide, or a variant, such as a B- domain deleted variant of Factor VIE, including the VEI:SQ variant.
- the immunoaffinity columns useful in the present invention may be any industrially acceptable column and resin, to which is adsorbed one or more monoclonal or polyclonal antibodies which are capable of binding to a Factor VIE polypeptide and in which the Factor
- VIE polypeptide may later be released using standard methods and reagents. Suitable monoclonal antibodies, for example, are disclosed in Fass et al., Blood, 59:594-600 (1982).
- desorbing substance is any non-polar agent, such as ethylene glycol, dioxane, propylene glycol and polyethylene glycol, or any appropriate low ionic strength, low polarity buffered solution.
- non-polar agent such as ethylene glycol, dioxane, propylene glycol and polyethylene glycol, or any appropriate low ionic strength, low polarity buffered solution.
- the mixtures containing Factor VIE polypeptides may also include detergents and/or solvents, such as polyoxyethyl detergents, including Triton X-100, Tween 80.
- the Factor VIE polypeptide containing solution may include buffering substances, such as histidine.
- DG44 ⁇ Y was electroporated with an expression vector containing the r-VIE SQ gene and an expression vector containing the dihydrofolate reductase gene.
- the conditioned medium (containing fetal calf serum) was clarified and then concentrated by tangential flow filtration.
- the solution was loaded onto an SP Sepharose Fast Flow cation exchange resin, wherein the r-VIE SQ binds selectively to the resin through electrostatic forces.
- the r-VIE SQ is eluted from the column at elevated ionic strength by flowing elution solution (0.8 M NaCl, 3 mM EDTA, 0.02% (v/v) surfactant [Octoxynol 9], 0.1 MNH 4 Ac, 5 mM CaCl 2 , 1M Sorbitol, pH 6.3 ⁇ 0.2) and is collected as a single UN adsorbing peak.
- the r-VIE SQ is then put through a virus inactivation step employing the solvent/detergent method using T ⁇ BP [Tri-(n-butyl)phosphate] and surfactant [such as Octoxynol 9, Triton X-100).
- the r-VIE SQ is next loaded onto an immunoaffinity chromatography gel, where the ligand was a monoclonal antibody (mAb, named 8A4) directed towards the heavy chain of factor VIE.
- the factor VIE was eluted with a buffer containing 0.05 M histidine, 0.05 M CaCl 2 and 50% ethylene glycol and 0.02% Octoxynol 9 (Tween), pH 6.6.
- the mAb eluate was loaded onto an anion exchange column, Q Sepharose® FF sold by Pharmacia AB of Uppsala, Sweden.
- the FVIE SQ was eluted with a Q elution buffer containing 0.05 M histidine, 4 mM CaCl 2 0.4 M ⁇ aCl, pH 6.3.
- the solution used to equilibrate the Q-column (the same as the desorption buffer of the monoclonal antibody column) prior to loading onto die ion exchange column) comprises approximately the following composition: 0.05 M histidine
- the eluate was diluted from about 3-fold to about 5-fold with Q-equilibration buffer containing NaCl in the range of about 7 to about 20 mM.
- Dilution generally produces a significant reduction in the amount of murine IgG recovered from the eluate prior to placing on the ion exchange column.
- NaCl also increased recovery of FVIE activity. This increase in recovery was sufficient to offset the loss in recovery resulting from dilution.
- the best results were observed in 3-fold to 5-fold dilutions with NaCl in the range of about 10 to about 17 mM NaCl.
- the best recovery yields of FVIE activity were obtained with dilutions of about 3- fold with about 15 mM NaCl. Dilution with less than about 7 mM NaCl or greater than about 20 mM NaCl resulted in a loss of final recovery of FVIE activity.
- the eluate is diluted from about 1.5-fold to about 3-fold with Q Equilibration Buffer that does not contain ethylene glycol, resulting in final ethylene glycol content varying from about 50% (v/v) in the Q Equilibration Buffer down to as low as about 17% (v/v) in the 3-fold dilution without ethylene glycol.
- Q Equilibration Buffer that does not contain ethylene glycol
- a 1.5-fold dilution without ethylene glycol resulted in about a 33% (v/v) final ethylene glycol concentration.
- Q Equilibration Buffer containing about 50% (v/v) ethylene glycol.
- a suitable downstream purification process for Factor VELSQ as produced in Example 1 may consist of five chromatographic steps: cationic exchange (SP Sepharose FF), immunoaffinity (mAb Sepharose FF), anionic exchange (Q Sepharose FF), hydrophobic interaction (EEC, butyl Sepharose FF), and gel permeation chromatography (Superdex 200 pg).
- SP Sepharose FF cationic exchange
- mAb Sepharose FF immunoaffinity
- Q Sepharose FF anionic exchange
- EEC butyl Sepharose FF
- Superdex 200 pg gel permeation chromatography
- Q-Sepharose FF resin was packed in a 79x5 mm ID Pharmacia HR column. All buffers employed in this study were prepared by CTS by established procedures. The mAb peak pool from the purification process were obtained frozen at -80 °C from P&U,
- the Q-Sepharose FF column was initially equilibrated with 10 CV of buffer at a flow rate of 0.5 ml/min. Subsequently, the mAb peak pool was diluted with the appropriate dilution buffer and loaded onto the Q-Sepharose FF column at a flow rate of 0.2 ml min.
- the total activity units loaded in all the experiments was 48,350 U/ml of the resin, and is close to the upper limit specified in the PLA.
- the activity of the mAb peak pool used to perform these experiments was 2860 IU/ml.
- the load volume in the 3-fold and 5-fold dilution experiments was therefore 78.6 mis and 131 mis respectively.
- the column was washed with 40 CVs of a buffer containing 150 mM NaCl, 4 mM CaC12, 50 mM Histidine, pH 6.6, at a flow rate of 0.32 ml/min (wash 2).
- the bound protein was then eluted with a buffer containing 400 mM NaCl, 4 mM CaC12, 50 mM Histidine, pH 6.3 at a flow rate of 0.05 ml min. Wash 2 and elution in all the experiments were performed at a flow rate of 0.05 ml min. Wash 2 and elution in all the experiments were performed in the reverse direction.
- the column effluent during the various operations was collected and assayed for activity.
- a 1.6 cv fraction was pooled during elution beginning at the upward drift in the absorbance at 280 nm and is termed the peak pool.
- the load and peak pool samples were assayed for mouse IgG and HCP levels by performing ELISA (P&U,
- the anion exchange column was regenerated, following each scale down run, by passing five column volumes each of 2.0 M NaCl, 100 mM sodium phosphate (monobasic), pH 3.0 and 2.0M NaCl, 100 nM sodium phosphate (dibasic), pH 11.0
- the mAB peak pool was diluted different fold with (I) mAb elution buffer and (ii) mAb elution buffer containing 40 mM NaCl, and incubated at room temperature. The activity in these samples was then assayed at different time points.
- the mAB peak pool was diluted 2-fold, 3-fold and 5-fold with mAB elution buffer and incubated at room temperature. The drop in activity of these samples was monitored as a function of time. A modest drop of 20% in activity was observed over the course of 24 hours. The loss in activity was negligible at the end of 4 hours, and less than 10% at the end of 8 hours. Further, percentage drop in activity was observed to be independent of the extent of dilution of the mAb peak pool and hence independent of the solution concentration of FVIE in the mAb elution buffer. Similar results were obtained upon dilution of the mAb peak pool with mAb elution buffer containing 40 mM NaCl. Q-Sepharose scale down experiments:
- Table 1 Dilution with mAb Elution Buffer Dilution Challenge Load Loading Load Flow Fold (ICJ ml (IU/ml) Time Activity at Through
- NaCl concentration was as high as 20% at an NaCl concentration of 20 mM. This is expected since the protein migrates farther down the column during loading and subsequently takes longer to elute when the flow is reversed.
- mice IgG data on the peak and post-peak pools for all 3- and 5-fold dilution experiments are shown in Table 3:
- the IgG values in the peak pool for the 3-fold dilution runs varied from 0.5 to 1.2 ng/KIU and 0.5 to 0.8 ng/KIU for the 5-fold dilution runs.
- dilution of the mAb peak pool with mAb elution buffer, with or without NaCl prior to loading reduced the IgG levels in the Q- Sepharose peak pool.
- This effect beyond the mere dilution of IgG levels, may be the result of a given association constant for formation of IgG-FVTE:SQ complex.
- lowering the concentrations of IgG and FVELSQ lowers the concentration of the complex, thereby allowing greater removal of IgG across the ion exchanger.
- HCP Host Cell Protein
- the optimum concentration for the 3-fold and 5-fold dilution runs appears to be in the 7 to 20 mM concentration, more particularly about 15 mM.
- Diluting the mAb peak pool with mAb elution buffer also reduced the IgG and HCP levels in the Q-Sepharose peak pool.
- HCP levels in the Q-Sepharose peak pool decreased with increasing NaCl concentrations in the load.
- a combination of dilution of the mAb peak pool and adding NaCl at concentrations of 7 to 20 mM resulted in improved purification without significant yield loss.
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Abstract
Description
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Priority Applications (7)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP99912427A EP1062244B1 (en) | 1998-03-12 | 1999-03-11 | Improved methods for producing factor viii proteins |
AU30803/99A AU747274C (en) | 1998-03-12 | 1999-03-11 | Improved methods for producing factor VIII proteins |
DE69902639T DE69902639T2 (en) | 1998-03-12 | 1999-03-11 | IMPROVED METHOD FOR PRODUCING FACTOR VIII |
DK99912427T DK1062244T3 (en) | 1998-03-12 | 1999-03-11 | Improved Methods for Preparation of Factor VIII Proteins |
AT99912427T ATE222925T1 (en) | 1998-03-12 | 1999-03-11 | IMPROVED METHOD FOR PRODUCING FACTOR VIII |
JP2000535675A JP4405668B2 (en) | 1998-03-12 | 1999-03-11 | Improved method for producing factor VIII protein |
CA002306820A CA2306820C (en) | 1998-03-12 | 1999-03-11 | Improved methods for producing factor viii proteins |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US7780298P | 1998-03-12 | 1998-03-12 | |
US60/077,802 | 1998-03-12 |
Publications (1)
Publication Number | Publication Date |
---|---|
WO1999046299A1 true WO1999046299A1 (en) | 1999-09-16 |
Family
ID=22140139
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/US1999/005329 WO1999046299A1 (en) | 1998-03-12 | 1999-03-11 | Improved methods for producing factor viii proteins |
Country Status (11)
Country | Link |
---|---|
US (2) | US6683159B2 (en) |
EP (1) | EP1062244B1 (en) |
JP (1) | JP4405668B2 (en) |
AT (1) | ATE222925T1 (en) |
AU (1) | AU747274C (en) |
CA (1) | CA2306820C (en) |
DE (1) | DE69902639T2 (en) |
DK (1) | DK1062244T3 (en) |
ES (1) | ES2182501T3 (en) |
PT (1) | PT1062244E (en) |
WO (1) | WO1999046299A1 (en) |
Cited By (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2008102923A1 (en) | 2007-02-23 | 2008-08-28 | Sk Chemicals Co., Ltd. | Process for producing and purifying factor viii and its derivatives |
US7485291B2 (en) | 2003-06-03 | 2009-02-03 | Cell Genesys, Inc. | Compositions and methods for generating multiple polypeptides from a single vector using a virus derived peptide cleavage site, and uses thereof |
US7498024B2 (en) | 2003-06-03 | 2009-03-03 | Cell Genesys, Inc. | Compositions and methods for enhanced expression of immunoglobulins from a single vector using a peptide cleavage site |
RU2500818C1 (en) * | 2012-07-19 | 2013-12-10 | Общество с ограниченной ответственностью "Международный биотехнологический центр "Генериум" ("МБЦ "Генериум") | RECOMBINANT PLASMID DNA pAP227 CODING POLYPEPTIDE OF RECOMBINANT FACTOR VIII OF HUMAN BLOOD COAGULABILITY, LINE OF CELLS Cricetulus griseus CHO, 2H5-PRODUCER OF RECOMBINANT FACTOR VIII OF HUMAN BLOOD COAGULABILITY, AND METHOD FOR OBTAINING POLYPEPTIDE HAVING ACTIVITY OF RECOMBINANT FACTOR VIII |
Families Citing this family (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US8183344B2 (en) | 1996-04-24 | 2012-05-22 | University Of Michigan | Inactivation resistant factor VIII |
DK1062244T3 (en) * | 1998-03-12 | 2002-12-30 | Inst Genetics Llc | Improved Methods for Preparation of Factor VIII Proteins |
US7820796B2 (en) * | 1998-03-12 | 2010-10-26 | Genetics Institute, Llc. | Methods for producing Factor VIII proteins |
US20050060775A1 (en) * | 1998-05-14 | 2005-03-17 | Hooker Brian S. | Production of human coagulation factor VIII from plant cells and whole plants |
US20080070251A1 (en) * | 2006-06-30 | 2008-03-20 | Kaufman Randal J | Method of Producing Factor VIII Proteins by Recombinant Methods |
US20090203077A1 (en) * | 2006-06-30 | 2009-08-13 | The Regents Of The University Of Michigan | Method of producing factor viii proteins by recombinant methods |
WO2012082933A1 (en) | 2010-12-15 | 2012-06-21 | Baxter International, Inc. | Eluate collection using conductivity gradient |
JP2014138614A (en) * | 2014-04-09 | 2014-07-31 | Sk Chemicals Co Ltd | Process for producing and purifying factor viii and its derivatives |
Citations (2)
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WO1986004486A1 (en) * | 1985-02-01 | 1986-08-14 | New York University | Method for purifying antihemophilic factor |
EP0286323A2 (en) * | 1987-03-31 | 1988-10-12 | BAXTER INTERNATIONAL INC. (a Delaware corporation) | An ultrapurification process for factor VIII:C |
Family Cites Families (9)
Publication number | Priority date | Publication date | Assignee | Title |
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US4312935A (en) * | 1979-08-09 | 1982-01-26 | International Business Machines Corporation | Class of E-beam resists based on conducting organic charge transfer salts |
JPS5764648A (en) * | 1980-10-06 | 1982-04-19 | Takeda Chem Ind Ltd | N-substituted derivative of valienamine, its preparation, and alpha-glucosidase inhibitor |
US4757006A (en) | 1983-10-28 | 1988-07-12 | Genetics Institute, Inc. | Human factor VIII:C gene and recombinant methods for production |
CA1341174C (en) | 1985-04-12 | 2001-01-30 | John J. Toole Jr. | Procoagulant proteins derived from factor viii: c |
WO1987004187A1 (en) | 1986-01-03 | 1987-07-16 | Genetics Institute, Inc. | METHOD FOR PRODUCING FACTOR VIII:c-TYPE PROTEINS |
US5451521A (en) | 1986-05-29 | 1995-09-19 | Genetics Institute, Inc. | Procoagulant proteins |
US4912040A (en) | 1986-11-14 | 1990-03-27 | Genetics Institute, Inc. | Eucaryotic expression system |
CA1331157C (en) | 1987-04-06 | 1994-08-02 | Randal J. Kaufman | Method for producing factor viii:c-type proteins |
DK1062244T3 (en) * | 1998-03-12 | 2002-12-30 | Inst Genetics Llc | Improved Methods for Preparation of Factor VIII Proteins |
-
1999
- 1999-03-11 DK DK99912427T patent/DK1062244T3/en active
- 1999-03-11 CA CA002306820A patent/CA2306820C/en not_active Expired - Lifetime
- 1999-03-11 ES ES99912427T patent/ES2182501T3/en not_active Expired - Lifetime
- 1999-03-11 AU AU30803/99A patent/AU747274C/en not_active Expired
- 1999-03-11 AT AT99912427T patent/ATE222925T1/en active
- 1999-03-11 WO PCT/US1999/005329 patent/WO1999046299A1/en active IP Right Grant
- 1999-03-11 EP EP99912427A patent/EP1062244B1/en not_active Expired - Lifetime
- 1999-03-11 DE DE69902639T patent/DE69902639T2/en not_active Expired - Lifetime
- 1999-03-11 PT PT99912427T patent/PT1062244E/en unknown
- 1999-03-11 JP JP2000535675A patent/JP4405668B2/en not_active Expired - Lifetime
-
2000
- 2000-12-29 US US09/752,280 patent/US6683159B2/en not_active Expired - Lifetime
-
2003
- 2003-12-03 US US10/728,242 patent/US20040087776A1/en not_active Abandoned
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO1986004486A1 (en) * | 1985-02-01 | 1986-08-14 | New York University | Method for purifying antihemophilic factor |
EP0286323A2 (en) * | 1987-03-31 | 1988-10-12 | BAXTER INTERNATIONAL INC. (a Delaware corporation) | An ultrapurification process for factor VIII:C |
US5470954A (en) * | 1987-03-31 | 1995-11-28 | Baxter International Inc. | Ultrapurification process for factor VIII |
US5470954C1 (en) * | 1987-03-31 | 2001-02-06 | Baxter Travenol Lab | Ultrapurification process for factor viii |
Cited By (11)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US7485291B2 (en) | 2003-06-03 | 2009-02-03 | Cell Genesys, Inc. | Compositions and methods for generating multiple polypeptides from a single vector using a virus derived peptide cleavage site, and uses thereof |
US7498024B2 (en) | 2003-06-03 | 2009-03-03 | Cell Genesys, Inc. | Compositions and methods for enhanced expression of immunoglobulins from a single vector using a peptide cleavage site |
US7662623B2 (en) | 2003-06-03 | 2010-02-16 | Biosante Pharmaceuticals, Inc. | Compositions and methods for enhanced expression of recombinant polypeptides from a single vector using a peptide cleavage site |
US7709224B2 (en) | 2003-06-03 | 2010-05-04 | Biosante Pharmaceuticals, Inc. | Compositions and methods for enhanced expression of recombinant polypeptides from a single vector using a peptide cleavage site |
WO2008102923A1 (en) | 2007-02-23 | 2008-08-28 | Sk Chemicals Co., Ltd. | Process for producing and purifying factor viii and its derivatives |
EP2126106A1 (en) * | 2007-02-23 | 2009-12-02 | SK Chemicals, Co., Ltd. | Process for producing and purifying factor viii and its derivatives |
EP2126106A4 (en) * | 2007-02-23 | 2012-11-28 | Sk Chemicals Co Ltd | Process for producing and purifying factor viii and its derivatives |
US9441030B2 (en) | 2007-02-23 | 2016-09-13 | Sk Chemicals Co., Ltd. | Process for producing and purifying factor VIII and its derivatives |
EP3112471A1 (en) * | 2007-02-23 | 2017-01-04 | Sk Chemicals Co., Ltd. | Process for producing and purifying factor viii and its derivatives |
US9926361B2 (en) | 2007-02-23 | 2018-03-27 | Sk Chemicals Co., Ltd. | Process for producing and purifying factor VIII and its derivatives |
RU2500818C1 (en) * | 2012-07-19 | 2013-12-10 | Общество с ограниченной ответственностью "Международный биотехнологический центр "Генериум" ("МБЦ "Генериум") | RECOMBINANT PLASMID DNA pAP227 CODING POLYPEPTIDE OF RECOMBINANT FACTOR VIII OF HUMAN BLOOD COAGULABILITY, LINE OF CELLS Cricetulus griseus CHO, 2H5-PRODUCER OF RECOMBINANT FACTOR VIII OF HUMAN BLOOD COAGULABILITY, AND METHOD FOR OBTAINING POLYPEPTIDE HAVING ACTIVITY OF RECOMBINANT FACTOR VIII |
Also Published As
Publication number | Publication date |
---|---|
AU747274C (en) | 2004-03-18 |
US6683159B2 (en) | 2004-01-27 |
EP1062244B1 (en) | 2002-08-28 |
ATE222925T1 (en) | 2002-09-15 |
DK1062244T3 (en) | 2002-12-30 |
JP4405668B2 (en) | 2010-01-27 |
US20020115832A1 (en) | 2002-08-22 |
JP2002506081A (en) | 2002-02-26 |
AU747274B2 (en) | 2002-05-09 |
EP1062244A1 (en) | 2000-12-27 |
US20040087776A1 (en) | 2004-05-06 |
DE69902639D1 (en) | 2002-10-02 |
CA2306820C (en) | 2008-09-09 |
DE69902639T2 (en) | 2003-04-24 |
PT1062244E (en) | 2002-12-31 |
AU3080399A (en) | 1999-09-27 |
ES2182501T3 (en) | 2003-03-01 |
CA2306820A1 (en) | 1999-09-16 |
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