WO2017119272A1 - マクロライド系免疫抑制剤の高分子誘導体 - Google Patents
マクロライド系免疫抑制剤の高分子誘導体 Download PDFInfo
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- WO2017119272A1 WO2017119272A1 PCT/JP2016/087774 JP2016087774W WO2017119272A1 WO 2017119272 A1 WO2017119272 A1 WO 2017119272A1 JP 2016087774 W JP2016087774 W JP 2016087774W WO 2017119272 A1 WO2017119272 A1 WO 2017119272A1
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- C—CHEMISTRY; METALLURGY
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- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
- C08G69/00—Macromolecular compounds obtained by reactions forming a carboxylic amide link in the main chain of the macromolecule
- C08G69/40—Polyamides containing oxygen in the form of ether groups
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
- A61K31/436—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a six-membered ring having oxygen as a ring hetero atom, e.g. rapamycin
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
- A61K47/59—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes
- A61K47/60—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
- A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
- A61K47/645—Polycationic or polyanionic oligopeptides, polypeptides or polyamino acids, e.g. polylysine, polyarginine, polyglutamic acid or peptide TAT
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
- C08G69/00—Macromolecular compounds obtained by reactions forming a carboxylic amide link in the main chain of the macromolecule
- C08G69/02—Polyamides derived from amino-carboxylic acids or from polyamines and polycarboxylic acids
- C08G69/08—Polyamides derived from amino-carboxylic acids or from polyamines and polycarboxylic acids derived from amino-carboxylic acids
- C08G69/10—Alpha-amino-carboxylic acids
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
- C08G69/00—Macromolecular compounds obtained by reactions forming a carboxylic amide link in the main chain of the macromolecule
- C08G69/48—Polymers modified by chemical after-treatment
Definitions
- the present invention relates to a polymer derivative of a macrolide compound, its production method and its use.
- the macrolide compound used in the present invention has an affinity for FKBP type immunophilin and has a common activity of inhibiting peptidyl-prolyl isomerase and / or rotamase enzyme activity.
- macrolide compounds include tricyclo compounds including rapamycin, tacrolimus (FK506), ascomycin and the like.
- a macrolide compound or a pharmaceutically acceptable salt thereof has excellent immunosuppressive action, antibacterial activity, and other pharmacological activities. Therefore, rejection of organ or tissue transplantation, graft-versus-host reaction,
- Patent Document 1 describes that it is useful for the treatment and prevention of autoimmune diseases and infectious diseases.
- Tacrolimus is widely used to treat rejection after organ transplantation, graft-versus-host disease after bone marrow transplantation, refractory active ulcerative colitis, etc.
- tacrolimus is poorly soluble in water (2.4 to 3.6 ⁇ M, room temperature) and has low bioavailability when administered orally.
- the therapeutic range of tacrolimus is narrow, and the pharmacokinetics between individuals and within individuals are large, making it difficult to control blood concentration.
- Known factors for tacrolimus pharmacokinetics include low solubility in water, the expression level of P-glycoprotein and drug metabolizing enzyme CYP3A in the gastrointestinal mucosa, and individual differences in genotype (Non-patent Document 1, 2).
- nephrotoxicity is caused by a decrease in blood flow and glomerular filtration rate due to the vasoconstrictive action of renal arterioles, and in addition, a lack of nutritional supply to tubular cells.
- Pancreatic toxicity is the onset of abnormal glucose tolerance due to suppression of insulin production from pancreatic ⁇ cells mainly based on insulin mRNA transcription inhibition of pancreatic ⁇ cells. These toxicities are expressed depending on the plasma concentration of tacrolimus.
- Non Patent Literature 3 As a side effect of the central nervous system of tacrolimus, reversible posterior leukoencephalopathy syndrome, hypertensive encephalopathy, etc. have been reported in humans (including 0.1-0.5% including post-marketing surveillance). In addition, intravenous administration of tacrolimus to rats has been observed to induce mild respiratory distress, decreased locomotor activity, prone position, and stereotypical behavior. (Non Patent Literature 3)
- the bonding mode of the polymer and the drug includes a case where the polymer and the drug are covalently bonded and a case where the polymer and the drug are physically adsorbed.
- the drug covalently bonded to the polymer is released from the polymer by a hydrolysis reaction or the like in the body.
- the drug physically adsorbed on the polymer is gradually released from the polymer in the body regardless of a chemical reaction such as a hydrolysis reaction. In either case, the enzyme is not involved in the release of the drug from the polymer, but the difference in the binding mode between the polymer and the drug is thought to cause a difference in the release mechanism of the drug.
- Patent Document 2 and Patent Document 3 describe a polymer derivative obtained from a copolymer composed of polyethylene glycols and polyaspartic acid and a drug.
- a copolymer and a drug are physically adsorbed.
- the sustained release property of the polymer derivative is due to the gradual dissociation of the drug from the copolymer, and selectively exhibits a medicinal effect on the affected area and has few side effects.
- Patent Document 4 discloses a method of obtaining a compound in which tacrolimus is physically adsorbed on a polymer composed of alkyl substituted polylactide (MPEG-hexPLA). However, there is no description regarding the specific transfer of blood concentration or tacrolimus to the affected area.
- MPEG-hexPLA alkyl substituted polylactide
- Non-Patent Document 4 reports a PEGylated tacrolimus synthesized by chemically binding tacrolimus to a PEG polymer.
- PEGylated tacrolimus has not been more effective than tacrolimus on inflammatory disease model animals such as adjuvant arthritic mice and lupus nephritic mice.
- Non-Patent Document 5 has a report on micelles synthesized by physically adsorbing tacrolimus on a poly (ethylene glycol) ester-poly caprolactone (PEG-PCL) polymer.
- PEG-PCL tacrolimus micelles are more effective in improving inflammation in mice with DSS-induced ulcerative colitis compared to tacrolimus, such as suppression of weight loss, suppression of colonic shortening, colonic bleeding, and loss of crypt cells. It has been revealed.
- PEG-PCL tacrolimus micelles are administered once a day for 12 consecutive days in order to show an inflammation-improving effect, so this compound is considered unable to maintain a long blood concentration.
- Non-Patent Documents 6 and 7 have reports on micelles synthesized by physically adsorbing tacrolimus on a polymer composed of poly caprolactone-b-poly (ethylene oxide) (PCL-b-PEO).
- PCL-b-PEO tacrolimus micelles are described to be gradually taken up into cells compared to tacrolimus. It has also been shown that PCL-b-PEO tacrolimus micelle improves spontaneous motility in rats with sciatic nerve injury model by administering 5 mg / kg three times in the tail vein at 6-day intervals.
- PCL-b-PEO tacrolimus micelles are specialized in functions as neuroprotective agents rather than functions as immunosuppressants. it is conceivable that.
- Non-Patent Document 8 reports a nanoparticle synthesized by physically adsorbing tacrolimus on a polymer composed of poly (lactic-co-glycic acid) (PLGA) or pH-sensitive Eudragit P-4135F.
- the nanoparticles exhibit a higher inflammation-improving effect than tacrolimus on collagen-induced arthritis and DSS-induced colitis mice.
- PLGA poly (lactic-co-glycic acid)
- pH-sensitive Eudragit P-4135F pH-sensitive Eudragit P-4135F
- An object of the present invention is to provide a novel immunosuppressive agent or anti-inflammatory agent that accumulates a drug at the site of inflammation, has a higher effect at a low dose, and maintains a target blood concentration to reduce a long administration interval and toxicity. It is to provide an agent.
- the present inventor has found that a polymer derivative of tacrolimus in which a polyethylene glycol segment and an alcoholic hydroxyl group of tacrolimus are bonded to a carboxy group of a side chain of a polyamino acid derivative solves the problems of the present invention.
- the present invention relates to the following [1] to [14].
- [1] A polymer derivative of tacrolimus in which a polyethylene glycol segment and an alcoholic hydroxyl group of tacrolimus are bonded to a side chain carboxy group of a polyaspartic acid derivative.
- [2] The polymer derivative of tacrolimus according to the above [1] represented by the following general formula (1).
- R 1 represents a hydrogen atom, a group selected from the group consisting of alkyl groups and polyethylene glycol segments of carbon atoms (C1 ⁇ C8)
- acyl R 2 is a hydrogen atom, the number of carbon atoms (C1 ⁇ C8)
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 is a polyethylene glycol segment
- R 5 is —N (R 6 ) CONH (R 7 ) (R 6 , R 7 may be the same or different and may be substituted with a cyclic alkyl group having 3 to 6 carbon atoms or a tertiary amino group)
- X 1 is a bond or a linking group
- a and b are each an integer of 1 to 299
- c, d and e are each 0 or 298 or less.
- R 1 represents a hydrogen atom, a group selected from the group consisting of alkyl groups and polyethylene glycol segments of carbon atoms (C1 ⁇ C8)
- acyl R 2 is a hydrogen atom, the number of carbon atoms (C1 ⁇ C8)
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 is a polyethylene glycol segment
- R 5 is —N (R 6 ) CONH (R 7 ) (R 6 , R 7 may be the same or different and may be substituted with a cyclic alkyl group having 3 to 6 carbon atoms or a tertiary amino group)
- X 1 is a bond or a linking group
- f, g, h and i are each 0 or an integer of 299 or less
- R 9 are each independently hydrogen atom or a carbon atoms (C1 ⁇ C8)
- R 10 is amino group which may have a substituent carbon atoms (C1 ⁇ C20) linear, branched or cyclic alkylamino groups, and optionally substituted (C7 to C20) linear, branched or cyclic aralkylamino groups and substituents
- R 11 is a hydrogen atom or a linear, branched or cyclic alkyl group having a carbon number (C1 to C6) which may have a substituent
- R 12 is a carbon number (C2 to C6) is an alkylene group
- X 2 is a functional group capable of binding to a carboxy group in the side chain of the polyaspartic acid derivative
- o is an integer of 5 to 11,500.
- R 1 is an alkyl group having a carbon number (C1 to C6) or a polyethylene glycol segment represented by the following general formula (6). Polymer derivative of tacrolimus.
- R 13 is a hydrogen atom or a linear, branched or cyclic alkyl group having a carbon number (C1 to C6) which may have a substituent, and R 14 is a carbon number (C2 to C6).
- C6) is an alkylene group, and p is an integer of 5 to 11,500.
- R 2 is an acyl group having a carbon number (C1 to C6)
- o is an integer of 10 to 3000
- (a + b + c + d + e) or (f + g + h + i + j + k + l + m + n) is an integer of 4 to 250
- the polymer derivative of tacrolimus according to [9].
- the polymer derivative of tacrolimus according to the present invention is characterized in that a polyethylene glycol segment and an alcoholic hydroxyl group of tacrolimus are bonded to a side chain carboxy group of a polyaspartic acid derivative directly or via a bonding group.
- This polymer derivative is stable in vivo, can release tacrolimus in an enzyme-independent manner, and exhibits excellent blood retention, so that it is excellent in therapeutic effect at a low dose.
- the release of physiologically active substances independent of enzymes and the maintenance of blood concentration make it unnecessary to control the dose based on blood kinetics (blood trough concentration) and significantly improve safety. There is expected.
- the ratio of the amount of released tacrolimus to the total amount of tacrolimus in Example 1 and Comparative Example 1 in Test Example 1 is shown.
- the ratio of the amount of released tacrolimus to the total amount of tacrolimus in Examples 1, 2, 5, 6 and 8 in Test Example 1 is shown.
- the ratio of the amount of released tacrolimus to the total amount of tacrolimus in Examples 3, 4, 7 and 9 in Test Example 1 is shown.
- the changes in rat blood concentrations of Examples 1, 2, 8 and Comparative Example 1 and tacrolimus in Test Example 2 are shown.
- the change in blood concentration in rats of Examples 3, 7 and 9 in Test Example 3 is shown.
- the anti-inflammatory effect with respect to the rat collagen arthritis of the comparative example 1 in the test example 4 is shown.
- the anti-inflammatory effect with respect to the rat collagen arthritis of Example 1 in the test example 4 is shown.
- the anti-inflammatory effect with respect to the rat collagen arthritis of Examples 2 and 8 in Test Example 4 is shown.
- the anti-inflammatory effect with respect to the rat collagen arthritis of Examples 3, 7 and 9 in Test Example 4 is shown.
- the polymer derivative of tacrolimus according to the present invention is characterized in that a polyethylene glycol segment and an alcoholic hydroxyl group of tacrolimus are bonded to a side chain carboxy group of a polyaspartic acid derivative directly or via a bonding group. The details will be described below.
- the polymer derivative of tacrolimus of the present invention is a polymer compound having a polyaspartic acid derivative of a plurality of units having a polymer main chain structure. That is, a polyaspartic acid derivative having a plurality of side chain carboxy groups is used as the main chain structure of the polymer carrier, and the side chain carboxy groups are chemically functionalized with a polyethylene glycol segment and a macrolide immunosuppressant. It is a polymerized derivative.
- the polyaspartic acid derivative having a side chain carboxy group used in the polymer derivative of tacrolimus of the present invention may be an ⁇ -amide bond type polymer or an amide bond type polymer with a side chain carboxy group. , ⁇ -amide bond polymer, or a mixture thereof, preferably ⁇ -amide bond polymer, ⁇ -amide bond polymer and ⁇ -amide bond polymer. .
- the polymer derivative of tacrolimus of the present invention has, for example, a structure represented by the following general formula (1) or general formula (2).
- R 1 is a group selected from the group consisting of a hydrogen atom, an alkyl group having a carbon number (C1 to C8) and a polyethylene glycol segment
- R 2 is a hydrogen atom, carbon A group selected from the group consisting of an acyl group having a number (C1 to C8) and an alkoxycarbonyl group having a carbon number (C1 to C8)
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 is a polyethylene group A glycol segment
- R 5 is —N (R 6 ) CONH (R 7 ) (R 6 , R 7 may be the same or different, and a cyclic alkyl group having 3 to 6 carbon atoms or a tertiary group
- X 1 is a bond or a linking group (which is an alkyl group having a carbon number (C1 to C5) which may be substituted with an amino group).
- a, b, c, d, and e are all average values. a and b are each an integer of 1 to 299. C, d and e are each an integer of 0 or 298 or less, and a + b + c + d + e is an integer of 2 to 300.
- f, g, h, i, j, k, l, m, and n are all average values.
- f, g, h, and i are each an integer of 0 or 299 or less.
- F + g and h + i are each an integer of 1 to 299, j, k, l, m and n are each an integer of 0 or 298 or less, and f + g + h + i + j + k + l + m + n is an integer of 2 to 300.
- each repeating unit is not particularly limited, and may be randomly arranged, and is not limited to those described in the general formula (1) and the general formula (2).
- the terminal groups of the polyaspartic acid derivative are both N terminal groups (R 2 in the general formula (1) and general formula (2)) and C terminal groups (R 1 in the general formula (1) and general formula (2)). It is not particularly limited, and may be an unprotected free amino group and a free carboxylic acid, and salts thereof, and may be an appropriate modification of the N-terminal group and C-terminal group.
- modified N-terminal group of the polyaspartic acid derivative examples include an acylamide modified, an alkoxycarbonylamide modified (urethane modified), and an alkylaminocarbonylamide modified (urea modified). Can do.
- the modified form of R 2 that is the N terminal group in the general formula (1) and the general formula (2) is an acylamide-type modified body or an alkoxycarbonylamide-type modified body (urethane-type modified body).
- examples of the modified C-terminal group of the polyaspartic acid derivative include ester-type modified products, amide-type modified products, and thioester-type modified products.
- the modified form of R 1 which is the C terminal group in the general formula (1) and the general formula (2) is an amide type modified body.
- the modifying group for the N-terminal group and C-terminal group of the polyaspartic acid derivative may be any modifying group.
- a linear, branched or cyclic alkyl group having a carbon number (C1 to C6) which may have a substituent via an appropriate bonding group bonded to the N-terminal group and the C-terminal group.
- a terminal modifying group such as an aromatic group having carbon atoms (C6 to C18) which may have a substituent, an aralkyl group having carbon atoms (C7 to C20) which may have a substituent.
- it may be a polyethylene glycol segment capable of imparting water solubility, and may be a terminal modification group bonded via an appropriate bonding group bonded to the N terminal group and the C terminal group.
- the N-terminal group is preferably a suitable acylamide-type modified product or alkoxycarbonylamide-type modified product (urethane-type modified product), and has the above-described substituent via a carbonyl group or a carbonyloxy group.
- An aralkyl group having a carbon number (C7 to C20) may be preferable.
- the C-terminal group is preferably a suitable amide type substituent or ester type substituent, and the number of carbon atoms (C1 to C8) optionally having the above substituent via an amide group or ester group.
- Linear, branched or cyclic alkyl group, an aromatic group having a carbon number (C6 to C18) which may have a substituent, and a carbon number which has a substituent (C7 to C7) C20) is preferably an aralkyl group or a polyethylene glycol segment.
- the polyethylene glycol segment of R 1 in the general formulas (1) and (2), which is the C terminal group, is a segment having a repeating structure of an ethyleneoxy group: (CH 2 CH 2 O) unit.
- a polyethylene glycol chain having an average number of ethyleneoxy group repeating units (degree of polymerization) of 5 to 11500, more preferably a unit of polymerization degree of 10 to 3000, and particularly preferably a unit of polymerization degree of 20 to 1500 Is a segment structure.
- the polyethylene glycol segment has an average molecular weight corresponding to polyethylene glycol of 200 daltons to 500 kilodaltons.
- the molecular weight is preferably 500 to 150 kilodaltons, and more preferably 1000 to 70 kilodaltons. In particular, the molecular weight is preferably 1000 daltons to 50 kilodaltons.
- the structure of the polyethylene glycol segment of the R 1 include the structures represented by the following general formula (6). That is, a structure in which an oxygen atom of an ethyleneoxy group unit and an alkylene group having a carbon number (C1 to C8) are ether-bonded is preferable.
- p is an average value and is an integer of about 5 to 11,500, preferably about 10 to 3000, particularly preferably about 20 to 1500.
- the terminal group (R 13 ) of the polyethylene glycol segment of R 1 is not particularly limited, and may be a hydrogen atom, a straight chain having a carbon number (C1 to C6) optionally having a substituent, a branched chain or Examples thereof include a cyclic alkyl group, an optionally substituted alkynyl group having a carbon number (C2 to C6), and an optionally substituted carbon number (C7 to C20) aralkyl group.
- the substituent in the alkyl group, alkynyl group, and aralkyl group include a hydroxyl group, an amino group, a formyl group, and a carboxyl group.
- Examples of the linear alkyl group which may have a substituent in the terminal group of the polyethylene glycol segment of R 1 include, for example, methyl group, ethyl group, n-propyl group, n-butyl group, n-pentyl group, n -Hexyl group and the like can be mentioned.
- Examples of the branched alkyl group which may have a substituent include isopropyl group, isobutyl group, t-butyl group, isopentyl group, 2-methylbutyl group, neopentyl group, 1-ethylpropyl group and 4-methylpentyl group.
- 3-methylpentyl group 2-methylpentyl group, 1-methylpentyl group, 3,3-dimethylbutyl group, 2,2-dimethylbutyl group, 1,1-dimethylbutyl group, 1,2-dimethylbutyl group 1,3-dimethylbutyl group, 2,3-dimethylbutyl group, 2-ethylbutyl group and the like.
- Examples of the cyclic alkyl group which may have a substituent include a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, and a cyclohexyl group.
- Examples of the substituent that the linear, branched or cyclic alkyl group may have in the terminal group of the polyethylene glycol segment of R 1 include a thiol group, a hydroxyl group, a halogen atom, a nitro group, a cyano group, and an alkylthio group.
- an optionally substituted carbon number (C2-C6) alkynyl group includes, for example, 2-propynyl, 3-butynyl group, 4-heptynyl group, 5- A hexynyl group etc. are mentioned.
- an optionally substituted carbon number (C7 to C20) aralkyl group is a straight chain in which any one hydrogen atom is substituted with an aryl group or It is a branched alkyl group.
- Examples include benzyl group, 2-phenylethyl group, 4-phenylbutyl group, 3-phenylbutyl group, 5-phenylpentyl group, 6-phenylhexyl group, 8-phenyloctyl group and the like.
- a benzyl group, a 4-phenylbutyl group, and an 8-phenyloctyl group are preferable.
- the bonding side terminal group (R 14 ) with the polyaspartic acid main chain is not particularly limited as long as it is a suitable linking group to the polyaspartic acid derivative.
- An alkylene group having a carbon number (C1 to C8) which may have a substituent is preferable.
- a methylene group, ethylene group, trimethylene group, butylene group, hexamethylene group, octamethylene group and the like can be mentioned.
- R 14 in the general formula (6) is preferably a trimethylene group.
- R 2 in the general formulas (1) and (2) is a substituent selected from the group consisting of a hydrogen atom, an acyl group having carbon atoms (C1 to C8) and an alkoxycarbonyl group having carbon atoms (C1 to C8). It is a group.
- the carbon group (C1 to C8) acyl group is a linear, branched or cyclic acyl group (C1 to C8). Examples include formyl group, acetyl group, propionyl group, butyroyl group, cyclopropylcarbonyl group, cyclopentanecarbonyl group and the like.
- the carbon number (C1 to C8) alkoxycarbonyl group is a linear, branched or cyclic alkoxycarbonyl group (C1 to C8).
- the residue of the alcoholic hydroxyl group of tacrolimus which is R 3 in the general formula (1) and the general formula (2) is a side chain carboxy group of the polyaspartic acid derivative, directly or via a bonding group.
- Tacrolimus is an alcoholic hydroxyl group of tacrolimus having an ester bond.
- Tacrolimus is represented by the following formula (I). There are a plurality of alcoholic hydroxyl groups of tacrolimus, but the substitution position is not limited as long as it is an alcoholic hydroxyl group.
- the polyethylene glycol segment bonded to the side chain carboxy group of the polyaspartic acid derivative directly or via a bonding group is the same as the polyethylene glycol segment of R 1 in the general formula (1) and the general formula (2). have a repeating structure, the degree of polymerization and average molecular weight is the same as polyethylene glycol segments of R 1.
- R 4 in the general formula (1) and the general formula (2) is a polyethylene glycol segment bonded to the side chain carboxy group of the polyaspartic acid derivative directly or via a bonding group.
- the polyethylene glycol segment bonded to the side chain carboxy group of the polyaspartic acid derivative directly or via a bonding group has 1 to 200 units bonded to one molecule of the polyaspartic acid derivative. That is, the polymer derivative of tacrolimus has a plurality of units of polyethylene glycol segments. Preferably it has 1 to 150 units of polyethylene glycol segments, more preferably 1 to 100 units.
- R 4 in the general formula (1) and the general formula (2) include a structure represented by the following general formula (5).
- R 11 has a hydrogen atom or a linear, branched or cyclic alkyl group having a carbon number (C1 to C6) which may have a substituent, and a substituent. Or a group selected from the group consisting of an alkynyl group having a good carbon number (C2 to C6) and an aralkyl group having a carbon number (C7 to C20) which may have a substituent.
- R 12 is an alkylene group having a carbon number (C2 to C6).
- X 2 is a functional group that can be bonded to the carboxy group in the side chain of the polyaspartic acid derivative.
- o is an average value, an integer of 5 to 11,500, preferably 10 to 3000, and particularly preferably 20 to 1500.
- R 11 is not particularly limited, and has a hydrogen atom, a linear, branched or cyclic alkyl group having a carbon number (C1 to C6) which may have a substituent, and a substituent. And an alkynyl group having a carbon number (C2 to C6), an aralkyl group having a carbon number (C7 to C20) which may have a substituent, and the like.
- Examples of the substituent in the alkyl group, alkynyl group, and aralkyl group include a hydroxyl group, an amino group, a formyl group, and a carboxyl group.
- Examples of the linear alkyl group which may have a substituent in R 11 include a methyl group, an ethyl group, an n-propyl group, an n-butyl group, an n-pentyl group, and an n-hexyl group. be able to.
- Examples of the branched alkyl group which may have a substituent include isopropyl group, isobutyl group, t-butyl group, isopentyl group, 2-methylbutyl group, neopentyl group, 1-ethylpropyl group and 4-methylpentyl group.
- 3-methylpentyl group 2-methylpentyl group, 1-methylpentyl group, 3,3-dimethylbutyl group, 2,2-dimethylbutyl group, 1,1-dimethylbutyl group, 1,2-dimethylbutyl group 1,3-dimethylbutyl group, 2,3-dimethylbutyl group, 2-ethylbutyl group and the like.
- Examples of the cyclic alkyl group which may have a substituent include a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, and a cyclohexyl group.
- the substituent that the linear, branched or cyclic alkyl group may have is a thiol group, a hydroxyl group, a halogen atom, a nitro group, or a cyano group.
- the optionally substituted carbon number (C2-C6) alkynyl group includes, for example, a 2-propynyl group, a 3-butynyl group, a 4-heptynyl group , 5-hexynyl group and the like.
- the aralkyl group having a carbon number (C7 to C20) which may have a substituent has any one hydrogen atom substituted with an aryl group.
- a linear or branched alkyl group examples thereof include benzyloxy group, 2-phenylethyl group, 4-phenylbutyl group, 3-phenylbutyl group, 5-phenylpentyl group, 6-phenylhexyl group, and 8-phenyloctyl group.
- a benzyl group, a 4-phenylbutyl group, and an 8-phenyloctyl group are preferable.
- R 11 is preferably a hydrogen atom or a linear, branched or cyclic alkyl group having a carbon number (C1 to C6) which may have a substituent.
- terminal group (R 12 ) on the side bonded to the side chain carboxy group of the polyaspartic acid derivative of R 4 is a functional group capable of binding to the side chain carboxy group of the polyaspartic acid derivative, It is not particularly limited. In the present invention, it is an alkylene group having a carbon number (C2 to C6), and examples thereof include an ethylene group, a trimethylene group, a butylene group and the like, and a trimethylene group is particularly preferable.
- X 2 in the general formula (5) is not particularly limited as long as it is a functional group that can be bonded to the carboxy group of the side chain of the polyaspartic acid derivative.
- —NH— -O- or -S- is preferred.
- R 5 in the general formula (1) and the general formula (2) may be —N (R 6 ) CONH (R 7 ).
- R 6 and R 7 may be the same or different, and may be a cyclic alkyl group having a carbon number (C3 to C6) or an alkyl group having a carbon number (C1 to C5) optionally substituted with a tertiary amino group. is there.
- Examples of the cyclic alkyl having a carbon number (C3 to C6) include a cyclohexyl group.
- alkyl group having a carbon number (C1 to C5) which may be substituted with a tertiary amino group examples include an ethyl group, an isopropyl group, and a 3-dimethylaminopropyl group.
- the tertiary amino group of the alkyl group having a carbon number (C1 to C5) which may be substituted with a tertiary amino group is a dimethylamino group, a diethylamino group, or the like.
- the residue of the alcoholic hydroxyl group of the tacrolimus, the polyethylene glycol segment, and R 5 in the general formula (1) and the general formula (2) are the same as the X 1 in the general formula (1) and the general formula (2), respectively.
- X 1 in the general formula (1) and the general formula (2) may be a linking group that bonds the R 3 , R 4, and R 5 to the side chain carbonyl group of the polyaspartic acid main chain, or It may be simply a bond, and when X 1 is a linking group, it can bind to the binding functional group of R 3 , R 4 and R 5 and the side chain carboxy group of the polyaspartic acid derivative. If it has a functional group in both ends, it will not specifically limit.
- the terminal binding functional group on the R 3 , R 4 and R 5 side is preferably a carboxy group, an oxycarboxy group or an aminocarboxy group. Since R 3 , R 4 and R 5 have an amino group, a hydroxyl group and / or a thiol group in the molecule, these binding functional groups have an amide bond or an ester bond with the amino group, hydroxyl group and / or thiol group. Thioester bond, urethane bond, carbonate bond and urea bond.
- the other end-bonding functional group on the side chain carboxy group side of X 1 is preferably an amino group, a hydroxyl group or a thiol group.
- These binding functional groups can form a side chain carboxy group and an amide bond, an ester bond, or a thioester bond.
- X 1 may have a substituent in which one end group is a carboxy group, an oxycarboxy group or an aminocarboxy group, and the other end group is an amino group, a hydroxyl group or a thiol group ( A C1-C8) alkylene group or alkenylene group is preferred.
- X 1 is a linking group
- X 1 is a linking group
- a hydrogen atom may be modified with an appropriate substituent.
- substituents include a hydroxyl group, an amino group, a halogen atom, an alkyl group having a carbon number (C1 to C8), an alkylcarbonylalkoxy group having a carbon number (C1 to C8), and an alkylcarbonylamide group having a carbon number (C1 to C8).
- X 1 is preferably —CO— (CH 2 ) y —NH— or —CO— (CH 2 ) y —O—.
- —CO— (CH 2 having a carboxy group capable of forming an amide bond, an ester bond or a thioester bond with R 3 , R 4 or R 5 and an amino group capable of forming an amide bond with the side chain carboxy group.
- Y —NH—.
- X 1 may be used amino acid derivatives.
- X 1 is an amino acid derivative
- the amino group at the N-terminal of the amino acid is amide-bonded to the side chain carboxy group
- the carboxy group at the C-terminal is the amino group, hydroxyl group or thiol group of the R 3 , R 4 or R 5
- An amide bond, an ester bond or a thioester bond is formed.
- Amino acids used as X 1 can be a naturally occurring amino acid or unnatural amino acid, L body, it can be used without being limited particularly either D-form.
- hydrocarbon amino acids such as glycine, ⁇ -alanine, alanine, leucine and phenylalanine
- acidic amino acids such as aspartic acid and glutamic acid
- basic amino acids such as lysine, arginine and histidine
- the amino acid derivative as X 1 is preferably an aspartic acid derivative.
- the aspartic acid derivative is an aspartic acid derivative in which an ⁇ -carboxy group is bonded to the R 3 , R 4 or R 5 and the ⁇ -carboxy group is an amide.
- an aspartic acid derivative in which a ⁇ -carboxy group is bonded to the R 3 , R 4 or R 5 and the ⁇ -carboxy group is an amide may be used.
- R 3 , R 4, or R 5 When the other carboxy group that is not involved in the bonding of R 3 , R 4, or R 5 is an amide, it may have a substituent (C1-20) alkylamide, substituent An aromatic amide having a carbon number (C5 to C20) which may have a substituent, an aralkylamide having a carbon number (C7 to C20) which may have a substituent, or an amino acid binding residue in which a carboxy group is protected, etc. Is mentioned.
- alkylamide having a carbon number (C1-20) which may have a substituent of the aspartic acid derivative examples include, for example, methylamide, ethylamide, isopropylamide, t-butylamide, cyclohexylamide, dodecylamide, octadecylamide and the like. Can be mentioned.
- Examples of the aromatic amide having a carbon number (C5 to C20) which may have a substituent of the aspartic acid derivative include phenylamide, 4-methoxyphenylamide, 4-dimethylaminophenylamide, 4-hydroxyphenyl. Examples include amides. Examples of the aralkyl amide having a carbon number (C7 to C20) which may have a substituent of the aspartic acid derivative include benzylamide, 2-phenylethylamide, 4-phenylbutyramide, 8-phenyloctylamide and the like. Is mentioned.
- amino acid binding residue in which the carboxy group of the aspartic acid derivative is protected examples include glycinyl-methyl ester groups, glycinyl-ethyl ester groups, glycinyl represented by the following formulas (7-1) to (7-4): -Propyl ester group, glycinyl-benzyl ester group, alaninyl-methyl ester group, alaninyl-ethyl ester group, alaninyl-propyl ester group, alaninyl-benzyl ester represented by the following formulas (8-1) to (8-4) Groups, valinyl-methyl ester groups, valinyl-ethyl ester groups, valinyl-propyl ester groups, valinyl-benzyl ester groups represented by the following formulas (9-1) to (9-4), Leucinyl-methyl ester group, leucinyl-ethyl ester group,
- X 1 may be an aspartic acid derivative or a maleic acid derivative represented by the following general formula (3) or general formula (4).
- R 8 and R 9 are each independently a hydrogen atom or an alkyl group having a carbon number (C1 to C8).
- R 10 is an amino group, an optionally substituted linear (C1-C20) linear, branched or cyclic alkylamino group, an optionally substituted carbon number (C7 To C20) linear, branched or cyclic aralkylamino groups, optionally substituted aromatic (C5-C20) aromatic amino groups, and carboxy-group-protected amino acid residues.
- CX—CY is C ⁇ C (double bond) in the CH—CH or Z configuration.
- the alkyl group having carbon atoms (C1 to C8) in R 8 and R 9 is a linear, branched or cyclic alkyl group having carbon atoms (C1 to C8).
- the linear alkyl group include a methyl group, an ethyl group, an n-propyl group, an n-butyl group, and an n-hexyl group.
- the branched alkyl group include isopropyl group, t-butyl group, 1-methyl-propyl group, 2-methyl-propyl group, 2,2-dimethylpropyl group and the like.
- Examples of the cyclic alkyl group include a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, a cyclohexyl group, and the like.
- the linear, branched or cyclic alkylamino group having a carbon number (C1 to C20) which may have a substituent is, for example, a methylamino group, an ethylamino group, an isopropylamino group, Examples thereof include t-butylamino group, cyclohexylamino group, n-octylamino group, dodecylamino group, and octadecylamino group.
- Examples of the linear, branched or cyclic aralkylamino group having a carbon number (C7 to C20) which may have a substituent include, for example, benzylamino group, 2-phenylethylamino group, 4-phenylbutyl An amino group, 8-phenyloctylamino group, etc. are mentioned.
- Examples of the aromatic amino group having a carbon number (C5 to C20) which may have a substituent include an anilino group, a 4-methoxyanilino group, a 4-dimethylaminoanilino group, and a 4-hydroxyanilino group. Etc.
- R 10 may be an amino acid binding residue in which a carboxy group is protected.
- amino acid-bonded residues in which the carboxy group is protected include glycinyl-methyl ester groups, glycinyl-ethyl ester groups, glycinyl-propyl ester groups represented by the following formulas (7-1) to (7-4), Glycinyl-benzyl ester group, alaninyl-methyl ester group, alaninyl-ethyl ester group, alaninyl-propyl ester group, alaninyl-benzyl ester group represented by the following formulas (8-1) to (8-4): 9-1) to (9-4) Valinyl-methyl ester group, valinyl-ethyl ester group, valinyl-propyl ester group, valinyl-benzyl ester group, represented by the following formulas (10-1) to (10-4) ) Leucinyl-methyl este
- the average value of the total number of ⁇ -aspartic acids in the polymer derivative of tacrolimus represented by the general formula (1) is represented by a + b + c + d + e, which is about 2 to 300, preferably about 4 to 250, particularly preferably. Is 8 to 200.
- the ratio of tacrolimus-bound aspartic acid number (a) to the total ⁇ -aspartic acid number (a + b + c + d + e) is 1 to 99%, preferably 1 to 80%, more preferably 1 to 60%. Since it is necessary to coexist with the polyethylene glycol segment-bound aspartic acid unit, it is preferably 1 to 50%, more preferably 1 to 30%, and particularly preferably 5 to 30% in consideration of the balance.
- the ⁇ -aspartic acid number (a) is 1 to 200, preferably about 2 to 150, particularly preferably about 2 to 100, or about 2 to 40.
- the ratio of the number (b) of aspartic acid bonded to the polyethylene glycol segment to the total number of ⁇ -aspartic acids (a + b + c + d + e) is 1 to 99%, preferably 1 to 80%, more preferably 1 to 60%. Since it is necessary to coexist with the tacrolimus-bound aspartic acid unit, it is preferably 1 to 40%, particularly 2 to 40% in consideration of the balance.
- the ⁇ -aspartic acid number (b) is 1 to 200, preferably about 1 to 150, particularly preferably about 1 to 100, or about 2 to 40.
- the average value of the total aspartic acid number in the polymer derivative of tacrolimus represented by the general formula (2) is represented by f + g + h + i + j + k + l + m + n and is about 2 to 300, preferably about 4 to 250, particularly preferably 8 ⁇ 200.
- the ratio of tacrolimus-bound aspartic acid (f + g) to the total aspartic acid number (f + g + h + i + j + k + l + m + n) is 1 to 99%, preferably 1 to 80%, more preferably 1 to 60%. Since it is necessary to coexist with the polyethylene glycol segment-bound aspartic acid unit, it is preferably 1 to 50%, more preferably 1 to 30%, and particularly preferably 5 to 30% in consideration of the balance.
- the number of aspartic acids (f + g) bound to tacrolimus is 1 to 200, preferably about 2 to 150, particularly preferably about 2 to 100, or about 2 to 40.
- the ratio of the number of aspartic acid bonded to the polyethylene glycol segment (h + i) to the total number of aspartic acid (f + g + h + i + j + k + l + m + n) is 1 to 99%, preferably 1 to 80%, more preferably 1 to 60%. Since it is necessary to coexist with the tacrolimus-bound aspartic acid unit, it is preferably 1 to 40%, particularly 2 to 40% in consideration of the balance.
- the number of aspartic acids (h + i) bound to tacrolimus is 1 to 200, preferably about 1 to 150, particularly preferably about 1 to 100, or about 2 to 40.
- the ratio of ⁇ -aspartic acid (f + h + j + l) to the total aspartic acid number (f + g + h + i + j + k + l + m + n) is 1 to 80%, preferably 1 to 50%. This ratio can be changed as appropriate by, for example, selecting deprotection conditions for the protecting group of polyaspartic acid.
- the molecular weight of the polymer derivative of tacrolimus of the present invention a calculated value obtained by adding the constituent molecular weights of the above-mentioned constituent parts is adopted as the “molecular weight of the polymer derivative of tacrolimus”. That is, (1) the molecular weight of the polyaspartic acid main chain, (2) the total molecular weight of the polyethylene glycol segment multiplied by the molecular weight of the polyethylene glycol segment, and (3) the molecular weight of the binding residue of tacrolimus multiplied by the number of bonds.
- the total molecular weight of tacrolimus was, (4) X 1 attached to any of the total molecular weight of X 1 residues multiplied by the number of bonds on the molecular weight of X 1 residues bonded to polyethylene glycol segment, and (5) any tacrolimus the total molecular weight of X 1 residues multiplied by the number of bonds to residues molecular weight, a calculated value obtained by summing the the molecular weight.
- the molecular weight of the polyaspartic acid derivative is required to be regulated by the accuracy in kilodalton units. Therefore, the analysis method of each component is not particularly limited as long as it is an analysis method with sufficient accuracy in measuring the molecular weight of the polyaspartic acid derivative in kilodalton units, and various analysis methods are appropriately selected. You can do it. Below, the preferable analysis method in each component is listed.
- the molecular weight of the (1) polyaspartic acid main chain is a calculated value obtained by multiplying the molecular weight of the polymerization monomer unit of the main chain by the polymerization number.
- the number of polymerizations the number of polymerizations calculated from the integral value of 1 H-NMR, the number of polymerizations calculated by amino acid analysis, or the number of polymerizations calculated by neutralization titration can be used.
- the total molecular weight of the polyethylene glycol segment is a calculated value obtained by multiplying the molecular weight of the polyethylene glycol segment by the number of bonds.
- the molecular weight of the polyethylene glycol segment an average molecular weight determined by the peak top molecular weight of the polyethylene glycol segment structural compound to be used, which is measured by a GPC method based on a polyethylene glycol standard product, is employed.
- the number of bonds of the polyethylene glycol segment can be determined by cleaving the polyethylene glycol segment from a polymer derivative of tacrolimus and quantitatively analyzing the released polyethylene glycol segment. Alternatively, a method of calculating from the consumption rate of the polyethylene glycol segment in the reaction of introducing the polyethylene glycol segment into the polyaspartic acid main chain may be used.
- the total molecular weight of tacrolimus is a calculated value obtained by multiplying the binding residue molecular weight of tacrolimus by the number of bonds.
- the binding number of the tacrolimus is determined by hydrolyzing the polymer derivative of the tacrolimus, derivatizing the liberated tacrolimus or its degradation product as necessary, and quantitatively analyzing it by high performance liquid chromatography (HPLC). Is mentioned. Alternatively, it may be a method of calculating from the consumption rate of tacrolimus in the reaction of introducing tacrolimus into the polyaspartic acid main chain.
- the total molecular weight of X 1 residues bonded to any polyethylene glycol segment of the (4) is a calculated value obtained by multiplying the number of bonds on the molecular weight of X 1 residues.
- In the reaction of introducing the X 1 to the polyaspartic acid backbone may be a method of calculating the consumption rate of X 1.
- the total molecular weight of X 1 residue attached to any of tacrolimus of the (5) is a calculated value obtained by multiplying the number of bonds on the molecular weight of X 1 residues.
- a method of calculating from the consumption rate of X 1 may be used.
- the polymer derivative of tacrolimus of the present invention is characterized in that the mass content of the polyethylene glycol segment in the polymer derivative is from 30% by mass to 95% by mass.
- the mass content of the polyethylene glycol segment can be calculated from the content ratio of the total molecular weight of the polyethylene glycol segment of (2) above to the molecular weight of the polymer derivative of the tacrolimus described above. That is, the mass content of the polyethylene glycol segment is calculated by the following formula.
- PEG mass content (%) PEG total molecular weight / TAC polymer derivative molecular weight ⁇ 100 Note) PEG: Polyethylene glycol segment Note) TAC: Tacrolimus
- the more preferable range of the mass content of the polyethylene glycol segment is 30% by mass to 95% by mass, and particularly preferably 40% by mass to 95% by mass.
- the polymer derivative of tacrolimus of the present invention preferably has a mass content of tacrolimus in the polymer derivative of 2% to 50% by mass.
- the mass content of the macrolide immunosuppressant in the polyamino acid derivative can be calculated from the content ratio of the total molecular weight of the (3) tacrolimus to the molecular weight of the polyaspartic acid derivative. That is, the mass content of tacrolimus is calculated by the following formula.
- TAC mass content (%) TAC total molecular weight / TAC polymer derivative molecular weight ⁇ 100 Note) TAC: Tacrolimus
- a more preferable range of the mass content of the tacrolimus is 2% by mass or more and 50% by mass or less.
- the tacrolimus content is particularly preferably 2% by mass or more and 40% by mass or less.
- the polymer derivative of tacrolimus according to the present invention comprises an ester bond, an amide bond and / or a thioester group using a dehydration condensing agent in a side chain carboxy group of a polyaspartic acid derivative and the alcoholic hydroxyl group and polyethylene glycol segment of tacrolimus in an organic solvent. This manufacturing method is also included in the present invention.
- DMF formamide
- DI 1,3-dimethyl-2-imidazolidinone
- NMP N-methylpyrrolidone
- DCC diisopropylcarbodiimide
- EDCI 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride
- EDCI 1-ethoxycarbonyl-2-ethoxy-1,2-dihydroxyquinolinone
- DMAP N, N-dimethylaminopyridine
- a polymer derivative of tacrolimus in which R 5 is a —N (R 6 ) CONH (R 7 ) group can also be obtained using the above carbodiimides as a condensing agent.
- the side chain carboxy group of the polyaspartic acid derivative is activated by the above method and then the amount of the corresponding tacrolimus alcohol to be added.
- a method of reacting a basic hydroxyl group and a polyethylene glycol segment under basic conditions, a method of activating the corresponding tacrolimus and polyethylene glycol segment and then reacting with a side chain carboxy group of a polyaspartic acid derivative are also possible.
- tacrolimus may be introduced and then polyethylene glycol segment may be introduced, or polyethylene glycol segment may be introduced and then tacrolimus may be introduced. , May be simultaneous.
- unreacted carboxy groups in the polyamino acid derivative can be reactivated by the same reaction. May be introduced.
- the production method of the polymer derivative of tacrolimus of the present invention is not limited to the above method.
- the polymer derivative of tacrolimus of the present invention has a property of gradually releasing tacrolimus after administration into a living body, and has a use as a medicine containing the tacrolimus as an active ingredient.
- the use of the polymer derivative of tacrolimus of the present invention as a pharmaceutical is not particularly limited as long as it is a disease having a therapeutic effect by the tacrolimus.
- it is suitable for pharmaceuticals used for treatment of autoimmune diseases, inflammatory diseases, allergic diseases, suppression of rejection in organ transplantation and bone marrow transplantation, and the like.
- Particularly preferred is a medicament for the treatment of autoimmune diseases or inflammatory diseases.
- the autoimmune disease include rheumatoid arthritis, systemic lupus erythematosus, and ulcerative colitis.
- the inflammatory disease include interstitial pneumonia.
- the medicament containing the polymer derivative of tacrolimus of the present invention may have other additives that are usually accepted as pharmaceuticals.
- additives include excipients, extenders, fillers, binders, wetting agents, disintegrants, lubricants, surfactants, dispersants, buffers, preservatives, solubilizers, preservatives, flavoring agents. Agents, soothing agents, stabilizers, tonicity agents and the like.
- the medicament containing the polymer derivative of tacrolimus of the present invention may be prepared as a pharmaceutical preparation for treatment.
- the preparation can be administered by any method such as oral, injection, intrarectal administration, intraportal administration, mixing with organ perfusate, and local administration to the affected organ, preferably parenteral administration.
- Intravenous administration by injection, intraarterial administration or local administration to the affected organ is more preferable, and usually, for example, water, physiological saline, 5% glucose or mannitol solution, water-soluble organic solvent (eg, glycerol, ethanol, dimethyl) Sulfoxide, N-methylpyrrolidone, polyethylene glycol, cremophor and the like, and a mixture thereof) and a mixture of water and the water-soluble organic solvent are used.
- water-soluble organic solvent eg, glycerol, ethanol, dimethyl
- the dose of the tacrolimus polymer derivative of the present invention can be naturally changed depending on the sex, age, physiological condition, pathological condition, etc. of the patient, but parenterally, usually 0.01 to 500 mg / m 2 , preferably 0.1 to 250 mg / m 2 is administered. Administration by injection is performed in veins, arteries, affected areas (inflamed areas) and the like.
- the polymer derivative of tacrolimus of the present invention accumulates in the affected area and has a higher effect than tacrolimus alone at a low dose.
- the polymer derivative of tacrolimus of the present invention is an immunosuppressive agent or anti-inflammatory agent useful for the treatment and prevention of rejection of organs or tissues, graft-versus-host reaction, autoimmune diseases, and infectious diseases. is there.
- Synthesis example 1 Synthesis of polyethylene glycol- ⁇ -polyaspartic acid block copolymer (polyethylene glycol molecular weight 12000, polyaspartic acid polymerization number 95) (compound 1)
- One-terminal methoxy group and one-terminal 3-aminopropyl group polyethylene glycol (SUNBRIGHT MEPA-12T, NOF Corporation, average molecular weight 12 kilodaltons, 10.0 g) was dissolved in dimethyl sulfoxide (DMSO, 130 mL), and then ⁇ - Benzyl-L-aspartic acid-N-carboxylic anhydride (BLA-NCA, 13.8 g, 200 equivalents) was added, and the mixture was stirred at 32.0 ° C.
- DMSO dimethyl sulfoxide
- BLA-NCA Benzyl-L-aspartic acid-N-carboxylic anhydride
- the reaction solution was dropped into a mixed solvent of ethanol (755 mL) and diisopropyl ether (3101 L) over 1 hour and stirred at room temperature for 1 hour.
- the precipitate was collected by filtration and dried in vacuo to obtain a solid (41.8 g).
- the obtained solid (40.0 g) was dissolved in N, N-dimethylformamide (DMF, 800 mL), acetic anhydride (1.26 mL) was added, and the mixture was stirred at 32.5 ° C.
- the reaction solution was dropped into a mixed solvent of ethanol (800 mL) and diisopropyl ether (7200 mL), and stirred at room temperature.
- the precipitate was collected by filtration and dried in vacuo to obtain a solid (32.2 g).
- the obtained solid (30 g) was suspended in acetonitrile (MeCN, 1200 mL), 0.2N aqueous sodium hydroxide solution (1200 mL) and purified water (180 mL) were added, and hydrolysis was performed at 23 ° C. It was. After 2N hydrochloric acid was added to the reaction solution for neutralization, acetonitrile was removed by concentration under reduced pressure, and the concentrated solution was washed three times with ethyl acetate (1800 mL).
- the aqueous layer was concentrated under reduced pressure, the pH of the solution was adjusted to 10.7 with 1N aqueous sodium hydroxide solution, sodium chloride (60.8 g) was added, and then partitioned adsorption resin column chromatography and ion exchange resin column chromatography. The solution was purified and desalted using chromatography, and the eluted solution was concentrated under reduced pressure and lyophilized to obtain Compound 1 (15.1 g). The polymerization number of aspartic acid in one molecule of this compound based on the titration value using a 0.1 N aqueous potassium hydroxide solution was 94.9.
- Synthesis example 2 Synthesis of polyethylene glycol- ⁇ -polyaspartic acid block copolymer (polyethylene glycol molecular weight 12000, polyaspartic acid polymerization number 43) (compound 2) According to the method described in Synthesis Example 1, the title compound 2 was obtained by using 52.2 equivalents of BLA-NCA to polyethylene glycol having one end methoxy group and one end 3-aminopropyl group. The polymerization number of aspartic acid in one molecule of this compound based on a titration value using a 0.1 N aqueous potassium hydroxide solution was 43.2.
- Synthesis example 3 Synthesis of polyethylene glycol- ⁇ -polyaspartic acid block copolymer (polyethylene glycol molecular weight 12000, polyaspartic acid polymerization number 73) (compound 3)
- One end methoxy group and one end 3-aminopropyl group polyethylene glycol (SUNBRIGHT MEPA-12T, NOF Corporation, average molecular weight 12 kilodalton, 10.0 g) was dissolved in DMSO (190 mL), and then BLA-NCA (8 .89 g, 129 equivalents) was added and stirred at 32 ° C. overnight.
- the reaction solution was dropped into a mixed solvent of ethanol (400 mL) and diisopropyl ether (1600 mL) over 1 hour and stirred at room temperature for 1 hour.
- the precipitate was collected by filtration and dried in vacuo to obtain a solid (26.5 g).
- the obtained solid (23.0 g) was dissolved in acetonitrile (460 mL), acetic anhydride (724 mL) was added, and the mixture was stirred at 35 ° C. for 3 hr, and then cooled to 23 ° C.
- 0.2 N sodium hydroxide (539 mL) was added dropwise to the reaction solution, and hydrolysis was performed at 23 ° C.
- the reaction solution was neutralized by adding 2N hydrochloric acid, and after removing acetonitrile by concentration under reduced pressure, the concentrated solution was washed three times with ethyl acetate (720 mL). Concentrate the aqueous layer under reduced pressure, adjust the pH of the solution to 10.6 with 1N aqueous sodium hydroxide solution, add sodium chloride (57.5 g), then partition adsorption resin column chromatography, followed by ion exchange resin. After purification using column chromatography, the eluted solution was concentrated under reduced pressure and lyophilized to obtain Compound 3 (13.0 g). The polymerization number of aspartic acid in 1 molecule of this compound based on the titration value using 0.1 N potassium hydroxide aqueous solution was 72.8.
- Synthesis example 4 Synthesis of ⁇ -polyaspartic acid polymer (polyaspartic acid polymerization number 78) (compound 4) n-Butylamine (Tokyo Kasei Co., Ltd., 18.3 mg) was dissolved in DMSO (30 mL), BLA-NCA (5.61 g, 90 equivalents) was added, and the mixture was stirred at 30 ° C. overnight. The reaction solution was dropped into a mixed solvent of ethanol (120 mL) and diisopropyl ether (480 mL), and stirred at room temperature. The precipitate was collected by filtration and dried in vacuo to obtain a solid.
- 1,3-Dimethyl-2-imidazolidinone (DMI, 35 mL) was added to the total amount of the obtained solid, dissolved at 78 ° C., and then 2.5 mL of acetic anhydride was added at 70 ° C. After stirring for 3 hours, the reaction solution was dropped into a mixed solution of ethyl acetate (175 mL) and diisopropyl ether (700 mL), and the mixture was stirred overnight at room temperature. The precipitate was collected by filtration and dried in vacuo to obtain a solid (3.74 g).
- Synthesis example 5 Synthesis of ⁇ -polyaspartic acid polymer (polyaspartic acid polymerization number 112) (compound 5) n-Butylamine (manufactured by Tokyo Chemical Industry, 15.8 mg) was dissolved in DMSO (30 mL), BLA-NCA (7.26 g, 135 equivalents) was added, and the mixture was stirred at 30 ° C. overnight. The reaction solution was dropped into a mixed solvent of ethanol (120 mL) and diisopropyl ether (480 mL), and stirred at room temperature. The precipitate was collected by filtration and dried in vacuo to give a solid (5.37 g).
- Synthesis Example 6 Synthesis of polyethylene glycol- ⁇ -polyaspartic acid block copolymer (polyethylene glycol molecular weight 12000, polyaspartic acid polymerization number 87) (Compound 6) Polyethylene glycol having one end methoxy group and one end 3-aminopropyl group (SUNBRIGHT MEPA-12T, manufactured by NOF Corporation, average molecular weight 12 kilodalton, 6.0 g) was dissolved in DMSO (114 mL), and then BLA-NCA (15 0.0 g, 120 equivalents) and stirred at 30 ° C. overnight.
- DMSO DMSO
- BLA-NCA 15 0.0 g, 120 equivalents
- the reaction solution was dropped into a mixed solvent of ethanol (228 mL) and diisopropyl ether (912 mL), and the mixture was stirred at room temperature.
- the precipitate was collected by filtration and dried in vacuo to obtain a solid (23.8 g).
- the total amount of the obtained solid was dissolved in DMF (200 mL), acetic anhydride (0.75 mL) was added, and the mixture was stirred at 30 ° C. for 3 hr.
- the reaction solution was dropped into a mixed solvent of ethyl acetate (200 mL) and diisopropyl ether (1800 mL), and the mixture was stirred at room temperature.
- the precipitate was collected by filtration and dried in vacuo to obtain a solid (15.8 g).
- the obtained solid (8.72 g) was dissolved in 1-methyl-2-pyrrolidone (NMP, 140 mL), 10% palladium-carbon (0.87 g) was added, and hydrogenolysis was performed overnight at 35 ° C. went.
- Activated carbon was added to the reaction solution and stirred for 1 hour, and then 10% palladium-carbon was filtered off.
- the filtrate was added dropwise into a mixed solvent of ethyl acetate (200 mL) and diisopropyl ether (1000 mL), and stirred overnight at room temperature.
- the precipitate was collected by filtration and dried in vacuo to give a solid (4.18 g).
- Synthesis example 7 Synthesis of polyethylene glycol- ⁇ -polyaspartic acid block copolymer (polyethylene glycol molecular weight 12000, polyaspartic acid polymerization number 40) (compound 7) According to the method described in Synthesis Example 6, 51.3 equivalents of BLA-NCA was used with respect to polyethylene glycol having one end methoxy group and one end 3-aminopropyl group to obtain the title compound 7. At this time, DMF was used instead of NMP. The number of aspartic acids polymerized in one molecule of this compound based on a titration value using a 0.1 N aqueous potassium hydroxide solution was 40.2.
- Synthesis example 8 Synthesis of polyethylene glycol- ⁇ -polyaspartic acid block copolymer (polyethylene glycol molecular weight 12000, polyaspartic acid polymerization number 90) (compound 8) Polyethylene glycol having one end methoxy group and one end 3-aminopropyl group (SUNBRIGHT MEPA-12T, manufactured by NOF Corporation, average molecular weight 12 kilodalton, 10.0 g) was dissolved in DMSO (200 mL), and then BLA-NCA (28 0.7 g, 140 equivalents) was added and stirred at 32.5 ° C. overnight.
- DMSO 200 mL
- BLA-NCA 28 0.7 g, 140 equivalents
- the reaction solution was dropped into a mixed solvent of ethanol (400 mL) and diisopropyl ether (1600 mL), and stirred at room temperature. The precipitate was collected by filtration and dried in vacuo to obtain a solid (28.6 g).
- the obtained solid (18.1 g) was dissolved in NMP (181 mL), acetic anhydride (0.57 mL) was added, and the mixture was stirred at 30 ° C. overnight.
- the reaction solution was dropped into a mixed solvent of diisopropyl ether (2000 mL) and stirred at room temperature. The precipitate was collected by filtration and dried in vacuo to obtain a solid.
- R 1 has the structure of general formula (6)
- R 2 is an acetyl group
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 has the structure of general formula (5).
- R 5 is an isopropylaminocarbonylisopropylamino group
- R 11 and R 13 are methyl groups
- R 12 and R 14 are trimethylene groups
- X 1 is a bond
- X 2 is an —NH— group
- the average value of o and p is 272, a high molecular derivative having an average value of f + g + h + i + j + k + l + m + n of 43.2, an average value of f + g of 7.0, and an average value of h + i of 5.2 (compound 9)
- Compound 2 (245 mg), tacrolimus (250 mg), one-end methoxy group and one-end 3-aminopropyl group polyethylene glycol (SUNBRIGHT
- R 1 has the structure of general formula (6)
- R 2 is an acetyl group
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 has the structure of general formula (5).
- R 5 is an isopropylaminocarbonylisopropylamino group
- R 11 and R 13 are methyl groups
- R 12 and R 14 are trimethylene groups
- X 1 is a bond
- X 2 is an —NH— group
- the average value of o and p is 272, f + g + h + i + j + k + l + m + n mean value 72.8, f + g mean value 14.1 and h + i mean value 5.0 (compound 10)
- Compound 3 130 mg
- tacrolimus 240 mg
- one-end methoxy group and one-end 3-aminopropyl group polyethylene glycol SUVNBRIGHT MEPA-12T, manufactured by NOF Corporation, average molecular weight 12
- R 1 has the structure of general formula (6)
- R 2 is an acetyl group
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 has the structure of general formula (5).
- R 5 is an isopropylaminocarbonylisopropylamino group
- R 11 and R 13 are methyl groups
- R 12 and R 14 are trimethylene groups
- X 1 is a bond
- X 2 is an —NH— group
- the average value of o and p is 272
- a high molecular derivative (compound 11) having an average value of f + g + h + i + j + k + l + m + n of 94.9, an average value of f + g of 7.4, and an average value of h + i of 12.8
- Compound 1 (5.30 g)
- the reaction solution was added dropwise to diisopropyl ether (13 L) and stirred.
- the precipitate was collected by filtration and dried in vacuo to obtain a solid (41.5 g).
- the obtained solid (41.0 g) was dissolved in DMF (575 mL), an ion exchange resin (130 mL) was added, and the mixture was stirred for 1 hour, and then the ion exchange resin was filtered off.
- the filtrate was added dropwise to diisopropyl ether (18.5 L) and stirred.
- the precipitate was collected by filtration and vacuum dried to obtain a solid (39.5 g).
- the obtained solid (37.0 g) was dissolved in acetonitrile (370 mL), purified water (370 mL) was added, and the mixture was concentrated under reduced pressure and lyophilized to obtain compound 11 (37.2 g).
- the tacrolimus content of Compound 11 was calculated to be 3.22%.
- R 1 has the structure of general formula (6)
- R 2 is an acetyl group
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 has the structure of general formula (5).
- R 5 is an isopropylaminocarbonylisopropylamino group
- R 11 and R 13 are methyl groups
- R 12 and R 14 are trimethylene groups
- X 1 is a bond
- X 2 is an —NH— group
- the average value of o and p is 272
- a polymer derivative (compound 12) having an average value of 72.8, an average value of f + g + h + i + j + k + l + m + n, an average value of f + g of 5.5, and an average value of h + i of 9.1
- Compound 3 (4.50 g)
- tacrolimus (4.13 g)
- polyethylene glycol having one end methoxy group and one end 3-
- the reaction solution was added dropwise to diisopropyl ether (6.0 L) and stirred.
- the precipitate was collected by filtration, and the solid (28.8 g) obtained by vacuum drying was dissolved in DMF (404 mL).
- an ion exchange resin (95.1 mL) and stirring for 1 hour, the ion exchange resin was filtered off. did.
- the filtrate was added dropwise to diisopropyl ether (13.0 L) and stirred.
- the precipitate was collected by filtration, the solid obtained by vacuum drying was dissolved in acetonitrile (294 mL), purified water (294 mL) was added, and the mixture was concentrated under reduced pressure and lyophilized to give compound 12 (27.3 g).
- the tacrolimus content of Compound 12 was calculated to be 3.24%.
- Example 5 In the general formula (2), R 1 is an n-butyl group, R 2 is an acetyl group, R 3 is a residue of an alcoholic hydroxyl group of tacrolimus, R 4 has the structure of the general formula (5), and R 5 is isopropyl Aminocarbonylisopropylamino group, R 11 is methyl group, R 12 is trimethylene group, X 1 is bonded, X 2 is —NH— group, average value of o and p is 272, average value of f + g + h + i + j + k + l + m + n is 78, average of f + g Polymer derivative (compound 13) having a value of 7.4 and an average value of h + i of 10.1 Compound 4 (83.2 mg) and tacrolimus (150 mg) were dissolved in NMP (7.5 mL), DMAP (44.1 mg) and DIPCI (223 ⁇ L) were added at 25 ° C., and the mixture was
- Example 6 In the general formula (2), R 1 is an n-butyl group, R 2 is an acetyl group, R 3 is a residue of an alcoholic hydroxyl group of tacrolimus, R 4 has the structure of the general formula (5), and R 5 is isopropyl Aminocarbonylisopropylamino group, R 11 is methyl group, R 12 is trimethylene group, X 1 is bonded, X 2 is —NH— group, average value of o and p is 272, average value of f + g + h + i + j + k + l + m + n is 112, average of f + g Polymer derivative (compound 14) having a value of 10.4 and an average value of h + i of 14.6 Compound 5 (85.8 mg) and tacrolimus (150 mg) were dissolved in NMP (7.5 mL), DMAP (45.6 mg) and DIPCI (230 ⁇ L) were added at 25 ° C., and the mixture
- R 1 has the structure of general formula (6)
- R 2 is an acetyl group
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 has the structure of general formula (5).
- R 5 is an isopropylaminocarbonylisopropylamino group
- R 11 and R 13 are methyl groups
- R 12 and R 14 are trimethylene groups
- X 1 is a bond
- X 2 is an —NH— group
- the average value of o and p is 272
- polymer derivative (compound 15) having an average value of a + b + c + d + e of 40.2, an average value of a of 5.9, and an average value of b of 5.0
- Compound 7 (3.30 g)
- polyethylene glycol having one end methoxy group and one end 3-aminopropyl group (SUNBRIGHT MEPA-12T, manufactured by NOF Corporation, average mole
- the reaction solution was added dropwise to diisopropyl ether (4.8 L) and stirred.
- the precipitate was collected by filtration and dried in vacuo to give a solid (15.1 g).
- the obtained solid (15.0 g) was dissolved in DMF (210 mL), an ion exchange resin (50 mL) was added, and the mixture was stirred for 1 hour, and then the ion exchange resin was filtered off.
- the filtrate was added dropwise to diisopropyl ether (6.8 L) and stirred.
- the precipitate was collected by filtration and dried in vacuo to give a solid (14.2 g).
- the obtained solid (13.7 g) was dissolved in acetonitrile (275 mL), purified water (275 mL) was added, and the mixture was concentrated under reduced pressure and lyophilized to give compound 15 (12.4 g).
- the tacrolimus content of Compound 16 was calculated to be 5.74%.
- R 1 has the structure of general formula (6)
- R 2 is an acetyl group
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 has the structure of general formula (5).
- R 5 is an isopropylaminocarbonylisopropylamino group
- R 11 and R 13 are methyl groups
- R 12 and R 14 are trimethylene groups
- X 1 is a bond
- X 2 is an —NH— group
- the average value of o and p is 272
- polymer derivative (compound 16) having an average value of a + b + c + d + e of 87, an average value of a of 19.4, and an average value of b of 5.0
- Compound 6 157 mg
- tacrolimus 375 mg
- polyethylene glycol having one end methoxy group and one end 3-aminopropyl group
- SUNBRIGHT MEPA-12T manufactured by NOF Corporation, average molecular weight 12 k
- R 1 has the structure of general formula (6)
- R 2 is an acetyl group
- R 3 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 4 has the structure of general formula (5).
- R 5 is an isopropylaminocarbonylisopropylamino group
- R 11 and R 13 are methyl groups
- R 12 and R 14 are trimethylene groups
- R 13 is a methyl group
- R 14 is a trimethylene group
- X 1 is bonded
- X 2 is —NH— group
- polymer derivative having an average value of o and p of 272, an average value of a + b + c + d + e of 90.0, an average value of a of 21.6, and an average value of b of 5.3
- Compound 8 (3.0 g), tacrolimus (7.27 g), polyethylene glycol having one end methoxy group and one end 3-aminopropyl group (SUNBRIGHT MEPA-12T, manufactured
- the average value of t is 272
- the average value of q + r + s + u + v + w + x is 42.8
- the average value of q + r is 10.0
- R 15 is a residue of an alcoholic hydroxyl group of tacrolimus
- R 16 is an isopropylaminocarbonylisopropylamino group. is there.
- the tacrolimus content of the compound in the present invention was calculated from the consumption rate of tacrolimus in the reaction solution measured under the following HPLC conditions.
- Test example 1 Drug release test in phosphate buffered saline
- the compounds of Examples 1 to 9 and Comparative Example 1 were dissolved in phosphate buffered saline (pH 7.4) to a concentration of 1.0 mg / mL, and the temperature was adjusted to 37 ° C. And left at a constant temperature.
- the amount of released tacrolimus was measured over time by HPLC, and the ratio of the amount of released tacrolimus to the total amount of tacrolimus in the used compound was determined.
- the results of Example 1 and Comparative Example 1 are shown in FIG. 1, the results of Examples 1, 2, 5, 6 and 8 are shown in FIG. 2, and the results of Examples 3, 4, 7 and 9 are shown in FIG.
- Test example 2 Rat blood concentration transition (1) Tacrolimus or 5 mg / kg of the compounds of Examples 1, 2, 8 and Comparative Example 1 were administered to 8 week-old female SD rats (Nippon Charles River Co., Ltd.) in a single tail vein in each group. At 5 minutes, 1, 6, 24, and 72 hours after administration, 0.3 mL of blood was collected from the jugular vein under isoflurane anesthesia, and the tacrolimus concentration in the collected blood was measured. The results of Examples 1, 2, 8 and Comparative Example 1 and tacrolimus are shown in FIG. Table 2 shows the blood concentration parameters of each compound. However, the results of Examples 1, 2, 8 and Comparative Example 1 are the concentration and parameters of tacrolimus cut out from micelles.
- Examples 1, 2, 8 and Comparative Example 1 showed a half-life in blood concentration and MRTinf. It is clear that the retention in blood is improved by polymerizing tacrolimus. Further, in Examples 1, 2 and 8, compared to Comparative Example 1, the blood concentration half-life and MRTinf. showed that.
- Test example 3 Rat blood concentration transition (2) Eight-week-old female SD rats (Nippon Charles River Co., Ltd.) were administered a single tail vein intravenously with 15 mg / kg of the compounds of Examples 3, 7 and 9 in each group. Five minutes after administration, 1, 6, 24, and 72 hours later, 0.2 mL of blood was collected over time from the jugular vein under isoflurane anesthesia, and the tacrolimus concentration in the collected blood was measured. The results of Examples 3, 7, and 9 are shown in FIG. Table 3 shows the blood concentration parameters of each compound. However, the results of Examples 3, 7 and 9 are the concentration and parameters of tacrolimus cut out from micelles.
- Test example 4 Anti-inflammatory effect on rat collagen arthritis (1) Collagen arthritis was induced by intradermal administration of 0.3 mg of bovine articular cartilage-derived type II collagen (immune grade: Collagen Technical Training Co., Ltd.) to the back of 9-week-old female DA rats (Japan SLC, Inc.) . Type II collagen sensitization day and 7 days, 14 days, and 21 days after sensitization, physiological saline (5 mg / kg) of the compound of Comparative Example 1, Examples 1, 2 and 8 in each group 5 mice Administered. A non-administered group was set as a control. Arthritis was judged by visual scoring. The result of the compound of Comparative Example 1 is shown in FIG. 6, the result of the compound of Example 1 is shown in FIG. 7, and the results of the compounds of Examples 2 and 8 are shown in FIG. Table 4 shows the average arthritis score on the 28th or 29th day after collagen sensitization.
- Test Example 5 Anti-inflammatory effect on rat collagen-induced arthritis (2) Collagen arthritis was induced in DA rats in the same manner as in Test Example 4. On the day of type II collagen sensitization and 14 days after the sensitization, physiological saline (15 mg / kg) of the compounds of Examples 3, 7 and 9 was administered into the tail vein of 5 mice in each group. A non-administered group was set as a control. Arthritis was judged by visual scoring. The results are shown in FIG. Table 4 shows the average arthritis score on the 28th day after collagen sensitization.
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Abstract
Description
[1]ポリアスパラギン酸誘導体の側鎖カルボキシ基に、ポリエチレングリコールセグメント及びタクロリムスのアルコール性水酸基が結合しているタクロリムスの高分子誘導体。
[2]下記一般式(1)で表される前記[1]に記載のタクロリムスの高分子誘導体。
[3]下記一般式(2)で表される前記[1]に記載のタクロリムスの高分子誘導体。
[4]X1が結合である前記[2]又は[3]に記載のタクロリムスの高分子誘導体。
[5]X1がアスパラギン酸誘導体である前記[2]又は[3]に記載のタクロリムスの高分子誘導体。
[6]X1が、下記一般式(3)又は一般式(4)である前記[2]又は[3]に記載のタクロリムスの高分子誘導体。
[7]R8、R9が共に水素原子であり、及びCY-CZがCH-CHである前記[6]に記載のタクロリムスの高分子誘導体。
[8]R4のポリエチレングリコールセグメントが、下記一般式(5)である前記[2]乃至[7]の何れかの一項に記載のタクロリムスの高分子誘導体。
[9]R1が炭素数(C1~C6)のアルキル基又は下記一般式(6)で表されるポリエチレングルコールセグメントである前記[2]乃至[8]の何れかの一項に記載のタクロリムスの高分子誘導体。
[10]R2が炭素数(C1~C6)のアシル基であり、oが10~3000の整数であり、及び(a+b+c+d+e)又は(f+g+h+i+j+k+l+m+n)が4~250の整数である前記[8]又は[9]に記載のタクロリムスの高分子誘導体。
[11]R2が炭素数(C1~C3)のアシル基であり、oが20~1500の整数であり、及び(a+b+c+d+e)又は(f+g+h+i+j+k+l+m+n)が8~200の整数である前記[8]又は[9]に記載のタクロリムスの高分子誘導体。
[12]R1がメチル基であり、及びR2がアセチル基である前記[2]乃至[11]の何れか一項に記載のタクロリムスの高分子誘導体。
[13]ポリアスパラギン酸誘導体の側鎖のカルボキシ基に、タクロリムスのアルコール性水酸基及びポリエチレングリコールセグメントを、有機溶媒中、脱水縮合剤を用いて結合させることを特徴とする前記[1]乃至[12]の何れか一項に記載のタクロリムスの高分子誘導体の製造方法。
[14]前記[1]乃至[12]の何れか一項に記載のタクロリムスの高分子誘導体を有効成分とするマクロライド系免疫抑制剤。
直鎖状アルキル基としては、例えば、メチル基、エチル基、n-プロピル基、n-ブチル基、n-へキシル基等を挙げることができる。
分岐鎖状アルキル基としては、例えば、イソプロピル基、t-ブチル基、1-メチル-プロピル基、2-メチル-プロピル基、2,2-ジメチルプロピル基等が挙げられる。
環状アルキル基としては、例えば、シクロプロピル基、シクロブチル基、シクロペンチル基、シクロヘキシル基等が挙げられる。
置換基を有していても良い炭素数(C7~C20)の直鎖状、分岐鎖状又は環状のアラルキルアミノ基としては、例えば、ベンジルアミノ基、2-フェニルエチルアミノ基、4-フェニルブチルアミノ基、8-フェニルオクチルアミノ基等が挙げられる。
置換基を有していても良い炭素数(C5~C20)の芳香族アミノ基としては、例えば、アニリノ基、4-メトキシアニリノ基、4-ジメチルアミノアニリノ基、4-ヒドロキシアニリノ基等が挙げられる。
又、α-アスパラギン酸数(a)として1~200個、好ましくは2~150個程度、特に好ましくは2~100個程度、若しくは2~40個程度である。
又、α-アスパラギン酸数(b)として1~200個、好ましくは1~150個程度、特に好ましくは1~100個程度、若しくは2~40個程度である。
又、タクロリムスの結合したアスパラギン酸数(f+g)は1~200個、好ましくは2~150個程度、特に好ましくは2~100個程度、若しくは2~40個程度である。
又、タクロリムスの結合したアスパラギン酸数(h+i)は1~200個、好ましくは1~150個程度、特に好ましくは1~100個程度、若しくは2~40個程度である。
PEGの質量含有率(%)=PEG総分子量/TACの高分子誘導体分子量×100
注)PEG:ポリエチレングリコールセグメント
注)TAC:タクロリムス
TACの質量含有率(%)=TACの総分子量/TACの高分子誘導体分子量×100
注)TAC:タクロリムス
ポリエチレングリコール-αβ-ポリアスパラギン酸ブロック共重合体(ポリエチレングリコール分子量12000、ポリアスパラギン酸重合数95)の合成(化合物1)
片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、10.0g)をジメチルスルホキシド(DMSO、130mL)に溶解後、γ-ベンジル-L-アスパラギン酸-N-カルボン酸無水物(BLA-NCA、13.8g、200当量)を加え、32.0℃にて一夜攪拌した。反応液を、エタノール(755mL)及びジイソプロピルエーテル(3101L)の混合溶媒中に1時間かけて滴下し、室温にて1時間攪拌した。沈殿物を濾取後、真空乾燥し固形物(41.8g)を得た。
得られた固形物(40.0g)をN,N-ジメチルホルムアミド(DMF、800mL)に溶解し、無水酢酸(1.26mL)を加えて32.5℃にて撹拌した。反応液を、エタノール(800mL)及びジイソプロピルエーテル(7200mL)の混合溶媒中に滴下し、室温にて攪拌した。沈殿物を濾取後、真空乾燥し固形物(32.2g)を得た。得られた固形物(30g)をアセトニトリル(MeCN、1200mL)に懸濁後、0.2規定の水酸化ナトリウム水溶液(1200mL)及び精製水(180mL)を加えて、23℃にて加水分解を行った。反応液に2規定の塩酸を加えて中和したのち、減圧濃縮にてアセトニトリルを除去後、酢酸エチル(1800mL)を用い濃縮液を3回洗浄した。水層を減圧濃縮後、1規定の水酸化ナトリウム水溶液にて溶解液のpHを10.7に調整し、食塩(60.8g)を添加後、分配吸着樹脂カラムクロマトグラフィー及びイオン交換樹脂カラムクロマトグラフィーを用いて精製・脱塩し、溶出した溶液を減圧濃縮したのち、凍結乾燥し、化合物1(15.1g)を得た。0.1規定の水酸化カリウム水溶液を用いた滴定値に基づく本化合物1分子中のアスパラギン酸の重合数は94.9であった。
ポリエチレングリコール-αβ-ポリアスパラギン酸ブロック共重合体(ポリエチレングリコール分子量12000、ポリアスパラギン酸重合数43)の合成(化合物2)
合成例1記載の方法に準じ、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコールに対してBLA-NCAを52.2当量用いることにより、表記化合物2を得た。0.1規定の水酸化カリウム水溶液を用いた滴定値に基づく本化合物1分子中のアスパラギン酸の重合数は43.2であった。
ポリエチレングリコール-αβ-ポリアスパラギン酸ブロック共重合体(ポリエチレングリコール分子量12000、ポリアスパラギン酸重合数73)の合成(化合物3)
片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、10.0g)をDMSO(190mL)に溶解後、BLA-NCA(8.89g、129当量)を加え、32℃にて一夜攪拌した。反応液を、エタノール(400mL)及びジイソプロピルエーテル(1600mL)の混合溶媒中に1時間かけて滴下し、室温にて1時間攪拌した。沈殿物を濾取後、真空乾燥し固形物(26.5g)を得た。
得られた固形物(23.0g)をアセトニトリル(460mL)に溶解し、無水酢酸(724mL)を加えて35℃にて3時間撹拌した後23℃まで冷却した.反応液に0.2規定の水酸化ナトリウム(539mL)を滴下し、23℃にて加水分解を行った。反応液に2規定の塩酸を加えて中和したのち、減圧濃縮にてアセトニトリルを除去後、酢酸エチル(720mL)を用い濃縮液を3回洗浄した。水層を減圧濃縮後、1規定の水酸化ナトリウム水溶液にて溶解液のpHを10.6に調製し、食塩(57.5g)を添加後、分配吸着樹脂カラムクロマトグラフィー、続いてイオン交換樹脂カラムクロマトグラフィーを用いて精製し、溶出した溶液を減圧濃縮したのち、凍結乾燥し、化合物3(13.0g)を得た。0.1規定の水酸化カリウム水溶液を用いた滴定値に基づく本化合物1分子中のアスパラギン酸の重合数は72.8であった。
αβ-ポリアスパラギン酸重合体(ポリアスパラギン酸重合数78)の合成(化合物4)
n-ブチルアミン(東京化成製、18.3mg)をDMSO(30mL)に溶解後、BLA-NCA(5.61g、90当量)を加え、30℃にて一夜攪拌した。反応液を、エタノール(120mL)及びジイソプロピルエーテル(480mL)の混合溶媒中に滴下し、室温にて攪拌した。沈殿物を濾取後、真空乾燥し固形物を得た。得られた固形物全量に1,3-ジメチル-2-イミダゾリジノン(DMI、35mL)を加え、78℃にて溶解後、70℃にて無水酢酸2.5mLを加えた。3時間撹拌後、反応液を酢酸エチル(175mL)及びジイソプロピルエーテル(700mL)の混合溶液中に滴下し、室温にて一夜撹拌した.沈析物を濾取後、真空乾燥し固形物(3.74g)を得た。
得られた固形物(3.74g)をアセトニトリル(30mL)に溶解し、0.2規定の水酸化ナトリウム水溶液(138mL)を添加し、室温にて加水分解を行った。反応後、減圧濃縮にてアセトニトリルを除去後、酢酸エチル(420mL)を用い濃縮液を3回洗浄した。水層を減圧濃縮後、イオン交換樹脂カラムクロマトグラフィーを用いて精製し、凍結乾燥を行い、化合物4(2.29g)を得た。NMRに基づく本化合物1分子中のアスパラギン酸の重合数は78であった。
αβ-ポリアスパラギン酸重合体(ポリアスパラギン酸重合数112)の合成(化合物5)
n-ブチルアミン(東京化成製、15.8mg)をDMSO(30mL)に溶解後、BLA-NCA(7.26g、135当量)を加え、30℃にて一夜攪拌した。反応液を、エタノール(120mL)及びジイソプロピルエーテル(480mL)の混合溶媒中に滴下し、室温にて攪拌した。沈殿物を濾取後、真空乾燥し固形物(5.37g)を得た。得られた固形物(5.37g)にDMI(35mL)を加え、80℃にて溶解後、68℃にて無水酢酸2.5mLを加えた。3時間撹拌後、反応液を酢酸エチル(200mL)及びジイソプロピルエーテル(800mL)の混合溶液中に滴下し、室温にて一夜撹拌した.沈析物を濾取後、真空乾燥し固形物(5.16g)を得た。
得られた固形物(5.16g)をアセトニトリル(22mL)に溶解し、0.2規定の水酸化ナトリウム水溶液(167mL)を添加し、室温にて加水分解を行った。反応後、減圧濃縮にてアセトニトリルを除去後、酢酸エチル(510mL)を用い濃縮液を3回洗浄した。水層を減圧濃縮後、イオン交換樹脂カラムクロマトグラフィーを用いて精製し、凍結乾燥を行い、化合物5(3.00g)を得た。NMRに基づく本化合物1分子中のアスパラギン酸の重合数は112であった。
ポリエチレングリコール-α-ポリアスパラギン酸ブロック共重合体(ポリエチレングリコール分子量12000、ポリアスパラギン酸重合数87)の合成(化合物6)
片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、6.0g)をDMSO(114mL)に溶解後、BLA-NCA(15.0g、120当量)を加え、30℃にて一夜攪拌した。反応液を、エタノール(228mL)及びジイソプロピルエーテル(912mL)の混合溶媒中に滴下し、室温にて攪拌した。沈殿物を濾取後、真空乾燥し固形物(23.8g)を得た。得られた固形物全量をDMF(200mL)に溶解し、無水酢酸(0.75mL)を加えて30℃にて3時間撹拌した。反応液を、酢酸エチル(200mL)及びジイソプロピルエーテル(1800mL)の混合溶媒中に滴下し、室温にて攪拌した。沈殿物を濾取後、真空乾燥し固形物(15.8g)を得た。
得られた固形物(8.72g)を1-メチル-2-ピロリドン(NMP、140mL)に溶解後、10%パラジウム-炭素(0.87g)を加えて、35℃にて一夜加水素分解を行った。反応液に活性炭を加えて1時間撹拌したのち、10%パラジウム-炭素を濾別した。濾液を、酢酸エチル(200mL)及びジイソプロピルエーテル(1000mL)の混合溶媒中に滴下し、室温にて一夜攪拌した。沈殿物を濾取後、真空乾燥し固形物(4.18g)を得た。固形物全量を精製水(418mL)に溶解し、食塩(20.9g)を添加、溶解し、2規定の水酸化ナトリウム水溶液にて溶解液のpHを11.0に調製後、分配吸着樹脂カラムクロマトグラフィー、続いてイオン交換樹脂カラムクロマトグラフィーを用いて精製し、溶出した溶液を減圧濃縮したのち、凍結乾燥することによって、化合物6(760mg)を得た。NMRに基づく本化合物1分子中のアスパラギン酸の重合数は87であった。
ポリエチレングリコール-α-ポリアスパラギン酸ブロック共重合体(ポリエチレングリコール分子量12000、ポリアスパラギン酸重合数40)の合成(化合物7)
合成例6記載の方法に準じ、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコールに対してBLA-NCAを51.3当量用いることにより、表記化合物7を得た。この際、NMPの代わりにDMFを使用した。0.1規定の水酸化カリウム水溶液を用いた滴定値に基づく本化合物1分子中のアスパラギン酸の重合数は40.2であった。
ポリエチレングリコール-α-ポリアスパラギン酸ブロック共重合体(ポリエチレングリコール分子量12000、ポリアスパラギン酸重合数90)の合成(化合物8)
片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、10.0g)をDMSO(200mL)に溶解後、BLA-NCA(28.7g、140当量)を加え、32.5℃にて一夜攪拌した。反応液を、エタノール(400mL)及びジイソプロピルエーテル(1600mL)の混合溶媒中に滴下し、室温にて攪拌した。沈殿物を濾取後、真空乾燥し固形物(28.6g)を得た。得られた固形物(18.1g)をNMP(181mL)に溶解し、無水酢酸(0.57mL)を加えて30℃にて一夜撹拌した。反応液を、ジイソプロピルエーテル(2000mL)の混合溶媒中に滴下し、室温にて攪拌した。沈殿物を濾取後、真空乾燥し固形物を得た。
得られた固形物(15.3g)をNMP(155mL)に溶解後、10%パラジウム-炭素(3.46g)を加えて、35℃にて一夜加水素分解を行った。反応液に活性炭を加えて30分間撹拌したのち、10%パラジウム-炭素を濾別した。濾液を、ジイソプロピルエーテル(3700mL)の混合溶媒中に滴下し、室温にて攪拌した。沈殿物を濾取後、真空乾燥し固形物(5.64g)を得た。得られた固形物(4.53g)を精製水(453mL)に溶解し、水酸化ナトリウム水溶液にて溶解液のpHを11.0に調製後、陰イオン交換樹脂カラムクロマトグラフィー、続いて陽イオン交換樹脂カラムクロマトグラフィーを用いて精製し、溶出した溶液を減圧濃縮したのち、凍結乾燥することによって、化合物8(3.43g)を得た。0.1規定の水酸化カリウム水溶液を用いた滴定値に基づく本化合物1分子中のアスパラギン酸の重合数は90.0であった。
一般式(2)においてR1として一般式(6)の構造を有し、R2がアセチル基、R3がタクロリムスのアルコール性水酸基の残基、R4として一般式(5)の構造を有し、R5がイソプロピルアミノカルボニルイソプロピルアミノ基、R11及びR13がメチル基、R12及びR14がトリメチレン基、X1が結合、X2が-NH-基、o及びpの平均値が272、f+g+h+i+j+k+l+m+nの平均値が43.2、f+gの平均値が7.0、h+iの平均値が5.2である高分子誘導体(化合物9)
化合物2(245mg)、タクロリムス(250mg)、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、862mg)をNMP(6.2mL)に溶解し、25℃にてN,N-ジメチルアミノピリジン(DMAP、37.9mg)、N,N’-ジイソプロピルカルボジイミド(DIPCI、192μL)を加えて一夜撹拌した。反応液をジイソプロピルエーテル(186mL)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により得られた固形物をアセトニトリル(7mL)に溶解後、精製水(7mL)及びイオン交換樹脂(7mL)を加え、撹拌した後、イオン交換樹脂を濾別した。濾液を減圧濃縮した後、凍結乾燥することによって、化合物9(1.12g)を得た。化合物9のタクロリムス含量は6.84%と計算された。
一般式(2)においてR1として一般式(6)の構造を有し、R2がアセチル基、R3がタクロリムスのアルコール性水酸基の残基、R4として一般式(5)の構造を有し、R5がイソプロピルアミノカルボニルイソプロピルアミノ基、R11及びR13がメチル基、R12及びR14がトリメチレン基、X1が結合、X2が-NH-基、o及びpの平均値が272、f+g+h+i+j+k+l+m+nの平均値が72.8、f+gの平均値が14.1、h+iの平均値が5.0である高分子誘導体(化合物10)
化合物3(130mg)、タクロリムス(240mg)、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、382mg)をNMP(4.6mL)に溶解し、25℃にてDMAP(28.3mg)、DIPCI(143μL)を加えて一夜撹拌した。反応液をジイソプロピルエーテル(139mL)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により得られた固形物をアセトニトリル(5mL)に溶解後、精製水(5mL)及びイオン交換樹脂(5mL)を加え、撹拌した後、イオン交換樹脂を濾別した。濾液を減圧濃縮した後、凍結乾燥することによって、化合物10(528mg)を得た。化合物10のタクロリムス含量は12.3%と計算された。
一般式(2)においてR1として一般式(6)の構造を有し、R2がアセチル基、R3がタクロリムスのアルコール性水酸基の残基、R4として一般式(5)の構造を有し、R5がイソプロピルアミノカルボニルイソプロピルアミノ基、R11及びR13がメチル基、R12及びR14がトリメチレン基、X1が結合、X2が-NH-基、o及びpの平均値が272、f+g+h+i+j+k+l+m+nの平均値が94.9、f+gの平均値が7.4、h+iの平均値が12.8である高分子誘導体(化合物11)
化合物1(5.30g)、タクロリムス(12g)、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、34.7g)をNMP(430mL)に溶解し、25℃にてDMAP(1.31g)、DIPCI(6.59mL)を加えて二夜撹拌した。反応液をジイソプロピルエーテル(13L)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により固形物(41.5g)を得た。得られた固形物(41.0g)をDMF(575mL)に溶解し、イオン交換樹脂(130mL)を加え1時間撹拌後、イオン交換樹脂を濾別した。濾液をジイソプロピルエーテル(18.5L)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により固形物(39.5g)を得た。得られた固形物(37.0g)をアセトニトリル(370mL)に溶解後、精製水(370mL)を加え、減圧濃縮した後、凍結乾燥することによって、化合物11(37.2g)を得た。化合物11のタクロリムス含量は3.22%と計算された。
一般式(2)においてR1として一般式(6)の構造を有し、R2がアセチル基、R3がタクロリムスのアルコール性水酸基の残基、R4として一般式(5)の構造を有し、R5がイソプロピルアミノカルボニルイソプロピルアミノ基、R11及びR13がメチル基、R12及びR14がトリメチレン基、X1が結合、X2が-NH-基、o及びpの平均値が272、f+g+h+i+j+k+l+m+nの平均値が72.8、f+gの平均値が5.5、h+iの平均値が9.1である高分子誘導体(化合物12)
化合物3(4.50g)、タクロリムス(4.13g)、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、24.1g)をNMP(229mL)に溶解し、25℃にてDMAP(979mg)、DIPCI(4.97mL)を加えて二夜撹拌した。反応液をジイソプロピルエーテル(6.0L)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により得られた固形物(28.8g)をDMF(404mL)に溶解し、イオン交換樹脂(95.1mL)を加え1時間撹拌後、イオン交換樹脂を濾別した。濾液をジイソプロピルエーテル(13.0L)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により得られた固形物をアセトニトリル(294mL)に溶解後、精製水(294mL)を加え、減圧濃縮した後、凍結乾燥することによって、化合物12(27.3g)を得た。化合物12のタクロリムス含量は3.24%と計算された。
一般式(2)においてR1がn-ブチル基、R2がアセチル基、R3がタクロリムスのアルコール性水酸基の残基、R4として一般式(5)の構造を有し、R5がイソプロピルアミノカルボニルイソプロピルアミノ基、R11がメチル基、R12がトリメチレン基、X1が結合、X2が-NH-基、o及びpの平均値が272、f+g+h+i+j+k+l+m+nの平均値が78、f+gの平均値が7.4、h+iの平均値が10.1である高分子誘導体(化合物13)
化合物4(83.2mg)、タクロリムス(150mg)をNMP(7.5mL)に溶解し、25℃にてDMAP(44.1mg)、DIPCI(223μL)を加えて1時間撹拌後、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、1.12g)を添加し35℃に昇温し一夜撹拌した。反応液をジイソプロピルエーテル(225mL)に滴下し、一夜撹拌した。沈殿物を濾取し、真空乾燥により得られた固形物をアセトニトリル(40mL)に溶解し、精製水(40mL)を加え、イオン交換樹脂(20mL)を加え、氷冷化で撹拌後、イオン交換樹脂を濾別した。濾液を減圧濃縮した後、凍結乾燥することによって、化合物13(1.16g)を得た。化合物13のタクロリムス含量は4.40%と計算された。
一般式(2)においてR1がn-ブチル基、R2がアセチル基、R3がタクロリムスのアルコール性水酸基の残基、R4として一般式(5)の構造を有し、R5がイソプロピルアミノカルボニルイソプロピルアミノ基、R11がメチル基、R12がトリメチレン基、X1が結合、X2が-NH-基、o及びpの平均値が272、f+g+h+i+j+k+l+m+nの平均値が112、f+gの平均値が10.4、h+iの平均値が14.6である高分子誘導体(化合物14)
化合物5(85.8mg)、タクロリムス(150mg)をNMP(7.5mL)に溶解し、25℃にてDMAP(45.6mg)、DIPCI(230μL)を加えて1時間撹拌後、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、1.12g)を添加し35℃に昇温し一夜撹拌した。反応液をジイソプロピルエーテル(225mL)に滴下し、一夜撹拌した。沈殿物を濾取し、真空乾燥により得られた固形物をアセトニトリル(40mL)に溶解し、精製水(40mL)を加え、イオン交換樹脂(20mL)を加え、氷冷化で撹拌後、イオン交換樹脂を濾別した。濾液を減圧濃縮した後、凍結乾燥することによって、化合物14(1.18g)を得た。化合物14のタクロリムス含量は4.41%と計算された。
一般式(1)においてR1として一般式(6)の構造を有し、R2がアセチル基、R3がタクロリムスのアルコール性水酸基の残基、R4として一般式(5)の構造を有し、R5がイソプロピルアミノカルボニルイソプロピルアミノ基、R11及びR13がメチル基、R12及びR14がトリメチレン基、X1が結合、X2が-NH-基、o及びpの平均値が272、a+b+c+d+eの平均値が40.2、aの平均値が5.9、bの平均値が5.0である高分子誘導体(化合物15)
化合物7(3.30g)、タクロリムス(4.81g)、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、11.9g)をNMP(160mL)に溶解し、35℃にてDMAP(500mg)、DIPCI(2.47mL)を加えて二夜撹拌した。反応液をジイソプロピルエーテル(4.8L)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により固形物(15.1g)を得た。得られた固形物(15.0g)をDMF(210mL)に溶解し、イオン交換樹脂(50mL)を加え1時間撹拌後、イオン交換樹脂を濾別した。濾液をジイソプロピルエーテル(6.8L)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により固形物(14.2g)を得た。得られた固形物(13.7g)をアセトニトリル(275mL)に溶解後、精製水(275mL)を加え、減圧濃縮した後、凍結乾燥することによって、化合物15(12.4g)を得た。化合物16のタクロリムス含量は5.74%と計算された。
一般式(1)においてR1として一般式(6)の構造を有し、R2がアセチル基、R3がタクロリムスのアルコール性水酸基の残基、R4として一般式(5)の構造を有し、R5がイソプロピルアミノカルボニルイソプロピルアミノ基、R11及びR13がメチル基、R12及びR14がトリメチレン基、X1が結合、X2が-NH-基、o及びpの平均値が272、a+b+c+d+eの平均値が87、aの平均値が19.4、bの平均値が5.0である高分子誘導体(化合物16)
化合物6(157mg)、タクロリムス(375mg)、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、425mg)をNMP(5.5mL)に溶解し、35℃にてDMAP(38.0mg)、DIPCI(191μL)を加えて二夜撹拌した。反応液をジイソプロピルエーテル(165mL)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により得られた固形物をアセトニトリル(13mL)に溶解し、精製水(13mL)を加え、イオン交換樹脂(6.5mL)を加え、氷冷化で撹拌後、イオン交換樹脂を濾別した。濾液を減圧濃縮した後、凍結乾燥することによって、化合物16(630mg)を得た。化合物16のタクロリムス含量は14.9%と計算された。
一般式(1)においてR1として一般式(6)の構造を有し、R2がアセチル基、R3がタクロリムスのアルコール性水酸基の残基、R4として一般式(5)の構造を有し、R5がイソプロピルアミノカルボニルイソプロピルアミノ基、R11及びR13がメチル基、R12及びR14がトリメチレン基、R13がメチル基、R14がトリメチレン基、X1が結合、X2が-NH-基、o及びpの平均値が272、a+b+c+d+eの平均値が90.0、aの平均値が21.6、bの平均値が5.3である高分子誘導体(化合物17)
化合物8(3.0g)、タクロリムス(7.27g)、片末端メトキシ基及び片末端3-アミノプロピル基のポリエチレングリコール(SUNBRIGHT MEPA-12T、日油社製、平均分子量12キロダルトン、8.50g)をNMP(80.3mL)に溶解し、35℃にてDMAP(733mg)、DIPCI(3.72mL)を加えて二夜撹拌した。反応液をジイソプロピルエーテル(3150mL)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により得られた固形物(14.0g)をDMF(196mL)に溶解し、イオン交換樹脂(46mL)を加え撹拌後、イオン交換樹脂を濾別した。濾液をジイソプロピルエーテル(6.3L)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥することによって、化合物17(12.9g)を得た。化合物17のタクロリムス含量は16.6%と計算された。
タクロリムス結合ポリエチレングリコール-αβ-ポリアスパラギン酸ブロック共重合体(化合物18)
化合物2(350mg)、タクロリムス(366mg)をDMF(6.0mL)に溶解し、15℃にてDMAP(10.9mg)、DIPCI(315μL)を加えて一夜撹拌した。反応液をジイソプロピルエーテル(180mL)に滴下し、撹拌した。沈殿物を濾取し、真空乾燥により得られた固形物をアセトニトリル(17mL)に溶解後、精製水(17mL)及びイオン交換樹脂(7mL)を加え、氷冷化で3時間撹拌後、イオン交換樹脂を濾別した。濾液を減圧濃縮した後、凍結乾燥することによって、下記式(12)で表される化合物18(499mg)を得た。化合物18のタクロリムス含量は33.2%と計算された。
カラム :Shim-pack XR-ODSIII、2.0φ×200mm
カラム温度:40℃
溶離液 A液:0.1%リン酸水溶液、B液:アセトニトリル
A液/B液=80/20
流 速 :0.5mL/分
検出器(検出波長):UV(254nm)
カラム :Shim-pack XR-ODSIII、2.0φ×200mm
カラム温度:40℃
溶離液 A液:0.1%リン酸水溶液、B液:アセトニトリル
A液/B液=80/20から10/90へのグラジエント溶出
流 速 :0.5mL/分
検出器(検出波長):蛍光(励起波長:450nm、蛍光波長:590nm)
リン酸緩衝生理食塩水中の薬剤放出性試験
実施例1~9、比較例1の化合物をリン酸緩衝生理食塩水(pH7.4)に1.0mg/mLとなるように溶解し、37℃にて定温放置した。放出されたタクロリムス量をHPLCにて経時的に測定し、使用した化合物中の全タクロリムス量に対する放出されたタクロリムス量の割合を求めた。実施例1及び比較例1の結果を図1に、実施例1、2、5、6及び8の結果を図2に、実施例3、4、7及び9の結果を図3に示す。
ラット血中濃度推移(1)
8週齢雌性SDラット(日本チャールズ・リバー株式会社)にタクロリムスまたは実施例1、2、8及び比較例1の化合物5mg/kgを各群2匹ずつ単回尾静脈内投与した。投与後5分、1、6、24及び72時間にイソフルラン麻酔下で頸静脈から継時的に0.3mLずつ採血し、採取血液中のタクロリムス濃度を測定した。実施例1、2、8、比較例1及びタクロリムスの結果を図4に示す。また、各化合物の血中濃度パラメータを表2に示す。ただし実施例1、2、8及び比較例1の結果はミセルから切り出されたタクロリムスの濃度及びパラメータである。
また、比較例1と比べ実施例1、2及び8では、より長い血中濃度半減期及びMRTinf.を示した。
ラット血中濃度推移(2)
8週齢雌性SDラット(日本チャールズ・リバー株式会社)に実施例3、7及び9の化合物15mg/kgを各群2匹ずつ単回尾静脈内投与した。投与後5分、1、6、24及び72時間後にイソフルラン麻酔下で頸静脈から継時的に0.2mLずつ採血し、採取血液中のタクロリムス濃度を測定した。実施例3、7及び9の結果を図5に示す。また、各化合物の血中濃度パラメータを表3に示す。ただし、実施例3、7及び9の結果はミセルから切り出されたタクロリムスの濃度及びパラメータである。
ラットコラーゲン関節炎に対する抗炎症効果(1)
ウシ関節軟骨由来タイプIIコラーゲン(免疫グレード:コラーゲン技術研修会有限会社)0.3mgを9週齢雌性DAラット(日本エスエルシー株式会社)の背部に皮内投与することにより、コラーゲン関節炎を誘発した。タイプIIコラーゲン感作日及び感作後7日、14日、21日目に比較例1、実施例1、2及び8の化合物の生理食塩水(5mg/kg)を各群5匹ずつ尾静脈内に投与した。対照としては未投与群を設定した。関節炎の判定は、目視によるスコア化により行った。比較例1の化合物の結果を図6に、実施例1の化合物の結果を図7に、実施例2及び8の化合物の結果を図8にそれぞれ示す。また、コラーゲン感作後28又は29日目の平均関節炎スコアを表4に示す。
ラットコラーゲン誘導関節炎に対する抗炎症効果(2)
試験例4と同様の方法で、DAラットにコラーゲン関節炎を誘発した。タイプIIコラーゲン感作日及び感作後14日目に実施例3、7及び9の化合物の生理食塩水(15mg/kg)を各群5匹ずつ尾静脈内に投与した。対照としては未投与群を設定した。関節炎の判定は、目視によるスコア化により行った。結果を図9に示す。また、コラーゲン感作後28日目の平均関節炎スコアを表4に示す。
Claims (14)
- ポリアスパラギン酸誘導体の側鎖カルボキシ基に、ポリエチレングリコールセグメント及びタクロリムスのアルコール性水酸基が結合しているタクロリムスの高分子誘導体。
- 下記一般式(1)で表される請求項1に記載のタクロリムスの高分子誘導体。
- 下記一般式(2)で表される請求項1に記載のタクロリムスの高分子誘導体。
- X1が結合である請求項2又は請求項3に記載のタクロリムスの高分子誘導体。
- X1がアスパラギン酸誘導体である請求項2又は請求項3に記載のタクロリムスの高分子誘導体。
- X1が、下記一般式(3)又は一般式(4)である請求項2又は請求項3に記載のタクロリムスの高分子誘導体。
- R8、R9が共に水素原子であり、及びCY-CZがCH-CHである請求項6に記載のタクロリムスの高分子誘導体。
- R2が炭素数(C1~C6)のアシル基であり、oが10~3000の整数であり、及び(a+b+c+d+e)又は(f+g+h+i+j+k+l+m+n)が4~250の整数である請求項8又は9に記載のタクロリムスの高分子誘導体。
- R2が炭素数(C1~C3)のアシル基であり、oが20~1500の整数であり、及び(a+b+c+d+e)又は(f+g+h+i+j+k+l+m+n)が8~200の整数である請求項8又は9に記載のタクロリムスの高分子誘導体。
- R1がメチル基であり、及びR2がアセチル基である請求項2乃至11の何れか一項に記載のタクロリムスの高分子誘導体。
- ポリアスパラギン酸誘導体の側鎖のカルボキシ基に、タクロリムスのアルコール性水酸基及びポリエチレングリコールセグメントを、有機溶媒中、脱水縮合剤を用いてエステル結合、アミド結合及び/又はチオエステル結合させることを特徴とする請求項1乃至12の何れか一項に記載のタクロリムスの高分子誘導体の製造方法。
- 請求項1乃至12の何れか一項に記載のタクロリムスの高分子誘導体を有効成分とするマクロライド系免疫抑制剤。
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Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2000502109A (ja) * | 1995-12-21 | 2000-02-22 | ドイチェス クレブスフォルシュンクスツェントルム スチフトゥング デス エッフェントリヒェン レヒツ | 作用物質、ポリペプチドおよびポリエーテルを含有してなるコンジュゲート |
WO2005079861A2 (en) * | 2004-02-13 | 2005-09-01 | Safeway Investments Ltd. | Polymeric water soluble prodrugs |
WO2007111211A1 (ja) * | 2006-03-28 | 2007-10-04 | Nippon Kayaku Kabushiki Kaisha | タキサン類の高分子結合体 |
WO2010131675A1 (ja) * | 2009-05-15 | 2010-11-18 | 日本化薬株式会社 | 水酸基を有する生理活性物質の高分子結合体 |
WO2016021407A1 (ja) * | 2014-08-04 | 2016-02-11 | 日本化薬株式会社 | 核酸代謝拮抗剤が結合したポリアミノ酸誘導体 |
Family Cites Families (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5247076A (en) | 1991-09-09 | 1993-09-21 | Merck & Co., Inc. | Imidazolidyl macrolides having immunosuppressive activity |
KR100244164B1 (ko) | 1997-07-15 | 2000-03-02 | 김용옥 | 수용성 고분자-타크로리무스 접합체 화합물 및 그의 제조 방법 |
JP4462928B2 (ja) | 2001-06-20 | 2010-05-12 | 日本化薬株式会社 | 不純物含有量の低減したブロック共重合体、高分子担体及び高分子医薬製剤並びにその製造方法 |
PL1604687T3 (pl) | 2003-03-20 | 2011-04-29 | Nippon Kayaku Kk | Preparat micelarny zawierający słabo rozpuszczalny w wodzie lek przeciwnowotworowy oraz nowy kopolimer blokowy |
US20060134166A1 (en) * | 2004-05-25 | 2006-06-22 | Luthra Ajay K | Absorbable biocompatible materials |
CN101218280B (zh) | 2005-05-11 | 2012-06-27 | 日本化药株式会社 | 胞苷代谢拮抗剂的聚合衍生物 |
JP5548364B2 (ja) | 2006-10-03 | 2014-07-16 | 日本化薬株式会社 | レゾルシノール誘導体の高分子結合体 |
AR090736A1 (es) | 2012-04-20 | 2014-12-03 | Sucampo Ag | Conjugado de compuesto triciclo-polimero |
AU2016224760B2 (en) | 2015-02-23 | 2021-01-28 | Nippon Kayaku Kabushiki Kaisha | Block copolymer conjugate of physiologically active substance |
JP6851977B2 (ja) | 2015-11-18 | 2021-03-31 | 日本化薬株式会社 | マクロライド系免疫抑制剤の高分子誘導体 |
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Patent Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2000502109A (ja) * | 1995-12-21 | 2000-02-22 | ドイチェス クレブスフォルシュンクスツェントルム スチフトゥング デス エッフェントリヒェン レヒツ | 作用物質、ポリペプチドおよびポリエーテルを含有してなるコンジュゲート |
WO2005079861A2 (en) * | 2004-02-13 | 2005-09-01 | Safeway Investments Ltd. | Polymeric water soluble prodrugs |
WO2007111211A1 (ja) * | 2006-03-28 | 2007-10-04 | Nippon Kayaku Kabushiki Kaisha | タキサン類の高分子結合体 |
WO2010131675A1 (ja) * | 2009-05-15 | 2010-11-18 | 日本化薬株式会社 | 水酸基を有する生理活性物質の高分子結合体 |
WO2016021407A1 (ja) * | 2014-08-04 | 2016-02-11 | 日本化薬株式会社 | 核酸代謝拮抗剤が結合したポリアミノ酸誘導体 |
Non-Patent Citations (3)
Title |
---|
CHOI, D. ET AL.: "Effects of water-soluble tacrolimus-PEG conjugate on insulin-dependent diabetes mellitus and systemic lupus erythematosus", ARCHIVES OF PHARMACAL RESEARCH,, vol. 34, no. 8, 11 September 2011 (2011-09-11), pages 1301 - 1310, XP019950689 * |
CHUNG, Y. ET AL.: "Preparation of highly water soluble tacrolimus derivatives: poly(ethylene glycol) esters as potential prodrugs", ARCH. PHARM. RES., vol. 27, no. 8, August 2004 (2004-08-01), pages 878 - 883, XP055383555 * |
See also references of EP3400945A4 * |
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