WO2005079835A1 - Blアンジオスタチンを含む制がん剤 - Google Patents
Blアンジオスタチンを含む制がん剤 Download PDFInfo
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- WO2005079835A1 WO2005079835A1 PCT/JP2004/002124 JP2004002124W WO2005079835A1 WO 2005079835 A1 WO2005079835 A1 WO 2005079835A1 JP 2004002124 W JP2004002124 W JP 2004002124W WO 2005079835 A1 WO2005079835 A1 WO 2005079835A1
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- WO
- WIPO (PCT)
- Prior art keywords
- angiostatin
- buffer
- cancer
- anticancer agent
- cells
- Prior art date
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Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/43—Enzymes; Proenzymes; Derivatives thereof
- A61K38/46—Hydrolases (3)
- A61K38/48—Hydrolases (3) acting on peptide bonds (3.4)
- A61K38/482—Serine endopeptidases (3.4.21)
- A61K38/484—Plasmin (3.4.21.7)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
Definitions
- the present invention relates to an anticancer agent containing BL angiostatin and a method for producing the same.
- Angiostatin is known as one such substance.
- Angiostatin is a protein with a molecular weight of about 40,000 obtained by degrading the fibrinolytic factor plasminogen present in the blood. It has been reported that it has a significant effect (MS O'Reilly et al., Cell, 79, 315-328 (1994)).
- Angiostatin includes (1) plasminogen hydrolyzed with elastase (MS O'Reilly et al., Nature Medicine, 2, 689-692 (1996)) and (2) gene recombination technology. Two types were known, which were produced directly by yeast using the method (BK Sim et al., Cancer Research 57, 1329-1334 (1997)).
- angiostin is difficult to selectively produce angiostatin from plasminogen due to low by-product substrate specificity of elastase. There was a disadvantage that the activity of the statin was low. Also,
- amino acid sequence of human plasminogen a signal peptide - for those containing (Me t 1 G ly 19)
- amino acid sequence of the mature polypeptide that is, Glu 1 —A sn 791 .
- Recombinant angiostatin also varies in physical properties and activity compared to its native counterpart, which can be attributed to changes in conformation due to differences in protein folding during expression in heterologous organisms, sugar chains, It is thought that the difference is the difference (Cao, Y., Int
- Bashiroraishin MA cleaves plasminogen very selectively, fragments having the amino acid sequence of S er 441 from G 1 u 1 as BL-en Jiosutachin (major product, as a minor product, S er 441 from P he 75 , G lu 1 from V a 1 44 9, and P he 75 V a 1 fragment consisting of amino acids 449) to allowed to live Ji from (JP 2002- 272453 JP).
- plasminogen which develops Bashiroraishin MA immobilized reactor (Afuietito wrap reactor), as a main component fragment having the amino acid sequence of S er 441 from G 1 u 1 at one step of the step from human plasma
- plasminogen which develops Bashiroraishin MA immobilized reactor (Afuietito wrap reactor)
- plasminogen which develops Bashiroraishin MA immobilized reactor (Afuietito wrap reactor
- Recombinant angiostatin is an internal fragment of plasminogen consisting of amino acids from Le 74 to Le 451 (Sim, BK et al. Cancer Res 57, 1329-34 (1997)), whereas BL angiostatin Is angios as described above It has a different structure from that of statins, but has the same anti-vascular endothelial cell action as angiostatin.
- fragment having the Amino acid sequence of S er 4 4 1 from G 1 u 1 is the main product, the fragments further having the Amino acid sequence of V a 1 4 4 9 from G 1 u 1, plasminogen one Gen This includes the N-terminal side of
- plasminogen N-terminal peptide is rich in hydrophilic amino acids, the difference in the presence or absence of N-terminal base peptide between the BL Angi O statin and Anjiosutachi emissions, solubility It is thought that there will be differences not only in physical properties such as, but also in activity and kinetics in vivo.
- BL angiostatin actually has an anticancer effect in vivo, and it is inexpensive, highly soluble in water, and has excellent anticancer properties Development of anticancer agents and methods for producing them is awaited.
- BL angiostatin has an extremely excellent anticancer effect and Because of its high solubility, it was discovered that a particular anticancer effect was exhibited even by intravenous administration, and the present invention was completed.
- the present invention relates to an anticancer agent containing BL angiostatin as an active ingredient and a method for producing the same.
- FIG. 1 shows BL angiostatin purified by an affinity trap reactor.
- Lane 1 is the resulting BL angiostatin.
- Lane 2 contains 100 mM NaCl, vasirolysin MA (5 nM) and plasminogen (3 M). 0. 0 1% Tw een 8 0 , 1 mM C a C 1 2 5 0 mM Tris-HCl containing (pH 7. 4) After incubation 1 0 1 in 37, 60 minutes, ⁇ ⁇ ⁇ of sample buffer A solution (125 mM Tris-HCl buffer, pH 6.8 containing 4% SDS, 10% 2-mercaptoethanol, 20% sucrose, and 0.004% bromophenol blue) was added, and analysis was performed in the same manner as in Lane 1.
- sample buffer A solution 125 mM Tris-HCl buffer, pH 6.8 containing 4% SDS, 10% 2-mercaptoethanol, 20% sucrose, and 0.004% bromophenol blue
- FIG. 2 shows the effect of BL angiostatin on the growth of subcutaneously transplanted Lewis lung cancer (change in tumor volume).
- X-axis is the number of days (days) from when transplanted Lewis lung carcinoma
- Y axis is tumor volume (Yuzuru 3).
- ⁇ represents the control group
- ⁇ , ⁇ , X and ⁇ represent the BL angiostatin 0.3, 1, 3, and 1 Omg / kg / day treatment groups, respectively.
- the average value of the example represents the standard deviation.
- a statistically significant difference compared with the control group is marked with *.
- FIG. 3 shows the effect of BL angiostatin on the growth of subcutaneously transplanted Lewis lung cancer (tumor wet weight).
- the X-axis is the dose of BL angiostatin (mg / kg / day) and the Y-axis is the tumor wet weight (mg).
- Each value represents the mean soil standard error of 8 cases in each group. In the Dunnett's multiple comparison test, statistically significant differences compared to the control group are marked with *. * (p ⁇ 0.05), * * (p ⁇ 0.01)
- FIG. 4 shows the effect of BL angiostatin on body weight of Lewis lung cancer subcutaneously transplanted animals.
- the X axis is the number of days since transplantation of Lewis lung cancer (days), and the Y axis is body weight (g).
- Ginseng is a control (saline) treatment group, ⁇ is angiostatin
- FIG. 5 shows a histopathological image of the tumor and a Von Wi11 ebrand factor immunostaining image.
- Control group tumor major axis 8.2 mm, minor axis 7.8 mm
- A, B, C, D 3 mg / kg administration group
- tumor major axis 7.0, minor axis 5.6 s administration group
- E, F , G, H 10 mgZkg dose groups
- D, H, and L are immunostained for Von Willebrand factor, and the others are hematoxylin and eosin.
- Magnification 5 times (A, E, 1), 25 -fold (B, F, J), 1 00 -fold (C, G, K), and 50-fold (D, H, L) 0 invention for carrying out the Best form of
- the BL angiostatin of the present invention can be obtained by treating plasminogen with a protease, in particular, bacillolysin MA (BLMA), which is an enzyme produced by Bacillus megaterium A 9542.
- BLMA bacillolysin MA
- Bacillus megaterium A 9542 was deposited at the Patent Organism Depositary of the National Institute of Advanced Industrial Science and Technology, Institute of Biotechnology, on March 21, 2001 under the accession number FERM P-18268.
- the amino acid sequence of BLMA is disclosed in JP-A-2002-272453.
- BLMA is human plasminogen S er 441 - V a 1 442 , L eu 74 - cut between P he 75 and V a 1 449 -L eu 450, G li ⁇ - S er 441 G 1 u 1 - Va l 449, P he 75 - S er 441, P he 75 - it is known to produce a fragment with a four ⁇ Njiosutachin like activity of V a 1 449.
- angiostatin-like fragments can be used alone or in combination.
- Plasminogen is known to be highly conserved in its amino acid sequence in mammals, particularly in humans. Accordingly, as the plasminogen as a raw material for producing the BL angiostatin of the present invention, any plasminogen can be used as long as it is derived from mammals. Is preferred.
- BL angiostatin of the present invention can also be produced by chemical synthesis or gene recombination technology. Production by genetic recombination technology is preferred because relatively easy operation and mass production are possible.
- BL Angios of the present invention In order to produce tatatin, a target protein can be produced by preparing a DNA having a nucleotide sequence encoding the protein and introducing it into a suitable expression system.
- the DNA having the base sequence of BL angiostatin of the present invention is obtained by using a human-derived cDNA library (for example, derived from HepG2 cells) as a base for the base sequence information encoding the amino acid sequence of BL angiostatin. It can be obtained by screening using a suitable primer or probe designed based on the above. Screening can be performed by plaque hybridization or the like. Alternatively, a human-derived cDNA library (for example, derived from HepG2 cells) is used as type III, and a suitable primer designed based on the above nucleotide sequence information is used.
- a human-derived cDNA library for example, derived from HepG2 cells
- a suitable primer designed based on the above nucleotide sequence information is used.
- the target gene can also be cloned directly.
- Expression systems for expressing the recombinant protein are known to those skilled in the art.
- To express DNA in a host cell first, the DNA is inserted downstream of a promoter in an expression vector, and then the recombinant expression vector is introduced into a host cell suitable for the expression vector.
- Examples of expression vectors for bacteria include pGEMEX_l (Promega), pQE-9 (QIAGEN), pQE-30 (QIAGEN), pRSET (Invitrogen), pLEX (Invitrogen), and animal cells.
- expression vectors include pc DNA I, pc DM8 (Funakoshi), pc DNA I / AmP (Invitrogen), pREP4 (Invitrogen), and expression vectors for producing recombinant viruses, such as pM
- promoters that can be used in bacterial expression vectors include promoters derived from Escherichia coli and phage, such as the trp promoter, T7 promoter, and 1ac promoter.
- Examples of a promoter that can be used in an expression vector for yeast include a PH05 promoter and a GAP promoter.
- Examples of promoters that can be used in expression vectors for animal cells include, for example, the promoter of the IE (immediate early) gene of cytomegalovirus (human CMV), the early promoter of SV40, the promoter of retroinoles, and the promoter of adenowinores.
- Examples include the tarothionine promoter and the heat shock promoter.
- the host cell is not particularly limited as long as it can express the target protein, and includes bacteria, yeast, animal cells, insect cells, and the like.
- Methods for introducing a recombinant vector into a host include, for example, the calcium phosphate method, the protoplast method, the electroporation method, the spheroblast method, the lithium acetate method, and the lipofection method, etc., depending on the type of host cell. It can be selected as appropriate.
- a conventional protein isolation and purification method can be used. For example, if the recombinant protein is expressed in a lysed state in the cells, after the culture is completed, the cells are collected by centrifugation, suspended in an aqueous buffer, crushed, and the cell-free extract is collected. obtain.
- an ordinary protein isolation and purification method that is, a solvent extraction method, a salting-out method using ammonium sulfate, a desalting method, a precipitation method using an organic solvent
- Purification can be carried out using an ion exchange chromatography method, a gel filtration method, and an affinity mouth matdaraphy method alone or in combination.
- the present invention provides the following steps:
- the present invention also relates to a method for producing BL angiostatin, comprising a step of elution with 6-aminohexanoic acid, wherein the buffer does not contain isopropyl alcohol. Preferably, all operations are performed at 4 ° C. Thereby, BL angiostatin can be obtained in high yield.
- the anticancer agent of the present invention is generally provided in the form of a pharmaceutical composition containing BL angiostatin as an active ingredient and pharmaceutical additives such as a carrier and an excipient.
- the anticancer agent of the present invention can be administered as a medicine to mammals including humans.
- the route of administration of the drug of the present invention is not particularly limited, and is orally or parenterally administered (eg, For example, intramuscular administration, intravenous administration, subcutaneous administration, intraperitoneal administration, mucosal administration to the nasal cavity or the like, or inhalation administration may be used.
- BL angiostatin unlike prior art angiostatin, has a high solubility in water (20 mg / ml or more), so that the anticancer agent of the present invention can be administered by intravenous administration. It has the feature of.
- the form of the anticancer agent of the present invention is not particularly limited, and examples of preparations for oral administration include tablets, capsules, fine granules, powders, granules, liquids, and syrups.
- preparations for parenteral administration include injections, drops, suppositories, inhalants, transmucosal absorbents, transdermal absorbents, nasal drops, ear drops and the like.
- Those skilled in the art can appropriately select the form of the drug of the present invention, the additive for the preparation to be used, the method for producing the preparation, and the like.
- the dose of the drug of the present invention can be appropriately selected in consideration of the gender, age or weight of the patient, the severity of symptoms, the administration purpose such as prevention or treatment, or the presence or absence of other complications. .
- the dose is generally, 0. lmg / kg body weight / day ⁇ 1 0 Om g / kg body weight / day, Ru preferably 1 mg / kg body weight / day ⁇ 1 Omg / kg body weight / day der.
- the anticancer agent of the present invention can be used for the treatment of breast cancer, lung cancer, pharyngeal cancer, stomach cancer, rotoma, liver cancer, colon cancer, uterine cancer, ovarian cancer, and the like.
- Basirolysin MA was isolated from Bacillus megaterium A9542 strain (deposited at the Patent Depositary of Biotechnology, Industrial Technology Research Institute, National Institute of Advanced Industrial Science and Technology under the deposit number F ERM P-18268) by the following method. Purified.
- Example 2 Preparation of an affinity trap activator of the present invention in which basilolysin MA and lysine are fixed (reactor 10 ml is prepared)
- An immobilization reaction was performed by preparing a 84 mg / ml bacillolysin MA solution with the same buffer A, adding 17.6 ml of the solution to the column, and stirring at room temperature for 2 hours. After completion of the reaction, remove the solution by suction, add 20 ml of 5% isopropyl alcohol aqueous solution (pH 8.0) containing 0.2 M L-lysine hydrochloride, and stir at room temperature for 2 hours to perform the lysine immobilization reaction.
- pH 8.0 isopropyl alcohol aqueous solution
- buffer B (2 OraM MES (2- [morpholino] ethanesulfonic acid) -NaOH) containing 5% isopropyl alcohol. And stored at 4 ° C. in buffer B of the above composition containing 0.02% sodium azide.
- Example 3 One-step purification process of BL angiostatin from human plasma using the bacillolysin MA lysine reactor
- the used reactor was washed at 4 ° C. with a buffer C 5 Oml having the above composition containing 1 M NaCl and 200 mM 6-aminohexanoic acid. Then, it was stored at 4 ° C. in buffer C containing 0.02% sodium azide. The reactor washed and stored in this state could be used repeatedly for more than two months.
- Example 4 One-step purification process of BL angiostatin from human plasma using the bacillolysin MA / lysine reactor (isopropyl alcohol-free method)
- the used reactor was washed with 2.5 ml of buffer C containing 1 M NaCl and 200 mM 6-aminohexanoic acid. Then, it was stored at 4 ° C in buffer C containing 0.02% sodium azide. In this state, the saved reactor is It could be used repeatedly for more than two months.
- the resulting BL angiopathy O to 4 g and 1 Omicron mu 1 was dissolved in purified water of statins, this ⁇ ⁇ ⁇ sample buffer (4% SDS, 1 0% 2- mercaptoethanol, 20% sucrose , 125 mM Tris-HCl buffer, pH 6.8) containing 0.004% bromophenol blue, and 10 ⁇ l of this was fractionated by electrophoresis using a 7.5% polyacrylamide gel. Stained with Coomassie Brilliant Blue R250 ( Figure 1).
- Lewis lung carcinoma cells were subcutaneously implanted into C57B LZ6 mice and maintained for passage.
- the tumor was excised and the cancer cells were suspended in phosphate buffered saline.
- the ventral skin of BDF i mice were preliminarily reared then under 1 week SPF environment, a cell suspension of 0. 05 ml containing 1 X 1 0 5 cells per mouse It was transplanted. All subsequent experiments were performed in the SPF environment.
- the BL angiostatin prepared by the method of Example 3 was lyophilized and dialyzed against purified water three times for 2 hours.
- the obtained solution was freeze-dried and dissolved in physiological saline to a concentration of 20 mg / ml.
- This solution was sterilized by filtration using a filter having a pore size of 0.22 / m.
- This solution was further diluted with sterile saline to make solutions of 0.06 mg / ml, 0.2 mg / ml, 0.6 mg / ml and 2 mg / ml.
- BL angiostatin solution was intravenously administered at a rate of 5 ml / kg once daily for 14 days. This resulted in respective doses of 0.3, 1, 3, and 10 mg / kg.
- the control group received intravenous saline at the same volume. The number of animals in each group was 10.
- the size of the tumor (the tumor volume was calculated from the minor axis and major axis as follows: square of minor axis X major axis X 0.52) and body weight were measured daily.
- the day after the end of the administration the mice were sacrificed under ether anesthesia, and the tumors were excised and their wet weight was measured. Data for a total of 8 animals were statistically processed, excluding the maximum and minimum values for each group. Significant difference The Dunnett's multiple comparison test was used for the test, and a significance level of 0.05% or less was regarded as a significant difference.
- control group 3 mg / kg administration group and 1 Omg / kg administration group, three representative examples each of tumors obtained from three individuals, measurement of major axis and minor axis, cell proliferation, hemorrhage, necrosis, histology of dividing cells Observation and immunostaining of vascular endothelial cells using anti-VonWi11 ebrand factor antibody.
- the tumor volume was significantly reduced in the 3 and 10 mg / kg BL angiostatin-administered groups on day 10 after Lewis lung cancer transplantation (FIG. 2).
- the tumor volume was less than or equal to 12 in the control group at all observation periods.
- Tumor growth was also suppressed in the 0.3 and lmg / kg groups, but a statistically significant difference was observed in the 0.3 mg / kg group on days 14 and 15 in the lmg / kg group. It was day 14 in the kg group (Fig. 2). On the 15th day after transplantation, the tumor was removed and its wet weight was measured. ).
- Table 1 summarizes the results of histopathological examination of the removed tumor.
- cell proliferation was high (Fig. 5A)
- bleeding was mild (Fig. 5B)
- necrosis was slight (Fig. 5B)
- moderately split cells were observed (Fig. 5C).
- immunostaining of vascular endothelial cells with an anti-vw WnIll ebrand factor antibody revealed a mild to moderate positive cell ratio (FIG. 5D).
- Moderate cell proliferation Figure 5E
- mild bleeding Figure 5F
- mild necrosis Figure 5F
- moderately split cells in the BL angiostatin 3 mg / kg group Figure 5G
- the percentage of von Willebrand factor-positive cells was found to be mild to moderate (Figure 5H).
- the anticancer agent of the present invention can be used for treating breast cancer, lung cancer, pharyngeal cancer, stomach cancer, knee cancer, liver cancer, colon cancer, uterine cancer, ovarian cancer, prostate cancer, etc. .
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Abstract
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Priority Applications (5)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
PCT/JP2004/002124 WO2005079835A1 (ja) | 2004-02-24 | 2004-02-24 | Blアンジオスタチンを含む制がん剤 |
CA002557154A CA2557154A1 (en) | 2004-02-24 | 2004-02-24 | Anticancer agent containing bl-angiostatin |
US10/590,342 US20110092442A1 (en) | 2004-02-24 | 2004-02-24 | Anticancer agent containing bl-angiostatin |
JP2006510141A JPWO2005079835A1 (ja) | 2004-02-24 | 2004-02-24 | Blアンジオスタチンを含む制がん剤 |
AU2004316095A AU2004316095A1 (en) | 2004-02-24 | 2004-02-24 | Anticancer agent containing BL angiostatin |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
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PCT/JP2004/002124 WO2005079835A1 (ja) | 2004-02-24 | 2004-02-24 | Blアンジオスタチンを含む制がん剤 |
Publications (1)
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WO2005079835A1 true WO2005079835A1 (ja) | 2005-09-01 |
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PCT/JP2004/002124 WO2005079835A1 (ja) | 2004-02-24 | 2004-02-24 | Blアンジオスタチンを含む制がん剤 |
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Country | Link |
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US (1) | US20110092442A1 (ja) |
JP (1) | JPWO2005079835A1 (ja) |
AU (1) | AU2004316095A1 (ja) |
CA (1) | CA2557154A1 (ja) |
WO (1) | WO2005079835A1 (ja) |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2010070746A1 (ja) * | 2008-12-17 | 2010-06-24 | 株式会社ティムス | イヌアンジオスタチン様ポリペプチド |
JP2012050386A (ja) * | 2010-09-01 | 2012-03-15 | Kaneka Corp | 2種類以上の多孔質充填剤を充填するカラム |
JP2012062259A (ja) * | 2010-09-14 | 2012-03-29 | Kaneka Corp | クリングル配列を有する蛋白質またはペプチドを精製または除去するアフィニティ担体とその精製方法、除去方法。 |
Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO1996035774A2 (en) * | 1995-04-26 | 1996-11-14 | The Children's Medical Center Corporation | Angiostatin fragments and aggregate angiostatin and methods of use |
JPH09286798A (ja) * | 1996-04-19 | 1997-11-04 | Chemo Sero Therapeut Res Inst | 活性蛋白質組成物の製造方法 |
WO1998054217A1 (en) * | 1997-05-30 | 1998-12-03 | The Children's Medical Center Corporation | Angiostatin fragments and method of use |
JP2002272453A (ja) * | 2001-03-23 | 2002-09-24 | Japan Science & Technology Corp | 新規なアンジオスタチン変換酵素、該酵素の生産菌、該酵素の製造方法、該酵素の利用によるアンジオスタチンおよび活性型セリンプロテアーゼの製造 |
-
2004
- 2004-02-24 AU AU2004316095A patent/AU2004316095A1/en not_active Abandoned
- 2004-02-24 WO PCT/JP2004/002124 patent/WO2005079835A1/ja active Application Filing
- 2004-02-24 CA CA002557154A patent/CA2557154A1/en not_active Abandoned
- 2004-02-24 US US10/590,342 patent/US20110092442A1/en not_active Abandoned
- 2004-02-24 JP JP2006510141A patent/JPWO2005079835A1/ja active Pending
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO1996035774A2 (en) * | 1995-04-26 | 1996-11-14 | The Children's Medical Center Corporation | Angiostatin fragments and aggregate angiostatin and methods of use |
JPH09286798A (ja) * | 1996-04-19 | 1997-11-04 | Chemo Sero Therapeut Res Inst | 活性蛋白質組成物の製造方法 |
WO1998054217A1 (en) * | 1997-05-30 | 1998-12-03 | The Children's Medical Center Corporation | Angiostatin fragments and method of use |
JP2002272453A (ja) * | 2001-03-23 | 2002-09-24 | Japan Science & Technology Corp | 新規なアンジオスタチン変換酵素、該酵素の生産菌、該酵素の製造方法、該酵素の利用によるアンジオスタチンおよび活性型セリンプロテアーゼの製造 |
Non-Patent Citations (1)
Title |
---|
SHIMIZU K. ET AL, JAPAN SOCIETY FOR BIOSCIENCE, BIOTECHNOLOGY, AND AGROCHEMISTRY., vol. 2003, 2003, pages 258, XP002995815 * |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2010070746A1 (ja) * | 2008-12-17 | 2010-06-24 | 株式会社ティムス | イヌアンジオスタチン様ポリペプチド |
JP2012050386A (ja) * | 2010-09-01 | 2012-03-15 | Kaneka Corp | 2種類以上の多孔質充填剤を充填するカラム |
JP2012062259A (ja) * | 2010-09-14 | 2012-03-29 | Kaneka Corp | クリングル配列を有する蛋白質またはペプチドを精製または除去するアフィニティ担体とその精製方法、除去方法。 |
Also Published As
Publication number | Publication date |
---|---|
CA2557154A1 (en) | 2005-09-01 |
JPWO2005079835A1 (ja) | 2007-08-02 |
US20110092442A1 (en) | 2011-04-21 |
AU2004316095A1 (en) | 2005-09-01 |
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