WO2008023796A1 - Promoteur de la production de protéoglycane - Google Patents

Promoteur de la production de protéoglycane Download PDF

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Publication number
WO2008023796A1
WO2008023796A1 PCT/JP2007/066459 JP2007066459W WO2008023796A1 WO 2008023796 A1 WO2008023796 A1 WO 2008023796A1 JP 2007066459 W JP2007066459 W JP 2007066459W WO 2008023796 A1 WO2008023796 A1 WO 2008023796A1
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Prior art keywords
solvate
pharmaceutically acceptable
acceptable salt
compound according
added
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PCT/JP2007/066459
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English (en)
Japanese (ja)
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Shin-Ichiro Nishimura
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National University Corporation Hokkaido University
Shionogi & Co., Ltd.
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Priority claimed from JP2006227818A external-priority patent/JP2009274954A/ja
Priority claimed from JP2006227822A external-priority patent/JP2009274955A/ja
Application filed by National University Corporation Hokkaido University, Shionogi & Co., Ltd. filed Critical National University Corporation Hokkaido University
Publication of WO2008023796A1 publication Critical patent/WO2008023796A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H3/00—Compounds containing only hydrogen atoms and saccharide radicals having only carbon, hydrogen, and oxygen atoms
    • C07H3/06—Oligosaccharides, i.e. having three to five saccharide radicals attached to each other by glycosidic linkages
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/06—Dipeptides
    • C07K5/06008—Dipeptides with the first amino acid being neutral
    • C07K5/06017—Dipeptides with the first amino acid being neutral and aliphatic
    • C07K5/06026—Dipeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atom, i.e. Gly or Ala
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K9/00—Peptides having up to 20 amino acids, containing saccharide radicals and having a fully defined sequence; Derivatives thereof
    • C07K9/001—Peptides having up to 20 amino acids, containing saccharide radicals and having a fully defined sequence; Derivatives thereof the peptide sequence having less than 12 amino acids and not being part of a ring structure
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention relates to a fluorine-containing sugar chain and a novel derivative of a sugar peptide useful for selective biosynthesis of proteodarican. Furthermore, this invention relates to the pharmaceutical which uses this derivative as an active ingredient.
  • Proteodarican is a molecule that has a structure in which one or more glycosaminodarican (GAG) chains are bound to a protein called a core protein. Depending on the sugars that make up the GAG chain, It is broadly classified into chondroitin sulfate type proteodarican (CSPG) and heparan sulfate type proteodarican (HSPG).
  • GAG glycosaminodarican
  • CSPG chondroitin sulfate type proteodarican
  • HSPG heparan sulfate type proteodarican
  • CSPG CSPG vascular endothelial growth factor
  • FGF fibroblast growth factor
  • HGF vascular endothelial growth factor
  • Cartilage is a connective tissue composed of chondrocytes and a matrix surrounding the chondrocytes, and is present in joints, spinal discs, costal cartilage, pinna, external auditory canal, pubic connection, pharyngeal palate and the like.
  • Cartilage is composed of chondrocytes and cartilage matrix produced by chondrocytes, and the main components of cartilage matrix are proteodalycan and collagen (type II, type IX, etc.).
  • the joint disease is a disease whose main lesion is degeneration of articular cartilage, and the release of proteodarican from these cartilage tissues is promoted by various causes. Synthetic capacity begins to decline. At the same time, the release and activation of various types of metaloproteinases such as type I collagenase are enhanced and the collagen in the cartilage tissue is degraded. These series of reactions lead to the destruction of the cartilage tissue, and then the progression of the lesion causes synovial growth, subchondral bone destruction, cartilage hypertrophy or osteogenesis at the joint margin, and after joint deformation, In severe cases it leads to dysfunction.
  • the disease is osteoarthritis.
  • aging is considered to be one of the causes of cartilage degeneration, and there is concern that the number of patients will increase further in the future as society ages.
  • analgesic anti-inflammatory agents and hyaluronic acid preparations are used for the purpose of removing the pain associated with cartilage degeneration-subchondral bone destruction, both of which are used symptomatically However, it is not effective enough.
  • proteodarican production promoters are new! / And are attracting attention as therapeutic agents! /.
  • Nishimura et al. Proposed a glycopeptide that can be used as an initiator of GAG chain elongation that mimics the structure of proteodarican, suggesting its usefulness as a therapeutic agent for rheumatoid arthritis and osteoarthritis. (See Patent Document 1 below).
  • an initiator made of this polysaccharide peptide has not yet been confirmed to be selective for HSPG / CSPG biosynthesis.
  • Patent Literature l WO2004 / 076476
  • Non-Patent Document 1 JD Esko and Shi Zhang, Curr. Opin. Struct. Biol., 6, 663-670 (199 6)
  • Non-Patent Document 2 K. Sugahara, Y. Ohkita, Y. Shibata,. Yoshida, and K. Ikegami, J. Biol. Chem., 271, 7204-7212 (1995)
  • Non-Patent Document 3 H. itagawa, M. Oyama,. Masayama, Y. Yamaguchi, and K. Suga hara, Glycobiology., 7, 1175-1180 (1997)
  • Non-Patent Document 4 S. Yamada, M. Oyama, H. inugasa. T. Nakagawa, T. Kawasaki, S. Nagasawa, HR Morris, A. Dell, and K. Sugahara, Glycobiology., 5, 335-341 (199 5)
  • Non-Patent Document 5 S. Yamada, M. Oyama, Y. Yuki,. Ato, and K. Sugahara, Eur. J. Biochem., 233, 687-693 (1995)
  • Non-Patent Document 6 K. Sugahara, R. Tohnooka, S. Yamada, .H. Hoo, H. R. Morris, and A. Dell, Glycobiology., 4, 535-544 (1994)
  • An object of the present invention is to provide a compound having a function of promoting the production of proteodarican, and in particular, selectively promoting the biosynthesis of HSPG or C SPG.
  • the GAG chain of proteodalycan has a characteristic structure with repeating units of disaccharides of uronic acid and amino sugar, and a serine residue in the core protein via a common tetrasaccharide GlcA_Ga ⁇ Gato Xyl. Is bound to.
  • CSPG and HSPG are classified into two types according to the difference in the sugars constituting the GAG chain! /, As described above, force S, which is the same as the biosynthetic branch point of both types of PG. The mechanism of hexosamine transfer to sugar is still unknown.
  • the present invention provides the following.
  • R represents a hydrogen atom, an optionally substituted alkyl, an optionally substituted aryl, an optionally substituted heteroaryl, or a peptide residue comprising a natural or non-natural amino acid
  • X and Y represent a halogen atom, a hydroxyl group or an alkoxy group, provided that one of X and Y is a hydroxyl group, and X and Y are not a hydroxyl group at the same time.
  • R represents a peptide residue composed of a natural or non-natural amino acid, a pharmaceutically acceptable salt thereof, or a solvate thereof.
  • OR is a serine and / or threonine-containing oligopeptide, wherein an oxygen atom of OR is bound to serine and / or threonine, and a disaccharide unit is further bound to the amino acid.
  • FITC represents that the amino group is fluorescently labeled with fluorescein isothiocyanate.
  • a medicament comprising as an active ingredient the compound described in any one of (1) to (8) above, a pharmaceutically acceptable salt thereof, or a solvate thereof.
  • a heparan sulfate proteodarican production promoter comprising the compound described in (2) or (3) above, a pharmaceutically acceptable salt thereof, or a solvate thereof as an active ingredient.
  • a chondroitin sulfate proteodarican production promoter comprising the compound described in (4) or (5) above, a pharmaceutically acceptable salt thereof, or a solvate thereof as an active ingredient.
  • a wound healing promoter comprising the compound described in (2) or (3) above, a pharmaceutically acceptable salt thereof, or a solvate thereof as an active ingredient.
  • a prophylactic and / or therapeutic agent for osteoarthritis comprising the compound described in (4) or (5) above, a pharmaceutically acceptable salt thereof, or a solvate thereof as an active ingredient. .
  • the compound of the present invention has a function of promoting the production of proteodalycan, and in particular, can selectively promote the biosynthesis of HSPG or C SPG.
  • FIG. 1 shows the result of CHO-K1 cell culture after adding lGal-6F substitution product (57) and subjecting the supernatant to genomic filtration chromatography (A). The chromatographic result after digesting the eluted fraction with chondroitinase ABC is shown (B).
  • FIG. 2 shows the result of CHO-K1 cell culture after adding lGal-4F substitution product (58) and subjecting the supernatant to genomic filtration chromatography (A).
  • the chromatographic result after digesting the eluted fraction with chondroitinase ABC is shown (B).
  • FIG. 3 shows the results of CHO-K1 cell culture by adding the compound of the present invention (2Gal-6F substitution product (59)) and subjecting the supernatant to gel filtration chromatography (A). The chromatographic result after digesting the eluted fraction with chondroitinase ABC is shown (B).
  • FIG. 4 shows the results of CHO-K1 cell culture by adding the compound of the present invention (2Gal-4F substitution product (60)) and subjecting the supernatant to gel filtration chromatography (A). The chromatographic result after digesting the eluted fraction with chondroitinase ABC is shown (B).
  • FIG. 5 shows the results of CHO-K1 cell culture with the addition of a fluorine-free compound (61) and the supernatant subjected to gel filtration chromatography (A).
  • the eluted fraction is chondroitinase
  • FIG. 6 is a graph showing the amount of mucopolysaccharide produced when human normal chondrocytes are cultured with the compound of the present invention added.
  • FIG. 7 is a graph showing aggrecan produced by culturing human normal chondrocytes with the compound of the present invention added.
  • FIG. 8 shows the results of mass spectrometry spectrum of the compound of the present invention.
  • Gal-4F substitution (58), 2Gal-6F substitution (59), 2Gal-4F substitution (60), and fluorine-free substitution (61).
  • the PG production promoter of the present invention has the following formula (I) [Chemical 4]
  • R represents a hydrogen atom, an optionally substituted alkyl, an optionally substituted aryl, an optionally substituted heteroaryl, or a peptide residue comprising a natural or non-natural amino acid
  • Y and X represent a halogen atom, a hydrogen atom, a hydroxyl group or an alkoxy group, provided that one of X and Y is a hydroxyl group, and X and Y are not simultaneously a hydroxyl group. It is a pharmaceutically acceptable salt or a solvent thereof.
  • the PG production promoter of the present invention has the following formula (II)
  • R and Y are as defined above
  • a sugar chain derivative a pharmaceutically acceptable salt thereof, or a solvate thereof, and in particular, selecting biosynthesis of HSPG Promote.
  • the PG production promoter of the present invention has the following formula (III)
  • alkyl includes linear or branched alkyl having a carbon number;! To 10, preferably 1 to 6, more preferably 1 to 3, and methyl.
  • Ethyl, n-propinole, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentinole, isopentinole, neopentinole, hexinole, isohexinole, n-heptinole, isoheptyl, n-octyl Examples include isootatil, n-nonyl and n-decyl.
  • Examples of the substituent of "substituted! /, May! /, Alkyl” include halogen, hydroxy, alkoxy, carboxy, alkoxycarbonyl, and acyl.
  • Halogen includes F, Cl, Br, and I.
  • alkyl part of "alkoxy” and “alkoxycarbonyl” is the same as the above “alkyl", both of which are straight-chain or branched having a carbon number;! To 6 and more preferably having 1 to 3 carbon atoms Examples include methyl, ethyl, n-propyl, isopropyl, n-butinole, isobutinole, sec-butinole, tert-butinole, n-pentinole, isopentinole, neopentyl, hexyl, isohexyl and the like. The Particularly preferred is methyl or ethynole.
  • Asil includes aliphatic asil and Caroyl having 1 to 7 carbon atoms. Specific examples include honoreminole, acetyl, propionyl, butyryl, isobutyryl, valeryl, bivaloyl, hexanoyl, attalyloyl, propioroyl, methacryloyl, crotonol and benzoyl.
  • Heteroaryl includes an aromatic heterocyclic group having one or more heteroatoms in the ring in which ⁇ , S and N forces are also arbitrarily selected. Specifically, pyrrolyl, imidazolyl, pyrazol Ginole, Pi !; Ginole, Pi !; Dajini, Pi !; ; Azo! ; Nore, ⁇ !
  • Aryl refers to a monocyclic, bicyclic, or tricyclic aromatic carbocyclic ring having 6 to 16 carbon atoms, and includes, for example, phenyl, naphthyl, anthryl, phenanthryl, etc. Ninole is preferred.
  • Natural amino acids mean 20 kinds of L-a-amino acids universally used by organisms, and “unnatural amino acids” are natural isomers (D-forms) as well as natural isomers. This includes L-form or D-form amino acid derivatives that do not exist in amino acids and have side chains. The optical isomers (D-form) of natural amino acids are preferred!
  • Peptide residues include oligopeptides connected by 10 peptide bonds from these natural and / or non-natural amino acid strengths, and include one natural or non-natural amino acid. 1 to 6 peptides are preferred and 1 to 3 peptides are preferred. In addition, this peptide residue may be appropriately modified with a protecting group for peptide synthesis and / or a label generally used by those skilled in the art.
  • an acid addition salt for example, hydrochloride, sulfate, nitrate, Inorganic acid salts such as phosphate, carbonate, bicarbonate, perchlorate; for example, oxalate, acetate, propionate, lactate, maleate, fumarate, tartrate, malic acid
  • Organic salts such as salts, citrates and ascorbates; for example, sulfonates such as methanesulfonate, isethionate, benzenesulfonate, p-toluenesulfonate; for example aspartate, glutamate And the like, and the like.
  • alkali metal salts such as lithium, sodium and potassium
  • alkaline earth metal salts such as calcium and magnesium
  • basic amino acid salts such as arginine and lysine.
  • the compound of the present invention may be a solvate such as water, acetonitrile, ethyl acetate, methanol, ethanol and the like.
  • the solvation number of the solvate of the compound of the present invention can usually vary depending on the synthesis method, purification method, crystallization conditions, etc., but is, for example, in the range of 1 to 5 molecules per molecule.
  • the compound of the present invention can be produced as follows.
  • an unsubstituted trisaccharide unit (41) is synthesized according to the following scheme.
  • the silylene group was deprotected with HF and then benzylated to give (24).
  • the MP (p-methoxyphenyl) group of the disaccharide unit (24) is converted to an imidate form as described below, and this is glycosylated with a known compound, xylosylacceptor (36), to produce a trisaccharide. Unit (41).
  • the selection ratio of anomeric positions is about ⁇ : (1: 5) (NMR ratio), but only / ⁇ can be separated and purified by flash column chromatography.
  • the synthesis scheme of the 4-position fluorinated trisaccharide unit (40) is as follows.
  • trisaccharide unit 041 According to the synthesis of the trisaccharide unit 041) can be synthesized fluorinated trisaccharide unit (40) c
  • the trisaccharide unit l) according to the synthesis of, c can be synthesized fluorinated trisaccharide unit (39)
  • D Galactose (1) was arylated at the anomeric position by the Fisher method to give (2).
  • a benzylidene acetal was applied to the 4th and 6th positions, and the 2nd position was benzoylated as it was, and then the benzylidene acetal was deprotected without purification to give a known compound (3).
  • This was selectively fluorinated at the 6th position with DAST, and after simple purification, the 4th position was benzoylated as (4). After deprotecting the aryl group at the anomeric position, it was imidated to give a 6F fragment (5).
  • the compound of the present invention in which R is P-methoxyphenyl is obtained.
  • R is P-methoxyphenyl
  • it can be derived into various compounds by converting it to an imidate form according to a conventional method, coupling with HOR, and deprotecting the benzoyl group.
  • R is preferably an oligopeptide having 2 to 4 natural amino acids, and particularly preferably R-coupled with the side chain OH group of serine or threonine.
  • a labeled derivative is preferably used as long as the HSPG production promoting action is not impaired.
  • the compound of the present invention can be used, for example, for promoting wound healing.
  • it can be formulated for oral or parenteral administration as a wound healing agent.
  • the compound of the present invention can be used for the treatment or prevention of cartilage diseases such as rheumatoid arthritis and osteoarthritis.
  • cartilage diseases such as rheumatoid arthritis and osteoarthritis.
  • it can be formulated for oral or parenteral administration as an analgesic.
  • this drug is a normal formulation such as a solid preparation such as a tablet, powder, granule, capsule, etc .; a liquid preparation; an oily suspension; or a liquid preparation such as a syrup or elixir. It can also be used as a dosage form. When administered parenterally, it can be used as an aqueous or oily suspension or nasal solution.
  • aqueous or oily suspension or nasal solution In the preparation, conventional excipients, binders, lubricants, aqueous solvents, oil solvents, emulsifiers, suspending agents, preservatives, stabilizers and the like can be arbitrarily used.
  • formulations of the present invention are manufactured by combining (eg, mixing) a therapeutically effective amount of a compound with a pharmaceutically acceptable carrier or diluent, in which case the well-known and readily available ingredients Is manufactured by a known method.
  • the active ingredient is mixed with the carrier, or diluted with the carrier, or in the carrier in the form of a capsule, sash, paper, or other container. Be put in.
  • the carrier acts as a diluent
  • the carrier is a solid, semi-solid, or liquid material that acts as a medium, such as tablets, pills, powders, mouthpieces, elixirs, suspensions, emulsions, Can be made into solutions, syrups, aerosols (solids in liquid media), ointments, eg containing up to 10% active compound.
  • any suitable carrier known to those skilled in the art can be used for this formulation.
  • the carrier is a solid, liquid, or a mixture of solid and liquid.
  • intravenous injection Dissolve the active compound in 4% dextrose / 0.5% sodium quenate aqueous solution for spraying.
  • Solid formulations include powders, tablets and capsules.
  • a solid carrier is one or more substances that can also serve as a flavoring agent, lubricant, solubilizer, suspending agent, binder, tablet disintegrant, or capsule.
  • Tablets for oral administration consist of corn starch, alginic acid and other disintegrants, and / or binders such as gelatin, acacia, and lubricants such as magnesium stearate, stearic acid and talc, Contains suitable excipients such as ratatoses and calcium phosphate.
  • the carrier is a finely divided solid mixed with the finely divided active ingredient.
  • the active ingredients are mixed in the appropriate proportions with a carrier having the necessary binding properties and consolidated into the desired shape and size.
  • Powders and tablets contain from about 1 to about 99 weight percent of the active ingredient which is the novel compound of the present invention.
  • Suitable solid carriers are magnesium carbonate, magnesium stearate, talc, sugar, ratatose, pectin, dextrin, starch, gelatin, tragacanth gum, methylcellulose, sodium carboxymethylcellulose, a low melting wax, cocoa butter.
  • Liquid formulations include suspensions, emulsions, syrups, and elixirs.
  • the active ingredient can be dissolved or suspended in a pharmaceutically acceptable carrier such as sterile water, sterile organic solvent, or a mixture of both.
  • a pharmaceutically acceptable carrier such as sterile water, sterile organic solvent, or a mixture of both.
  • the active ingredient can often be dissolved in a suitable organic solvent, for example an aqueous propylene glycol solution.
  • Other compositions may be made by spraying the active ingredients with aqueous starch, sodium-powered l-poxymethylcellulose solution, or a suitable oil.
  • the dose of the compound in the present invention varies depending on the administration method, the patient's age, weight, condition and disease type. Usually, in the case of oral administration, about 0.1 mg to 7 OOOmg per day for an adult, Preferably, about 0.5 mg to 2000 mg may be administered in divided portions if necessary. In the case of parenteral administration, about 0.1 mg to 1000 mg, preferably about 0.5 mg to 500 mg, is administered per adult.
  • D-galactose (1) was converted into known compound (2), this was dissolved in DMF, benzaldehyde dimethyl acetal and CSA were added, and the mixture was stirred at 80 ° C for 3 hours. This was azeotroped with toluene to remove DMF, and then dissolved in CH C1. Pyridine, BzCl and DMAP were added and stirred overnight. Water was added to quench the reaction, and the organic layer was washed with saturated aqueous sodium hydrogen carbonate and brine and dried over MgSO. After filtration and concentration, the residue is dissolved in 80% acetic acid and stirred at 80 ° C for 3 hours.
  • the Ir catalyst was dissolved in THF and stirred for 2 hours under a hydrogen atmosphere. After substituting the inside of the flask with nitrogen, the compound (4) dissolved in THF was added and stirred at room temperature for 2 hours. After confirming the isomerization of the aryl group by TLC, saturated aqueous sodium hydrogen carbonate and I were added and stirred for 1 hour. The organic layer was washed with saturated aqueous sodium thiosulfate solution, saturated aqueous sodium hydrogen carbonate, and brine, and dried over MgSO. Filtration,
  • the commercially available (14) was derived from the known compound (15) according to the literature. (15) was dissolved in CH C1, t-Bu Si (OTf) and pyridine were added, and the mixture was stirred for 3 hours. The organic layer was washed with saturated aqueous sodium hydrogen carbonate and brine, and dried over MgSO. After filtration and concentration, the crude product was dissolved in CH C1, and pyridine, BzCl and DMAP were added, and the mixture was stirred at 60 ° C for 5 hours. After adding water to quench the reaction, the organic layer was washed with saturated aqueous sodium bicarbonate and brine and dried over MgSO. After filtration and concentration, flush
  • the Ir catalyst was dissolved in THF and stirred for 2 hours under a hydrogen atmosphere. After the atmosphere in the flask was replaced with nitrogen, (16) was added to THF, and the mixture was stirred at room temperature for 2 hours. After confirming the isomerization of the aryl group by TLC, TsOH′H 2 O was added and stirred for 1 hour. The organic layer was washed with saturated aqueous sodium hydrogen carbonate and brine, and dried over MgSO. Filtration, concentration, flash chromatography To give (17).
  • Glycosinore donor (5) obtained in Production Example 1 and glycosinore acceptor (17) obtained in Production Example 3 and MSAW300 were dissolved in CH CI / CH CN (1: 1) and nitrogen atmosphere The mixture was stirred at room temperature for 30 minutes. After cooling to 0 ° C., TMSOTf having a melting power in CH C1 was slowly added and stirred for 1 hour. Saturated aqueous sodium hydrogen carbonate was added to this solution to stop the reaction, followed by filtration through Celite, and the organic layer was washed with brine. After drying over MgSO, filtration and concentration, flash column chromatography
  • TMSOTf which had a solvent power, was slowly added to C1 and stirred for 1 hour. Saturated aqueous sodium hydrogen carbonate was added to the solution to stop the reaction, followed by Celite filtration, and the organic layer was washed with brine. Dry with MgSO
  • glycosyl donor (18), glycosyl acceptor (17) and MSAW300 which are known compounds, were dissolved in CH CI / CH CN (1: 1) and stirred at room temperature for 30 minutes in a nitrogen atmosphere. After cooling to 0 ° C., TMSOTf, which had been dissolved in CH C1, was slowly added and stirred for 1 hour. Saturated aqueous sodium bicarbonate solution was added to the solution to stop the reaction, followed by Celite filtration to remove the organic layer. Washed with brine. After drying over MgSO, filtration and concentration, flash column chromatography
  • Glycosyl donor (18), glycosyl acceptor (11), and MSAW300, which are known compounds, were dissolved in CHCI / CHCN (1: 1) and stirred at room temperature for 30 minutes in a nitrogen atmosphere. After cooling to 0 ° C., TMSOTf having a melting force in CH C1 was slowly added and stirred for 1 hour. Saturated aqueous sodium hydrogen carbonate was added to the solution to stop the reaction, followed by Celite filtration, and the organic layer was washed with brine. After drying over MgSO, filtration, concentration and purification by flash column chromatography
  • glycosyl donor (44), peptide acceptor (30), and MSAW300 were dissolved in CH C1 / CH CN (1: 1) and stirred at room temperature for 30 minutes under a nitrogen atmosphere. Cooled to 0 ° C
  • R 1 F
  • R 2 , R 3 , R 4 OBz: 65%
  • the glycosyl donor (31) was glycosylated with a known compound, xylosyl acceptor (36), to obtain a trisaccharide unit (37). Also, (3 2) reacted under the same conditions! /, (38) was obtained.
  • Atsey was performed with CHO-K1 cells.
  • CHO-K1 cells were seeded on a 24-well plate, and 10 L of the compounds synthesized in the examples (57, 58, 59, 60, 61) (adjusted to 30 mM) were added thereto. After culturing at 37 ° C. for 48 hours, the culture supernatant was filtered and the supernatant was gel filtered (S-labeled hadex G-10) for simple purification such as desalting. This was analyzed by gel filtration chromatography (TSK-GEL SuperSW3000). Whether or not the GAG chain was extended was determined by examining whether the initiator eluted from the excluded volume region of the gel filtration column.
  • the fraction eluted with the excluded volume region force was enzymatically digested with chondroitinase ABC, and the resulting product was analyzed by HPLC.
  • the conditions for enzymatic degradation are as follows.
  • HS / CS was estimated from the ratio of the peak not digested by chondroitinase ABC (HS) and the peak of the enzyme digestion product of CS.
  • the 2Gal-6F substitution product (59) was shown to selectively produce HSPG because the GAG chain was elongated and was not digested at all by chondroitinase ABC (Fig. 3).
  • all of the IGal-6F substitution (57), IGal-4F substitution (58), 2Gal-4F substitution (60) and fluorine-free substitution (61) showed CS chain extension (Fig. 1, 2, 4, 5). Based on these results This clearly suggests that sulfation of the 6th hydroxyl group (2Gal-6F) of the third sugar, Gal, is deeply involved in sorting control of CSPG / HSPG biosynthesis.
  • Test example 2 cartilage formation promotion test
  • NHAC-kn Normal human chondrocytes
  • CGM medium cell growth medium
  • Jung Co., Ltd .; 24 well Dilute each test substance to a final concentration of 1-100 ng / ml, add DMSO (base group) as a negative control, and BMP-2 (30, 100 ng / ml) as a positive control.
  • DMSO base group
  • BMP-2 30, 100 ng / ml
  • the cell mass was crushed by sonication and centrifuged to collect the lysate.
  • the amount of mucopolysaccharide and the amount of aggrecan contained in the cell mass lysate and the cell supernatant were quantified with an acidic mucopolysaccharide quantification kit (Hokudo, Primary Cell) and human adalican ELISA kit (Biosource).
  • the mucopolysaccharide production and the aggrecan production are shown in FIGS. 6 and 7, respectively.
  • both the fluorine-free (61) and 2Gal-4F-substituted (60) are 100 ng / Only in ml, mucopolysaccharide and aggrecan production was significantly enhanced. It was suggested that in addition to acting as a proteodalican initiator, it also activates the aggrecan production system.
  • the compound of the present invention is useful as a medicament, particularly a wound healing promoter, a regenerative medical material, osteoarthritis and other cartilage diseases.

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Abstract

La présente invention concerne un composé représenté par la formule (I) ci-dessous, qui accélère de manière sélective la biosynthèse d'un protéoglycane, en particulier un protéoglycane à sulfate de chondroïtine ou un protéoglycane à sulfate d'héparane. L'invention concerne également un produit pharmaceutique contenant un tel composé en tant qu'ingrédient actif. (I) (Dans la formule, R représente un atome d'hydrogène, un alkyle éventuellement substitué, un aryle éventuellement substitué, un hétéroaryle éventuellement substitué, ou un résidu peptidique composé d'un acide aminé naturel ou non naturel ; et X et Y représentent un atome d'halogène, un groupement hydroxy ou un groupement alcoxy, mais à la fois X et Y ne sont pas des groupements hydroxy au même moment).
PCT/JP2007/066459 2006-08-24 2007-08-24 Promoteur de la production de protéoglycane WO2008023796A1 (fr)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
JP2006227818A JP2009274954A (ja) 2006-08-24 2006-08-24 創傷治癒促進剤
JP2006-227818 2006-08-24
JP2006227822A JP2009274955A (ja) 2006-08-24 2006-08-24 変形性関節症治療薬
JP2006-227822 2006-08-24

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WO2008023796A1 true WO2008023796A1 (fr) 2008-02-28

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012065054A1 (fr) * 2010-11-12 2012-05-18 Los Alamos National Security, Llc Procédés et systèmes de détection d'infection, composés et compositions associés

Citations (3)

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Publication number Priority date Publication date Assignee Title
WO2004076476A1 (fr) * 2003-02-04 2004-09-10 Shionogi Co., Ltd. Glycopeptide et son utilisation comme amorceur du proteoglycane
JP2006063055A (ja) * 2004-08-30 2006-03-09 National Institute Of Advanced Industrial & Technology Ｏ−アセチル化糖アミノ酸
JP2006111618A (ja) * 2004-09-14 2006-04-27 National Institute Of Advanced Industrial & Technology Ｏ−結合型糖アミノ酸

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2004076476A1 (fr) * 2003-02-04 2004-09-10 Shionogi Co., Ltd. Glycopeptide et son utilisation comme amorceur du proteoglycane
JP2006063055A (ja) * 2004-08-30 2006-03-09 National Institute Of Advanced Industrial & Technology Ｏ−アセチル化糖アミノ酸
JP2006111618A (ja) * 2004-09-14 2006-04-27 National Institute Of Advanced Industrial & Technology Ｏ−結合型糖アミノ酸

Non-Patent Citations (1)

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Title
SHIMAWAKI K. ET AL.: "Proteoglycan Seigosei Kiko no Yuki Goseiteki Kaimei", POLYMER PREPRINTS, JAPAN, vol. 54, no. 1, 10 May 2005 (2005-05-10), pages 1PF190, XP003024666 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012065054A1 (fr) * 2010-11-12 2012-05-18 Los Alamos National Security, Llc Procédés et systèmes de détection d'infection, composés et compositions associés

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