WO2014114262A1 - Water soluble polymer-amino acid oligopeptide-medicine combination, preparation method therefore, and use thereof - Google Patents

Water soluble polymer-amino acid oligopeptide-medicine combination, preparation method therefore, and use thereof Download PDF

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Publication number
WO2014114262A1
WO2014114262A1 PCT/CN2014/071395 CN2014071395W WO2014114262A1 WO 2014114262 A1 WO2014114262 A1 WO 2014114262A1 CN 2014071395 W CN2014071395 W CN 2014071395W WO 2014114262 A1 WO2014114262 A1 WO 2014114262A1
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group
cancer
acid
conjugate
tumor
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PCT/CN2014/071395
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French (fr)
Chinese (zh)
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汪进良
赵宣
王振国
冯泽旺
贾健欢
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天津键凯科技有限公司
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Priority claimed from CN201310241907.2A external-priority patent/CN103965458B/en
Application filed by 天津键凯科技有限公司 filed Critical 天津键凯科技有限公司
Priority to US14/764,166 priority Critical patent/US9700633B2/en
Publication of WO2014114262A1 publication Critical patent/WO2014114262A1/en

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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
    • C08G65/00—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule
    • C08G65/02—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring
    • C08G65/32—Polymers modified by chemical after-treatment
    • C08G65/329—Polymers modified by chemical after-treatment with organic compounds
    • C08G65/333—Polymers modified by chemical after-treatment with organic compounds containing nitrogen
    • C08G65/33396—Polymers modified by chemical after-treatment with organic compounds containing nitrogen having oxygen in addition to nitrogen
    • 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
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08L—COMPOSITIONS OF MACROMOLECULAR COMPOUNDS
    • C08L2203/00—Applications
    • C08L2203/02—Applications for biomedical use

Definitions

  • the present application provides a water-soluble polymer-amino acid oligopeptide-drug conjugate and a pharmaceutical composition thereof, and a preparation method and use of the conjugate and the pharmaceutical composition.
  • the conjugate increases the therapeutic effect of the drug on the disease by increasing the drug loading of the active drug.
  • the stability of the combination is also greatly improved.
  • the present invention provides a water-soluble polymer-amino acid oligopeptide-drug combination having The structure shown by the following formula (I),
  • P is a water soluble polymer
  • X is a linking group, the linking group linking P and A 1 ;
  • ALA 2 and A 3 are each independently the same or different amino acid or amino acid analog residues
  • Di and D 2 are each independently the same or different drug molecule residues
  • a is 0 or 1
  • b is an integer from 2 to 12;
  • c is an integer from 0 to 7;
  • d is 0 or 1.
  • P is selected from the group consisting of polyethylene glycol, polypropylene glycol, polyglutamic acid, polyaspartic acid, polyvinylpyrrolidone, polyvinyl alcohol, polypropylene morpholine, Dextran, carboxymethylcellulose and analogues or copolymers thereof.
  • P is polyethylene glycol
  • the polyethylene glycol has a molecular weight of 300-60,000.
  • the polyethylene glycol has a molecular weight of 20,000-40. 000.
  • the polyethylene glycol has a molecular weight of 20,000, 21,000, 22,000, 23,000, 24,000, 25,000, 26,000, 27,000, 28,000, 29,000, 30,000, 31,000, 32,000, 33,000, 34,000, 35,000, 36,000, 37,000, 38,000, 39,000, 40,000.
  • the polyethylene glycol is a linear, Y-branched or multi-branched polyethylene glycol.
  • the polyethylene glycol has a structure represented by the following formula ( ⁇ ),
  • ⁇ is alkyl with 12, 12 D_ embankment heteroaryl group, or aryl alkyl with hydrogen, e is an integer of 10-1,500.
  • P has the structure shown by the following formula ( ⁇ ), (m)
  • R 2 and R 3 are each independently d— 12 fluorenyl, d— 12 heterofluorenyl, hydrogen, aryl fluorenyl;
  • R 4 and R 5 are each independently a C 1-12 fluorenyl group, a C 1-12 fluorenylcarbonyl group;
  • f and g are each independently an integer of from 10 to 1,500.
  • R 2 , R 3 in formula (III) are cyclopropyl, cyclobutyl, cyclohexyl
  • R 2 , R 3 in formula (III) are methyl, F is ethyl, and R 5 is
  • P has the structure shown by formula (IV) below,
  • R 6 is a residue of a hydroxyl group of pentaerythritol, methyl glucoside, sucrose, diethylene glycol, propylene glycol, glycerol or polyglycerol to remove hydrogen; i is 3, 4, 6 or 8; h is 10-1,500 The integer.
  • X is (CH 2 ) n , (CH 2 ) n CO, (CH 2 ) n OCO, (CH 2 ) n NHCO, -S -, -S0 2 -, -SO4 -; n is an integer from 1-12. In certain embodiments, X is CH 2 CO.
  • 1 is an amino acid or amino acid analog residue having at least two carboxylic acid groups and one amino group.
  • R 7 is a Cwo fluorenyl group or a Cwo heterofluorenyl group.
  • R 7 is methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, Cu embankment group, C 12 embankment Gan Gan ⁇ ⁇ Gan ⁇ Gan ⁇ Gan ⁇ Gan Lane, o ⁇ ⁇ 4 ⁇ Lane, i5 ⁇ lis ⁇ Lane, n ⁇ Lane, lis ⁇ Lane, ⁇ 9 ⁇ Lane, C 20 ⁇ base.
  • the carbonyl group in the formula (V) may be bonded to any one of the carbons of R 7 .
  • R 7 is a linear fluorenyl or heteroindolyl group.
  • R 7 is a branched indenyl or heteroindolyl. In certain embodiments, having the structure shown by the following formula (VI), (VI)
  • R 8 and R 9 are independently hydrogen, d — 12 fluorenyl, d— 12 heterofluorenyl, aryl, heteroaryl, aryl fluorenyl, heteroaryl fluorenyl, and are present in each repeating unit.
  • R 8 and R 9 may be the same or different; j is an integer of 1-10.
  • a 2 and A 3 independently have the structure shown by the following formula (VII), (VII)
  • R 1Q and R u are independently hydrogen, d 6 fluorenyl, d 6 heterofluorenyl, and R 1 ( ) , Ru in each repeating unit may be the same or different; h is 1-10 Integer.
  • R 10 , Ru are independently methyl, ethyl, propyl, butyl, pentyl, hexyl.
  • R 10 , Ru are independently linear fluorenyl or heteroindolyl.
  • R 1Q , Ru are independently branched Sulfhydryl or heteropoly.
  • a 2 , A 3 are independently glycine, alanine, leucine, isoleucine, valine, valine, phenylalanine, methionine, serine, Residues of threonine, cysteine and tyrosine. In certain embodiments, A 2 , A 3 are proline.
  • 0 1 and 1 ⁇ 4 are each independently a residue of an anti-tumor drug.
  • the residues can be antineoplastic agents or A 2 and A 3 form a peptide bond or an ester bond.
  • the anti-tumor drug is capable of forming a peptide bond or an ester bond with 2 or 3 by modification.
  • the anti-tumor drug is dasatinib, rapamycin, irinotecan, imatinib, erlotinib, gefitinib, lapatinib, sorafenib , sunitinib, paclitaxel, camptothecin, scutellaria, glycyrrhetinic acid, sorghum lactone.
  • the anti-tumor drug is dasatinib.
  • the present application provides a conjugate having the structure shown by formula (VIII) below,
  • R 7 is a C ⁇ o fluorenyl group.
  • Ru is independently hydrogen, d 6 fluorenyl and R 1Q , Ru in each repeating unit may be the same or different; h is an integer of 1-10.
  • P, X, a, b, c, d, Di, D 2 are as described above.
  • the present application provides a combination having the structure shown by the following formulas (IX), (X), (XI), c
  • e, f, and g are each independently an integer of from 10 to 1,500. As mentioned above.
  • the present application provides a pharmaceutical composition comprising the above combination and a pharmaceutically acceptable carrier or excipient.
  • the pharmaceutical composition is a tablet, a capsule, a pill, a granule, a dispersion Agents, suppositories, injections, solutions, suspensions, ointments, patches, lotions, drops, liniments, sprays.
  • the present application provides a medicament for the preparation of an anti-tumor, fungal infection, rheumatoid arthritis, multiple sclerosis, heart stenosis or pneumonia in the above combination and/or pharmaceutical composition. Application in .
  • the anti-tumor drug is applied to the following conditions: leukemia, acute myeloid leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia, acute lymphocytic leukemia, myelodysplasia, multiple Myeloma, Hodgkin's disease or non-Hodgkin's disease, small cell or non-small cell lung cancer, gastric cancer, colon cancer, esophageal cancer, colorectal cancer, prostate cancer, ovarian cancer, breast cancer, brain cancer, urinary Cancer, kidney cancer, bladder cancer, malignant melanoma, liver cancer, uterine cancer, pancreatic cancer, myeloma, endometrial cancer, head and neck cancer, pediatric tumor, sarcoma.
  • the present application provides a method of treating a tumor, a fungal infection, rheumatoid arthritis, multiple sclerosis, heart valve restenosis, or pneumonia in a subject, comprising administering to the subject An effective amount of the above combination or pharmaceutical composition.
  • the subject is a mammal. In certain embodiments, the subject is a human.
  • the above-described modes of administration of the combination and/or pharmaceutical composition include oral, mucosal, sublingual, ocular, topical, parenteral, rectal, cerebral, vaginal, peritoneal, bladder, Nasal administration.
  • the present application provides a method of preparing the above-described combination, comprising:
  • Figure 1 shows the degradation of compound DSR1-6 in 0.01 M PBS buffer.
  • DSR-1 is mPEG-dipeptide acid-dasatinib (20 ng
  • DSR-2 is mPEG-dipeptide glycine-dasatinib (20K)
  • DSR-3 is mPEG-dipeptide alanine- Dasatinib (20K)
  • DSR-4 is mPEG-dipeptide valine-dasatinib (20 ⁇ )
  • DSR-5 is ⁇ -PEG-dipeptide valine-dasatinib ( 30K)
  • DSR-6 is mPEG-tripeptide valine-dasatinib (40 ⁇ ).
  • the water-soluble polymer-amino acid oligopeptide-drug combination of the present invention has a structure represented by the following formula (I),
  • P is a water-soluble polymer
  • X is a linking group, and the linking group is bonded to P and
  • a A ; AA 2 and A 3 are each independently the same or different amino acid or amino acid analog residues; and D 2 are each independently the same or different drug molecule residues; a is 0 or 1; b is 2- An integer of 12; c is an integer from 0 to 7; d is 0 or 1.
  • a water-soluble polymer refers to a polymer formed by linking a compound containing a polar or charged functional group, which is soluble in water, that is, hydrophilic.
  • Water-soluble polymers include, but are not limited to: polyethylene glycol, polypropylene glycol, polyglutamic acid, polyaspartic acid, polyvinylpyrrolidone, polyvinyl alcohol, polypropylene morpholine, dextran, carboxymethyl Cellulose and its analogues or copolymers.
  • P is polyethylene glycol
  • the polyethylene glycol has a molecular weight of from 300 to 60,000.
  • the polyethylene glycol has a molecular weight of from 20,000 to 40,000.
  • the polyethylene glycol has a molecular weight of 20,000, 21,000, 22,000, 23,000, 24,000, 25,000, 26,000, 27,000, 28,000, 29,000, 30,000, 31,000, 32,000, 33,000, 34,000, 35,000, 36,000, 37,000, 38,000, 39,000, 40,000.
  • the polyethylene glycol is a linear, Y-branched or multi-branched polyethylene glycol.
  • the polyethylene glycol has a structure represented by the following formula ( ⁇ ),
  • the 12 fluorenyl group, the 12 hydrazinyl group, the hydrogen or the aryl fluorenyl group, and e is an integer of from 10 to 1,500.
  • e is an integer from 100 to 1,400.
  • e is an integer from 200 to 1,300.
  • e is an integer from 300 to 1,200.
  • e is an integer from 400 to 1,100.
  • e is an integer from 500 to 1,000.
  • e is 600, 700, 800, 900 or 1,000.
  • ⁇ is. ⁇ . ⁇ . More preferably, it is _ 8 fluorenyl. More preferably, it is _ 6 fluorenyl.
  • ⁇ is ⁇ ⁇ base. More preferably, ⁇ is d_ 4 fluorenyl. More preferably, h is the embankment d_ 3-yl. More preferably, R is a Cw fluorenyl group. In certain embodiments, it is methyl, ethyl, propyl, butyl, pentyl.
  • P has a structure represented by the following formula ( ⁇ ), (m)
  • R 2 and R 3 are each independently d— 12 fluorenyl, d— 12 heterofluorenyl, hydrogen, aryl fluorenyl; preferably, R 2 and R 3 are each independently Cw. ⁇ . More preferably, R 2 and R 3 are each independently a d 8 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 6 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 5 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 4 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 3 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 2 fluorenyl group. In certain embodiments, R 2 and R 3 are each independently methyl, ethyl, propyl, butyl, pentyl.
  • R 4 and R 5 are each independently a Cw 2 fluorenyl group and a C 2 fluorenylcarbonyl group.
  • R 4 and R 5 are each independently a Cwo fluorenyl group, a Cwo fluorenylcarbonyl group.
  • R 5 is independently d_ 8 fluorenyl, d 8 fluorenylcarbonyl.
  • R 5 is independently d- 6 fluorenyl, d 6 fluorenylcarbonyl.
  • R 4 and R 5 are each independently a d 5 fluorenyl group, a d 5 fluorenylcarbonyl group.
  • R 4 and R 5 are each independently a d 4 fluorenyl group, a d 4 fluorenylcarbonyl group. More preferably, R 4 and R 5 are each independently a d 3 fluorenyl group, a d 3 fluorenylcarbonyl group. More preferably, R 5 is independently d— 2 fluorenyl, d 2 fluorenylcarbonyl.
  • f and g are each independently an integer of from 10 to 1,500. In certain embodiments, f, g are each independently an integer from 100 to 1,400. In certain embodiments, f, g are each independently an integer from 200 to 1,300. In certain embodiments, f, g are each independently an integer from 300 to 1,200. In some embodiments, f and g are each independently an integer from 400 to 1,100. In certain embodiments, f, g are each independently an integer from 500 to 1,000. In certain embodiments, f, g are each independently 600, 700, 800, 900 or 1,000.
  • R 2 , R 3 in formula (III) are independently cyclopropyl, cyclobutyl, cyclohexyl or benzyl. In certain embodiments, R 2 , R 3 in formula (III) are methyl, ethyl, and R 5 is methylcarbonyl.
  • P has the structure shown by formula (IV) below, Formula (IV)
  • R 6 is a residue at which a hydroxyl group of pentaerythritol, methyl glucoside, sucrose, diethylene glycol, propylene glycol, glycerol or polyglycerol removes hydrogen; i is 3, 4, 6 or 8; h is 10-1,500 Integer.
  • h is an integer from 100 to 1,400.
  • h is an integer from 200 to 1,300.
  • h is an integer from 300 to 1,200.
  • h is an integer from 400 to 1,100.
  • h is an integer from 500 to 1,000.
  • h is 600, 700, 800, 900 or 1,000.
  • the linking group X in the formula (I) means a group which functions as a linking between the hydrophilic polymer and the amino acid oligopeptide. In certain embodiments, the linking group is introduced to modify the hydrophilic polymer to better attach to the amino acid oligopeptide.
  • X is (CH 2 ) n , (CH 2 ) n CO, (CH 2 ) n OCO, (CH 2 ) n NHCO, -S -, -S0 2 -, -S0 4 -; n is an integer from 1-12.
  • n is 1-6.
  • n is 1-3.
  • n is 1-2.
  • X is CH 2 CO.
  • amino acid or amino acid analog residues in formula (I) may comprise at least one amino group and one carboxylic acid group.
  • a 1 is an amino acid or amino acid analog residue having at least two carboxylic acid groups and one amino group.
  • R 7 is d_ 2 . ⁇ base, d_ 2 . Clay base.
  • ⁇ is -1() thiol, Cw. Clay base.
  • R 7 is d- 9 ⁇ , d- 9 heteroalkyl.
  • R 7 is an indenyl, d- 8 heterofluorenyl.
  • R 7 is d- 7 ⁇ , C ⁇ hetero.
  • R 7 is d- 6 fluorenyl, d- 6 heteropoly.
  • R 7 is d — 5 fluorenyl, d — 5 heterofluorenyl.
  • R 7 is d 4 alkyl, d 4 heteroaryl. In certain embodiments, R 7 is d — 3 fluorenyl, d — 3 heterofluorenyl. In certain embodiments, R 7 is methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, Cu
  • R 7 is a linear fluorenyl or heteroindolyl group. In certain embodiments, R 7 is a branched indenyl or heteroindolyl.
  • R 8 and R 9 are independently hydrogen, d — 12 fluorenyl, d— 12 heterofluorenyl, aryl, heteroaryl, aryl fluorenyl, heteroaryl fluorenyl, and in each repeating unit.
  • R 8 and R 9 may be the same or different; j is an integer of 1-10. In certain embodiments, j is an integer from 1 to 10. In certain embodiments, j is an integer from 1-9. In certain embodiments, j is an integer from 1-8. In certain embodiments, j is an integer from 1-7. In certain embodiments, j is an integer from 1 to 6. In certain embodiments, j is an integer from 1 to 5. In certain embodiments, j is an integer from 1 to 4. In certain embodiments, j is an integer from 1 to 3. In certain embodiments, j is an integer from 1 to 2.
  • 1 is a residue of aspartic acid or glutamic acid.
  • a 2 and A 3 in the formula (I) may independently be any amino acid or amino acid analog residue, and the amino acid or amino acid analog residue contains at least one amino group and one carboxylic acid group.
  • a 2 and A 3 independently have the structure shown by the following formula (VII), Formula (VII)
  • R 1Q and R u are independently hydrogen, d 6 fluorenyl, d 6 heterofluorenyl, and R 1 ( ) , Ru in each repeating unit may be the same or different; h is 1-10 Integer.
  • R 10 , Ru are independently methyl, ethyl, propyl, butyl, pentyl, hexyl.
  • R 10 , Ru are independently linear fluorenyl or heteroindolyl.
  • R 1Q , Ru is independently branched fluorenyl or heterofluorenyl.
  • h is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10.
  • a 2 , A 3 are independently glycine, alanine, leucine, isoleucine, valine, valine, phenylalanine, methionine, serine, Residues of threonine, cysteine and tyrosine.
  • the pharmaceutical compound in the water-soluble polymer-amino acid oligopeptide-drug conjugate of the present invention includes any drug molecule which can bind to an amino acid or an amino acid analog.
  • a drug molecule that can bind to an amino acid or amino acid analog comprises a functional group such as an amino group, a hydroxyl group, a phenol group, a thio group or a thiol group.
  • Examples of drug molecules containing an amino group, a hydroxyl group, a phenol group, a thiol group or a thiol group include, but are not limited to, acetaminophen, acyclovir ⁇ 2-amino-1,9-dihydro-9-[(2- Hydroxyethoxy)methyl] 6H-indol-6-one ⁇ , allopurinol, adenosine (6-amino-9-i3-D-furonucleoside nitrosourea-9-H-indole), dehydrocortic (retained) alcohol, prednisone, triamcinolone acetonide, cortisone (hydrocortisone), adenosine (6-amino-9- ⁇ -indole-furan nitrosourea-9-H- ⁇ ), Pine, estradiol, estrogen, estriol, 16-hydroxyestrene, equine estradiol, equine estrone,
  • a drug molecule that can bind to an amino acid or amino acid analog comprises a carboxyl group or a phosphate group/phosphonic acid group.
  • drug molecules containing a carboxyl group include, but are not limited to, mesalic acid, aminosalicylic acid, mesalic acid, aminosalicylic acid, baclofen, carbidopa, levodopa, aminobenzoic acid, bumami Tani, captopril [l-[(2S)-3-mercapto-2-methylpropionyl]-L-valine], cilastatin [(2)-7-[[( 3 ) -2-amino-2-carboxyethyl]thio]-2-[(S)-2,2-dimethylcyclopropanecarboxamide]-2-heptanoic acid], levothyroxine [D-3, 5, 3', 5'-tetraiodothyronine], amphotericin B, etret
  • 0 1 and 126 are each independently a residue of an anti-tumor drug.
  • the residues can be antineoplastic agents or A 2 and A 3 form a peptide bond or an ester bond.
  • the antitumor agent can be modified by a peptide bond or an ester bond with A 2 or A 3 is formed.
  • the anti-tumor drug is dasatinib, rapamycin, irinotecan, imatinib, erlotinib, gefitinib, lapatinib, sorafenib , sunitinib, paclitaxel, camptothecin, scutellaria, glycyrrhetinic acid, sorghum lactone.
  • the present application provides a conjugate having the structure shown by formula (VIII) below,
  • R 1( ) , R u are independently hydrogen, d 6 fluorenyl and R 10 , Rii in each repeating unit may be the same or different; h is an integer from 1 to 10.
  • R 7 , P, X, a, b, c, d, D 2 are as described above.
  • the present application provides a combination having the structure shown by the following formulas (IX), (X), (XI), Formula (IX)
  • e, f, and g are each independently an integer of from 10 to 1,500.
  • e, f, g are each independently an integer from 100 to 1,400.
  • e, f, g are each independently an integer from 200 to 1,300.
  • e, f, g are each independently an integer from 300 to 1,200.
  • e, f, g are each independently an integer from 400 to 1,100.
  • e, f, g are each independently an integer from 500 to 1,000.
  • e, f, g are each independently 600, 700, 800, 900 or 1,000. As mentioned above. In one embodiment, it is dasatinib.
  • the minimum and maximum values of the carbon atom content in the hydrocarbon group are represented by a prefix, for example, the prefix (C a _ b ) fluorenyl group means any fluorenyl group containing "a" to "b" carbon atoms.
  • (d_ 6 ) fluorenyl refers to a fluorenyl group containing from one to six carbon atoms.
  • the thiol group is branched or linear.
  • Negoxy refers to a straight or branched, monovalent, saturated aliphatic chain bonded to an oxygen atom, including but not limited to, for example, methoxy, ethoxy, propoxy, Butoxy, isobutoxy, tert-butoxy and other similar groups.
  • Mercapto refers to a straight or branched, monovalent, saturated aliphatic chain including, but not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, Pentyl, isopentyl, hexyl and the like.
  • Heteropoly means that one or more carbon atoms in the above fluorenyl group have been replaced by a hetero atom such as nitrogen, oxygen or sulfur. If the heterocyclic group contains more than one heteroatom, these heteroatoms may be the same or different.
  • Alkenyl means a straight or branched hydrocarbon bearing one or more double bonds including, but not limited to, vinyl, propenyl, and the like.
  • Aryl means a cyclic aromatic hydrocarbon including, but not limited to, phenyl, naphthyl, anthryl, phenanthryl and the like.
  • Cycloalkyl refers to a saturated monocyclic or polycyclic indenyl group, possibly fused to an aromatic hydrocarbon group. Cyclodecyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, indanyl, tetrahydronaphthyl, and the like.
  • Aryl fluorenyl means a group to which the above aryl group is covalently bonded to the above thiol group, such as, but not limited to, phenylmethyl, phenylethyl, phenylpropyl.
  • Heteroaryl means a monocyclic or polycyclic aromatic hydrocarbon in which one or more carbon atoms have been replaced by a hetero atom such as nitrogen, oxygen or sulfur. If a heteroaryl contains more than one heteroatom, these heteroatoms may be the same or different.
  • Heteroaryl groups include, but are not limited to, benzofuranyl, benzothienyl, benzimidazolyl, benzoxazolyl, benzothiazolyl, benzopyranyl, furyl, imidazolyl, oxazolyl, Pyridazinyl, fluorenyl, isobenzofuranyl, isodecyl, isoquinolyl, isothiazolyl, isoxazolyl, naphthyridinyl, oxadiazolyl, oxazinyl, oxazolyl, Pyridazinyl, pteridinyl, fluorenyl, pyranyl, pyrazinyl, pyrazolyl, pyridazinyl, pyridine [3,4-b]decyl, pyridyl, pyrimidinyl, pyrrolyl, quinolizine A quinolyl group, a quinolyl group, a
  • One cyclic group can be bonded to another group in a variety of ways. If the bonding method is not explicitly defined, it means that all possible ways are included. For example, “pyridyl” includes 2-, 3-, or 4-pyridyl, and “thienyl” includes 2- or 3-thienyl.
  • amino acid analog means an organic compound having a structure similar to that of an amino acid containing at least one amino group and at least one carboxyl group.
  • conjugates of the present invention can be synthesized according to the following scheme. This procedure is illustrative only and not limiting of other ways in which the combination may be made. In addition, the steps in the scheme are only for better description of the preparation method of the combination of the present invention, and the steps can be modified without departing from the scope of the invention described herein.
  • a method of preparing a water soluble polymer-amino acid oligopeptide-drug conjugate comprising:
  • the hydrophilic polymer P is linked to an amino acid via a linking group X to obtain a conjugate
  • the drug compound 0 1 is reacted with the amino acid A 2 to give the conjugate DA 2 .
  • drug compound 02 is reacted with an amino acid A 3 was obtained in conjunction with D 2 -A 3.
  • the hydrophilic polymer can be modified to introduce a reactive functional group that is bonded to the amino acid oligopeptide.
  • the introduction of a reactive functional group is achieved by a linking group X.
  • the reactive functional group introduced is a carboxylic acid group.
  • the conjugates can be formed DA 2 and / or D 2 -A 3 with an active functional group on the drug compound with an amino acid A 3 or A 2 reaction.
  • the reactive functional group on the drug compound can be a hydroxyl group, an amino group.
  • the drug compound and the amino acid are esterified or amidated.
  • the amides Di-As and I or D 2 -A 3 are amidated with the compound P_ ⁇ X ⁇ A1 to give a compound of formula (I).
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising a water-soluble polymer-amino acid oligopeptide-drug complex and a pharmaceutically acceptable carrier.
  • pharmaceutically acceptable carrier refers to a pharmaceutically acceptable substance, ingredient or medium, such as a liquid or solid filler, diluent, excipient, solvent or potting.
  • a pharmaceutically acceptable substance such as a liquid or solid filler, diluent, excipient, solvent or potting.
  • the pharmaceutically acceptable carrier can be a medium, diluent, excipient or other material which is not excessively toxic or side effects and which can be used to contact animal tissues.
  • Typical pharmaceutically acceptable carriers include sugars, starches, cellulosics, maltose, tragacanth, gelatin, Ringer's solution, alginic acid, physiological saline, buffers and the like.
  • Each pharmaceutically acceptable carrier should be compatible with the other ingredients, for example, with the conjugates provided herein, without excessive toxicity, irritation, allergic response, immunogen to biological living tissue or organs. Sexual or other problems or complications, and a reasonable ratio of benefits to risks.
  • Some pharmaceutically acceptable carrier materials include: (1) sugars such as lactose, glucose and sucrose; (2) starches such as corn starch and potato starch; (3) cellulose and its derivatives, such as Sodium carboxymethylcellulose, ethylcellulose, cellulose acetate; (4) scutellaria powder; (5) maltose; (6) gelatin; (7) talc; (8) excipients, such as cocoa Fats and suppository waxes; (9) Oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols such as propylene glycol; (11) polyols, such as Glycerin, sorbitol, mannitol and polyethylene glycol; (12) lipids such as ethyl oleate, ethyl laurate; (13) agarose; (14) buffers such as magnesium hydroxide and aluminum hydroxide; (15) Alginic acid; (16) starches
  • the pharmaceutical composition may include pharmaceutically acceptable excipients to mimic physiological conditions such as pH adjustment and buffering agents, toxicity modulators, etc., such as sodium acetate, sodium chloride, potassium chloride, calcium chloride, sodium lactate Wait.
  • pharmaceutically acceptable excipients to mimic physiological conditions such as pH adjustment and buffering agents, toxicity modulators, etc., such as sodium acetate, sodium chloride, potassium chloride, calcium chloride, sodium lactate Wait.
  • the pharmaceutical ingredient can be formulated into any suitable dosage form, such as a solid dosage form (e.g., tablets, capsules, powders, granules, etc.) and a liquid dosage form (e.g., aqueous solution, emulsion, elixirs, syrup, etc.).
  • a solid dosage form e.g., tablets, capsules, powders, granules, etc.
  • a liquid dosage form e.g., aqueous solution, emulsion, elixirs, syrup, etc.
  • Processes for the preparation of pharmaceutical compositions are well known and can be prepared according to conventional procedures, such as those found in Remington, The Science and Practice of Pharmacy (Gennaro ed. 20th edition, Williams & Wilkins PA, USA) (2000).
  • the compounds or pharmaceutical compositions provided herein can be formulated into a dosage form suitable for drug delivery, such as subcutaneous, intravenous, intramuscular, arterial, sheath, vesicles.
  • a dosage form suitable for drug delivery such as subcutaneous, intravenous, intramuscular, arterial, sheath, vesicles.
  • non-injectable routes eg, oral, intestinal, oral, nasal, intranasal, mucosal, epidermal, plaster, dermis, eye drops, lungs, tongue Subcutaneous, rectal, vaginal or epidermal administration).
  • Suitable dosage forms include, but are not limited to, dosage forms for injection use such as emulsions, solutions and suspensions, and dosage forms for oral use such as tablets, capsules, pills, dragees, powders and granules, topical or topical Formulations for skin absorption such as sprays, ointments, pastes, creams, lotions, gels, solutions, drug patches and inhalants, vaginal or rectal administration such as suppositories.
  • These dosage forms can be prepared according to the compound and suitable excipients under suitable conditions, and are well known, for example, by Remington: in The Science and Practice of Pharmacy (Gennaro ed. 20th edition, Williams & Wilkins PA, USA) (2000) ) provided.
  • One aspect of the invention is to increase the water solubility of a drug molecule by modification of a hydrophilic polymer, thereby prolonging the half-life of the drug.
  • the pharmaceutically active amount of the conjugate is increased as compared to the unmodified drug molecule, the drug loading, drug activity, stability, and water solubility.
  • Another aspect of the invention is to increase the stability of a drug conjugate by an amino acid oligopeptide, thereby reducing toxic side effects.
  • the increase in stability of the drug conjugate is achieved by selection of a suitable amino acid oligopeptide.
  • the pharmaceutical composition to increase the stability of the binding by selecting a suitable amino acid residue in combination with a pharmaceutical compound, i.e., the amino acid of formula A (I) 2 A 3, or implemented.
  • the amino acid residue bound to the drug compound is a proline residue.
  • the toxicity of the combination is reduced.
  • the conjugate further enhances the clinical effectiveness of the drug.
  • each hydrophilic polymer can be linked to a plurality of drug compounds by an amino acid oligopeptide, thereby greatly increasing the drug loading rate.
  • each hydrophilic polymer can be attached to at least two pharmaceutical compounds.
  • each The hydrophilic polymer can be attached to at least three drug compounds.
  • each hydrophilic polymer can be attached to at least four pharmaceutical compounds.
  • Another aspect of the invention provides the use of the above conjugates and/or pharmaceutical compositions for the preparation of a medicament and for the treatment of a disease.
  • the present invention provides a medicament for the preparation of an antitumor, fungal infection, rheumatoid arthritis, multiple sclerosis, heart stenosis or pneumonia in the above conjugate and/or pharmaceutical composition.
  • a medicament for the preparation of an antitumor, fungal infection, rheumatoid arthritis, multiple sclerosis, heart stenosis or pneumonia in the above conjugate and/or pharmaceutical composition.
  • the anti-tumor drug is applied to the following conditions: leukemia, acute myeloid leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia, acute lymphocytic leukemia, myelodysplasia, multiple Myeloma, Hodgkin's disease or non-Hodgkin's disease, small cell or non-small cell lung cancer, gastric cancer, colon cancer, esophageal cancer, colorectal cancer, prostate cancer, ovarian cancer, breast cancer, brain cancer, urinary Cancer, kidney cancer, bladder cancer, malignant melanoma, liver cancer, uterine cancer, pancreatic cancer, myeloma, endometrial cancer, head and neck cancer, pediatric tumor, sarcoma.
  • the present application provides a method of treating a tumor, a fungal infection, rheumatoid arthritis, multiple sclerosis, heart valve restenosis, or pneumonia in a subject, comprising administering to the subject An effective amount of the above combination or pharmaceutical composition.
  • the subject is a mammal.
  • the subject is a human, animal or pet.
  • the conjugate or pharmaceutical composition of the present invention may be administered to a living organism by any suitable route, such as by oral, intravenous, intranasal, topical, intramuscular, intradermal, transdermal or subcutaneous routes.
  • the present invention relates to a method of administration of a combination or pharmaceutical composition comprising oral, mucosal, sublingual, ocular, topical, parenteral, rectal, cerebral, vaginal, peritoneal, bladder, nasal Dosing.
  • the conjugates or pharmaceutical compositions of the present invention can be administered concurrently with a second active substance, such that a synergistic or even synergistic effect can be achieved in the organism.
  • a second active substance such that a synergistic or even synergistic effect can be achieved in the organism.
  • the compounds of the invention may be combined with the second active substance into a pharmaceutical composition, or may be administered simultaneously in separate compositions, or sequentially in separate compositions.
  • Second active substances which can be administered simultaneously with the compounds of the invention for the treatment of cancer include, but are not limited to: fluorouracil, doxorubicin, daunorubicin, tamoxifen, leuprolide, goserelin, fluoro Hemet, Nilemet, Finasteride, Dexamethasone, Amlodipine, Ampicillin, Anastrozole, Asparaginase, BCG, Bicalutamide, Bleomycin, Clinical, Busulfan, camptothecin, capecitabine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, colchicine, cyclophosphamide, drug, cyproterone, a Cytosine, dacarbazine, actinomycin d, gentamicin, diethestrol, diethylstilbestrol, docetaxel, doxorubicin, doxorubicin, epirubicin, estradio
  • the conjugates provided herein can be used in conjunction with non-chemical methods for cancer treatment. In certain embodiments, the conjugates provided herein can be performed concurrently with radiation therapy. In certain embodiments, the conjugates provided herein can be used in combination with surgery, tumor heat therapy, ultrasound focus therapy, cryotherapy, or several of the above.
  • the conjugates provided herein can be used concurrently with steroids.
  • Suitable steroids include, but are not limited to: Anxis, beclomethasone, betamethasone, budesonide, chlorinated Nisson, clobetasol, corticosterone, cortisone, hydroxyprednisolone, deshydroxymetasone, dexamethasone, diflurazon, difluoromethasone, difluprednate, glycyrrhetinic acid, fluza Fluke, flumetasone, flunisolide, flucline acetonide, acetonide acetate, fluocinolone acetonide, flubutylbutabutyl ester, fluconazole, acetonide, fluphene acetate, fluphene acetate Nitinidine, fluprednisolone, fluorohydrogenated, propionic acid fluoride, aldehyde-based shrinkage
  • the compounds provided herein can be used concurrently with an immunotherapeutic agent.
  • immunotherapeutic agents include: tumor cell multidrug resistance reversal agents (such as verapamil), rapamycin, mycophenolate mofetil, thalidomide, cyclophosphamide, cyclosporine, and Cloning of antibodies.
  • the beneficial effects of the conjugates described herein 1) The water soluble polymer-amino acid oligopeptide-drug conjugates described herein have a better therapeutic effect than drugs. At the same dosage, the conjugates described herein exhibit better anti-tumor and anti-cancer activities; 2) The water-soluble polymer-amino acid described in the present application can bind more drugs and increase the drug loading of the conjugate. , while the sustained release of drug can be avoided to achieve the sustained effective blood concentration administered to a patient repeatedly; 3) the conjugate reduces the toxicity of the drug; 4) by using the a and eight amino acid 2, the The stability of the drug conjugate is improved, facilitating the preparation, storage and administration of the drug.
  • Dasatinib used in the present embodiment was purchased from Nanjing Ange Pharmaceutical Chemical Co., Ltd., and L-(+)-glutamic acid was purchased from Beijing Chemical Reagent Co., Ltd., p-toluenesulfonic acid, benzyl alcohol and two Cyclohexylcarbodiimide (DCC) was purchased from Sinopharm Chemical Reagent Co., Ltd., 4-dimethylaminopyridine (DMAP) and 1-hydroxybenzotriazole (HOBt) were purchased from Shanghai Covalent Chemical Technology Co., Ltd.
  • DMAP 4-dimethylaminopyridine
  • HOBt 1-hydroxybenzotriazole
  • N,tert-butoxycarbonylglycine, N-tert-butoxycarbonylalanine, N-tert-butoxycarbonylproline and N-tert-butoxycarbonyl-L-glutamate-5-benzyl ester from Sichuan Acrylic Amino Acid Co., Ltd., monooxypolyethylene glycol acetate (20K), Y-type polyethylene glycol acetic acid (40 ⁇ ) is supplied by Beijing Keykai Technology Co., Ltd., and the rest are commercially available reagents (
  • Example 1 monomethoxy polyethylene glycol glutamic acid dipeptide acid (number average molecular weight 20000)
  • L-(+)-glutamic acid dibenzyl ester p-toluenesulfonate (Compound 1) 61go [00137] L-(+)-glutamic acid dibenzyl ester p-toluenesulfonate (compound 1) 30 g (0.06 mol) was dissolved in 500 mL of dichloromethane, and tert-butoxycarbonyl-L-glutamic acid was added. 5-benzyl ester 20.86 g (0.062 mol), DMAP 7.55 g (0.062 mol), HOBt 8.35 g (0.07 mol), a solution of 14.3 g of dichloromethane was added under nitrogen.
  • Example 2 Monomethoxypolyethylene glycol glutamic acid dipeptide glycine (number average molecular weight 20000) Preparation of a dasatinib conjugate (DSR-2)
  • N-tert-butoxycarbonyl glycine dasatinib (Compound 6) 1 g (1.6 mmol), dichloromethane (50 mL) was added to the reaction flask, then 25 mL of trifluoroacetic acid was added, and the reaction was stirred. After 3 hours, the solvent was evaporated under reduced pressure. dichloromethane was evaporated and evaporated to dryness, and then evaporated to dryness.
  • Example 3 Preparation of monomethoxypolyethylene glycol glutamic acid dipeptide alanine (number average molecular weight 20000) - Dasatinib conjugate (DSR-3).
  • N-tert-Butoxycarbonylalanine dasatinib (Compound 8) 1 g (1.6 mmol), dichloromethane (50 mL) was added to the reaction flask, then 25 mL of trifluoroacetic acid was added. After stirring for 3 hours, the solvent was evaporated under reduced pressure, dichloromethane was added to the residue, and evaporated to dryness under reduced pressure, and then repeated three times to obtain dasatinib alanine.
  • Trifluoroacetate (Compound 9) 1.3 g, directly reacted downward.
  • reaction solution was concentrated the next day, and the residue was recrystallized from isopropyl alcohol to give monomethoxy polyethylene glycol glutamic acid dipeptide alanine (20K) - dasatinib conjugate (DSR-3) 0.39 g .
  • Example 4 Monomethoxypolyethylene glycol glutamate dipeptide proline (number average molecular weight 20000) Preparation of a dasatinib conjugate (DSR-4).
  • valine dasatinib trifluoroacetate 140 mg (0.15 mmol), HOBt (3.4 mg, 0.025 mmo) and DMAP 24.4 mg (0.2 mmol) were added to the reaction flask. Dissolve in a mixed solvent of methylene chloride and N,N-dimethylformamide, cool in an ice bath under nitrogen atmosphere, and then add DCC 36.1 mg (0.175 mmol) in a solution of dichloromethane. The reaction was overnight. The reaction solution was concentrated the next day, and the residue was recrystallized from isopropanol to obtain monomethoxy polyethylene glycol glutamic acid dipeptide proline.
  • Example 5 Y-type polyethylene glycol glutamic acid dipeptide proline (number average molecular weight 40000)
  • N-tert-Butoxycarbonylglutamic acid benzyl ester dipeptide (Compound 2) 0.78 g (Example 1) Dissolved in 7 mL of dichloromethane, 3 mL of trifluoroacetic acid, and reacted at room temperature for 2 h. The solvent was removed, 100 mL of dichloromethane was added, and the pH was adjusted to 7-8 with a 5% sodium hydrogen carbonate solution. The extract was separated and the organic phase was washed twice with 5% sodium hydrogen carbonate solution and dried over anhydrous sodium sulfate.
  • valine dasatinib trifluoroacetate (Compound 11) was prepared according to the method described in Example 4.
  • Y-type polyethylene glycol glutamic acid dipeptide acid (Compound 13) (40K, 0.6g, 0.02mmol)
  • Dasatinib triacetate valine 112mg (0.12mmol)
  • HOBt 2.7 Mg, 0.02mmo
  • DMAP 24.4 mg (0.2mmol)
  • Example 6 Preparation of monomethoxypolyethylene glycol glutamic acid tripeptide valine (number average molecular weight 20000) - Dasatinib conjugate (DSR-6) [00159] N-tert-Butoxycarbonyl glutamic acid benzyl ester dipeptide (Compound 2) 6.47 g (0.1 mol) was dissolved in 15 mL of dichloromethane, 6 mL of trifluoroacetic acid was added, and reacted at room temperature for 2 h. The solvent was removed, 100 mL of dichloromethane was added, and the pH was adjusted to 7-8 with a 5% sodium hydrogen carbonate solution.
  • Compound 2 N-tert-Butoxycarbonyl glutamic acid benzyl ester dipeptide
  • the extract was separated and the organic phase was washed twice with 5% sodium hydrogen carbonate solution and dried over anhydrous sodium sulfate. Filtration, the filtrate was directly added to the reaction flask, under the protection of nitrogen, tert-butoxycarbonyl-L-glutamic acid-5-benzyl ester 3.37g (O.Olmol), DMAP 1.22g (O.Olmol), HOBt l. 35 g (O.Olmol), after completely dissolved, a solution of 2.39 g (O.Ol.lmol) of dichloromethane was added dropwise. After the dropwise addition, the closed system was reacted overnight. The reaction was monitored by TLC.
  • valine dasatinib trifluoroacetate (Compound 11) was prepared according to the method described in Example 4. Monomethoxy polyethylene glycol glutamic acid tripeptide (Compound 17) (20K, 0.6 g, 0.03 mmol), Valine dasatinib trifluoroacetate (compound ll) 224 mg (0.24 mmol), HOBt (4 mg, 0.03 mmol) and hydrazine DMAP 29.3 mg (0.24 mmol) were added to the reaction flask with dichloro Methyl hydrazine and N,N-dimethylformamide were dissolved in a mixed solvent, cooled in an ice bath under nitrogen protection, and then dropped into DCC 43.3 mg.
  • Example 8 Antitumor effects of different polyethylene glycol dasatinib conjugates in a K562 human chronic myeloid leukemia subcutaneous tumor model. [00166] Experimental method:
  • K562 cells were subcutaneously inoculated into the right side of NOD/SCID mice to establish a subcutaneous model of human chronic myeloid leukemia xenograft animals. When the average tumor volume reached 130 mm3, the experimental mice were divided into groups of 8 and administered intravenously twice a week. The efficacy was evaluated based on the relative tumor growth rate (T/C%).
  • K562 cell line is supplemented with 10% fetal bovine serum, L-glutamine (2 mM)
  • RPMI-1640 medium culture in vitro at 37 ° C in the presence of 5% CO 2 in air. Tumor cells were routinely passaged twice a week. Tumor cells in the exponential growth phase were collected and suspended in an equal volume of PBS: Matrigel mixture and placed on ice for tumor inoculation.
  • mice were subcutaneously inoculated with 5 x 106 K562 cells on the right side of the right side, and tumor growth was periodically observed. When the tumors were grown to an average of 130 mm3, the tumor size and the body weight of the mice were randomly grouped and treatment was started.
  • RTV refers to the relative tumor volume.
  • the T/C ratio is the percentage of the tumor volume relative to the treatment group and the control group at some point after the end of treatment, reflecting the antitumor efficacy of the different treatment groups.
  • the positive drug dasatinib 5mg/kg was statistically significantly different from the solvent control group (p ⁇ 0.001), and the average tumor volume was 311mm 3 , the relative tumor proliferation rate. (T/C%) was 12.4%; the drug effects of the test drugs DSR-4, DSR-5 and DSR-6 (5mg/kg) were statistically significantly different from the solvent control group (p ⁇ 0.001).
  • mean tumor volume were 39.2 mm 3, 178.6 mm 3, and 137mm 3, the relative tumor proliferation rate (T / C%) was 1.6%, 7.3% and 5.8%.
  • the average tumor volume of the dasatinib (5 mg/kg) group was 391 mm 3
  • the test drugs DSR-4, DSR-5 and DSR-6 were effective. in contrast there were significant differences (p ⁇ 0.001) statistically, the mean tumor volume were 176 mm 3, 203mm 3 and 258mm 3, the relative tumor proliferation rate (T / C%) were 45%, 52 % and 66%.
  • Relative tumor proliferation rate of the positive drug dasatinib (5 mg/kg), test drug DSR-4, DSR-5 and DSR-6 (5 mg/kg) compared with the solvent control group (T/ C%) were 12.4%, 1.6%, 7.3%, and 5.8%, respectively, indicating that all compounds had significant anti-K562 tumor growth effects (p ⁇ 0.001).
  • dasatinib 5 mg/kg
  • the antitumor effects of the same doses of DSR-4, DSR-5 and DSR-6 were all significantly more ⁇ 0.001).
  • Example 9 Pharmacodynamic study of different polyethylene glycol dasatinib conjugates on a PC-3 human prostate cancer subcutaneous tumor model.
  • the PC-3 cell line was cultured in vitro in Ham's F12K medium supplemented with 10% fetal calf serum, L-glutamine (2 mM) at 37 ° C in air containing 5% CO 2 . Tumor cells were routinely passaged twice a week. Tumor cells in the exponential growth phase were collected, suspended in an equal volume of PBS: Matrigel mixture, placed on ice for tumor inoculation.
  • mice were subcutaneously inoculated with 5 ⁇ 10 6 PC-3 cells on the right side of the right side, and tumor growth was observed periodically, and when the tumors were grown to an average of 160 mm 3 , the tumor size and the body weight of the mice were randomly grouped and treatment was started.
  • the body weight and tumor size of the mice were measured twice a week throughout the experiment.
  • Tumor size calculation formula: tumor volume (mm 3 ) 0.5 x (tumor long diameter X tumor short diameter 2 ).
  • the RTV and T/C ratios were calculated based on the tumor volume of the experimental and control groups.
  • RTV refers to the relative tumor volume.
  • T/C The ratio refers to the percentage of tumor volume in the treatment group and the control group at a certain point after the end of treatment, reflecting the antitumor efficacy of different treatment groups.
  • the tumors were recorded in the following two ways: 1. After each group of animals was euthanized by co 2 , the tumor-bearing side was facing upwards and photographed separately according to the group; 2. After the tumor was removed, weighed first. Tumor weight, T/C (% of the tumor weight of the treatment group and the control group) was calculated, and then the tumors of each group were simultaneously placed and photographed in order.
  • the drug efficacy of the positive drug dasatinib (10mg/kg) was statistically significantly different from the solvent control group (p ⁇ 0.01), and the average tumor volume was 1752mm 3 , the relative tumor proliferation rate. (T/C%) was 73%; the drug effects of the test drugs DSR-4, DSR-5 and DSR-6 (10 mg/kg) were statistically significantly different from the solvent control group (p ⁇ 0.001).
  • the average tumor volume of the dasatinib (10 mg/kg) group was 1868 mm 3
  • the test drugs DSR-4, DSR-5 and DSR-6 (10 mg/kg) were effective. were compared statistically significant difference (p ⁇ 0.05, p ⁇ 0.05 and p ⁇ 0.01), mean tumor volumes were 1200 mm 3 1406mm 3 and 1449mm 3, relative tumor proliferation rate, (T / C %) are 61%, 75% and 72% respectively.
  • Relative tumor growth rate (T/C%) of the positive drug dasatinib (10 mg/kg), test drugs DSR-4, DSR-5 and DSR-6 (10 mg/kg) compared to the solvent control ) were 73%, 47%, 57%, and 56%, respectively, indicating that all compounds had statistically significant anti-PC-3 tumor growth effects (p ⁇ 0.01).
  • the antitumor effects of the same doses of DSR-4, DSR-5 and DSR-6 (10 mg/kg) were more significant (p ⁇ 0.05) compared with dasatinib (10 mg/kg).
  • rapamycin used in the examples was purchased from Wuhan Yuancheng Co-creation Technology Co., Ltd., and t-butyl bromoacetate and triphenylphosphine were purchased from Sinopharm Chemical Reagent Co., Ltd.
  • Example 10 Preparation of monomethoxypolyethylene glycol glycine acetate (number average molecular weight 20000) - rapamycin conjugate (LPR-1)
  • tert-Butyl bromoacetate (5.82 g, 30 mmol) was added to a reaction flask, dissolved in acetone (80 mL), then sodium azide (4.55 g, 70 mmol) dissolved in water (40 mL) The solution was heated to reflux overnight. The reaction mixture was evaporated to dryness. EtOAc was evaporated. This liquid was dissolved in methanol (90 mL), then 1N sodium hydroxide solution (90 mL) was added, stirred and heated to reflux for 3 h. After cooling, the methanol was evaporated under reduced pressure, and the mixture was cooled with ice-cooled overnight.
  • Azidoacetic acid (Compound 18) (253 mg, 2.5 mmoL) and rapamycin (2.28 g, 2.5) mmoL) was added to the reaction flask, dissolved in dichloromethane, cooled in an ice bath, and then added with 4-dimethylaminopyridine (DMAP, 611 mg, 5 mmoL) and N,N-dicyclohexylcarbimide ( DCC, 1.03 g, 5 mmoL) was added to the reaction flask and stirring was continued at room temperature overnight. After the reaction mixture was concentrated, the residue was purified by column chromatography to give the titled product of the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the the
  • Azoxyacetic acid rapamycin ester (Compound 19) (0.7 g, 0.7 mmoL) and triphenylphosphine (0.37 g, 1.4 mmoL) were added to the reaction flask, followed by a mixture of tetrahydrofuran and water ( 5:1, 180 mL), heated to 50 ° C overnight, and the residue was concentrated with ethyl acetate. The extract was washed with saturated brine and dried. After concentration under reduced pressure, the residue was purified by column chromatography to yield EtOAc (yield: 20), yield: 70%, MS m/z: 994 [M+Na]+.
  • Example 1 Monomethoxypolyethylene glycol glutamic acid dipeptide glycine (number average molecular weight)
  • Monomethoxypolyethylene glycol glutamic acid dipeptide acid (Compound 5) was prepared according to the method described in Example 1.
  • Example 12 Y-type polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight 40000) Preparation of a rapamycin conjugate (LPR-3)
  • Y-type polyethylene glycol glutamic acid dipeptide acid (Compound 13) was prepared as described in the above examples.
  • Y-type polyethylene glycol glutamic acid dipeptide acid (Compound 13) (40K, 0.5g, 0.0125mmol), glycine rapamycin ester (Compound 20) 24.3 mg (0.025mmol), HOBt (1.7 Mg, 0.0125mmo) and DMAP 3 mg (0.025mmol) were added to the reaction flask, dissolved in dichloromethane, cooled in ice bath, and then added dropwise to DCC 4.1 mg (0.02 mmol) in dichloromethane solution. After that, it naturally rose to room temperature and reacted overnight.
  • reaction solution was concentrated the next day, and the residue was crystallized from isopropanol to obtain Y-type polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight: 40000) - rapamycin conjugate (LPR-3) 0.44 g.
  • Plc/prf/5 cells were cultured in vitro in a single layer in a MEM medium supplemented with 10% heat-inactivated fetal bovine serum and cultured in an incubator containing 5% CO2 air at 37 °C. Digestion was performed twice with trypsin-EDTA twice a week. When cells are exponentially growing, cells are harvested, counted, and vaccinated.
  • LPR1, 2, 3 anti-tumor efficacy evaluation indexes of plc/prf/5 subcutaneous xenograft model are shown in Table 8. Evaluation of anti-tumor effect of each treatment group
  • LPR-1 and LPR-2 use polyethylene glycol of the same structure and the same number average molecular weight, except that polyethylene glycol of LPR-1 is only bonded to rapamycin via glycine, polyethylene glycol molecule One end group is only bonded to one rapamycin molecule; and the polyethylene glycol of LPR-2 is bonded to rapamycin through glutamic acid dipeptide and glycine, and one terminal group of the polyethylene glycol molecule can be bonded Combine three rapamycin molecules.
  • the drug loading rate of LPR-2 is three times that of LPR-1, and its antitumor activity is also significantly higher than that of LPR-1.
  • the irinotecan hydrochloride used in the examples was purchased from Shanghai Longxiang Biomedical Development Co., Ltd.
  • Example 14 monomethoxy polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight 20000)
  • Monomethoxypolyethylene glycol glutamic acid dipeptide acid (Compound 5) was prepared according to the method described in Example 1.
  • Monomethoxypolyethylene glycol glutamic acid dipeptide acid (Compound 5) (molecular weight: 20,000) 5.0 g, enonotecan glycine (Compound 22) 1.02 g, N-hydroxysuccinimide (NHS) 115 mg , 4-dimethylaminopyridine (DMAP) 153mg dissolved in 50mL anhydrous dichloromethane, under the protection of nitrogen, add two Cyclohexylcarbodiimide (DCC) 309 mg. The reaction was stirred at room temperature overnight.
  • Example 15 Y-type polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight 40000) - Preparation of INOTECK conjugate (YNR-2)
  • Y-type polyethylene glycol glutamic acid dipeptide acid (compound 13) (molecular weight: 40,000) was prepared according to the method described in the above examples.
  • Y-type polyethylene glycol glutamic acid dipeptide acid (compound 13) (molecular weight 40,000) 10 g, quinolidine glycine (compound 22) 1.02 g, N-hydroxysuccinimide (NHS) 115 mg, 4- Dimethylaminopyridine (DMAP) 153 mg was dissolved in 50 mL of anhydrous dichloromethane, and 309 mg of dicyclohexylcarbodiimide (DCC) was added under nitrogen. The reaction was stirred at room temperature overnight.
  • DCC dicyclohexylcarbodiimide
  • Example 16 Monomethoxypolyethylene glycol glutamic acid tripeptide glycine (number average molecular weight 20000)
  • Monomethoxypolyethylene glycol glutamic acid tripeptide acid (molecular weight: 20,000) was prepared according to the method described in the above examples.
  • Monomethoxy polyethylene glycol glutamic acid tripeptide acid (compound 17) (molecular weight 20000) 5.0 g, ennotecan glycine ester (compound 22) 1.36 g, N-hydroxyl
  • <RTI ID 0.0>> Monomethoxypolyethylene glycol glutamic acid tripeptide glycine (number average molecular weight - INOTECO conjugate (YNR-3) 4 -NMR (DMSO-d6): .84.89 (m, 8H), 1.24(m, 9H),, 1.71-1.74(m, 23H), 2.12(m, 16H), 2.95-2.99(m, 19H), 3.50(m, 1800H), 4.22(m, 18H), 5.45(m 5H), 5.49 (m, 5H), 7.07 (m, 2H), 7.55-7.60 (m, 3H), 7.89 (m, 3H), 8.1 l (m, 4H), 8.20 (m, 3H).
  • Example 17 Growth Inhibition of Human Intestinal Carcinoma HCT-1 16 Nude Mice Transplanted with Polyethylene Glycosin Inocomb Conjugate
  • test substance YNR-1 is a monomethoxypolyethylene glycol acetate glycine-inonotecan conjugate.
  • CPT-1 1 Positive control drug enonotecan hydrochloride injection (CPT-1 1 ), 40mg/2ml, batch number is 8UL002-B, manufactured by Aventis Pharma (Dagenham). Dilute with physiological saline to the desired concentration at the time of use.
  • YNR-1 dose was set to 45 mg / kg (calculated according to the amount of INOTECK), administered intravenously, once a week for three weeks; CPT-1 1 dose was 45 mg / kg, weekly Administered once intravenously; and 15 mg/kg, administered intravenously three times a week for three consecutive weeks.
  • Animals BALB/cA nude mice, male, 5-6 weeks old, weighing 19 ⁇ 2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017.
  • the number of animals in each group 12 in the negative control group and 6 in the drug-administered group.
  • Human intestinal cancer HCT-1 16 cell line was purchased from ATCC. The cell strain was inoculated subcutaneously into the right axillary fossa of the nude mice, and the inoculation amount was 5 ⁇ 106/head, and the transplanted tumor was formed and then used in nude mice for 2 generations.
  • the tumor tissue in the vigorous growth period was cut into about 1.5 mm 3 and inoculated subcutaneously in the right axilla of the nude mice under aseptic conditions.
  • the diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm 3 .
  • YNR-1 was administered in the 45 mg/kg dose group and the control group was given the same amount of normal saline once a week for three consecutive weeks.
  • CPT-1 1 (15 mg/kg) was used as a positive control drug, administered intravenously three times a week for three weeks. After the end of the administration, continue to observe for one week.
  • the diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed.
  • V0 is the measured tumor volume at the time of sub-cage administration (i.e., d0)
  • Vt is the tumor volume at each measurement.
  • the experimental results are shown in Table 9.
  • YNR-1 45 mg/kg was administered intravenously once a week for three weeks, and the growth of subcutaneous xenografts of human intestinal cancer HCT-116 nude mice was significantly inhibited.
  • the percentage of T/C was 27.60%.
  • the effect is better than the same dose of CPT-1145mg/kg of the same dosage regimen.
  • the same treatment scheme CPT-11 can also inhibit the growth of HCT-116 subcutaneous xenografts to a certain extent, but the percentage of T/C is only 63.56%.
  • the positive control CPT-11 (15mg/kg) was administered intravenously three times a week for three weeks, which also significantly inhibited the growth of HCT-116 subcutaneous xenografts.
  • the T/C value was 39.84%.
  • the body weight of the mice decreased. Compared with the solvent control group, the weight loss of the nude mice in the CPT-11 45 mg/kg group was slightly stronger than that in the
  • Example 18 Growth Inhibition of Human Intestinal Carcinoma HT-29 Nude Mice Transplanted Tumor by Polyethylene Glycosin Integate Conjugate
  • test substance YNR-1 is a monomethoxy polyethylene glycol glutamic acid dipeptide glycine-inonotecan complex.
  • YNR-1 dose is set to 45mg / kg (calculated according to the amount of INOTEC), intravenous administration, once a week for three weeks; CPT-11 dose is 45mg / kg, weekly vein The drug was administered once; and 15 mg/kg, administered intravenously three times a week for three consecutive weeks.
  • BALB/cA nude mice male, 5-6 weeks old, weighing 18 ⁇ 2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017.
  • the number of animals in each group 12 in the negative control group and 6 in the drug-administered group.
  • Human intestinal cancer HT-29 cell line was purchased from ATCC. The cell strain was used to inoculate the right axilla of the nude mice subcutaneously, and the inoculation amount of the cells was 5 ⁇ 106/head, and the transplanted tumor was formed and then used in nude mice for 2 generations.
  • the tumor tissue in the vigorous growth period was cut into 1.5 mm 3 and inoculated subcutaneously in the right axilla of nude mice under aseptic conditions.
  • the diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm 3 .
  • YNR-1 and CPT-11 were given the same amount of normal saline in the 45 mg/kg dose group and the control group, intravenously once a week for three weeks.
  • CPT-11 (15 mg/kg) was administered as a positive control drug three times a week for three weeks. After the end of the administration, continue to observe for one week.
  • the diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed.
  • V0 is the measured tumor volume at the time of sub-cage administration (ie, d0)
  • vt is the tumor volume at each measurement.
  • T/C (%) (TRTV/CRTV) X100%, TRTV: treatment group RTV; CRTV: negative control group RTV.
  • mice in the experimental results were shown in Table 10.
  • YNR-1 45 mg/kg was administered intravenously once a week for three weeks, and the growth of subcutaneous xenografts of human intestinal cancer HT-29 nude mice was significantly inhibited.
  • mice in the experimental treatment group The increase in tumor volume is slowed down.
  • T/C the percentage of T/C was 41.08%.
  • the positive control CPT-1 1 (15 mg/kg) was administered intravenously three times a week for three consecutive weeks, which also significantly inhibited the growth of HT-29 subcutaneous xenografts with a T/C value of 27.27%.
  • the nude mice in each group grew well, and only the two different dose groups of CPT-1 1 had a decrease in body weight.
  • test substance YNR-1 is a monomethoxy polyethylene glycol glutamic acid dipeptide glycine-inonotecan complex.
  • CPT-1 1 The positive control drug inonotecan hydrochloride (CPT-1 1 ) 40 mg/2 ml, lot number 8UL002-B, was manufactured by Aventis Pharma (Dagenham). When used, it is diluted with physiological saline to the desired concentration. [00280] Dose setting
  • YNR-1 dose was set to 45 mg / kg (calculated according to the amount of INOTECK), administered intravenously, once a week for three weeks; CPT-1 1 dose was 45 mg / kg, weekly Intravenous administration And 15 mg/kg, administered intravenously three times a week for three consecutive weeks.
  • BALB/cA nude mice female, 5-6 weeks old, weighing 18 ⁇ 2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017.
  • the number of animals in each group 12 in the negative control group and 6 in the drug-administered group.
  • Human lung cancer A549 cell line was purchased from ATCC. The cell strain was used to inoculate the right axilla of the nude mice, and the inoculation amount was 5 ⁇ 106/head. After the transplanted tumor was formed, it was used in nude mice for 2 generations.
  • the tumor tissue in the vigorous growth period was cut into 1.5 mm 3 and inoculated subcutaneously in the right axilla of the nude mice under aseptic conditions.
  • the diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm 3 .
  • YNR-1 and CPT-11 were given the same amount of normal saline in the 45 mg/kg dose group and the control group, intravenously once a week for three weeks.
  • CPT-11 (15 mg/kg) was administered as a positive control drug three times a week for three weeks. After the end of the administration, continue to observe for one week.
  • the diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed.
  • T/C (%) (TRTV/CRTV) X100%
  • TRTV treatment group RTV
  • CRTV negative control group RTV.
  • YNR-1 and CPT-11 (45 mg/kg) were administered intravenously once a week for three weeks, which significantly inhibited the growth of subcutaneous xenografts in human lung cancer A549 nude mice.
  • the T/C percentage is 20.62%.
  • the anti-tumor effect of YNR-1 is much better than that of CPT-11 of the same dosage regimen.
  • the growth of subcutaneous xenografts in tumor-bearing mice was slowed after one week of YNR-1 treatment.
  • the positive control CPT-11 (15 mg/kg) was administered intravenously three times a week for three consecutive weeks, which also significantly inhibited the growth of A549 subcutaneous xenografts with a T/C value of 53.26%.
  • the nude mice in each treatment group grew well, and the weight gain was slightly slower than the solvent control group.
  • Example 20 Growth inhibition of ectopic conjugates of polyethylene glycol on human ovarian cancer SKOV-3 nude mice
  • test substance YNR-1 is a monomethoxy polyethylene glycol glutamic acid dipeptide glycine-inonotecan complex.
  • YNR-1 dose was set to 45 mg / kg (calculated according to the amount of INOTECK), administered intravenously, once a week for three weeks; CPT-11 dose was 45 mg / kg, weekly vein The drug was administered once; and 15 mg/kg, administered intravenously three times a week for three consecutive weeks. [00296] animals
  • BALB/cA nude mice female, 5-6 weeks old, weighing 19 ⁇ 2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017.
  • the number of animals in each group 12 in the negative control group and 6 in the drug-administered group.
  • Human ovarian cancer SKOV-3 cell strain was purchased from ATCC. The cell strain was inoculated subcutaneously into the right axilla of the nude mice, and the inoculation amount was 5 ⁇ 106/piece, and the transplanted tumor was formed and then used in nude mice for 2 generations.
  • the tumor tissue in the vigorous growth period was cut into 1.5 mm 3 and inoculated subcutaneously in the right axilla of nude mice under aseptic conditions.
  • the diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm3.
  • YNR-1 and CPT-11 were given the same amount of normal saline in the 45 mg/kg dose group and the control group, intravenously once a week for three weeks.
  • CPT-11 (15 mg/kg) was administered as a positive control drug three times a week for three weeks. After the end of the administration, continue to observe for one week.
  • the diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed.
  • T/C (%) (TRTV/CRTV) X100%
  • TRTV treatment group RTV
  • CRTV negative control group RTV.
  • CPT-1 1 (15 mg/kg) was administered intravenously three times a week for three consecutive weeks, which also significantly inhibited the growth of SKOV-3 subcutaneous xenografts with a T/C value of 55.69%.
  • the weight of mice in the solvent control group and CPT-1 1 treatment group decreased slightly; while the YNR-1 experimental treatment group had good growth and weight gain.
  • Example 21 Inhibition of growth of human intestinal cancer SW-620 nude mice xenografts by different formulations of irinotecan
  • test substance YNR-1 is a monomethoxy polyethylene glycol glutamic acid dipeptide glycine-inonotecan complex.
  • CPT-1 1 The positive control drug inonotecan hydrochloride (CPT-1 1 ) 40 mg / 2 ml, batch number 8UL002-B, manufactured by Aventis Pharma (Dagenham). Dilute with physiological saline to the desired concentration at the time of use. [00308] Dose setting
  • YNR-1 dose was set to 45 mg / kg (calculated according to the amount of INOTECK), administered intravenously, once a week for three weeks; CPT-11 dose was 45 mg / kg, weekly vein The drug was administered once; and 15 mg/kg, administered intravenously three times a week for three consecutive weeks.
  • BALB/cA nude mice male, 4-6 weeks old, weighing 19 ⁇ 2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017.
  • the number of animals in each group 12 in the negative control group and 6 in the drug-administered group.
  • Human intestinal cancer SW-620 cell line was purchased from ATCC. The cell strain was inoculated subcutaneously into the right axillary fossa of the nude mice, and the inoculation amount was 5 ⁇ 106/head, and the transplanted tumor was formed and then used in nude mice for 2 generations.
  • the tumor tissue in the vigorous growth period was cut into 1.5 mm 3 and inoculated subcutaneously in the right axilla of the nude mice under aseptic conditions.
  • the diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm 3 .
  • YNR-1 and CPT-11 were given the same amount of normal saline in the 45 mg/kg dose group and the control group, intravenously once a week for three weeks.
  • CPT-11 (15 mg/kg) was administered as a positive control drug three times a week for three weeks. After the end of the administration, continue to observe for one week.
  • the diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed.
  • YNR-1 and CPT-1 1 were administered intravenously once a week for three weeks, which significantly inhibited the growth of subcutaneous xenografts in human intestinal cancer SW-620 nude mice, and YNR-1.
  • the anti-tumor effect was better than the same dose of CPT-1 1 in the same dosage regimen.
  • two tumor-bearing mice in the YNR-1 treatment group had complete tumors. Regressed, there was no rebound after one week of withdrawal.
  • the positive control CPT-1 1 (15 mg/kg) was administered intravenously three times a week for three consecutive weeks, which also significantly inhibited the growth of SW-620 subcutaneous xenografts.
  • the T/C value was

Abstract

A water soluble polymer-amino acid oligopeptide-medicine combination as presented in formula (I), and a medicinal composition having the combination. In the combination, P is a water soluble polymer; X is a linking group that links P and A1; A1, A2, and A3 are independent residues of identical or different amino acids or amino acid analogs; D1 and D2 are independent residues of identical or different medicinal molecules; a is 0 or 1; b is an integer between 2 and 12; c is an integer between 0 and 7; d is 0 or 1; the combination can enhance the drug loading rate, water solubility, stability, and activity of the medicine.

Description

水溶性聚合物 -氨基酸寡肽 -药物结合物及其制备方法和用途  Water-soluble polymer-amino acid oligopeptide-drug conjugate, preparation method and use thereof
相关申请的交叉引用 Cross-reference to related applications
[0001] 本申请要求于 2013年 1月 28日提交的中国申请号 201310032635.5 的优先权， 于 2013年 6月 18日提交的中国申请号 201310241907.2的优先权， 以及 2013年 12月 2日提交的中国申请号 201310632830.1的优先权。三份申请 都在此全文引用并入。 发明领域  [0001] This application claims the priority of China Application No. 201310032635.5 filed on January 28, 2013, the priority of China Application No. 201310241907.2 filed on June 18, 2013, and China submitted on December 2, 2013. Priority of application number 201310632830.1. All three applications are hereby incorporated by reference in their entirety. Field of invention
[0002] 本申请提供了一种水溶性聚合物一氨基酸寡肽一药物结合物及其药 物组合物， 以及所述结合物和药物组合物的制备方法和用途。 所述结合物通过 增加活性药物的载药量， 提高药物对疾病的治疗效果。 此外， 所述结合物的稳 定性也得到了很大的提高。 技术背景  The present application provides a water-soluble polymer-amino acid oligopeptide-drug conjugate and a pharmaceutical composition thereof, and a preparation method and use of the conjugate and the pharmaceutical composition. The conjugate increases the therapeutic effect of the drug on the disease by increasing the drug loading of the active drug. In addition, the stability of the combination is also greatly improved. technical background
[0003] 如何将有效浓度的药物输送到作用位点并保证药物的稳定性和低毒 性是药学中的一个重要课题。 通过将药物偶联到水溶性聚合物， 例如， 聚乙二 醇 （PEG) 可以增加药物的水溶性， 但是普通聚乙二醇修饰技术的一个缺陷是 药物或其他功能基团一般只能连接到聚乙二醇分子的两端， 从而大大限制了聚 乙二醇载体的载药量。  [0003] How to deliver an effective concentration of a drug to a site of action and to ensure the stability and low toxicity of the drug is an important subject in pharmacy. By coupling a drug to a water soluble polymer, for example, polyethylene glycol (PEG) can increase the water solubility of the drug, but a drawback of conventional polyethylene glycol modification techniques is that the drug or other functional group is generally only attached to The two ends of the polyethylene glycol molecule greatly limit the drug loading of the polyethylene glycol carrier.
[0004] 因此， 如何找到一个可以提高药物水溶性， 增加稳定性， 降低毒副作 用， 同时具有较高载药量并且本身没有毒性的载体和给药技术仍然是亟待解决 的问题。 本发明就是针对解决这一问题。 发明简介 [0004] Therefore, how to find a carrier and drug delivery technology which can improve the water solubility of the drug, increase the stability, and reduce the side effects of the drug, while having a higher drug loading amount and not being toxic by itself, is still an urgent problem to be solved. The present invention is directed to solving this problem. Introduction to invention
[0005] 本发明提供了一种水溶性聚合物-氨基酸寡肽-药物的结合物， 具有如 下式 （I) 所示的结构， The present invention provides a water-soluble polymer-amino acid oligopeptide-drug combination having The structure shown by the following formula (I),
Figure imgf000004_0001
式 （I)
Figure imgf000004_0001
Formula (I)
[0006] 其中， P为水溶性聚合物； Wherein P is a water soluble polymer;
[0007] X为连接基团， 所述连接基团连接 P和 A1 ; [0007] X is a linking group, the linking group linking P and A 1 ;
[0008] ALA2和 A3各自独立的为相同或不同的氨基酸或氨基酸类似物残基；[0008] ALA 2 and A 3 are each independently the same or different amino acid or amino acid analog residues;
[0009] Di和 D2各自独立的为相同或不同的药物分子残基； [0009] Di and D 2 are each independently the same or different drug molecule residues;
[0010] a为 0或 1 ;  [0010] a is 0 or 1;
[0011] b为 2-12的整数； [0011] b is an integer from 2 to 12;
[0012] c为 0-7的整数；  [0012] c is an integer from 0 to 7;
[0013] d为 0或 1。  [0013] d is 0 or 1.
[0014] 在某些实施方式中， P选自下组： 聚乙二醇、 聚丙二醇， 聚谷氨酸， 聚天门冬氨酸， 聚乙烯吡咯垸酮， 聚乙烯醇， 聚丙烯吗啉， 葡聚糖， 羧甲基纤 维素及其类似物或共聚物。  [0014] In certain embodiments, P is selected from the group consisting of polyethylene glycol, polypropylene glycol, polyglutamic acid, polyaspartic acid, polyvinylpyrrolidone, polyvinyl alcohol, polypropylene morpholine, Dextran, carboxymethylcellulose and analogues or copolymers thereof.
[0015] 在某些实施方式中， P为聚乙二醇， 且所述聚乙二醇的分子量为 300-60,000 在某些实施方式中， 聚乙二醇的分子量为 20，000-40，000。 在某些 实施方式中， 聚乙二醇的分子量为 20,000、 21,000、 22,000、 23,000、 24,000、 25,000、 26,000、 27,000、 28,000、 29,000、 30,000、 31,000、 32,000、 33,000、 34,000、 35,000、 36,000、 37,000、 38,000、 39,000、 40,000。  [0015] In certain embodiments, P is polyethylene glycol, and the polyethylene glycol has a molecular weight of 300-60,000. In certain embodiments, the polyethylene glycol has a molecular weight of 20,000-40. 000. In certain embodiments, the polyethylene glycol has a molecular weight of 20,000, 21,000, 22,000, 23,000, 24,000, 25,000, 26,000, 27,000, 28,000, 29,000, 30,000, 31,000, 32,000, 33,000, 34,000, 35,000, 36,000, 37,000, 38,000, 39,000, 40,000.
[0016] 在某些实施方式中，聚乙二醇为直链、 Y型支链或多分支链聚乙二醇。  [0016] In certain embodiments, the polyethylene glycol is a linear, Y-branched or multi-branched polyethylene glycol.
[0017] 在某些实施方式中， 聚乙二醇具有如下式 （Π) 所示的结构，  [0017] In certain embodiments, the polyethylene glycol has a structure represented by the following formula (Π),
R1-0-^CH2CH20^- 式 （π) R 1 -0-^CH 2 CH 2 0^- Formula (π)
[0018] 其中所述 为^12垸基、 d_12杂垸基、氢或芳基垸基， e为 10-1,500 的整数。 [0018] wherein ^ is alkyl with 12, 12 D_ embankment heteroaryl group, or aryl alkyl with hydrogen, e is an integer of 10-1,500.
[0019] 在某些实施方式中， P具有如下式 （ΠΙ) 所示的结构，
Figure imgf000005_0001
（m) [0019] In some embodiments, P has the structure shown by the following formula (ΠΙ),
Figure imgf000005_0001
(m)
[0020】 其中 R2、 R3各自独立的为 d_12垸基、 d_12杂垸基、 氢、 芳基垸基； Wherein R 2 and R 3 are each independently d— 12 fluorenyl, d— 12 heterofluorenyl, hydrogen, aryl fluorenyl;
[0021】 R4、 R5各自独立的为 C1-12垸基、 C1-12垸基羰基； [0021] R 4 and R 5 are each independently a C 1-12 fluorenyl group, a C 1-12 fluorenylcarbonyl group;
[0022】 f、 g各自独立的为 10-1,500的整数。  [0022] f and g are each independently an integer of from 10 to 1,500.
[0023] 在某些实施方式中， 式 （III) 中的 R2、 R3为环丙基、 环丁基、 环己 In certain embodiments, R 2 , R 3 in formula (III) are cyclopropyl, cyclobutyl, cyclohexyl
[0024】 在某些实施方式中， 式 （III) 中的 R2、 R3为甲基， F 为乙基， R5为 In certain embodiments, R 2 , R 3 in formula (III) are methyl, F is ethyl, and R 5 is
[0025] 在某些实施方式中， P具有如下式 （IV) 所示的结构, [0025] In certain embodiments, P has the structure shown by formula (IV) below,
R6~^0十 CH2CH20- 式 （IV) R 6 ~^0 十 CH 2 CH 2 0- (IV)
[0026] 其中， R6为季戊四醇， 甲基葡萄糖苷， 蔗糖， 二甘醇， 丙二醇， 甘 油或聚甘油的羟基去除氢的残基； i为 3， 4， 6或 8; h为 10-1,500的整数。 Wherein R 6 is a residue of a hydroxyl group of pentaerythritol, methyl glucoside, sucrose, diethylene glycol, propylene glycol, glycerol or polyglycerol to remove hydrogen; i is 3, 4, 6 or 8; h is 10-1,500 The integer.
[0027] 在某些实施方式中， X为 (CH2)n， (CH2)nCO，（CH2)nOCO， (CH2)nNHCO， -S -， -S02-， -SO4-; n为 1-12的整数。 在某些实施方式中， X为 CH2CO。 In certain embodiments, X is (CH 2 ) n , (CH 2 ) n CO, (CH 2 ) n OCO, (CH 2 ) n NHCO, -S -, -S0 2 -, -SO4 -; n is an integer from 1-12. In certain embodiments, X is CH 2 CO.
[0028] 在某些实施方式中， 1为具有至少两个羧酸基和一个氨基的氨基酸 或氨基酸类似物残基。 In certain embodiments, 1 is an amino acid or amino acid analog residue having at least two carboxylic acid groups and one amino group.
[0029] 在某些实施方式中， 具有如下式 （V) 所示的结构， — NH— R7— C _ 式 （V) [0029] In certain embodiments, having the structure shown by the following formula (V), — NH— R 7 — C _ (V)
[0030] 其中， R7为 Cwo垸基、 Cwo杂垸基。在某些实施方式中， R7为甲基、 乙基、 丙基、 丁基、 戊基、 己基、 庚基、 辛基、 壬基、 癸基、 Cu垸基、 C12垸 甘 ^甘 ^甘 ^甘 ^甘 ^甘 ^甘 ^甘 巷、 o τη ΐ4 兀巷、 i5 τη lis 兀巷、 n 兀巷、 lis 兀巷、 ΐ9 兀巷、 C20垸基。 式 （V) 中的羰基可以与 R7的任何一个碳连接。 在某些实施方式中， R7为直链的垸基或杂垸基。 在某些实施方式中， R7为支链的垸基或杂垸基。 0031] 在某些实施方式中， 具有如下式 （VI) 所示的结构，
Figure imgf000006_0001
（VI) Wherein R 7 is a Cwo fluorenyl group or a Cwo heterofluorenyl group. In certain embodiments, R 7 is methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, Cu embankment group, C 12 embankment Gan Gan ^ ^ Gan ^ Gan ^ Gan ^ Gan ^ Gan ^ Gan Lane, o τη ΐ 4 兀 Lane, i5 τη lis 兀 Lane, n 兀 Lane, lis 兀 Lane, ΐ 9 兀 Lane, C 20垸 base. The carbonyl group in the formula (V) may be bonded to any one of the carbons of R 7 . In certain embodiments, R 7 is a linear fluorenyl or heteroindolyl group. In certain embodiments, R 7 is a branched indenyl or heteroindolyl. In certain embodiments, having the structure shown by the following formula (VI),
Figure imgf000006_0001
(VI)
[0032] 其中， R8、 R9独立的为氢、 d_12垸基、 d_12杂垸基、 芳基、 杂芳基、 芳垸基、 杂芳垸基、 且在每个重复单元中的 R8、 R9可以相同或不同； j为 1-10 的整数。 Wherein R 8 and R 9 are independently hydrogen, d — 12 fluorenyl, d— 12 heterofluorenyl, aryl, heteroaryl, aryl fluorenyl, heteroaryl fluorenyl, and are present in each repeating unit. R 8 and R 9 may be the same or different; j is an integer of 1-10.
[0033] 在某些实施方式中， 为天冬氨酸或谷氨酸的残基。  In certain embodiments, is a residue of aspartic acid or glutamic acid.
[0034] 在某些实施方式中， A2、 A3独立的具有如下式 （VII) 所示的结构，
Figure imgf000006_0002
(VII) [0034] In certain embodiments, A 2 and A 3 independently have the structure shown by the following formula (VII),
Figure imgf000006_0002
(VII)
[0035] 其中， R1Q、 Ru独立的为氢、 d_6垸基、 d_6杂垸基且在每个重复单 元中的 R1()、 Ru可以相同或不同； h为 1-10的整数。 在某些实施方式中， R10、 Ru独立的为甲基、 乙基、 丙基、 丁基、 戊基、 己基。 在某些实施方式中， R10、 Ru独立的为直链的垸基或杂垸基。 在某些实施方式中， R1Q、 Ru独立的为支链 的垸基或杂垸基。 Wherein R 1Q and R u are independently hydrogen, d 6 fluorenyl, d 6 heterofluorenyl, and R 1 ( ) , Ru in each repeating unit may be the same or different; h is 1-10 Integer. In certain embodiments, R 10 , Ru are independently methyl, ethyl, propyl, butyl, pentyl, hexyl. In certain embodiments, R 10 , Ru are independently linear fluorenyl or heteroindolyl. In certain embodiments, R 1Q , Ru are independently branched Sulfhydryl or heteropoly.
[0036] 在某些实施方式中， A2、 A3独立的为甘氨酸、 丙氨酸、 亮氨酸、 异 亮氨酸、 缬氨酸、 脯氨酸、 苯丙氨酸、 蛋氨酸、 丝氨酸、 苏氨酸、 半胱氨酸和 酪氨酸的残基。 在某些实施方式中， A2、 A3为缬氨酸。 In certain embodiments, A 2 , A 3 are independently glycine, alanine, leucine, isoleucine, valine, valine, phenylalanine, methionine, serine, Residues of threonine, cysteine and tyrosine. In certain embodiments, A 2 , A 3 are proline.
[0037] 在某些实施方式中， 01和1¾各自独立的为抗肿瘤药物的残基。 在某 些实施方式中， 抗肿瘤药物的残基能够与 A2或 A3形成肽键或酯键。 在某些实 施方式中， 抗肿瘤药物通过修饰从而能够与 2或 3形成肽键或酯键。 在某些 实施方式中， 抗肿瘤药物为达沙替尼、 雷帕霉素、 依诺替康、 伊马替尼、 厄洛 替尼、 吉非替尼、 拉帕提尼、 索拉非尼、 舒尼替尼、 紫杉醇、 喜树碱、 华蟾酥 毒基、 甘草次酸、 东莨菪内酯。 在某些实施方式中， 抗肿瘤药物为达沙替尼。 [0037] In certain embodiments, 0 1 and 1⁄4 are each independently a residue of an anti-tumor drug. In certain embodiments, the residues can be antineoplastic agents or A 2 and A 3 form a peptide bond or an ester bond. In certain embodiments, the anti-tumor drug is capable of forming a peptide bond or an ester bond with 2 or 3 by modification. In certain embodiments, the anti-tumor drug is dasatinib, rapamycin, irinotecan, imatinib, erlotinib, gefitinib, lapatinib, sorafenib , sunitinib, paclitaxel, camptothecin, scutellaria, glycyrrhetinic acid, sorghum lactone. In certain embodiments, the anti-tumor drug is dasatinib.
[0038] 在某些实施方式中， 本申请提供了具有如下式（VIII )所示的结构的 结合物， [0038] In certain embodiments, the present application provides a conjugate having the structure shown by formula (VIII) below,
Figure imgf000007_0001
式 （vm)
Figure imgf000007_0001
Formula (vm)
[0039] 其中， R7为 C^o垸基， 。、 Ru独立的为氢、 d_6垸基且在每个重 复单元中的 R1Q、 Ru可以相同或不同； h为 1-10的整数。 P、 X、 a、 b、 c、 d、 Di , D2如上所述。 Wherein R 7 is a C^o fluorenyl group. Ru is independently hydrogen, d 6 fluorenyl and R 1Q , Ru in each repeating unit may be the same or different; h is an integer of 1-10. P, X, a, b, c, d, Di, D 2 are as described above.
[0040] 在某些实施方式中， 本申请提供了具有如下式（IX) 、 (X) 、 (XI) 所示的结构的结合物， c[0040] In certain embodiments, the present application provides a combination having the structure shown by the following formulas (IX), (X), (XI), c
Figure imgf000008_0001
Figure imgf000008_0001
式 （X) Formula (X)
Figure imgf000008_0002
Figure imgf000008_0002
式 （XI )  Formula (XI)
[0041] 其中， e、 f、 g各自独立的为 10-1,500的整数。 如上所述。  Wherein e, f, and g are each independently an integer of from 10 to 1,500. As mentioned above.
[0042] 在某些实施方式中，本申请提供了包含上述结合物以及药学上可接受 的载体或赋形剂的药物组合物。  [0042] In certain embodiments, the present application provides a pharmaceutical composition comprising the above combination and a pharmaceutically acceptable carrier or excipient.
[0043] 在某些实施方式中， 药物组合物为片剂、 胶囊剂、 丸剂、 颗粒剂、 散 剂、 栓剂、 注射剂、 溶液剂、 混悬剂、 膏剂、 贴剂、 洗剂、 滴剂、 擦剂、 喷雾 剂。 [0043] In certain embodiments, the pharmaceutical composition is a tablet, a capsule, a pill, a granule, a dispersion Agents, suppositories, injections, solutions, suspensions, ointments, patches, lotions, drops, liniments, sprays.
[0044] 在某些实施方式中， 本申请提供了上述结合物和 /或药物组合物在制 备抗肿瘤、 真菌感染、 类风湿性关节炎、 多发性硬化症、 心瓣再狭窄或肺炎的 药物中的应用。  [0044] In certain embodiments, the present application provides a medicament for the preparation of an anti-tumor, fungal infection, rheumatoid arthritis, multiple sclerosis, heart stenosis or pneumonia in the above combination and/or pharmaceutical composition. Application in .
[0045] 在某些实施方式中， 所述抗肿瘤药物应用于以下病症： 白血病、急性 粒细胞性白血病、 慢性粒细胞性白血病、 慢性淋巴性白血病、 急性淋巴性白血 病、 脊髓发育不良、 多发性骨髓瘤、 何杰金氏病或非何杰金氏病、 小细胞或非 小细胞性肺癌、 胃癌、 肠癌、 食道癌、 结肠直肠癌、 前列腺癌、 卵巢癌、 乳腺 癌、 脑癌、 泌尿道癌、 肾癌、 膀胱癌、 恶性黑素瘤、 肝癌、 子宫癌、 胰腺癌、 骨髓瘤癌、 子宫内膜癌、 头颈癌、 小儿肿瘤、 肉瘤。  [0045] In certain embodiments, the anti-tumor drug is applied to the following conditions: leukemia, acute myeloid leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia, acute lymphocytic leukemia, myelodysplasia, multiple Myeloma, Hodgkin's disease or non-Hodgkin's disease, small cell or non-small cell lung cancer, gastric cancer, colon cancer, esophageal cancer, colorectal cancer, prostate cancer, ovarian cancer, breast cancer, brain cancer, urinary Cancer, kidney cancer, bladder cancer, malignant melanoma, liver cancer, uterine cancer, pancreatic cancer, myeloma, endometrial cancer, head and neck cancer, pediatric tumor, sarcoma.
[0046] 在某些实施方式中， 本申请提供了一种治疗主体中肿瘤、 真菌感染、 类风湿性关节炎、 多发性硬化症、 心瓣再狭窄或肺炎的方法， 包括向所述主体 给与有效量的上述结合物或药物组合物。 在某些实施方式中， 所述主体是哺乳 动物。 在某些实施方式中， 所述主体是人。  [0046] In certain embodiments, the present application provides a method of treating a tumor, a fungal infection, rheumatoid arthritis, multiple sclerosis, heart valve restenosis, or pneumonia in a subject, comprising administering to the subject An effective amount of the above combination or pharmaceutical composition. In certain embodiments, the subject is a mammal. In certain embodiments, the subject is a human.
[0047] 在某些实施方式中， 上述结合物和 /或药物组合物的给药方式包括口 腔、 粘膜、 舌下、 眼部、 局部、 肠道外、 直肠、 脑池、 阴道、 腹膜、 膀胱、 鼻 部给药。  [0047] In certain embodiments, the above-described modes of administration of the combination and/or pharmaceutical composition include oral, mucosal, sublingual, ocular, topical, parenteral, rectal, cerebral, vaginal, peritoneal, bladder, Nasal administration.
[0048] 在某些实施方式中，本申请提供了一种制备上述的结合物的方法，包 括：  [0048] In certain embodiments, the present application provides a method of preparing the above-described combination, comprising:
[0049] 使药物化合物与氨基酸发生酯化或酰胺化反应形成结合物 D A2和 / 或 D2-A3; [0049] causing a drug compound to be esterified or amidated with an amino acid to form a conjugate DA 2 and / or D 2 -A 3 ;
[0050] 使结合物!)1- 2和 /或 D2-A3
Figure imgf000009_0001
[0050] conjugates!) 1 - 2 and / or D 2 - A 3
Figure imgf000009_0001
得到式 （I) 所示化合物。 [0051] 本发明的其它特点和优势可见下面的详细叙述。以下实施例和具体实 施方式旨在清楚说明本发明的技术方案及其技术效果和优势， 并非限制本发明 的范围。 The compound of the formula (I) is obtained. [0051] Other features and advantages of the invention will be apparent from the following detailed description. The following examples and specific embodiments are intended to clarify the technical solutions of the present invention and the technical effects and advantages thereof, and do not limit the scope of the present invention.
附图说明 DRAWINGS
[0052] 图 1为化合物 DSR1— 6在 0.01 M的 PBS缓冲液中的降解情况。 其 中 DSR-1 为 mPEG-二肽酸一达沙替尼酯 (20ig，DSR-2 为 mPEG-二肽甘氨酸 一达沙替尼酯 (20K)， DSR-3 为 mPEG-二肽丙氨酸一达沙替尼酯 (20K)， DSR-4 为 mPEG-二肽缬氨酸一达沙替尼酯 (20Κ)， DSR-5 为 Υ— PEG-二肽缬氨酸一 达沙替尼酯 (30K)， DSR-6 为 mPEG-三肽缬氨酸一达沙替尼酯 (40Κ)。  Figure 1 shows the degradation of compound DSR1-6 in 0.01 M PBS buffer. Wherein DSR-1 is mPEG-dipeptide acid-dasatinib (20 ng, DSR-2 is mPEG-dipeptide glycine-dasatinib (20K), and DSR-3 is mPEG-dipeptide alanine- Dasatinib (20K), DSR-4 is mPEG-dipeptide valine-dasatinib (20Κ), DSR-5 is Υ-PEG-dipeptide valine-dasatinib ( 30K), DSR-6 is mPEG-tripeptide valine-dasatinib (40Κ).
[0053] 图 2是由 LPR-1、 LPR-2、 LPR-3及溶媒引起的人肝癌 plc/prf/5荷瘤 小鼠的体重变化结果。  2 is a result of changes in body weight of human liver cancer plc/prf/5 tumor-bearing mice caused by LPR-1, LPR-2, LPR-3 and a vehicle.
[0054] 图 3是 LPR-1、 LPR-2、 LPR-3及溶媒在人肝癌 plc/prf/5皮下移植肿 瘤模型中的抗癌活性结果。  3 is the result of anti-cancer activity of LPR-1, LPR-2, LPR-3 and a vehicle in a human liver cancer plc/prf/5 subcutaneous tumor model.
发明详述 Detailed description of the invention
[0055] 水溶性聚合物-氨基酸寡肽-药物的结合物的结构  Structure of a water-soluble polymer-amino acid oligopeptide-drug combination
[0056] 一方面， 本发明的水溶性聚合物-氨基酸寡肽-药物的结合物， 具有如 下式 （I) 所示的结构，  In one aspect, the water-soluble polymer-amino acid oligopeptide-drug combination of the present invention has a structure represented by the following formula (I),
Figure imgf000010_0001
式 （I)
Figure imgf000010_0001
Formula (I)
[0057] 其中， P为水溶性聚合物； X为连接基团， 所述连接基团连接 P和Wherein P is a water-soluble polymer; X is a linking group, and the linking group is bonded to P and
Ai ; A A2和 A3各自独立的为相同或不同的氨基酸或氨基酸类似物残基； 和 D2各自独立的为相同或不同的药物分子残基； a为 0或 1 ; b为 2-12的整数; c为 0-7的整数； d为 0或 1。 A A ; AA 2 and A 3 are each independently the same or different amino acid or amino acid analog residues; and D 2 are each independently the same or different drug molecule residues; a is 0 or 1; b is 2- An integer of 12; c is an integer from 0 to 7; d is 0 or 1.
[0058] 水溶性聚合物是指由包含极性或带电荷的官能团的化合物链接形成 的聚合物， 该聚合物能溶于水， 即具有亲水性。 水溶性聚合物包括但不限于： 聚乙二醇、 聚丙二醇， 聚谷氨酸， 聚天门冬氨酸， 聚乙烯吡咯垸酮， 聚乙烯醇， 聚丙烯吗啉， 葡聚糖， 羧甲基纤维素及其类似物或共聚物。  [0058] A water-soluble polymer refers to a polymer formed by linking a compound containing a polar or charged functional group, which is soluble in water, that is, hydrophilic. Water-soluble polymers include, but are not limited to: polyethylene glycol, polypropylene glycol, polyglutamic acid, polyaspartic acid, polyvinylpyrrolidone, polyvinyl alcohol, polypropylene morpholine, dextran, carboxymethyl Cellulose and its analogues or copolymers.
[0059] 在本发明的某些实施方式中， P为聚乙二醇， 且所述聚乙二醇的分子 量为 300-60,000。 在某些实施方式中， 聚乙二醇的分子量为 20，000-40，000。 在 某些实施方式中，聚乙二醇的分子量为 20,000、 21,000、 22,000、 23,000、 24,000、 25,000、 26,000、 27,000、 28,000、 29,000、 30,000、 31,000、 32,000、 33,000、 34,000、 35,000、 36,000、 37,000、 38,000、 39,000、 40,000。  In certain embodiments of the invention, P is polyethylene glycol, and the polyethylene glycol has a molecular weight of from 300 to 60,000. In certain embodiments, the polyethylene glycol has a molecular weight of from 20,000 to 40,000. In certain embodiments, the polyethylene glycol has a molecular weight of 20,000, 21,000, 22,000, 23,000, 24,000, 25,000, 26,000, 27,000, 28,000, 29,000, 30,000, 31,000, 32,000, 33,000, 34,000, 35,000, 36,000, 37,000, 38,000, 39,000, 40,000.
[0060] 在某些实施方式中，聚乙二醇为直链、 Y型支链或多分支链聚乙二醇。  [0060] In certain embodiments, the polyethylene glycol is a linear, Y-branched or multi-branched polyethylene glycol.
[0061] 在某些实施方式中， 聚乙二醇具有如下式 （Π) 所示的结构，  [0061] In certain embodiments, the polyethylene glycol has a structure represented by the following formula (Π),
R1-0-^CH2CH20^ ~ R 1 -0-^CH 2 CH 2 0^ ~
e 式 （Π)  e type (Π)
[0062] 其中所述 为 12垸基、 12杂垸基、氢或芳基垸基， e为 10-1,500 的整数。 在某些实施方式中， e为 100-1,400的整数。 在某些实施方式中， e为 200-1,300的整数。 在某些实施方式中， e为 300-1,200的整数。 在某些实施方 式中， e为 400-1,100的整数。 在某些实施方式中， e为 500-1,000的整数。 在 某些实施方式中， e为 600、 700、 800、 900或 1,000。优选的， ^为。^。垸基。 更优选的， 为 _8垸基。 更优选的， 为 _6垸基。 更优选的， ^为 ^ 垸基。 更优选的， ^为 d_4垸基。 更优选的， ^为 d_3垸基。 更优选的， R 为 Cw垸基。 在某些实施方式中， 为甲基、 乙基、 丙基、 丁基、 戊基。 Wherein the 12 fluorenyl group, the 12 hydrazinyl group, the hydrogen or the aryl fluorenyl group, and e is an integer of from 10 to 1,500. In certain embodiments, e is an integer from 100 to 1,400. In certain embodiments, e is an integer from 200 to 1,300. In certain embodiments, e is an integer from 300 to 1,200. In certain embodiments, e is an integer from 400 to 1,100. In certain embodiments, e is an integer from 500 to 1,000. In certain embodiments, e is 600, 700, 800, 900 or 1,000. Preferably, ^ is. ^.垸基. More preferably, it is _ 8 fluorenyl. More preferably, it is _ 6 fluorenyl. More preferably, ^ is ^ 垸 base. More preferably, ^ is d_ 4 fluorenyl. More preferably, h is the embankment d_ 3-yl. More preferably, R is a Cw fluorenyl group. In certain embodiments, it is methyl, ethyl, propyl, butyl, pentyl.
[0063] 在某些实施方式中， P具有如下式 （ΠΙ) 所示的结构，
Figure imgf000012_0001
（m) [0063] In some embodiments, P has a structure represented by the following formula (ΠΙ),
Figure imgf000012_0001
(m)
[0064] 其中 R2、 R3各自独立的为 d_12垸基、 d_12杂垸基、 氢、 芳基垸基； 优选的， R2、 R3各自独立的为 Cw。垸基。更优选的， R2、 R3各自独立的为 d_8 垸基。 更优选的， R2、 R3各自独立的为 d_6垸基。 更优选的， R2、 R3各自独立 的为 d_5垸基。 更优选的， R2、 R3各自独立的为 d_4垸基。 更优选的， R2、 R3 各自独立的为 d_3垸基。 更优选的， R2、 R3各自独立的为 d_2垸基。 在某些实 施方式中， R2、 R3各自独立的为甲基、 乙基、 丙基、 丁基、 戊基。 Wherein R 2 and R 3 are each independently d— 12 fluorenyl, d— 12 heterofluorenyl, hydrogen, aryl fluorenyl; preferably, R 2 and R 3 are each independently Cw.垸基. More preferably, R 2 and R 3 are each independently a d 8 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 6 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 5 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 4 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 3 fluorenyl group. More preferably, R 2 and R 3 are each independently a d 2 fluorenyl group. In certain embodiments, R 2 and R 3 are each independently methyl, ethyl, propyl, butyl, pentyl.
[0065] R4、 R5各自独立的为 Cw2垸基、 Cw2垸基羰基。 优选的， R4、 R5各 自独立的为 Cwo垸基、 Cwo垸基羰基。 更优选的， 、 R5各自独立的为 d_8 垸基、 d_8垸基羰基。 更优选的， 、 R5各自独立的为 d_6垸基、 d_6垸基羰 基。 更优选的， R4、 R5各自独立的为 d_5垸基、 d_5垸基羰基。 更优选的， R4、 R5各自独立的为 d_4垸基、 d_4垸基羰基。更优选的， R4、 R5各自独立的为 d_3 垸基、 d_3垸基羰基。 更优选的， 、 R5各自独立的为 d_2垸基、 d_2垸基羰 基。 R 4 and R 5 are each independently a Cw 2 fluorenyl group and a C 2 fluorenylcarbonyl group. Preferably, R 4 and R 5 are each independently a Cwo fluorenyl group, a Cwo fluorenylcarbonyl group. More preferably, R 5 is independently d_ 8 fluorenyl, d 8 fluorenylcarbonyl. More preferably, R 5 is independently d- 6 fluorenyl, d 6 fluorenylcarbonyl. More preferably, R 4 and R 5 are each independently a d 5 fluorenyl group, a d 5 fluorenylcarbonyl group. More preferably, R 4 and R 5 are each independently a d 4 fluorenyl group, a d 4 fluorenylcarbonyl group. More preferably, R 4 and R 5 are each independently a d 3 fluorenyl group, a d 3 fluorenylcarbonyl group. More preferably, R 5 is independently d— 2 fluorenyl, d 2 fluorenylcarbonyl.
[0066] f、 g各自独立的为 10-1,500的整数。 在某些实施方式中， f、 g各自 独立的为 100-1,400的整数。 在某些实施方式中， f、 g各自独立的为 200-1,300 的整数。 在某些实施方式中， f、 g各自独立的为 300-1,200的整数。 在某些实 施方式中， f、 g各自独立的为 400-1,100的整数。 在某些实施方式中， f、 g各 自独立的为 500-1,000的整数。在某些实施方式中， f、 g各自独立的为 600、 700、 800、 900或 1,000。  [0066] f and g are each independently an integer of from 10 to 1,500. In certain embodiments, f, g are each independently an integer from 100 to 1,400. In certain embodiments, f, g are each independently an integer from 200 to 1,300. In certain embodiments, f, g are each independently an integer from 300 to 1,200. In some embodiments, f and g are each independently an integer from 400 to 1,100. In certain embodiments, f, g are each independently an integer from 500 to 1,000. In certain embodiments, f, g are each independently 600, 700, 800, 900 or 1,000.
[0067] 在某些实施方式中， 式（III) 中的 R2、 R3独立的为环丙基、环丁基、 环己基或苄基。 [0068】 在某些实施方式中， 式 （III) 中的 R2、 R3为甲基， 为乙基， R5为 甲基羰基。In certain embodiments, R 2 , R 3 in formula (III) are independently cyclopropyl, cyclobutyl, cyclohexyl or benzyl. In certain embodiments, R 2 , R 3 in formula (III) are methyl, ethyl, and R 5 is methylcarbonyl.
0069] 在某些实施方式中， P具有如下式 （IV) 所示的结构，
Figure imgf000013_0001
式 （IV) In certain embodiments, P has the structure shown by formula (IV) below,
Figure imgf000013_0001
Formula (IV)
[0070] 其中， R6为季戊四醇， 甲基葡萄糖苷， 蔗糖， 二甘醇， 丙二醇， 甘 油或聚甘油的羟基去除氢的残基； i为 3， 4， 6或 8; h为 10-1,500的整数。 在 某些实施方式中， h为 100-1,400的整数。 在某些实施方式中， h为 200-1,300 的整数。 在某些实施方式中， h为 300-1,200的整数。 在某些实施方式中， h为 400-1,100的整数。 在某些实施方式中， h为 500-1,000的整数。 在某些实施方 式中， h为 600、 700、 800、 900或 1,000。 Wherein R 6 is a residue at which a hydroxyl group of pentaerythritol, methyl glucoside, sucrose, diethylene glycol, propylene glycol, glycerol or polyglycerol removes hydrogen; i is 3, 4, 6 or 8; h is 10-1,500 Integer. In certain embodiments, h is an integer from 100 to 1,400. In certain embodiments, h is an integer from 200 to 1,300. In certain embodiments, h is an integer from 300 to 1,200. In certain embodiments, h is an integer from 400 to 1,100. In certain embodiments, h is an integer from 500 to 1,000. In some embodiments, h is 600, 700, 800, 900 or 1,000.
[0071] 式 （I) 中的连接基团 X是指在亲水性聚合物和氨基酸寡肽之间起连 接作用的基团。 在某些实施方式中， 引入连接基团是为了将亲水性聚合物改性 以使其更好的与氨基酸寡肽连接。 在某些实施方式中， X为 (CH2)n， (CH2)nCO, (CH2)nOCO， （CH2)nNHCO， -S -， -S02-， -S04-; n为 1-12的整数。 优选的， n 为 1-6。优选的， n为 1-3。优选的， n为 1-2。在某些实施方式中， X为 CH2CO。 The linking group X in the formula (I) means a group which functions as a linking between the hydrophilic polymer and the amino acid oligopeptide. In certain embodiments, the linking group is introduced to modify the hydrophilic polymer to better attach to the amino acid oligopeptide. In certain embodiments, X is (CH 2 ) n , (CH 2 ) n CO, (CH 2 ) n OCO, (CH 2 ) n NHCO, -S -, -S0 2 -, -S0 4 -; n is an integer from 1-12. Preferably, n is 1-6. Preferably, n is 1-3. Preferably, n is 1-2. In certain embodiments, X is CH 2 CO.
[0072】 式 （I) 中的 可以是任意氨基酸或氨基酸类似物残基， 所述氨基 酸或氨基酸类似物残基包含至少一个氨基和一个羧酸基。 在某些实施方式中， A1为具有至少两个羧酸基和一个氨基的氨基酸或氨基酸类似物残基。 Any of the amino acid or amino acid analog residues in formula (I) may comprise at least one amino group and one carboxylic acid group. In certain embodiments, A 1 is an amino acid or amino acid analog residue having at least two carboxylic acid groups and one amino group.
[0073】 在某些实施方式中， 具有如下式 （V) 所示的结构，  [0073] In certain embodiments, having the structure shown by the following formula (V),
— NH— R7— C _ 式 （V) [0074】 其中， R7为 d_2。垸基、 d_2。杂垸基。 在某些实施方式中， ^为 — 1() 垸基、 Cw。杂垸基。 在某些实施方式中， R7为 d_9垸基、 d_9杂垸基。 在某些 实施方式中， R7为 垸基、 d_8杂垸基。在某些实施方式中， R7为 d_7垸基、 C^杂垸基。 在某些实施方式中， R7为 d_6垸基、 d_6杂垸基。 在某些实施方 式中， R7为 d_5垸基、 d_5杂垸基。 在某些实施方式中， R7为 d_4垸基、 d_4 杂垸基。 在某些实施方式中， R7为 d_3垸基、 d_3杂垸基。 在某些实施方式中， R7为甲基、 乙基、 丙基、 丁基、 戊基、 己基、 庚基、 辛基、 壬基、 癸基、 Cu— NH— R 7 — C _ (V) Wherein R 7 is d_ 2 .垸 base, d_ 2 . Clay base. In certain embodiments, ^ is -1() thiol, Cw. Clay base. In certain embodiments, R 7 is d- 9垸, d- 9 heteroalkyl. In certain embodiments, R 7 is an indenyl, d- 8 heterofluorenyl. In certain embodiments, R 7 is d- 7垸, C^hetero. In certain embodiments, R 7 is d- 6 fluorenyl, d- 6 heteropoly. In certain embodiments, R 7 is d — 5 fluorenyl, d — 5 heterofluorenyl. In certain embodiments, R 7 is d 4 alkyl, d 4 heteroaryl. In certain embodiments, R 7 is d — 3 fluorenyl, d — 3 heterofluorenyl. In certain embodiments, R 7 is methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, Cu
^甘 ^甘 ^甘 ^甘 ^甘 ^甘 ^甘 ^甘 兀巷、 Ci2 τη Co τη ΐ4 兀巷、 lis 兀巷、 ΐ6 兀巷、 兀巷、 lis 兀巷、 C19垸基、 C2。垸基。 式 （V) 中的羰基可以与 R7的任何一个碳连接。 在某些实 施方式中， R7为直链的垸基或杂垸基。 在某些实施方式中， R7为支链的垸基或 杂垸基。 ^甘^甘^甘^甘^甘^甘^甘^甘兀巷, Ci2 τη Co τη ΐ4 兀 Lane, lis 兀 Lane, ΐ6 兀 Lane, 兀 Lane, lis Lane, C 19垸, C 2 .垸基. The carbonyl group in the formula (V) may be bonded to any one of the carbons of R 7 . In certain embodiments, R 7 is a linear fluorenyl or heteroindolyl group. In certain embodiments, R 7 is a branched indenyl or heteroindolyl.
[0075] 在某些实施方式中， 具有如下式 （VI) 所示的结构，
Figure imgf000014_0001
（vi) [0075] In some embodiments, having the structure shown by the following formula (VI),
Figure imgf000014_0001
(vi)
[0076] 其中， R8、 R9独立的为氢、 d_12垸基、 d_12杂垸基、 芳基、 杂芳基、 芳垸基、 杂芳垸基、 且在每个重复单元中的 R8、 R9可以相同或不同； j为 1-10 的整数。 在某些实施方式中， j为 1-10的整数。 在某些实施方式中， j为 1-9的 整数。 在某些实施方式中， j为 1-8的整数。 在某些实施方式中， j为 1-7的整 数。 在某些实施方式中， j为 1-6的整数。 在某些实施方式中， j为 1-5的整数。 在某些实施方式中， j为 1-4的整数。 在某些实施方式中， j为 1-3的整数。 在 某些实施方式中， j为 1-2的整数。 Wherein R 8 and R 9 are independently hydrogen, d — 12 fluorenyl, d— 12 heterofluorenyl, aryl, heteroaryl, aryl fluorenyl, heteroaryl fluorenyl, and in each repeating unit. R 8 and R 9 may be the same or different; j is an integer of 1-10. In certain embodiments, j is an integer from 1 to 10. In certain embodiments, j is an integer from 1-9. In certain embodiments, j is an integer from 1-8. In certain embodiments, j is an integer from 1-7. In certain embodiments, j is an integer from 1 to 6. In certain embodiments, j is an integer from 1 to 5. In certain embodiments, j is an integer from 1 to 4. In certain embodiments, j is an integer from 1 to 3. In certain embodiments, j is an integer from 1 to 2.
[0077] 在某些实施方式中， 1为天冬氨酸或谷氨酸的残基。 [0078] 式 （I) 中的 A2、 A3独立的可以是任意氨基酸或氨基酸类似物残基， 所述氨基酸或氨基酸类似物残基包含至少一个氨基和一个羧酸基。 在某些实施 方式中， A2、 A3独立的具有如下式 （VII) 所示的结构，
Figure imgf000015_0001
式 （VII) In certain embodiments, 1 is a residue of aspartic acid or glutamic acid. A 2 and A 3 in the formula (I) may independently be any amino acid or amino acid analog residue, and the amino acid or amino acid analog residue contains at least one amino group and one carboxylic acid group. In certain embodiments, A 2 and A 3 independently have the structure shown by the following formula (VII),
Figure imgf000015_0001
Formula (VII)
[0079] 其中， R1Q、 Ru独立的为氢、 d_6垸基、 d_6杂垸基且在每个重复单 元中的 R1()、 Ru可以相同或不同； h为 1-10的整数。 在某些实施方式中， R10、 Ru独立的为甲基、 乙基、 丙基、 丁基、 戊基、 己基。 在某些实施方式中， R10、 Ru独立的为直链的垸基或杂垸基。 在某些实施方式中， R1Q、 Ru独立的为支链 的垸基或杂垸基。 在某些实施方式中， h为 1、 2、 3、 4、 5、 6、 7、 8、 9、 10。 Wherein R 1Q and R u are independently hydrogen, d 6 fluorenyl, d 6 heterofluorenyl, and R 1 ( ) , Ru in each repeating unit may be the same or different; h is 1-10 Integer. In certain embodiments, R 10 , Ru are independently methyl, ethyl, propyl, butyl, pentyl, hexyl. In certain embodiments, R 10 , Ru are independently linear fluorenyl or heteroindolyl. In certain embodiments, R 1Q , Ru is independently branched fluorenyl or heterofluorenyl. In certain embodiments, h is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10.
[0080] 在某些实施方式中， A2、 A3独立的为甘氨酸、 丙氨酸、 亮氨酸、 异 亮氨酸、 缬氨酸、 脯氨酸、 苯丙氨酸、 蛋氨酸、 丝氨酸、 苏氨酸、 半胱氨酸和 酪氨酸的残基。 In certain embodiments, A 2 , A 3 are independently glycine, alanine, leucine, isoleucine, valine, valine, phenylalanine, methionine, serine, Residues of threonine, cysteine and tyrosine.
[0081] 本发明的水溶性聚合物-氨基酸寡肽-药物的结合物中的药物化合物 包括任意可以与氨基酸或氨基酸类似物结合的药物分子。 在某些实施方式中， 可以与氨基酸或氨基酸类似物结合的药物分子包含一个官能团如氨基、 羟基、 酚基、 硫基或胍基。 含有氨基、 羟基、 酚基、 硫醇基或胍基的药物分子的例子 包括但不限于：醋羟胺酸、阿昔洛韦 {2-氨基 -1， 9-二氢 -9-[(2-羟乙氧基）甲基] 6H- 嘌呤 -6-酮}、 别嘌呤醇、 腺苷 （6-氨基 -9-i3-D-呋核亚硝脲 -9-H-嘌呤） 、 脱氢皮 质 （留）醇、 泼尼松、 曲安奈德、 考的索（氢化可的松） 、 腺苷（6-氨基 -9-β-ϋ- 呋核 亚硝脲 -9-H-嘌呤） 、 可的松、 雌二醇、 雌性素、 雌三醇、 16-羟雌固酮、 马烯雌 （留） 酮、 马萘雌酮、 双烯雌酚、 己雌酚、 己烯雌酚、 苯雌酚、 4-羟雄 留烯二酮、 ICI 164384、氨鲁米特、 ICI 182780、 7-氨基苯基硫代雄 _4_烯 -3、 17- 二酮、 甲地孕酮、 氯化孕酮、 甲基炔诺酮、 炔雌烯醇、 美雄酮、 米非司酮、 奥 那司酮、 达那唑、 美替诺龙、 司坦唑、 阿米卡星 （D-链霉胺） 、 9_氨基吖唆、 氨吖 唆、 阿托伐醌、 氯苯氨丁酸、 骨化二醇、 骨化三醇、 苯丙醇胺、 卡托普利 {l-[(2S)-3-巯基 -2-甲基丙酰基] -L-脯氨酸 }、仲丁巴比妥、卡马西 平、卡比多巴、 茶碱、 左旋多巴、 假麻黄瓰、 氯霉素、 二氯羟喹、 氯碘羟喹、 氯二甲苯酚、 氯 苯甘油氨酯、 氯噻酮、 苯丙醇胺、 氯压定 [2-(2， 6-二氯苯胺 基） -2-咪唑啉]、 克拉屈滨、 盐酸苯丙醇胺、 氯硝西泮、 阿糖胞苷 [4-氨基 -l-β -D-阿拉伯呋喃基 -2-(1Η)_嘧啶酮]、 达那唑、 右泛醇、 愈创甘油醚、 正定霉素、 阿霉素、 去甲氧 正定霉素、右旋甲状腺素 [D-3， 5， 3'，5'-四碘甲腺原氨酸]、地达诺新、地佐辛、 多巴胺、 二氢速留醇、 双香豆素、 屈大麻酚、 二羟丙茶碱、 依诺沙星、 恩纳普 利 [(S)-1_[N-(1-羧基 -3-苯丙基） -L-丙胺酰 -L-脯氨酸]、双烯雌酚、卡泊三烯 [(5Z， 7E， 22E， 24S) -24-环丙基 -9， 10-开环胆甾_5， 7， 10 (19)， 22-四烯 -1 α， 3 β， 24-三醇]、钙化甾醇 [9， 10-secoergsta-5, 7, 10 (19)， 22-四烯 -3-醇， （3 β， 5Ζ， 7Ε， 22Ε)], 左炔诺孕酮、 甲基炔诺酮、 炔诺酮、 甲基苄胼、 泛西洛维、 费乐地平、 肟炔诺酮、 氟尿苷、 疱疹净、 依托泊苷、 莫诺苯宗、 磷酸氟达拉滨、 二氢 速留 醇、 非那司提、 氟康唑、 氟达拉滨、 氟尿嘧啶、 氟胞嘧啶、 乙氯维诺、 氟米龙、 卤贝他索、 莫米松、 氟伏沙明、 氟氢缩松、 更昔洛韦、 氟地松、 去氧孕烯、 乙 炔 雌二醇、 炔雌醇、 炔雌醇甲醚、 去羟米松、 地塞米松、 庆大霉素、 羟孕酮、 甲孕酮、 吲达胺、 左多巴、 甲基多巴、 胼苯哒嗪、 二氢氯噻、 氢氟噻嗪、 双碘 喹啉、卡那 徽素、洛弗斯塔特因、马丙考、劳拉西泮、去甲羟基安定、 甲羟松、 甲基苯巴比妥、 甲苯喹唑酮、 美他沙酮、 美索巴莫、 甲氯噻嗪、 甲硝哒唑、 巯 嘌呤、 甲硫咪唑、 甲氨蝶呤、 米利酮、 南诺龙、 萘唑啉、 美西律、 呋喃妥英、 氯硝柳胺、 硝苯吡啶、 尼莫地平、 去甲肾上腺素、 新生霉素、 奥美拉唑、 氧雄 龙、苯异妥英、潘他米丁、 羟甲烯龙、奥美拉唑、氧雄龙、去甲二氢愈创木酸、 扎鲁司特、 Banzel (卢非酰胺） ， 苯乙酰脲、 苯乙胼、 苯偶氮毗胺、 苯巴比妥、 硫代异恶唑、 酚妥拉明、 苯妥英、 普达非洛、 甲基苄胼、 泊利噻嗪、 三氯噻嗪、 普里米酮、 普罗布考、 异丙酚、 丙基硫尿嘧啶、 甲基苄胼、 甲基苄胼、 磺胺多 辛、 喹乙宗、 丙基硫尿嘧啶、 三氮唑核甙、 链脲霉素、 双甲丙酰龙、 斯伐他汀、 卡林酰胺、司坦唑、磺胺甲恶 唑、磺胺甲恶唑、硫代异恶唑、磺胺、磺胺嘧啶、 柳氮磺吡啶、 替马西泮、 特拉唑嗪、 他克林、 噻苯咪唑、 戊硫代巴比妥、 苯甲 唑啉、硫鸟嘌呤、 奥美沙坦酯 [(5-甲基 -2-酮 -1， 3-二氧 -4-基） 甲基 -5-G-羟基小 甲基-乙基） -2-丙基 -3-[ W-[2-QH-四唑 -5-基） -苯基]-苯基]甲基] -3H-咪唑 -4-甲酸 酯]、 替尼泊甙、 托塞米、 氨苯蝶啶、 三氟胸苷、 甲氧苄氨嘧啶、 曲美沙特、 乌 拉莫司汀、 托品酰胺、 阿糖腺苷、 华法林、 扎西他宾、 齐多呋定、 氟替卡松糠 酸盐、 Ro 46-2005， 波生坦、 克拉生坦、 替唑生坦、 拉替拉韦 {N-[(4-氟苯基） 甲基] -1， 6-二氢 -5-羟基小甲基 -2-[1-甲基 -1[[(5- 甲基 -1， 3， 4-恶二唑 -2-基） 羰 基]氨基]乙基] -6-酮 -4-嘧啶甲酰胺 }、 阿利吉仑 (2S，4S， 5S， 7R) -5-氨基 -N- (2- 氨基甲酰基 _2_甲基-丙基） _4_羟基 -7-114-甲氧基 -3- (甲氧基丙氧基) -苯基]甲 基 8_甲基 -2-丙基 -2-基-壬酰胺、 依非韦伦、 右旋安非他命、 非那司提、 阿莫 达非尼、 阿尼芬净、 地瑞纳韦、 替拉那韦、 安泼那韦、 贝卡那韦、 替比夫定、 来那度胺、 擦里多米德、 恩替卡韦、 考尼伐 坦、 索拉非尼 （多吉美） 、 恩替 卡韦 （博路定） 、 氮杂胞嘧啶 （维达扎） 、 培美曲塞 （阿灵达） 、 雷美替胺、 依泽替米贝、 克罗拉滨 （氯法拉滨） 、 奈拉滨、 埃罗替尼 （得舒缓） 、 他达拉 非 （希爱力）、 安泼那韦、 阿扎那韦（瑞塔滋）、依泽替米贝、对乙酰氨基酚、 格列波脲、 依曲韦林、 阿巴卡韦 （Ziagen)、 N-[1-[(2R, 3R， 4S， 5R) -3,4- 二 羟基 -5-甲基四氢呋喃 _2_基] _5_氟 -2-酮嘧啶 -4-基] 胺、 替诺福韦、 伏立康唑、 双氢氯噻嗪、 唑来膦酸、 褪黑激素、 3-氨基丙垸磺酸、 氟维司群、 伏立康唑、 白藜芦醇、 洛伐它丁、 替诺福韦酯、 替诺福韦、 辛伐他 汀、 戊垸基 N-[1-K2R， 3R， 4S， 5R) -3,4- 二羟基 -5-甲基四氢呋喃 -2-基] -5-氟 -2-酮嘧啶 -4-基]氨基甲酸 酯 （ （卡培他滨） 、 钙化醇 （维生素 D2)、 胆钙化醇 （维生素 D3)、 1， 25-二 羟胆钙化醇、 拉米夫定、 度骨化醇（l et-羟基维生素 D2)、 二氢速留醇（维生素 D4)、 洛匹那韦、 3-[4-(4_氯苯基)环己基 ]-4-羟基萘 -1， 2-二酮、 西多福韦、 利托 那韦、 恩他卡 朋、 他达拉非 （希爱力） 、 非那司提、 齐留通、 退黑激素、 特 敏福 （奥司他韦） 、 帕立骨化醇、 甲硝哒唑、 二氟尼柳、 阿司匹林、 昔康、 吉 鲁威尔 （西他列汀)、 恩曲他滨 5-氟 -1- (2R， 5S) - [2-羟甲基 -1， 3-氧硫环 -5-基] 胞嘧啶、 异丙酚、 维生素 A类似物、 依维莫司 （1R， 9S， 12S， 15R， 16E， 18R， 19R， 21R， 23S , 24E, 26E, 28E, 30S, 32S, 35R)-1 , 18- 二羟基 -12- {(IR) -2- [ (18，3 ，411) -4- (2-轻基乙氧基)_3_ 甲氧基环己基 ]-1-甲基乙基}-19， 30-二甲氧 基 -15， 17， 21， 23， 四， 35-六甲基 -1 1， 36- 二氧杂 -4-氮杂 -三环 [30. 3. 1. 04， 9]三十六 -16， 24， 26， 28-四烯 -2，3，10，14， 20-五酮]、 姜黄素、 替拉那韦、 依曲 韦林、 他达拉非、 托伐普坦、 多肽、 DNAs，RNAs、 腺嘌呤、 鸟嘌呤、 胞核嘧 啶、 胞核嘧啶和尿嘧啶。 The pharmaceutical compound in the water-soluble polymer-amino acid oligopeptide-drug conjugate of the present invention includes any drug molecule which can bind to an amino acid or an amino acid analog. In certain embodiments, a drug molecule that can bind to an amino acid or amino acid analog comprises a functional group such as an amino group, a hydroxyl group, a phenol group, a thio group or a thiol group. Examples of drug molecules containing an amino group, a hydroxyl group, a phenol group, a thiol group or a thiol group include, but are not limited to, acetaminophen, acyclovir {2-amino-1,9-dihydro-9-[(2- Hydroxyethoxy)methyl] 6H-indol-6-one}, allopurinol, adenosine (6-amino-9-i3-D-furonucleoside nitrosourea-9-H-indole), dehydrocortic (retained) alcohol, prednisone, triamcinolone acetonide, cortisone (hydrocortisone), adenosine (6-amino-9-β-indole-furan nitrosourea-9-H-嘌呤), Pine, estradiol, estrogen, estriol, 16-hydroxyestrene, equine estradiol, equine estrone, diethestrol, cresyl, diethylstilbestrol, estrone, 4 -hydroxyandrostenedione, ICI 164384, aminoglutethimide, ICI 182780, 7-aminophenylthio male-4-ene-3, 17- Diketone, megestrol acetate, progesterone, methaqualone, ethinyl estradiol, medroxacin, mifepristone, ol's ketone, danazol, minotinol, stanozol, Amikacin (D-streptavidin), 9-aminoguanidine, aminoguanidine, atovaquone, chlorpheniric acid, calcifediol, calcitriol, phenylpropanolamine, Cato Puli {l-[(2S)-3-mercapto-2-methylpropanoyl]-L-valine}, sec-butrazine, carbamazepine, carbidopa, theophylline, levodopa , pseudoephedrine, chloramphenicol, dichloroquine, clioquinol, chloroxylenol, chlorophenyl glyceryl urethane, chlorthalidone, phenylpropanolamine, clonidine [2-(2, 6- Dichloroanilino)-2-imidazoline], cladribine, phenylpropanolamine hydrochloride, clonazepam, cytarabine [4-amino-l-β-D-arabinofuran-2-(1Η )_pyrimidinone], danazol, dexpanthenol, guaifenesin, gentamicin, doxorubicin, noroxyneggiomycin, dextrothyroxine [D-3, 5, 3', 5' -tetraiodothyronine], darnoxine, dextrozine, dopamine, dihydrocohol, dicoumarin Dronabinol, hydroxypropyl theophylline, enoxacin, enalapril [(S)-1_[N-(1-carboxy-3-phenylpropyl)-L-alaninyl-L-proline ], diethylstilbestrol, calcipotriene [(5Z, 7E, 22E, 24S) -24-cyclopropyl-9, 10-open ring cholesterium_5, 7, 10 (19), 22-tetraene -1 α, 3 β, 24-triol], calcified sterol [9, 10-secoergsta-5, 7, 10 (19), 22-tetraen-3-ol, (3 β, 5Ζ, 7Ε, 22Ε) ], levonorgestrel, norgestrel, norethisterone, procarbazine, panciclovir, felodipine, norethisterone, fluorouridine, herpes net, etoposide, mono Benzoin, fludarabine phosphate, dihydrofuranol, finasteride, fluconazole, fludarabine, fluorouracil, flucytosine, chlorophene, flumethonol, halobetasol, mo Rice pine, fluvoxamine, hydrofluoric acid, ganciclovir, fluticasone, desogestrel, ethinyl estradiol, ethinyl estradiol, ethinyl estradiol methyl ether, deshydroxymetasone, dexamethasone, Qing Daramycin, hydroxyprogesterone, medroxyprogesterone, indamine, levodopa, methyldopa, hydralazine, dihydrochlorothiazide Hydrofluorothiazide, bis-iodoquinoline, carbamazepine, lovastatin, mazinone, lorazepam, noroxydamine, hydroxyxanthone, methylphenobarbital, toloxazolone, Metaxalone, mesobaramo, methyl chlorothiazide, metronidazole, guanidine, methimazole, methotrexate, milidone, nanolone, naphtholine, mexiletine, nitrofurantin , niclosamide, nifedipine, nimodipine, norepinephrine, novobiocin, omeprazole, oxhonol, phenytoin, peptamide, oxymetholone, omeprazole Oxazole, oxo-dragon, nordihydroguaiaretic acid, Zaluzast, Banzel (Lufamide), Phenylaceazurea, Phenylacetamidine, Phenazoquinazoline, Phenobarbital, Thioisoxazole, Phentolamine, Phenytoin, Pudafilol, A Benzylidene, policoazine, trichlorothiazide, primidone, probucol, propofol, propylthiouracil, procarbazine, procarbazine, sulfadoxine, quinidine , propyl thiouracil, triazole ribavirin, streptozotocin, dipropionyl hydrazide, simvastatin, carinamide, stanozol, sulfamethoxazole, sulfamethoxazole, thioisoxine Azole, sulfonamide, sulfadiazine, sulfasalazine, temazepam, terazosin, tacrine, thiabendazole, pentothiobarbital, benzazoline, thioguanine, olmesartan medoxomil [( 5-methyl-2-keto-1,3-dioxy-4-yl)methyl-5-G-hydroxysuccinymethyl-ethyl)-2-propyl-3-[ W-[2-QH -tetrazol-5-yl)-phenyl]-phenyl]methyl]-3H-imidazole-4-carboxylate], teniposide, torsemide, triamterene, trifluorothymidine, Trimethoprim, trimesat, uraramustine, tropamide, arabin Adenosine, warfarin, zalcitabine, zidovudine, fluticasone furoate, Ro 46-2005, bosentan, clastatin, tazoxan, latiravir {N-[(4 -fluorophenyl)methyl]-1,6-dihydro-5-hydroxysuccinic methyl-2-[1-methyl-1[[(5-methyl-1,3,4-oxadiazole- 2-yl)carbonyl]amino]ethyl]-6-one-4-pyrimidinecarboxamide}, aliskiren (2S,4S, 5S, 7R) -5-amino-N- (2-carbamoyl-2 _Methyl-propyl) _4_hydroxy-7-114-methoxy-3-(methoxypropoxy)-phenyl]methyl 8-methyl-2-propyl-2-yl-oxime Amide, efavirenz, dextroamphetamine, finasteride, amodafinil, anifenyl, darunavir, telanavir, amprenavir, bekanavir, telbif Dent, lenalidomide, rubidide, entecavir, tonic valtan, sorafenib (Norgie), entecavir (Boluding), aza-cytosine (Vidaza), pemetrexed ( Alinga, rametamine, ezetimibe, clolapabine (clofarabine), naribine, erlotinib (soothing), tadala (Xi-li), amprenavir, atazanavir (Ritazi), ezetimibe, acetaminophen, glibenclamide, etravirine, biacavir, N-[1-[(2R, 3R, 4S, 5R)-3,4-dihydroxy-5-methyltetrahydrofuran-2-yl]-5-fluoro-2-ketopyrimidin-4-yl]amine, teno Fowe, voriconazole, hydrochlorothiazide, zoledronic acid, melatonin, 3-aminopropionine, fulvestrant, voriconazole, resveratrol, lovastatin, tenofovir, teno Fowey, simvastatin, pentamidine N-[1-K2R, 3R, 4S, 5R) -3,4-dihydroxy-5-methyltetrahydrofuran-2-yl]-5-fluoro-2-one pyrimidin-4-yl]carbamate ((capecitabine), Calcified alcohol (vitamin D2), cholecalciferol (vitamin D3), 1,25-dihydroxycholecalciferol, lamivudine, calciferol (l et-hydroxyvitamin D2), dihydrofuranol (vitamin) D4), lopinavir, 3-[4-(4-chlorophenyl)cyclohexyl]-4-hydroxynaphthalene-1,2-dione, cidofovir, ritonavir, entacapone , tadalafil (Xi Li Li), phenacetin, zileuton, melatonin, tamiflu (oseltamivir), paricalcitol, metronidazole, diflunisal, Aspirin, oxicam, girruvir (sitagliptin), emtricitabine 5-fluoro-1-(2R, 5S)-[2-hydroxymethyl-1, 3-oxothio-5-yl Cytosine, propofol, vitamin A analogues, everolimus (1R, 9S, 12S, 15R, 16E, 18R, 19R, 21R, 23S, 24E, 26E, 28E, 30S, 32S, 35R)-1 , 18-dihydroxy-12- {(IR) -2- [(18,3 ,411) -4-(2-light ethoxy)_3_methoxycyclohexyl]-1-methylethyl} -19, 30-two Oxyl-15, 17, 21, 23, tetra, 35-hexamethyl-1 1, 36-dioxa-4-aza-tricyclo[30. 3. 1. 04, 9]36- 16, 24, 26, 28-tetraene-2,3,10,14,20-pentanone], curcumin, telanavir, etravirine, tadalafil, tolvaptan, peptide, DNAs, RNAs, adenine, guanine, cytosine, cytosine and uracil.
[0082] 在某些实施方式中，可以与氨基酸或氨基酸类似物结合的药物分子包 含一个羧基或一个磷酸基 /膦酸基。含有羧基的药物分子的例子包括，但不限于： 美沙雌酸、 氨基水杨酸、 美沙雌酸、 氨基 水杨酸、 巴氯芬、 卡别多巴、 左旋多 巴、氨基苯甲酸、布美他尼、卡托普利 [l-[(2S)-3-巯基 -2-甲基丙酰基] -L-脯氨酸]、 西司他丁 [(2)-7-[[(3)-2-氨基 -2-羧乙基]硫代] -2- [ (S) -2， 2-二甲基环丙甲酰胺 基] -2-庚酸]、 左甲状腺素 [D-3， 5， 3'， 5'-四碘甲腺原氨酸]、 两性霉素 B、 依 曲替酯、 依氟鸟氨酸、 10-十一烯酸、 西诺沙星、 氯卓酸、 环丙氟啶酸 [1 -环丙基 -6-M-1 , 4- 二氢 -4-酮 -7-(1-哌嗪基） -3-喹啉酸]、 色甘酸钠、 去氢胆酸、 依那普 利 [(S)-1_[N-(1-羧基 -3-苯丙基） -L-丙胺酰] -L-脯氨 酸]、 依诺沙星、 利尿酸、 利 尿磺胺、 吉非罗奇、十八烯酸、 2-[4-(4_氯苯甲酰基） _苯氧基 ]-2-甲基 -丙酸（非 诺贝特酸)、 7-[ (IS, 3R， 7S， 8S,8aR)-l-(2S) -2-甲基丁酰氧基 _3， 7- 二甲基 -1， 2， 3， 7， 8， 8a_六氢萘小基] [(3R， 5R)-3 , 4-二羟基庚酸]、 加巴喷丁、 福辛普利、 普伐他汀、 阿加曲班、 7-茶碱乙酸、 碘番酸、 碘塞罗宁、 碘酞酸、 洛度酰胺 [N， N' -(2-氯 -5-氰基 -m-亚苯基）二草酸]、丙磺舒、赖诺普利 [(S)-1-[N- (1-羧基 -3-苯丙基） -L-赖氨酰] -L-脯氨酸]、 甲氨蝶呤、 乙酰基氨基丙垸磺酸盐、 奈多罗米、 硫代水杨酸、 喹那普利、 雷米普利、 氟哌酸、 碘克酸、 柳氮磺胺吡 啶、 帕伐他丁、丙戊酸、 奥美沙坦、安贝生坦（Ietairis)、达卢生坦、 壬二酸（杜 鹃花酸） 、 熊去氧胆酸、 氧氟沙星、 TAK-044{: 环 [D-门冬氨酰 -3-[(4-苯基哌 嗪小基）羰基： |_L-丙胺基 -L-门冬氨酰 -D-2-(2-噻吩基）氨基乙酰基 -L-亮氨酰 -D- 色氨酰 ]}、 BQ123 (环 [D-色氨酸 -D-天冬氨酸-脯氨酸 -D-缬氨酸-亮氨酸 {cyclo [D-Trp-D-Asp-Pro-D-Val-Leu]} , 阿伐他汀 （立普妥）、氟替卡松糠、 鲁比前列 素 （鲁比前列酮） 、 普加巴林 -13(LyriCa)、 培美曲塞 （力比泰） 、 曲前列环素、 罗苏伐他汀 （冠脂妥） 、 甲基多巴、 缬沙坦、 替米沙坦、 （E)-5-[[-4_(2-羧乙 基）氨甲酰]苯基]氮杂] -2-羟基苯甲酸、依普罗沙坦、依普罗沙坦、氟伐他汀（来 适可） 、 （E)-5-[-4-(2- 羧基）氨甲酰]苯基]氮杂] -2-羟基苯甲酸、 天门冬酰胺- 丙氨酸-脯氨酸-缬氨酸 -色氨酸-异亮氨酸 -脯氨酸-谷氨酰胺 （Asn-Al a-Pro-Val-Ser-IIe-Pro-Gln) , 2-萘乙酸、 舒洛芬、 3- (2-噻吩基羰基） -苯乙酸、 依 布洛芬、 氟比洛芬、 阿司匹林、 卡洛芬、 普拉洛芬、 阿明洛芬、 苯惡洛芬、 吲 哚洛芬、 己洛芬、 10， 11-二氢 -10-酮-二苯并 [b， f]硫杂卓 -2-羧酸、 W-(2-酮环- 戊基） -甲基]苯甲酸、 [5-苯基 -(2-噻吩） ]-羧酸、 （3-苯氧基苯基） 乙酸、 4- (4- 氯苯基） -2-苯基 -5-噻唑乙酸、 4- (2， 5- 二氢吡咯 -1-基） -苯乙酸、 4， 5-联苯 -2-噁唑丙酸、 [4-2-酮环戊垸） -甲基]苯乙酸、 10， 11- 二氢 -10-酮-二苯并 [b， f]硫杂卓 -2-羧酸、 5-环己基 -2， 3- 二氢 -IH茚基 -1-羧酸、 5-苯基 -2-呋喃丙酸、 Y -酮 -(1， 1 '-联苯） -4- 丁酸、 5-苯甲酰 -2， 3- 二氢 -IH-吡咯羧酸、 苯亚甲基 -IH- 茚 -3-乙酸、 1-苯甲酰 -5-甲氧基 -2-甲基 -IH-吲哚 -3-乙酸、 4-苯甲酰 -IH-吡咯 -2-乙 酸、 1，3，4，9-四氢吡喃 -[3，4-b]吲哚 -1-乙酸、 3-苯氨基-苯乙酸、 2-苯氨基 -苯乙 酸、 3-(4-氯苯基） -1-苯基 -IH-吡唑 -4-乙酸、 4-(2-丙烯氧基）苯乙酸、 2-苯基 -5- 噻唑-乙酸、 4- (6-甲氧基 -2-萘基 -3-丙酸、 乙酰水杨酸、 3-苯基苯甲酸、 二聚水 杨酸酯、 [(1-苄基 -IH-吲唑 -3-基） 氧]乙酸、 三聚水杨酸酯、 柳氮磺胺吡 啶、 2- 苯氨基吡啶 -3-羧酸、 艾曲波帕 （eltrambopag) 孟鲁司特、 苯达莫司汀 （达莫 司汀） 、 前列腺素 E2、 前列腺素 F2a、 卡前列腺素 （15-甲基前列腺素 F2a)、 前列腺素 D2、 前列腺素 E1 (前列地尔)、 前列腺素 Fl a、 (Z) -7- [ (1R, 2R， 3R， 5S) -3, 5- 二羟基 -2- [ (E， 3S) -3-羟基 -5-苯基 + 戊烯基]环戊基 ]-5-庚烯酸、 (E) -7- [ (1R， 2R， 3R， 5S) -3， 5- 二羟基 -2- [ (3R) -3-羟基 -5-苯基戊基]环戊基 ]-5- 庚烯酸、 前列腺素 (； 12(环前列腺素） 、 （Z)-7-[(lR， 2R， 3R， 5S)-3， 5-二羟 基 -2-[(E， 3R)-3-羟基 -4-[3- (三氟甲基）苯氧基]丁烯基]环戊基] _5_庚烯酸、 （E) -7-[ (1R, 2R， 3R， 5S) -3, 5- 二羟基 -2- (-3-酮癸)环戊基 ]5-庚烯酸、 米索前列 醇、 吉美前列素、 7-[3_羟基-2-3 (3-羟基 -4-苯氧基 -1- 丁烯基） -5-酮环戊基] _5_ 庚烯酸、 芬前列林、 前列腺素 Al、 前列腺素 A2、 前列腺素 Bi、 前列腺素 A2、 视黄酸、 蓓萨罗丁、 9-顺式维甲酸 （阿利维 A 酸） 、 视黄酸类似物、 13-顺式 视黄酸 （异维 A酸） 、 蓓萨罗丁类似物、 蓓萨罗丁类似物、 青霉素 G、 苯氧基 甲基青霉素、 甲氧苯青霉素、 苯唑西林、 哌拉西 林、 美洛西林、 羧苄青霉素、 a -替卡西林、 氨苄西林、 甲亚胺青霉素、 头孢菌素、 头孢吡硫、 头孢唑啉、 头 孢羟氨苄、 头孢拉定、 头孢尼西、 头孢孟多、 头孢呋辛、 头孢西丁、 头孢雷特、 头孢替 坦、 头孢呋肟、 氯碳头孢、 头孢噻肟、 头孢曲松、 头孢哌酮、 拉氧头 孢、 LIVALO (匹伐他汀）、 Tyvaso (曲前列素）、 R)lotyn (；普拉曲沙）、达菲（奥 司他韦） 、 贝前列素。 [0083] 在某些实施方式中， 01和1¾各自独立的为抗肿瘤药物的残基。 在某 些实施方式中， 抗肿瘤药物的残基能够与 A2或 A3形成肽键或酯键。 在某些实 施方式中， 抗肿瘤药物通过修饰从而能够与 A2或 A3形成肽键或酯键。 在某些 实施方式中， 抗肿瘤药物为达沙替尼、 雷帕霉素、 依诺替康、 伊马替尼、 厄洛 替尼、 吉非替尼、 拉帕提尼、 索拉非尼、 舒尼替尼、 紫杉醇、 喜树碱、 华蟾酥 毒基、 甘草次酸、 东莨菪内酯。 In certain embodiments, a drug molecule that can bind to an amino acid or amino acid analog comprises a carboxyl group or a phosphate group/phosphonic acid group. Examples of drug molecules containing a carboxyl group include, but are not limited to, mesalic acid, aminosalicylic acid, mesalic acid, aminosalicylic acid, baclofen, carbidopa, levodopa, aminobenzoic acid, bumami Tani, captopril [l-[(2S)-3-mercapto-2-methylpropionyl]-L-valine], cilastatin [(2)-7-[[( 3 ) -2-amino-2-carboxyethyl]thio]-2-[(S)-2,2-dimethylcyclopropanecarboxamide]-2-heptanoic acid], levothyroxine [D-3, 5, 3', 5'-tetraiodothyronine], amphotericin B, etretinate, etflurosine, 10-undecenoic acid, cinoxacin, clozapine, ring Propoferic acid [1-cyclopropyl-6-M-1, 4-dihydro-4-keto-7-(1-piperazinyl)-3-quinolinic acid], sodium cromoglycate, dehydrocholin Acid, enalapril [(S)-1_[N-(1-carboxy-3-phenylpropyl)-L-alaninyl]-L-valine], enoxacin, diuretic acid, diurea , Gemfibrozil, oleic acid, 2-[4-(4-chlorobenzoyl)-phenoxy]-2-methyl-propionic acid (non- Norbert acid), 7-[ (IS, 3R, 7S, 8S, 8aR)-l-(2S)-2-methylbutyryloxy_3, 7-dimethyl-1, 2, 3, 7, 8, 8a_hexahydronaphthalene] [(3R, 5R)-3, 4-dihydroxyheptanoic acid], gabapentin, fosinopril, pravastatin, argatroban, 7-theophylline acetate , iopanoic acid, liothyronine, iodonium, lordamide [N, N'-(2-chloro-5-cyano-m-phenylene) dioxalic acid], probenecid, lynop [(S)-1-[N-(1-carboxy-3-phenylpropyl)-L-lysyl]-L-proline], methotrexate, acetylaminopropane sulfonate , nedocromil, thiosalicylic acid, quinapril, ramipril, norfloxacin, iodoic acid, sulfasalazine, pravastatin, valproic acid, olmesartan, ambergen Itaiiris, dalusentan, azelaic acid (rhodoic acid), ursodeoxycholic acid, ofloxacin, TAK-044{: ring [D-aspartyl-3-[(4- Phenylpiperazine small base)carbonyl: |_L-propylamino-L-aspartyl-D-2-(2-thienyl)aminoacetyl-L-leucyl-D-tryptoyl], BQ123 (cyclo [D-tryptophan-D-aspartate-valine-D-valine-bright Acid {cyclo [D-Trp-D-Asp-Pro-D-Val-Leu]} , atorvastatin (Lipitor), fluticasone, lubiprost (lubiprostone), pregabalin-13 (Lyri C a), pemetrexed (Libitai), treprostinil, rosuvastatin (crown lactide), methyldopa, valsartan, telmisartan, (E)-5-[ [-4_(2-carboxyethyl)carbamoyl]phenyl]aza]-2-hydroxybenzoic acid, eprosartan, eprosartan, fluvastatin (Approx), (E)- 5-[-4-(2-carboxy)carbamoyl]phenyl]aza]-2-hydroxybenzoic acid, asparagine-alanine-valine-valine-tryptophan-isoluminescence -proline-glutamine (Asn-Al a-Pro-Val-Ser-IIe-Pro-Gln), 2-naphthylacetic acid, suloprofen, 3-(2-thienylcarbonyl)-phenylacetic acid, Ibuprofen, flurbiprofen, aspirin, carprofen, pranoprofen, aminprofen, phenoxaprofen, ibuprofen, hexprofen, 10, 11-dihydro-10-keto- Dibenzo[b,f]thiazepine-2-carboxylic acid, W-(2-ketocyclopentyl)-methyl]benzoic acid, [5-phenyl-(2-thiophene)]-carboxylic acid (3-phenoxybenzene Acetate, 4-(4-chlorophenyl)-2-phenyl-5-thiazoleacetic acid, 4-(2,5-dihydropyrrol-1-yl)-phenylacetic acid, 4, 5-biphenyl- 2-oxazopropionate, [4-2-ketocyclopentanyl)-methyl]phenylacetic acid, 10, 11-dihydro-10-keto-dibenzo[b,f]thiazepine-2-carboxylate Acid, 5-cyclohexyl-2,3-dihydro-IH-decyl-1-carboxylic acid, 5-phenyl-2-furanopropionic acid, Y-keto-(1, 1 '-biphenyl)-4- Butyric acid, 5-benzoyl-2,3-dihydro-IH-pyrrolecarboxylic acid, benzylidene-IH- Indole-3-acetic acid, 1-benzoyl-5-methoxy-2-methyl-IH-indole-3-acetic acid, 4-benzoyl-IH-pyrrole-2-acetic acid, 1,3, 4,9-tetrahydropyran-[3,4-b]indole-1-acetic acid, 3-phenylamino-phenylacetic acid, 2-phenylamino-phenylacetic acid, 3-(4-chlorophenyl)-1 -phenyl-IH-pyrazole-4-acetic acid, 4-(2-propenyloxy)phenylacetic acid, 2-phenyl-5-thiazole-acetic acid, 4-(6-methoxy-2-naphthyl- 3-propionic acid, acetylsalicylic acid, 3-phenylbenzoic acid, dimerized salicylate, [(1-benzyl-IH-indazol-3-yl)oxy]acetic acid, trimeric salicylate , sulfasalazine, 2-phenylaminopyridine-3-carboxylic acid, eltrambopag, montelukast, bendamustine (damlastatin), prostaglandin E2, prostaglandin F2a, card Prostaglandins (15-methyl prostaglandin F2a), prostaglandin D2, prostaglandin E1 (pro-lindil), prostaglandin Fl a, (Z) -7- [ (1R, 2R, 3R, 5S) -3, 5 - Dihydroxy-2-[(E, 3S)-3-hydroxy-5-phenyl+pentenyl]cyclopentyl]-5-heptenoic acid, (E) -7- [ (1R, 2R, 3R , 5S) -3, 5-dihydroxy-2-[(3R)-3-hydroxy-5-phenylpentyl]cyclopentyl]-5-g Oleic acid, prostaglandin (; 12 (cyclic prostaglandin), (Z)-7-[(lR, 2R, 3R, 5S)-3, 5-dihydroxy-2-[(E, 3R)-3-hydroxyl -4-[3-(Trifluoromethyl)phenoxy]butenyl]cyclopentyl] _5_heptenoic acid, (E) -7-[ (1R, 2R, 3R, 5S) -3, 5 - Dihydroxy-2-(-3-one oxime)cyclopentyl]5-heptenoic acid, misoprostol, gemeprost, 7-[3_hydroxy-2-3 (3-hydroxy-4-benzene) Oxy-1-butenyl)-5-ketocyclopentyl] _5_heptenoic acid, fenproline, prostaglandin Al, prostaglandin A2, prostaglandin Bi, prostaglandin A2, retinoic acid, valsaratin , 9-cis retinoic acid (aliviric acid), retinoic acid analogue, 13-cis retinoic acid (iso-A acid), bismuthrone analog, saludin analog, penicillin G , phenoxymethyl penicillin, methicillin, oxacillin, piperacillin, mezlocillin, carbenicillin, a-tencacillin, ampicillin, methamidomycin, cephalosporin, cefotaxime , cefazolin, cefadroxil, cefradine, cefonicid, cefmenome, cefuroxime, head Sidding, cefretide, cefotetan, cefuroxime, chlorocarbon cephalosporin, cefotaxime, ceftriaxone, cefoperazone, cephalosporin, LIVALO (pitavastatin), Tyvaso (treprostinil), R ) lotyn (; prazil), Tamiflu (oseltamivir), beraprost. In certain embodiments, 0 1 and 126 are each independently a residue of an anti-tumor drug. In certain embodiments, the residues can be antineoplastic agents or A 2 and A 3 form a peptide bond or an ester bond. In certain embodiments, the antitumor agent can be modified by a peptide bond or an ester bond with A 2 or A 3 is formed. In certain embodiments, the anti-tumor drug is dasatinib, rapamycin, irinotecan, imatinib, erlotinib, gefitinib, lapatinib, sorafenib , sunitinib, paclitaxel, camptothecin, scutellaria, glycyrrhetinic acid, sorghum lactone.
[0084] 在某些实施方式中， 本申请提供了具有如下式（VIII )所示的结构的 结合物，  [0084] In certain embodiments, the present application provides a conjugate having the structure shown by formula (VIII) below,
Figure imgf000021_0001
式 （vm)
Figure imgf000021_0001
Formula (vm)
[0085] 其中， R1()、 Ru独立的为氢、 d_6垸基且在每个重复单元中的 R10、 Rii可以相同或不同； h为 1-10的整数。 R7、 P、 X、 a、 b、 c、 d、 D2如上 所述。 Wherein R 1( ) , R u are independently hydrogen, d 6 fluorenyl and R 10 , Rii in each repeating unit may be the same or different; h is an integer from 1 to 10. R 7 , P, X, a, b, c, d, D 2 are as described above.
[0086] 在某些实施方式中， 本申请提供了具有如下式（IX) 、 (X) 、 (XI) 所示的结构的结合物，
Figure imgf000022_0001
式（IX) [0086] In certain embodiments, the present application provides a combination having the structure shown by the following formulas (IX), (X), (XI),
Figure imgf000022_0001
Formula (IX)
cc
Figure imgf000022_0002
Figure imgf000022_0002
Figure imgf000022_0003
式（XI)
Figure imgf000022_0003
Formula (XI)
[0087] 其中， e、 f、 g各自独立的为 10-1,500的整数。 在某些实施方式中， e、 f、 g各自独立的为 100-1,400的整数。 在某些实施方式中， e、 f、 g各自独 立的为 200-1 ,300的整数。 在某些实施方式中， e、 f、 g各自独立的为 300-1 ,200 的整数。 在某些实施方式中， e、 f、 g各自独立的为 400-1,100的整数。 在某些 实施方式中， e、 f、 g各自独立的为 500-1,000的整数。 在某些实施方式中， e、 f、 g各自独立的为 600、 700、 800、 900或 1,000。 如上所述。 在某一实施方 式中， 为达沙替尼。 Wherein e, f, and g are each independently an integer of from 10 to 1,500. In certain embodiments, e, f, g are each independently an integer from 100 to 1,400. In certain embodiments, e, f, g are each independently an integer from 200 to 1,300. In certain embodiments, e, f, g are each independently an integer from 300 to 1,200. In certain embodiments, e, f, g are each independently an integer from 400 to 1,100. In certain embodiments, e, f, g are each independently an integer from 500 to 1,000. In some embodiments, e, f, g are each independently 600, 700, 800, 900 or 1,000. As mentioned above. In one embodiment, it is dasatinib.
[0088] 本申请化合物及其衍生物是按照 IUPAC (国际纯粹与应用化学联合 会） 或 CAS (化学文摘服务社， 位于俄亥俄州哥伦布市） 命名系统命名的。  The compounds of the present application and their derivatives are named according to the IUPAC (International Union of Pure and Applied Chemistry) or CAS (Chemical Abstracts Service, Columbus, Ohio) naming system.
[0089] 碳氢基团中碳原子含量的最小值和最大值通过前缀表示，例如，前缀 (Ca_b)垸基表示任何含 "a"至 "b"个碳原子的垸基。 因此， 例如， （d_6)垸基是 指包含一至六个碳原子的垸基。 所述的垸基是支链的或直链的。 The minimum and maximum values of the carbon atom content in the hydrocarbon group are represented by a prefix, for example, the prefix (C a _ b ) fluorenyl group means any fluorenyl group containing "a" to "b" carbon atoms. Thus, for example, (d_ 6 ) fluorenyl refers to a fluorenyl group containing from one to six carbon atoms. The thiol group is branched or linear.
[0090] "垸氧基"是指与一氧原子键合的直链或带有支链的， 单价的， 饱和 脂肪链， 包括但不限于如甲氧基、 乙氧基、 丙氧基、 丁氧基、 异丁氧基、 叔丁 氧基以及其它类似基团。 "Negoxy" refers to a straight or branched, monovalent, saturated aliphatic chain bonded to an oxygen atom, including but not limited to, for example, methoxy, ethoxy, propoxy, Butoxy, isobutoxy, tert-butoxy and other similar groups.
[0091] "垸基"是指直链或带有支链的， 单价的， 饱和脂肪链， 包括但不限 于如甲基、 乙基、 丙基、 异丙基、 丁基、 异丁基、 戊基、 异戊基、 己基以及其 它类似基团。  "Mercapto" refers to a straight or branched, monovalent, saturated aliphatic chain including, but not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, Pentyl, isopentyl, hexyl and the like.
[0092] "杂垸基"是指上述垸基中的一个或多个碳原子已被如氮、氧或硫等 杂原子取代。 如果杂垸基含有不止一个杂原子， 则这些杂原子可能是相同， 也 可能是不同的。  "Heteropoly" means that one or more carbon atoms in the above fluorenyl group have been replaced by a hetero atom such as nitrogen, oxygen or sulfur. If the heterocyclic group contains more than one heteroatom, these heteroatoms may be the same or different.
[0093] "烯基"是指带有一个或多个双键的直链或支链烃，包括但不限于如 乙烯基、 丙烯基以及其它类似基团。  "Alkenyl" means a straight or branched hydrocarbon bearing one or more double bonds including, but not limited to, vinyl, propenyl, and the like.
[0094] "芳基"是指一种环状的芳香烃，包括但不限于如苯基、萘基、蒽基、 菲基以及其它类似基团。  "Aryl" means a cyclic aromatic hydrocarbon including, but not limited to, phenyl, naphthyl, anthryl, phenanthryl and the like.
[0095] "环垸基"是指饱和的单环或多环垸基， 可能与一芳烃基团稠合。环 垸基包括但不限于如环丙基、 环丁基、 环戊基、 环己基、 环庚基、 茚满基、 四 氢化萘基以及其它类似基团。 [0096] "芳基垸基"是指上述芳基与上述垸基通过共价结合的基团，例如但 不限于苯基甲基、 苯基乙基、 苯基丙基。 "Cycloalkyl" refers to a saturated monocyclic or polycyclic indenyl group, possibly fused to an aromatic hydrocarbon group. Cyclodecyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, indanyl, tetrahydronaphthyl, and the like. "Aryl fluorenyl" means a group to which the above aryl group is covalently bonded to the above thiol group, such as, but not limited to, phenylmethyl, phenylethyl, phenylpropyl.
[0097] "杂芳基"是指单环或多环芳香烃， 其中的一个或多个碳原子已被 如氮、 氧或硫等杂原子取代。 如果杂芳基含有不止一个杂原子， 则这些杂原子 可能是相同， 也可能是不同的。 杂芳基包括但不限于如苯并呋喃基、 苯并噻吩 基、 苯并咪唑基、 苯并恶唑基、 苯并噻唑基、 苯并吡喃基、 呋喃基、 咪唑基、 吲唑基、 吲嗪基、 吲哚基、 异苯并呋喃基、 异吲哚基、 异喹啉基、 异噻唑基、 异恶唑基、 萘啶基、 噁二唑基、 噁嗪基、 噁唑基、 酞嗪基、 蝶啶基、 嘌呤基、 吡喃基、 吡嗪基、 吡唑基、 哒嗪基、 吡啶 [3，4-b]吲哚基、 吡啶基、 嘧啶基、 吡 咯基、 喹嗪基、 喹啉基、 喹喔啉基、 噻二唑基、 噻三唑基、 噻唑基、 噻吩基、 三嗪基、 三唑基、 咕吨基以及其它类似基团。  "Heteroaryl" means a monocyclic or polycyclic aromatic hydrocarbon in which one or more carbon atoms have been replaced by a hetero atom such as nitrogen, oxygen or sulfur. If a heteroaryl contains more than one heteroatom, these heteroatoms may be the same or different. Heteroaryl groups include, but are not limited to, benzofuranyl, benzothienyl, benzimidazolyl, benzoxazolyl, benzothiazolyl, benzopyranyl, furyl, imidazolyl, oxazolyl, Pyridazinyl, fluorenyl, isobenzofuranyl, isodecyl, isoquinolyl, isothiazolyl, isoxazolyl, naphthyridinyl, oxadiazolyl, oxazinyl, oxazolyl, Pyridazinyl, pteridinyl, fluorenyl, pyranyl, pyrazinyl, pyrazolyl, pyridazinyl, pyridine [3,4-b]decyl, pyridyl, pyrimidinyl, pyrrolyl, quinolizine A quinolyl group, a quinolyl group, a thiadiazolyl group, a thiatriazole group, a thiazolyl group, a thienyl group, a triazinyl group, a triazolyl group, a fluorenyl group, and the like.
[0098] 一个环状基团可通过多种方式与另一基团键合。 如果未明确键合方 式， 则表示包括所有可能的方式。 例如， "吡啶基"包括 2-、 3-、 或 4-吡啶基， 而 "噻吩基"包括 2-或 3-噻吩基。  [0098] One cyclic group can be bonded to another group in a variety of ways. If the bonding method is not explicitly defined, it means that all possible ways are included. For example, "pyridyl" includes 2-, 3-, or 4-pyridyl, and "thienyl" includes 2- or 3-thienyl.
[0099] "氨基酸类似物"是指含有至少一个氨基和至少一个羧基的，与氨基 酸结构类似的有机化合物。  "Amino acid analog" means an organic compound having a structure similar to that of an amino acid containing at least one amino group and at least one carboxyl group.
[00100】 水溶性聚合物-氨基酸寡肽-药物的结合物制备方法  Water-soluble polymer-amino acid oligopeptide-drug conjugate preparation method
[00101】 本发明涉及的结合物可以按以下流程合成。该流程只是例证说明而非 限制其它可能制备该结合物的方法。 此外， 流程中的步骤只为更好的说明本发 明结合物的制备方法， 根据实际需要， 所述步骤可以在不脱离本申请所述发明 的范围内进行修改。  [00101] The conjugates of the present invention can be synthesized according to the following scheme. This procedure is illustrative only and not limiting of other ways in which the combination may be made. In addition, the steps in the scheme are only for better description of the preparation method of the combination of the present invention, and the steps can be modified without departing from the scope of the invention described herein.
[00102】 一种制备水溶性聚合物-氨基酸寡肽-药物结合物的方法， 包括：  [00102] A method of preparing a water soluble polymer-amino acid oligopeptide-drug conjugate, comprising:
[00103】 使亲水性聚合物 P 通过连接基团 X与氨基酸 连接得到结合物 [00104] 使药物化合物 01与氨基酸 A2发生反应得到结合物 D A2。 [00103] The hydrophilic polymer P is linked to an amino acid via a linking group X to obtain a conjugate [00104] The drug compound 0 1 is reacted with the amino acid A 2 to give the conjugate DA 2 .
[00105] 使药物化合物 02与氨基酸 A3发生反应得到结合物 D2-A3。 [00105] drug compound 02 is reacted with an amino acid A 3 was obtained in conjunction with D 2 -A 3.
[00106】 使结合物 Di-As和 /或 D2-A3
Figure imgf000025_0001
[00106] Combining Di-As and/or D 2 -A 3
Figure imgf000025_0001
所示结合物。 The combination shown.
[00107】 在某些实施方式中，可以对亲水性聚合物进行改性，引入活性官能团， 与氨基酸寡肽键合。在某些实施方式中，引入活性官能团是通过连接基团 X实 现的。 在某些实施方式中， 引入的活性官能团是羧酸基。  [00107] In certain embodiments, the hydrophilic polymer can be modified to introduce a reactive functional group that is bonded to the amino acid oligopeptide. In certain embodiments, the introduction of a reactive functional group is achieved by a linking group X. In certain embodiments, the reactive functional group introduced is a carboxylic acid group.
[00108】 在某些实施方式中， 可以利用药物化合物上的活性官能团与氨基酸 A2或 A3反应形成结合物 D A2和 /或 D2-A3。 在某些实施方式中， 药物化合物 上的活性官能团可以是羟基、 氨基。 在某些实施方式中， 药物化合物与氨基酸 发生的是酯化或酰胺化反应。 [00108] In certain embodiments, the conjugates can be formed DA 2 and / or D 2 -A 3 with an active functional group on the drug compound with an amino acid A 3 or A 2 reaction. In certain embodiments, the reactive functional group on the drug compound can be a hydroxyl group, an amino group. In certain embodiments, the drug compound and the amino acid are esterified or amidated.
[00109】 在某些实施方式中， 结合物 Di-As 和 I或 D2-A3 与化合物 P_^X^A1 发生酰胺化反应得到式 （I) 所示化合物。 In certain embodiments, the amides Di-As and I or D 2 -A 3 are amidated with the compound P_ ^ X ^ A1 to give a compound of formula (I).
[00110】 水溶性聚合物 氨基酸寡肽 药物结合物药物组合物  [00110] water soluble polymer amino acid oligopeptide drug conjugate pharmaceutical composition
[00111】 另一方面，本发明还提供了包括水溶性聚合物一氨基酸寡肽一药物结 合物及药学上可接受的载体的药物组合物。  In another aspect, the present invention provides a pharmaceutical composition comprising a water-soluble polymer-amino acid oligopeptide-drug complex and a pharmaceutically acceptable carrier.
[00112】 在此提到的术语"药学上可接受的载体"是指一种药学上可接受的物 质， 成分或者介质， 比如液体或固体填充剂， 稀释剂， 赋形剂， 溶剂或灌封材 料， 其参与将本发明涉及的化合物从某一位置， 体液， 组织， 器官 （内部或外 部） ， 或身体部分装载或传递到另一位置， 体液， 器官 （内部或外部） ， 或身 体部分。 药学上可接受的载体可以是介质， 稀释剂， 赋形剂或者其它没有过度 毒性或者副作用并能用于接触动物组织的材料。 典型的药学上可接受的载体包 括糖类， 淀粉， 纤维素类， 麦芽糖， 黄蓍胶， 明胶， 林格氏溶液， 海藻酸， 生 理盐水， 缓冲剂等。 [00113】 每种药学上可接受的载体应该与其它组成成分相容，例如与本发明中 提供的结合物形成制剂， 对生物活体组织或者器官没有过度毒性、 剌激、 过敏 性反应、 免疫原性或其它问题或并发症， 且有较合理的效益风险比。 [00112] The term "pharmaceutically acceptable carrier" as used herein refers to a pharmaceutically acceptable substance, ingredient or medium, such as a liquid or solid filler, diluent, excipient, solvent or potting. A material that is involved in loading or transferring a compound of the invention from a location, body fluid, tissue, organ (internal or external), or body part to another location, body fluid, organ (internal or external), or body part. The pharmaceutically acceptable carrier can be a medium, diluent, excipient or other material which is not excessively toxic or side effects and which can be used to contact animal tissues. Typical pharmaceutically acceptable carriers include sugars, starches, cellulosics, maltose, tragacanth, gelatin, Ringer's solution, alginic acid, physiological saline, buffers and the like. [00113] Each pharmaceutically acceptable carrier should be compatible with the other ingredients, for example, with the conjugates provided herein, without excessive toxicity, irritation, allergic response, immunogen to biological living tissue or organs. Sexual or other problems or complications, and a reasonable ratio of benefits to risks.
[00114】 一些药学上可接受的载体的物质包括： （1 ) 糖类， 比如乳糖， 葡萄 糖和蔗糖； （2)淀粉， 比如玉米淀粉和马铃薯淀粉； （3 )纤维素和其衍生物， 比如羧甲基纤维素钠， 乙基纤维素， 醋酸纤维素； （4) 西黄蓍胶粉； （5 ) 麦 芽糖； （6) 明胶； （7) 滑石粉； （8) 赋形剂， 比如可可脂和栓剂蜡； （9) 油类， 比如花生油，棉籽油， 红花油， 芝麻油， 橄榄油， 玉米油和大豆油； （10) 二醇类， 比如丙二醇； （11 ) 多元醇类， 比如甘油， 山梨醇， 甘露醇和聚乙二 醇； （12)脂类， 比如油酸乙酯， 月桂酸乙酯； （13 )琼脂胶； （14) 缓冲剂， 比如氢氧化镁和氢氧化铝； （15) 海藻酸； （16) 灭菌无热原水； （17) 生理盐 水； （18) 林格氏溶液； （19) 醇类， 比如乙醇和丙醇； （20) 磷酸缓冲液； (21 ) 其它在药物剂型中无毒性可相容的物质， 比如丙酮。 [00114] Some pharmaceutically acceptable carrier materials include: (1) sugars such as lactose, glucose and sucrose; (2) starches such as corn starch and potato starch; (3) cellulose and its derivatives, such as Sodium carboxymethylcellulose, ethylcellulose, cellulose acetate; (4) scutellaria powder; (5) maltose; (6) gelatin; (7) talc; (8) excipients, such as cocoa Fats and suppository waxes; (9) Oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols such as propylene glycol; (11) polyols, such as Glycerin, sorbitol, mannitol and polyethylene glycol; (12) lipids such as ethyl oleate, ethyl laurate; (13) agarose; (14) buffers such as magnesium hydroxide and aluminum hydroxide; (15) Alginic acid; (16) Sterilized pyrogen-free water; (17) Saline; (18) Ringer's solution; (19) Alcohols such as ethanol and propanol; (20) Phosphate buffer; Other non-toxic compatible substances in pharmaceutical dosage forms, Such as acetone.
[00115】 药物组合物可能包括药学上可接受的辅料， 以模拟生理条件， 比如 pH调节和缓冲剂， 毒性调节剂等等， 如乙酸钠， 氯化钠， 氯化钾， 氯化钙， 乳 酸钠等。  [00115] The pharmaceutical composition may include pharmaceutically acceptable excipients to mimic physiological conditions such as pH adjustment and buffering agents, toxicity modulators, etc., such as sodium acetate, sodium chloride, potassium chloride, calcium chloride, sodium lactate Wait.
[00116】 药物成分可制成任何合适的剂型， 如固体剂型（例如片剂， 胶囊， 粉 末， 颗粒等） 和液体剂型 （例如水溶液， 乳浊液， 酏剂， 糖浆等） 。 药物组合 物的制备方法工艺已众所周知，可根据常规工艺进行制备， 比如在 Remington, The Science and Practice of Pharmacy (Gennaro ed. 20th edition, Williams & Wilkins PA, USA) (2000)中提供。  [00116] The pharmaceutical ingredient can be formulated into any suitable dosage form, such as a solid dosage form (e.g., tablets, capsules, powders, granules, etc.) and a liquid dosage form (e.g., aqueous solution, emulsion, elixirs, syrup, etc.). Processes for the preparation of pharmaceutical compositions are well known and can be prepared according to conventional procedures, such as those found in Remington, The Science and Practice of Pharmacy (Gennaro ed. 20th edition, Williams & Wilkins PA, USA) (2000).
[00117] 在某些实施方式中，本发明中提供的化合物或药物组合物可以被制成 适于药物释放的剂型， 通过注射途径给药（如皮下， 静脉， 肌肉， 动脉， 鞘膜， 囊内， 框内， 心脏内， 真皮内， 腹膜内， 经气管， 表皮， 关节内， 囊下， 蛛网 膜下， 脊柱内， 胸骨内， 和 /或输液）和非注射途径给药（如口服， 肠道， 口腔， 鼻， 鼻内， 粘膜， 表皮， 贴膏剂， 真皮， 眼药， 肺部， 舌下， 直肠， 阴道或表 皮局部给药）。 [00117] In certain embodiments, the compounds or pharmaceutical compositions provided herein can be formulated into a dosage form suitable for drug delivery, such as subcutaneous, intravenous, intramuscular, arterial, sheath, vesicles. Inside, in-frame, intracardiac, intradermal, intraperitoneal, transtracheal, epidermis, intra-articular, subcapsular, cobweb Sub-membrane, intraspinal, intrasternal, and/or infusion) and non-injectable routes (eg, oral, intestinal, oral, nasal, intranasal, mucosal, epidermal, plaster, dermis, eye drops, lungs, tongue Subcutaneous, rectal, vaginal or epidermal administration).
[00118】 合适的剂型包括（但不局限于）注射用途的剂型比如乳状液， 溶液和 混悬液， 口服用途的剂型如片剂， 胶囊， 丸剂， 糖衣丸， 粉末和颗粒， 局部用 药或经皮肤吸收的剂型如喷剂， 软膏， 糊剂， 乳霜， 洗剂， 凝胶， 溶液， 药物 贴片和吸入剂， 阴道或直肠给药的剂型如栓剂。 这些剂型可根据化合物以及合 适的赋形剂在合适条件下制备， 制备方法及工艺众所周知， 比如由 Remington: 在 The Science and Practice of Pharmacy (Gennaro ed. 20th edition, Williams & Wilkins PA, USA) (2000) 提供。  [00118] Suitable dosage forms include, but are not limited to, dosage forms for injection use such as emulsions, solutions and suspensions, and dosage forms for oral use such as tablets, capsules, pills, dragees, powders and granules, topical or topical Formulations for skin absorption such as sprays, ointments, pastes, creams, lotions, gels, solutions, drug patches and inhalants, vaginal or rectal administration such as suppositories. These dosage forms can be prepared according to the compound and suitable excipients under suitable conditions, and are well known, for example, by Remington: in The Science and Practice of Pharmacy (Gennaro ed. 20th edition, Williams & Wilkins PA, USA) (2000) ) provided.
[00119】 水溶性聚合物一氨基酸寡肽一药物结合物和 I或药物组合物的应用  [00119] Application of water-soluble polymer-amino acid oligopeptide-drug conjugate and I or pharmaceutical composition
[00120】 本发明的一方面是通过亲水性聚合物的修饰增加药物分子的水溶性， 延长药物的半衰期。 在某些实施方式中， 所述结合物的药物活性与未修饰的药 物分子相比， 药物的载药量、 药物的活性、 稳定性以及水溶性都提高了。  [00120] One aspect of the invention is to increase the water solubility of a drug molecule by modification of a hydrophilic polymer, thereby prolonging the half-life of the drug. In certain embodiments, the pharmaceutically active amount of the conjugate is increased as compared to the unmodified drug molecule, the drug loading, drug activity, stability, and water solubility.
[00121】 本发明的另一个方面是通过氨基酸寡肽增加药物结合物的稳定性，进 而降低毒副作用。 在某些实施方式中， 所述药物结合物稳定性的增加通过选择 合适的氨基酸寡肽实现。 在某些实施方式中， 所述药物结合物稳定性的增加通 过选择合适的与药物化合物结合的氨基酸残基， 即， 式 （I) 中的 氨基酸 A2 或 A3实现。 优选的， 所述与药物化合物结合的氨基酸残基为缬氨酸残基。 在 某些实施方式中， 所述结合物的毒性有所降低。 在某些实施方式中， 所述结合 物进一步提高了药物的临床效果。 [00121] Another aspect of the invention is to increase the stability of a drug conjugate by an amino acid oligopeptide, thereby reducing toxic side effects. In certain embodiments, the increase in stability of the drug conjugate is achieved by selection of a suitable amino acid oligopeptide. ] In certain embodiments, the pharmaceutical composition to increase the stability of the binding by selecting a suitable amino acid residue in combination with a pharmaceutical compound, i.e., the amino acid of formula A (I) 2 A 3, or implemented. Preferably, the amino acid residue bound to the drug compound is a proline residue. In certain embodiments, the toxicity of the combination is reduced. In certain embodiments, the conjugate further enhances the clinical effectiveness of the drug.
[00122】 本发明的另一个方面是每一个亲水性聚合物通过氨基酸寡肽可以与 多个药物化合物相连， 从而大大体高药物的负载率。 在某些实施方式中， 每一 个亲水性聚合物可以与至少两个药物化合物相连。 在某些实施方式中，每一个 亲水性聚合物可以与至少三个药物化合物相连。 在某些实施方式中，每一个亲 水性聚合物可以与至少四个药物化合物相连。 [00122] Another aspect of the present invention is that each hydrophilic polymer can be linked to a plurality of drug compounds by an amino acid oligopeptide, thereby greatly increasing the drug loading rate. In certain embodiments, each hydrophilic polymer can be attached to at least two pharmaceutical compounds. In some embodiments, each The hydrophilic polymer can be attached to at least three drug compounds. In certain embodiments, each hydrophilic polymer can be attached to at least four pharmaceutical compounds.
[00123】 本发明的另一个方面提供了上述结合物和 /或药物组合物在制备药物 以及治疗疾病中的应用。  [00123] Another aspect of the invention provides the use of the above conjugates and/or pharmaceutical compositions for the preparation of a medicament and for the treatment of a disease.
[00124】 在某些实施方式中， 本发明提供了上述结合物和 /或药物组合物在制 备抗肿瘤、 真菌感染、 类风湿性关节炎、 多发性硬化症、 心瓣再狭窄或肺炎的 药物中的应用。  [00124] In certain embodiments, the present invention provides a medicament for the preparation of an antitumor, fungal infection, rheumatoid arthritis, multiple sclerosis, heart stenosis or pneumonia in the above conjugate and/or pharmaceutical composition. Application in .
[00125】 在某些实施方式中， 所述抗肿瘤药物应用于以下病症： 白血病、急性 粒细胞性白血病、 慢性粒细胞性白血病、 慢性淋巴性白血病、 急性淋巴性白血 病、 脊髓发育不良、 多发性骨髓瘤、 何杰金氏病或非何杰金氏病、 小细胞或非 小细胞性肺癌、 胃癌、 肠癌、 食道癌、 结肠直肠癌、 前列腺癌、 卵巢癌、 乳腺 癌、 脑癌、 泌尿道癌、 肾癌、 膀胱癌、 恶性黑素瘤、 肝癌、 子宫癌、 胰腺癌、 骨髓瘤癌、 子宫内膜癌、 头颈癌、 小儿肿瘤、 肉瘤。  [00125] In certain embodiments, the anti-tumor drug is applied to the following conditions: leukemia, acute myeloid leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia, acute lymphocytic leukemia, myelodysplasia, multiple Myeloma, Hodgkin's disease or non-Hodgkin's disease, small cell or non-small cell lung cancer, gastric cancer, colon cancer, esophageal cancer, colorectal cancer, prostate cancer, ovarian cancer, breast cancer, brain cancer, urinary Cancer, kidney cancer, bladder cancer, malignant melanoma, liver cancer, uterine cancer, pancreatic cancer, myeloma, endometrial cancer, head and neck cancer, pediatric tumor, sarcoma.
[00126】 在某些实施方式中， 本申请提供了一种治疗主体中肿瘤、 真菌感染、 类风湿性关节炎、 多发性硬化症、 心瓣再狭窄或肺炎的方法， 包括向所述主体 给与有效量的上述结合物或药物组合物。 在某些实施方式中， 所述主体是哺乳 动物。 在某些实施方式中， 所述主体是人、 牲畜或宠物。  [00126] In certain embodiments, the present application provides a method of treating a tumor, a fungal infection, rheumatoid arthritis, multiple sclerosis, heart valve restenosis, or pneumonia in a subject, comprising administering to the subject An effective amount of the above combination or pharmaceutical composition. In certain embodiments, the subject is a mammal. In certain embodiments, the subject is a human, animal or pet.
[00127】 本发明涉及的结合物或药物组合物可通过任何合适的途径进入生物 体内， 比如通过口服， 静脉注射， 鼻内， 外用， 肌注， 真皮内注射， 经皮给药 或皮下途径。 在某些实施方式中， 本发明涉及的结合物或药物组合物的给药方 式包括口腔、 粘膜、 舌下、 眼部、 局部、 肠道外、 直肠、 脑池、 阴道、 腹膜、 膀胱、 鼻部给药。 [00127] The conjugate or pharmaceutical composition of the present invention may be administered to a living organism by any suitable route, such as by oral, intravenous, intranasal, topical, intramuscular, intradermal, transdermal or subcutaneous routes. In certain embodiments, the present invention relates to a method of administration of a combination or pharmaceutical composition comprising oral, mucosal, sublingual, ocular, topical, parenteral, rectal, cerebral, vaginal, peritoneal, bladder, nasal Dosing.
[00128】 在某些实施方式中，本发明涉及的结合物或药物组合物可以与第二活 性物质同时进行施用， 这样能在生物体内达到叠加甚至协同的作用。 例如， 本 发明涉及的化合物可以和第二活性物质组合成一个药物组合物， 或者以单独的 组合物同时施用，或者以单独的组合物依次施用。能与本发明化合物同时施用、 用于治疗癌症的第二活性物质包括但不局限于： 氟尿嘧啶， 阿霉素， 柔红霉素， 它莫西芬， 亮丙瑞林， 戈舍瑞林， 氟他米特， 尼鲁米特， 非那雄胺， 地塞米松， 氨鲁米特， 安吖啶、 阿那曲唑， 天冬酰胺酶， 卡介苗， 比卡鲁胺、 博来霉素、 临床、 白消安， 喜树碱， 卡培他滨， 卡铂， 卡莫司汀， 苯丁酸氮芥， 顺铂， 克 拉屈滨， 秋水仙碱， 环磷酰胺、 药物、 环丙孕酮、 阿糖胞苷、 达卡巴嗪， 放线 菌素 d， 正定霉素， 双烯雌酚、 己烯雌酚、 多西紫杉醇、 阿霉素， 亚德里亚霉 素、 表柔比星、 雌二醇， 雌氮芥、 依托泊苷、 依西美坦， 非格司亭， 氟达拉滨， 氟氢可的松、 氟尿嘧啶、 氟甲睾酮、 氟他米特， 吉西他滨， 染料木黄酮， 戈舍 瑞林， 三苯氧胺、 替尼泊苷、 睾酮、 二氯化二茂钛， 拓普泰康， 曲妥单抗、 维 甲酸、 长春花碱、 羟基脲， 伊达比星， 异环磷酰胺、 伊马替尼、 干扰素、 伊立 替康， 伊立替康、 来曲唑、 甲酰四氢叶酸， 喷司他丁， 光神霉素、 甲基苄肼、 雷替曲塞卟菲尔， 利妥昔链脲菌素， 苏拉明， 亮丙瑞林、 左旋咪唑， 环己亚硝 脲、 氮芥、 甲羟孕酮、 甲地孕酮， 美法仑、 巯嘌呤、 巯乙磺酸钠、 甲氨蝶呤、 丝裂霉素、 米托坦、 米托蒽醌， 尼鲁米特， 诺考达唑、 奥曲肽、 铂、 紫杉醇、 帕米磷酸， 硫鸟嘌呤， 三胺硫磷、 氯甲垸、 拓扑替康二茂钛， 曲妥单抗， 维甲 酸、 长春花碱， 长春新碱、 长春地辛， 长春瑞斌。 [00128] In certain embodiments, the conjugates or pharmaceutical compositions of the present invention can be administered concurrently with a second active substance, such that a synergistic or even synergistic effect can be achieved in the organism. For example, this The compounds of the invention may be combined with the second active substance into a pharmaceutical composition, or may be administered simultaneously in separate compositions, or sequentially in separate compositions. Second active substances which can be administered simultaneously with the compounds of the invention for the treatment of cancer include, but are not limited to: fluorouracil, doxorubicin, daunorubicin, tamoxifen, leuprolide, goserelin, fluoro Hemet, Nilemet, Finasteride, Dexamethasone, Amlodipine, Ampicillin, Anastrozole, Asparaginase, BCG, Bicalutamide, Bleomycin, Clinical, Busulfan, camptothecin, capecitabine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, colchicine, cyclophosphamide, drug, cyproterone, a Cytosine, dacarbazine, actinomycin d, gentamicin, diethestrol, diethylstilbestrol, docetaxel, doxorubicin, doxorubicin, epirubicin, estradiol, estra Mustard, etoposide, exemestane, filgrastim, fludarabine, fludrocortisone, fluorouracil, fluorotestosterone, flutamide, gemcitabine, genistein, goserelin, tamoxifen , teniposide, testosterone, titanium dichloride, Topote , trastuzumab, retinoic acid, vinblastine, hydroxyurea, idarubicin, ifosfamide, imatinib, interferon, irinotecan, irinotecan, letrozole, formyltetrahydrogen Folic acid, pentastatin, mithramycin, procarbazine, raltezine, rituximab, rituximab, suramin, leuprolide, levamisole, cyclohexyl nitrosourea , nitrogen mustard, medroxyprogesterone, megestrol acetate, melphalan, guanidine, sodium sulfonate, methotrexate, mitomycin, mitoxantrone, mitoxantrone, nilutamide , nocodazole, octreotide, platinum, paclitaxel, pamidronate, thioguanine, triamine, methotrexate, topotecan, titanium, trastuzumab, retinoic acid, vinblastine, vincristine Changchun Dixin, Changchun Ruibin.
[00129】 在某些实施方式中，本发明提供的结合物可与非化学方法同时使用进 行癌症治疗。在某些实施方式中，本发明提供的结合物可与放射疗法同时进行。 在某些实施方式中， 本发明提供的结合物可与外科手术， 肿瘤热治疗， 超声聚 焦疗法， 冷冻疗法或以上几种疗法结合使用。  [00129] In certain embodiments, the conjugates provided herein can be used in conjunction with non-chemical methods for cancer treatment. In certain embodiments, the conjugates provided herein can be performed concurrently with radiation therapy. In certain embodiments, the conjugates provided herein can be used in combination with surgery, tumor heat therapy, ultrasound focus therapy, cryotherapy, or several of the above.
[00130】 在某些实施方式中，本发明提供的结合物可与类固醇同时使用。合适 的类固醇包括但不局限于： 安西缩松、 倍氯米松， 倍他米松， 布地奈德， 氯泼 尼松， 氯倍他索， 皮质 酮， 可的松， 羟泼尼缩松， 去羟米松， 地塞米松， 双 氟拉松， 双氟米松， 二氟孕 丁酯， 甘草次酸， 氟扎可松， 氟米松， 氟尼缩松， 氟氯奈德， 肤轻松醋酸酯， 氟轻松醋酸酯， 氟可丁丁酯， 氟可龙， 丙酮缩氟氢 羟龙， 醋酸氟培龙， 醋酸氟泼尼定， 氟泼尼龙， 氟氢缩松， 丙酸氟、 醛基缩松， 丙酸氯倍他索， 哈西缩松， 卤米松， 氢化可的松， 氯替泼诺碳酸乙酯， 甲哌地 强龙， 甲羟松， 甲泼尼松， 6-甲氢化泼尼松， 在任糠酸盐， 帕拉米松， 泼尼松 龙， 地塞米松， 和 25-二乙胺醋强的松龙。 [00130] In certain embodiments, the conjugates provided herein can be used concurrently with steroids. Suitable steroids include, but are not limited to: Anxis, beclomethasone, betamethasone, budesonide, chlorinated Nisson, clobetasol, corticosterone, cortisone, hydroxyprednisolone, deshydroxymetasone, dexamethasone, diflurazon, difluoromethasone, difluprednate, glycyrrhetinic acid, fluza Fluke, flumetasone, flunisolide, flucline acetonide, acetonide acetate, fluocinolone acetonide, flubutylbutabutyl ester, fluconazole, acetonide, fluphene acetate, fluphene acetate Nitinidine, fluprednisolone, fluorohydrogenated, propionic acid fluoride, aldehyde-based shrinkage, clobetasol propionate, Hasixine, halomethasone, hydrocortisone, loteprednol ethyl carbonate, A Piperidone, hydroxybutazone, methylprednisolone, 6-methylprednisolone, in decanoate, palmitone, prednisolone, dexamethasone, and 25-diethylamine acesulfame Dragon.
[00131】 在某些实施方式中， 本发明提供的化合物可与免疫治疗剂同时使用。 合适的免疫治疗剂包括： 肿瘤细胞多药耐药性逆转剂（比如维拉帕米）， 雷帕霉 素， 霉酚酸酯， 沙利度胺， 环磷酰胺， 环孢霉素， 和单克隆抗体类。  [00131] In certain embodiments, the compounds provided herein can be used concurrently with an immunotherapeutic agent. Suitable immunotherapeutic agents include: tumor cell multidrug resistance reversal agents (such as verapamil), rapamycin, mycophenolate mofetil, thalidomide, cyclophosphamide, cyclosporine, and Cloning of antibodies.
[00132】 本申请所述结合物的有益效果： 1 )本申请所述水溶性聚合物-氨基酸 寡肽 -药物结合物与药物相比具有更好的治疗效果。在相同剂量下， 本申请所述 结合物展现了更好的抗肿瘤、 抗癌活性； 2) 本申请所述水溶性聚合物-氨基酸 可以结合更多的药物， 提高了结合物的载药量， 同时使药物能够缓释释放， 避 免了为持续达到有效血药浓度而向病人多次给药； 3 )所述结合物降低了药物的 毒性； 4)通过使用了氨基酸 A和八2， 所述药物结合物的稳定性提高了， 便于 药物的制备、 保存及给药。 [00132] The beneficial effects of the conjugates described herein: 1) The water soluble polymer-amino acid oligopeptide-drug conjugates described herein have a better therapeutic effect than drugs. At the same dosage, the conjugates described herein exhibit better anti-tumor and anti-cancer activities; 2) The water-soluble polymer-amino acid described in the present application can bind more drugs and increase the drug loading of the conjugate. , while the sustained release of drug can be avoided to achieve the sustained effective blood concentration administered to a patient repeatedly; 3) the conjugate reduces the toxicity of the drug; 4) by using the a and eight amino acid 2, the The stability of the drug conjugate is improved, facilitating the preparation, storage and administration of the drug.
[00133】 实施例 -有关达沙替尼的结合物 [00133] Example - Combination of Dasatinib
[00134】 本实施例中所用的达沙替尼从南京安格医药化工有限公司购得， L-(+)-谷氨酸从北京化学试剂公司购得， 对甲苯磺酸、 苯甲醇及二环己基碳二 亚胺 （DCC) 从国药集团化学试剂有限公司购得， 4-二甲基氨基吡啶 （DMAP) 及 1-羟基苯并三氮唑 （HOBt) 从上海共价化学科技有限公司购得， N-叔丁氧 羰基甘氨酸， N-叔丁氧羰基丙氨酸， N-叔丁氧羰基缬氨酸及 N-叔丁氧羰基 -L-谷 氨酸 -5-苄酯从四川同晟氨基酸有限公司购得， 单氧基聚乙二醇乙酸 （20K) ， Y—型聚乙二醇乙酸 （40Κ) 由北京键凯科技有限公司提供， 其余为市售试剂( [00134] Dasatinib used in the present embodiment was purchased from Nanjing Ange Pharmaceutical Chemical Co., Ltd., and L-(+)-glutamic acid was purchased from Beijing Chemical Reagent Co., Ltd., p-toluenesulfonic acid, benzyl alcohol and two Cyclohexylcarbodiimide (DCC) was purchased from Sinopharm Chemical Reagent Co., Ltd., 4-dimethylaminopyridine (DMAP) and 1-hydroxybenzotriazole (HOBt) were purchased from Shanghai Covalent Chemical Technology Co., Ltd. N,tert-butoxycarbonylglycine, N-tert-butoxycarbonylalanine, N-tert-butoxycarbonylproline and N-tert-butoxycarbonyl-L-glutamate-5-benzyl ester from Sichuan Acrylic Amino Acid Co., Ltd., monooxypolyethylene glycol acetate (20K), Y-type polyethylene glycol acetic acid (40Κ) is supplied by Beijing Keykai Technology Co., Ltd., and the rest are commercially available reagents (
[00135] 买施例 1 : 单甲氧基聚乙二醇谷氨酸二肽酸（数均分子量 20000) [00135] Example 1 : monomethoxy polyethylene glycol glutamic acid dipeptide acid (number average molecular weight 20000)
-1 ) 的制备  -1 ) preparation
Figure imgf000031_0001
Figure imgf000031_0001
Figure imgf000031_0002
Figure imgf000031_0002
[00136] L-(+)-谷氨酸 29.4g ( 0.2mol)、 对甲苯磺酸 40g ( 0.23mol)、 苯甲醇 80mL溶于甲苯 500mL中， 氮气保护下回流分出 l lmL水， 继续回流 3h， 蒸除 150mL。 冷却到 50°C， 将反应液倒入盛有 600 mL石油醚的烧杯中， 搅拌 lh， 过滤收集沉淀。 滤饼用 95%乙醇 280 mL加热溶解后， 停止加热， 冷却过夜。 过滤收集沉淀真空干燥， 制得 L-(+)-谷氨酸二苄酯对甲苯磺酸盐 （化合物 1) 61go [00137】 L-(+)-谷氨酸二苄酯对甲苯磺酸盐 （化合物 l) 30g (0.06mol) 溶于 500mL二氯甲垸中， 加入叔丁氧羰基 -L-谷氨酸 -5-苄酯 20.86g (0.062mol)， DMAP 7.55g (0.062mol)、 HOBt 8.35g (0.07mol)， 氮气保护下加入 DCC 14.3g 的二氯甲垸溶液。 滴毕， 密闭体系反应过夜。 TLC监测反应完毕。 过滤除去溶 剂后， 浓缩液中加入 20 mL乙酸乙酯， 滤除固体， 母液加入石油醚 400 mL沉 淀， 过滤得产品 N-叔丁氧羰基谷氨酸苄酯二肽 （化合物 2) 15.8g。 [00136] L-(+)-glutamic acid 29.4g (0.2mol), p-toluenesulfonic acid 40g (0.23mol), benzyl alcohol 80mL dissolved in toluene 500mL, refluxed under nitrogen to separate l lmL water, continue to reflux 3h, distilled off 150mL. After cooling to 50 ° C, the reaction solution was poured into a beaker containing 600 mL of petroleum ether, stirred for 1 h, and the precipitate was collected by filtration. After the filter cake was dissolved by heating in 280 mL of 95% ethanol, the heating was stopped and allowed to cool overnight. The precipitate was collected by filtration and dried under vacuum to obtain L-(+)-glutamic acid dibenzyl ester p-toluenesulfonate (Compound 1) 61go [00137] L-(+)-glutamic acid dibenzyl ester p-toluenesulfonate (compound 1) 30 g (0.06 mol) was dissolved in 500 mL of dichloromethane, and tert-butoxycarbonyl-L-glutamic acid was added. 5-benzyl ester 20.86 g (0.062 mol), DMAP 7.55 g (0.062 mol), HOBt 8.35 g (0.07 mol), a solution of 14.3 g of dichloromethane was added under nitrogen. After the dropwise addition, the closed system was reacted overnight. The reaction was monitored by TLC. After removing the solvent by filtration, 20 mL of ethyl acetate was added to the concentrate, and the solid was filtered, and the mother liquid was poured into a mixture of petroleum ether (400 mL), and filtered to obtain 15.8 g of the product N-tert-butoxycarbonylglutamic acid benzyl ester dipeptide (Compound 2).
[00138】 N-叔丁氧羰基谷氨酸苄酯二肽 （化合物 2) 0.78g 溶解在 7 mL二氯 甲垸中， 加入三氟乙酸 3mL， 室温反应 2h。 除去溶剂， 加入 lOOmL二氯甲垸， 用 5%碳酸氢钠溶液调节 pH=7-8。 萃取分液， 有机相用 5%碳酸氢钠溶液洗两 次， 无水硫酸钠干燥。 过滤， 滤液直接加到反应瓶中， 氮气保护下， 加入单甲 氧基聚乙二醇乙酸 (20K) 20.0g(l mmol)、 DMAP 245 mg(2 mmol)、 HOBt 135 mg(l mmol)。 全部溶解后， 加入 DCC 412mg(2 mmol)。 室温搅拌反应过夜。 过 滤， 旋转蒸发除去溶剂， 残余物中添加异丙醇 500mL， 过滤， 产物真空干燥。 将该产物溶于 200mL无水甲醇中， 加入钯炭 1.0g， 通入氢气室温反应过夜。过 滤除去钯炭， 旋转蒸发除去溶剂， 残余物中添加异丙醇 500mL， 过滤， 产物真 空干燥。 得到单甲氧基聚乙二醇谷氨酸二肽酸 （20K) (化合物 5) 13.4g。  [00138] 0.78 g of N-tert-butoxycarbonylglutamic acid benzyl ester dipeptide (Compound 2) was dissolved in 7 mL of dichloromethane, and 3 mL of trifluoroacetic acid was added thereto, and the mixture was reacted at room temperature for 2 hours. The solvent was removed, 100 mL of dichloromethane was added, and the pH was adjusted to 7-8 with a 5% sodium hydrogen carbonate solution. The extract was separated and the organic phase was washed twice with 5% sodium hydrogen carbonate solution and dried over anhydrous sodium sulfate. After filtration, the filtrate was directly added to a reaction flask, and under a nitrogen atmosphere, 20.0 g (1 mmol), DMAP 245 mg (2 mmol), and HOBt 135 mg (1 mmol) of monomethoxypolyethylene glycol acetic acid (20K) were added. After all was dissolved, DCC 412 mg (2 mmol) was added. The reaction was stirred at room temperature overnight. After filtration, the solvent was removed by rotary evaporation. EtOAc (EtOAc) This product was dissolved in 200 mL of anhydrous methanol, and 1.0 g of palladium on carbon was added thereto, and the mixture was reacted with hydrogen at room temperature overnight. The palladium on charcoal was removed by filtration, and the solvent was removed by rotary evaporation. <EMI ID=9.1> Monomethoxypolyethylene glycol glutamic acid dipeptide acid (20K) (Compound 5) 13.4 g was obtained.
[00139】 将单甲氧基聚乙二醇谷氨酸二肽酸 （化合物 5) (20K，0.5g， [00139] Monomethoxy polyethylene glycol glutamic acid dipeptide acid (Compound 5) (20K, 0.5g,
0.025mmol) ，达沙替尼 73 mg (0.15mmol)， 和 DMAP 24.4 mg(0.2mmol) 加到反 应瓶中，用二氯甲垸和 N，N-二甲基甲酰胺混合溶剂溶解， 氮气保护下冰浴冷却， 再滴入 DCC 41.3 mg (0.2mmol) 溶于二氯甲垸的溶液， 滴完后自然升到室温反 应过夜。 次日将反应液浓缩， 残分用异丙醇重结晶， 得单甲氧基聚乙二醇谷氨 酸二肽酸（20K)—达沙替尼结合物 （DSR-1) 0.37go 1H-NMR (DMSO-d6): 2.12(s, 9H)， 2.13 (m， 2H), 2.17 (m， 2H), 2.40 (t， 4H)， 2.44 (s， 9H)， 2.51 (t， 6H)， 3.30 (s， 3H), 3.44 (t， 12H), 3.50 (s， 2H), 3.54(m, 1800H), 3.60 (t， 6H)， 3.62 (t， 12H), 4.45 (t， 1H), 4.53 (t， 1H), 5.28 (s， 3H), 7.01 (m， 3H), 7.26 (m， 3H), 7.44 (m， 3H), 8.03 (s， 2H), 8.10 (s， 3H), 9.15 (s, 3H), 11.44 (s， 3H). 0.025 mmol), dasatinib 73 mg (0.15 mmol), and DMAP 24.4 mg (0.2 mmol) were added to the reaction flask, dissolved in a mixed solvent of dichloromethane and N,N-dimethylformamide, and protected with nitrogen. After cooling in an ice bath, a solution of 41.3 mg (0.2 mmol) of DCC dissolved in dichloromethane was added dropwise, and the mixture was allowed to react to room temperature overnight. The reaction solution was concentrated the next day, and the residue was recrystallized from isopropanol to give monomethoxypolyethylene glycol glutamic acid dipeptidic acid (20K)-dasatinib conjugate (DSR-1) 0.37go 1H- NMR (DMSO-d 6 ): 2.12 (s, 9H), 2.13 (m, 2H), 2.17 (m, 2H), 2.40 (t, 4H), 2.44 (s, 9H), 2.51 (t, 6H), 3.30 (s, 3H), 3.44 (t, 12H), 3.50 (s, 2H), 3.54 (m, 1800H), 3.60 (t, 6H), 3.62 (t, 12H), 4.45 (t, 1H), 4.53 (t, 1H), 5.28 (s, 3H), 7.01 (m, 3H), 7.26 (m, 3H), 7.44 (m, 3H), 8.03 (s, 2H), 8.10 (s, 3H) ), 9.15 (s, 3H), 11.44 (s, 3H).
[00140】 实施例 2: 单甲氧基聚乙二醇谷氨酸二肽甘氨酸 (数均分子量 20000) 一达沙替尼结合物 （DSR-2) 的制备  Example 2: Monomethoxypolyethylene glycol glutamic acid dipeptide glycine (number average molecular weight 20000) Preparation of a dasatinib conjugate (DSR-2)
Figure imgf000033_0001
Figure imgf000033_0001
[00141】 根据实施例 1所述的方法制备单甲氧基聚乙二醇谷氨酸二肽酸 （化 合物 5)。 [00141] Monomethoxypolyethylene glycol glutamic acid dipeptide acid (Compound 5) was prepared according to the method described in Example 1.
[00142】 将达沙替尼 1.95g (4 mmol), N-叔丁氧羰基甘氨酸 840 mg (4.8mmol) 禾口 DMAP 714 mg (4.8 mmol) 加到反应瓶中， 用 40 mLN，N-二甲基甲酰胺溶 解，氮气保护下冰浴冷却，再滴入 DCC 1.03 g C5mmol) 溶于 20 mL二氯甲垸的 溶液， 滴完后自然升到室温反应过夜。 TLC监测反应完全后减压蒸去溶剂， 残 分用柱层析分离， 得 N-叔丁氧羰基甘氨酸达沙替尼酯 （化合物 6) 1.5 g， 收率 58%。 1H-NMR (DMSO-d6): 1.32(s, 9H)， 2.15(s, 3H), 2.46(s, 3H), 2.5 l(m, 2H), 3.53(m, 10H),4.10(s, 2H), 5.88(s， 1H)， 7.29(m, 3H), 8.12(s, 1H)， 8.28(s, 1H)， 9.81(s, 1H), 11.25(s, 1H)。 [00142] Dasatinib 1.95g (4 mmol), N-tert-butoxycarbonylglycine 840 mg (4.8 mmol) and DMAP 714 mg (4.8 mmol) were added to the reaction flask, using 40 mL of N, N- The methylformamide was dissolved, cooled in an ice bath under nitrogen atmosphere, and then dropped into a solution of DCC 1.03 g C5 mmol) dissolved in 20 mL of dichloromethane. After the completion of the dropwise addition, the mixture was allowed to react to room temperature overnight. After the reaction was completely monitored by TLC, the solvent was evaporated under reduced pressure, and the residue was separated by column chromatography to yield 1.5 g of N-tert-butoxycarbonylglycine ss. 1H-NMR (DMSO-d 6 ): 1.32 (s, 9H), 2.15 (s, 3H), 2.46 (s, 3H), 2.5 l (m, 2H), 3.53(m, 10H), 4.10(s, 2H), 5.88(s, 1H), 7.29(m, 3H), 8.12(s, 1H), 8.28(s, 1H), 9.81(s, 1H), 11.25 (s, 1H).
[00143】 将 N-叔丁氧羰基甘氨酸达沙替尼酯 （化合物 6) 1 g (1.6mmol), 二氯 甲垸 50 mL， 加到反应瓶中， 再加入三氟乙酸 25 mL， 搅拌反应 3h后减压蒸去 溶剂， 残分中加入二氯甲垸， 减压蒸干， 如此重复三次， 得甘氨酸达沙替尼酯 三氟乙酸盐 （化合物 7) 1.2 g， 直接往下反应。  [00143] N-tert-butoxycarbonyl glycine dasatinib (Compound 6) 1 g (1.6 mmol), dichloromethane (50 mL) was added to the reaction flask, then 25 mL of trifluoroacetic acid was added, and the reaction was stirred. After 3 hours, the solvent was evaporated under reduced pressure. dichloromethane was evaporated and evaporated to dryness, and then evaporated to dryness.
[00144】 将单甲氧基聚乙二醇谷氨酸二肽酸 （化合物 5) (20K，0.5g，  [00144] Monomethoxy polyethylene glycol glutamic acid dipeptide acid (Compound 5) (20K, 0.5g,
0.025mmol) ，甘氨酸达沙替尼酯三氟乙酸盐（化合物 7) 100 mg (0.15mmol)， HOBt (3.4 mg, 0.025mmo) 和 DMAP 24.4 mg(0.2mmol) 加到反应瓶中， 用二氯 甲垸和 N，N-二甲基甲酰胺混合溶剂溶解， 氮气保护下冰浴冷却， 再滴入 DCC 36.1 mg (0.175mmol) 溶于二氯甲垸的溶液， 滴完后自然升到室温反应过夜。次 日将反应液浓缩， 残分用异丙醇重结晶， 得单甲氧基聚乙二醇谷氨酸二肽甘氨 酸（20K)—达沙替尼结合物（DSR-2) 0.39 g。 1H-NMR (DMSO-d6): 2.12(s, 9H)， 2.13 (m， 2H), 2.17 (m， 2H), 2.40 (t， 4H)， 2.44 (s， 9H)， 2.97 (t， 6H)， 3.30 (s， 3H),0.025mmol), dasatinib glycinate trifluoroacetate (compound 7) 100 mg (0.15mmol), HOBt (3.4 mg, 0.025mmo) and DMAP 24.4 mg (0.2mmol) were added to the reaction flask, using two The mixed solvent of chloroformam and N,N-dimethylformamide was dissolved, cooled in an ice bath under nitrogen atmosphere, and then dropped into DCC 36.1 mg (0.175 mmol) solution dissolved in dichloromethane, and naturally rose to room temperature after the completion of the dropwise addition. The reaction was overnight. The reaction solution was concentrated the next day, and the residue was recrystallized from isopropanol to give monomethoxy polyethylene glycol glutamic acid dipeptide glycine (20K) - dasatinib conjugate (DSR-2) 0.39 g. 1H-NMR (DMSO-d 6 ): 2.12 (s, 9H), 2.13 (m, 2H), 2.17 (m, 2H), 2.40 (t, 4H), 2.44 (s, 9H), 2.97 (t, 6H) ), 3.30 (s, 3H),
3.44 (t， 12H), 3.50 (s， 2H), 3.51(s, 6H)， 3.54(m,1800H), 3.62 (t， 12H), 4.35 (t， 6H)，3.44 (t, 12H), 3.50 (s, 2H), 3.51 (s, 6H), 3.54 (m, 1800H), 3.62 (t, 12H), 4.35 (t, 6H),
4.45 (t， 1H), 4.53 (t， 1H), 5.28 (s， 3H), 7.01 (m， 3H), 7.26 (m， 3H), 7.44 (m， 3H), 8.03 (s， 2H), 8.05 (s， 3H), 8.11 (s， 3H), 9.15 (s, 3H), 11.45 (s， 3H). 4.45 (t, 1H), 4.53 (t, 1H), 5.28 (s, 3H), 7.01 (m, 3H), 7.26 (m, 3H), 7.44 (m, 3H), 8.03 (s, 2H), 8.05 (s, 3H), 8.11 (s, 3H), 9.15 (s, 3H), 11.45 (s, 3H).
[00145】 实施例 3: 单甲氧基聚乙二醇谷氨酸二肽丙氨酸 (数均分子量 20000) 一达沙替尼结合物 （DSR-3) 的制备。
Figure imgf000035_0001
Example 3: Preparation of monomethoxypolyethylene glycol glutamic acid dipeptide alanine (number average molecular weight 20000) - Dasatinib conjugate (DSR-3).
Figure imgf000035_0001
DSR-3  DSR-3
[00146] 根据买施例 1所述的方法制备单甲氧基聚乙二醇谷氨酸二肽酸 （化 合物 5)。 [00146] Monomethoxypolyethylene glycol glutamic acid dipeptide acid (Compound 5) was prepared according to the method described in Preparation Example 1.
[00147] 将达沙替尼 1.95g (4 mmol), N-叔丁氧羰基丙氨酸 907 mg (4.8mmol) 禾口 DMAP 714 mg (4.8 mmol) 加到反应瓶中， 用 40 mL N，N-二甲基甲酰胺溶 解，氮气保护下冰浴冷却，再滴入 DCC 1.03 g C5mmol) 溶于 20 mL二氯甲垸的 溶液， 滴完后自然升到室温反应过夜。 TLC监测反应完全后减压蒸去溶剂， 残 分用柱层析分离， 得 N-叔丁氧羰基丙氨酸达沙替尼酯 （化合物 8) 1.6 g， 收率 61%。 1H-NMR (DMS0-d6): 1.33(s, 9H)， 1.49(s， 3H), 2.12(s, 3H), 2.45(s, 3H), 2.98(m, 2H), 3.46(m, 4H),3.63(m, 4H)， 4.22(m, 2H), 4.3 l(m, 1H), 5.97(s, 1H)， 7.29(m, 3H), 8.15(s, 1H)， 8.32(s, 1H)， 9.66(s, 1H)， 11.17(s, 1H)。 [00147] 1.72 g (4 mmol) of Dasatinib, N-tert-butoxycarbonylalanine 907 mg (4.8 mmol) and DMAP 714 mg (4.8 mmol) were added to the reaction flask with 40 mL of N, The N-dimethylformamide was dissolved, cooled in an ice bath under nitrogen atmosphere, and then dropped into a solution of DCC 1.03 g C5 mmol) dissolved in 20 mL of dichloromethane. After the dropwise addition, the mixture was allowed to react to room temperature overnight. After the reaction was completed by TLC, the solvent was evaporated under reduced pressure, and the residue was separated by column chromatography to afford 1.6 g of N-tert-butoxycarbonyl alanine (Compound 8) in a yield of 61%. 1H-NMR (DMS0-d 6 ): 1.33 (s, 9H), 1.49 (s, 3H), 2.12 (s, 3H), 2.45 (s, 3H), 2.98 (m, 2H), 3.46 (m, 4H) ), 3.63 (m, 4H), 4.22 (m, 2H), 4.3 l (m, 1H), 5.97 (s, 1H), 7.29 (m, 3H), 8.15 (s, 1H), 8.32 (s, 1H) ), 9.66(s, 1H), 11.17(s, 1H).
[00148] 将 N-叔丁氧羰基丙氨酸达沙替尼酯 （化合物 8) 1 g (1.6mmol), 二氯 甲垸 50 mL， 加到反应瓶中， 再加入三氟乙酸 25 mL， 搅拌反应 3h后减压蒸去 溶剂， 残分中加入二氯甲垸， 减压蒸干， 如此重复三次， 得丙氨酸达沙替尼酯 三氟乙酸盐 （化合物 9) 1.3 g， 直接往下反应。 [00148] N-tert-Butoxycarbonylalanine dasatinib (Compound 8) 1 g (1.6 mmol), dichloromethane (50 mL) was added to the reaction flask, then 25 mL of trifluoroacetic acid was added. After stirring for 3 hours, the solvent was evaporated under reduced pressure, dichloromethane was added to the residue, and evaporated to dryness under reduced pressure, and then repeated three times to obtain dasatinib alanine. Trifluoroacetate (Compound 9) 1.3 g, directly reacted downward.
[00149】 将单甲氧基聚乙二醇谷氨酸二肽酸 （化合物 5) (20K，0.5g，  [00149] Monomethoxy polyethylene glycol glutamic acid dipeptide acid (Compound 5) (20K, 0.5g,
0.025mmol) ，丙氨酸达沙替尼酯三氟乙酸盐（化合物 9) 102 mg (0.15mmol)， HOBt (3.4 mg, 0.025mmo) 和 DMAP 24.4 mg(0.2mmol) 加到反应瓶中， 用二氯 甲垸和 N，N-二甲基甲酰胺混合溶剂溶解， 氮气保护下冰浴冷却， 再滴入 DCC 36.1 mg (0.175mmol) 溶于二氯甲垸的溶液， 滴完后自然升到室温反应过夜。次 日将反应液浓缩， 残分用异丙醇重结晶， 得单甲氧基聚乙二醇谷氨酸二肽丙氨 酸（20K)—达沙替尼结合物（DSR-3) 0.39 g。 1H-NMR (DMSO-d6): 1.28 (s, 3H), 2.12(s, 9H)， 2.13 (m， 2H), 2.17 (m， 2H), 2.40 (t， 4H)， 2.44 (s， 9H)， 2.97 (t， 6H)， 3.30 (s， 3H), 3.44 (t， 12H), 3.50 (s， 2H), 3.54(m,1800H), 3.62 (t， 12H), 3.63 (m， 3H), 4.18 (t, 6H)， 4.45 (t, 1H), 4.53 (t, 1H), 5.28 (s, 3H), 7.01 (m， 3H), 7.26 (m， 3H), 7.44 (m， 3H), 8.03 (s, 2H), 8.05 (s, 3H), 8.10 (s, 3H), 9.15 (s, 3H), 11.44 (s, 3H). 0.025 mmol), dasatinib alanine trifluoroacetate (compound 9) 102 mg (0.15 mmol), HOBt (3.4 mg, 0.025 mmo) and DMAP 24.4 mg (0.2 mmol) were added to the reaction flask. Dissolve in a mixed solvent of methylene chloride and N,N-dimethylformamide, cool in an ice bath under nitrogen atmosphere, and then add DCC 36.1 mg (0.175 mmol) in dichloromethane solution. The reaction was allowed to reach room temperature overnight. The reaction solution was concentrated the next day, and the residue was recrystallized from isopropyl alcohol to give monomethoxy polyethylene glycol glutamic acid dipeptide alanine (20K) - dasatinib conjugate (DSR-3) 0.39 g . 1H-NMR (DMSO-d 6 ): 1.28 (s, 3H), 2.12 (s, 9H), 2.13 (m, 2H), 2.17 (m, 2H), 2.40 (t, 4H), 2.44 (s, 9H) ), 2.97 (t, 6H), 3.30 (s, 3H), 3.44 (t, 12H), 3.50 (s, 2H), 3.54 (m, 1800H), 3.62 (t, 12H), 3.63 (m, 3H) , 4.18 (t, 6H), 4.45 (t, 1H), 4.53 (t, 1H), 5.28 (s, 3H), 7.01 (m, 3H), 7.26 (m, 3H), 7.44 (m, 3H), 8.03 (s, 2H), 8.05 (s, 3H), 8.10 (s, 3H), 9.15 (s, 3H), 11.44 (s, 3H).
[00150】 实施例 4: 单甲氧基聚乙二醇谷氨酸二肽缬氨酸 (数均分子量 20000) 一达沙替尼结合物 （DSR-4) 的制备。  Example 4: Monomethoxypolyethylene glycol glutamate dipeptide proline (number average molecular weight 20000) Preparation of a dasatinib conjugate (DSR-4).
Figure imgf000036_0001
[00151】 根据实施例 1所述的方法制备单甲氧基聚乙二醇谷氨酸二肽酸 （化 合物 5)。
Figure imgf000036_0001
[00151] Monomethoxypolyethylene glycol glutamic acid dipeptide acid (Compound 5) was prepared according to the method described in Example 1.
[00152】 将达沙替尼 1.95g (4 mmol), N-叔丁氧羰基缬氨酸 1.1 lg (4.8mmol)和 DMAP 714 mg (4.8 mmol)加到反应瓶中， 用 40 mL N，N-二甲基甲酰胺溶解， 氮 气保护下冰浴冷却， 再滴入 DCC 1.03 g (5mmol) 溶于 20 mL二氯甲垸的溶液， 滴完后自然升温反应过夜。 TLC监测反应完全后减压蒸去溶剂， 残分用柱层析 分离， 得 N-叔丁氧羰基缬氨酸达沙替尼酯 （化合物 10) 1.7 g， 收率 62%。 [00152] 1.72 g (4 mmol) of dasatinib, 1.1 g (4.8 mmol) of N-tert-butoxycarbonylproline and 714 mg (4.8 mmol) of DMAP were added to the reaction flask with 40 mL of N,N - Dimethylformamide was dissolved, cooled in an ice bath under nitrogen atmosphere, and then DCC 1.03 g (5 mmol) dissolved in 20 mL of dichloromethane was added dropwise. After the reaction was completed by TLC, the solvent was evaporated under reduced pressure, and the residue was separated by column chromatography to give 1.7 g of N-tert-butoxycarbonyl valine valine (Compound 10) in a yield of 62%.
1H-NMR (DMSO-d6): 1.02 (d， 6H)， 1.21 (s， 9H)， 2.23(s, 3H), 2.49(s, 3H), 2.5 l(m, 4H)， 2.78 (m， 6H)， 3.53(m, 8H)， 6.04(s, 1H), 7.27(m, 2H), 7.40(m, 1H), 8.22(s, 1H)， 9.88(s， 1H)， 11.48(s, 1H)。 1H-NMR (DMSO-d 6 ): 1.02 (d, 6H), 1.21 (s, 9H), 2.23 (s, 3H), 2.49 (s, 3H), 2.5 l (m, 4H), 2.78 (m, 6H), 3.53(m, 8H), 6.04(s, 1H), 7.27(m, 2H), 7.40(m, 1H), 8.22(s, 1H), 9.88(s, 1H), 11.48(s, 1H) ).
[00153】 将 N-叔丁氧羰基缬氨酸达沙替尼酯 1 g G.5mmol)，二氯甲垸 50 mL, 三氟乙酸 25 mL加到反应瓶中， 搅拌下反应 3h后减压蒸去溶剂， 残分中加入 二氯甲垸， 减压蒸干， 如此重复三次， 得缬氨酸达沙替尼酯三氟乙酸盐 （化合 物 11) 1.4 g， 直接往下反应。  [00153] N-tert-butoxycarbonyl valine dasatinib 1 g G. 5mmol), dichloromethane 50 mL, 25 mL of trifluoroacetic acid was added to the reaction flask, and the reaction was carried out for 3 hours under stirring. The solvent was evaporated, dichloromethane was added to the residue, and the mixture was evaporated to dryness, and the mixture was evaporated to dryness, and the mixture was then reacted three times to obtain 1.4 g of dasatinib triacetate (Compound 11).
[00154】 将单甲氧基聚乙二醇谷氨酸二肽酸 （化合物 5) (20K，0.5g，  [00154] Monomethoxy polyethylene glycol glutamic acid dipeptide acid (Compound 5) (20K, 0.5g,
0.025mmol) ， 缬氨酸达沙替尼酯三氟乙酸盐（化合物 11)140 mg (0.15mmol)， HOBt (3.4 mg, 0.025mmo) 和 DMAP 24.4 mg(0.2mmol) 加到反应瓶中， 用二氯 甲垸和 N，N-二甲基甲酰胺混合溶剂溶解， 氮气保护下冰浴冷却， 再滴入 DCC 36.1 mg (0.175mmol) 溶于二氯甲垸的溶液， 滴完后自然升温反应过夜。次日将 反应液浓缩， 残分用异丙醇重结晶， 得单甲氧基聚乙二醇谷氨酸二肽缬氨酸0.025 mmol), valine dasatinib trifluoroacetate (Compound 11) 140 mg (0.15 mmol), HOBt (3.4 mg, 0.025 mmo) and DMAP 24.4 mg (0.2 mmol) were added to the reaction flask. Dissolve in a mixed solvent of methylene chloride and N,N-dimethylformamide, cool in an ice bath under nitrogen atmosphere, and then add DCC 36.1 mg (0.175 mmol) in a solution of dichloromethane. The reaction was overnight. The reaction solution was concentrated the next day, and the residue was recrystallized from isopropanol to obtain monomethoxy polyethylene glycol glutamic acid dipeptide proline.
(20K) 一达沙替尼结合物 (DSR-4) 0.32g。 1H-NMR (DMSO-d6): 0.91 (s, 18H), 2.12(s, 9H)， 2.13 (m， 2H), 2.17 (m， 2H), 2.40 (t， 4H)， 2.44 (s， 9H)， 2.67 (m， 3H), 2.97 (t， 6H)， 3.30 (s， 3H), 3.44 (t， 12H), 3.46 (d， 3H),3.50 (s， 2H), 3.54(m,1800H), 3.62 (t， 12H), 4.18 (t， 6H)， 4.45 (t， 1H), 4.53 (t， 1H), 5.28 (s， 3H), 7.01 (m， 3H), 7.26 (m， 3H), 7.44 (m， 3H), 8.03 (s, 2H), 8.05 (s, 3H), 8.10 (s, 3H), 9.15 (s, 3H), 11.44 (s, 3H). (20K) One dasatinib conjugate (DSR-4) 0.32g. 1H-NMR (DMSO-d 6 ): 0.91 (s, 18H), 2.12 (s, 9H), 2.13 (m, 2H), 2.17 (m, 2H), 2.40 (t, 4H), 2.44 (s, 9H) ), 2.67 (m, 3H), 2.97 (t, 6H), 3.30 (s, 3H), 3.44 (t, 12H), 3.46 (d, 3H), 3.50 (s, 2H), 3.54 (m, 1800H) , 3.62 (t, 12H), 4.18 (t, 6H), 4.45 (t, 1H), 4.53 (t, 1H), 5.28 (s, 3H), 7.01 (m, 3H), 7.26 (m, 3H), 7.44 (m, 3H), 8.03 (s, 2H), 8.05 (s, 3H), 8.10 (s, 3H), 9.15 (s, 3H), 11.44 (s, 3H).
[00155】 实施例 5: Y型聚乙二醇谷氨酸二肽缬氨酸（数均分子量 40000)  Example 5: Y-type polyethylene glycol glutamic acid dipeptide proline (number average molecular weight 40000)
Figure imgf000038_0001
Figure imgf000038_0001
DSR-5  DSR-5
[00156】 N-叔丁氧羰基谷氨酸苄酯二肽（化合物 2) 0.78g (实施例 1) 溶解在 7 mL二氯甲垸中， 加入三氟乙酸 3mL， 室温反应 2h。 除去溶剂， 加入 lOOmL 二氯甲垸， 用 5%碳酸氢钠溶液调节 pH=7-8。 萃取分液， 有机相用 5%碳酸氢 钠溶液洗两次， 无水硫酸钠干燥。 过滤， 滤液直接加到反应瓶中， 氮气保护下， 加入 Y型聚乙二醇乙酸 (40K) 40.0g、 DMAP 245 mg(2 mmol)、 HOBt 135 mg(l mmol)。 全部溶解后， 加入 DCC 412mg(2 mmol)。 室温搅拌反应过夜。 过滤， 旋转蒸发除去溶剂， 残余物中添加 500mL异丙醇， 过滤， 产物真空干燥。 将该 产物溶于 200mL无水甲醇中， 加入钯炭 1.0g， 通入氢气室温反应过夜。过滤除 去钯炭， 旋转蒸发除去溶剂， 残余物中添加 500mL异丙醇， 过滤， 产物真空干 燥。 得到 Y型聚乙二醇谷氨酸二肽酸 （40K) (化合物 13) 33.4g。 [00156] N-tert-Butoxycarbonylglutamic acid benzyl ester dipeptide (Compound 2) 0.78 g (Example 1) Dissolved in 7 mL of dichloromethane, 3 mL of trifluoroacetic acid, and reacted at room temperature for 2 h. The solvent was removed, 100 mL of dichloromethane was added, and the pH was adjusted to 7-8 with a 5% sodium hydrogen carbonate solution. The extract was separated and the organic phase was washed twice with 5% sodium hydrogen carbonate solution and dried over anhydrous sodium sulfate. Filtration, the filtrate was directly added to the reaction flask, under nitrogen protection, Y-type polyethylene glycol acetate (40K) 40.0 g, DMAP 245 mg (2 mmol), HOBt 135 mg (1 mmol) were added. After all was dissolved, DCC 412 mg (2 mmol) was added. The reaction was stirred at room temperature overnight. After filtration, the solvent was removed by rotary evaporation. EtOAc (EtOAc) This product was dissolved in 200 mL of anhydrous methanol, and 1.0 g of palladium on carbon was added thereto, and the mixture was reacted with hydrogen at room temperature overnight. The palladium on charcoal was removed by filtration, the solvent was removed by rotary evaporation, and 500 mL of isopropyl alcohol was added to the residue, which was filtered and dried. Y-type polyethylene glycol glutamic acid dipeptide acid (40K) (Compound 13) was obtained in an amount of 33.4 g.
[00157】 根据实施例 4所述的方法制备缬氨酸达沙替尼酯三氟乙酸盐 (化合物 11 ) 。 将 Y型聚乙二醇谷氨酸二肽酸 （化合物 13) (40K，0.6g， 0.02mmol) ，缬 氨酸达沙替尼酯三氟乙酸盐 112 mg (0.12mmol)， HOBt (2.7 mg， 0.02mmo) 禾口 DMAP 24.4 mg(0.2mmol) 加到反应瓶中，用二氯甲垸和 N，N-二甲基甲酰胺混合 溶剂溶解， 氮气保护下冰浴冷却， 再滴入 DCC 36.1 mg (0.175mmol) 溶于二氯 甲垸的溶液， 滴完后自然升温反应过夜。 次日将反应液浓缩， 残分用异丙醇重 结晶， 得 Y型聚乙二醇谷氨酸二肽缬氨酸 （40K) —达沙替尼结合物 （DSR-5) 0.39g. 1H-NMR (DMSO-d6): 0.91 (s， 18H), 2.12(s, 9H)， 2.13 (m， 2H), 2.17 (m， 2H), 2.40, (t， 4H)， 2.44 (s， 9H)， 2.67 (m， 3H), 2.97 (t， 6H)， 3.30 (s， 6H)， 3.37 (t， 2H), 3.44 (t, 12H), 3.46 (d, 3H),3.54 (m，1800H)， 3.62 (t, 12H), 3.76 (t, 2H), 4.09 (s, 2H), 4.18 (t， 6H)， 4.26 (s， 2H), 4.45 (t， 1H), 4.53 (t， 1H), 5.28 (s， 3H), 7.01 (m， 3H), 7.26 (m， 3H), 7.44 (m， 3H), 8.03 (s， 2H), 8.05 (s， 3H), 8.10 (s， 3H), 9.15 (s， 3H), 11.44 (s， 3H). [00157] The valine dasatinib trifluoroacetate (Compound 11) was prepared according to the method described in Example 4. Y-type polyethylene glycol glutamic acid dipeptide acid (Compound 13) (40K, 0.6g, 0.02mmol), Dasatinib triacetate valine 112mg (0.12mmol), HOBt (2.7 Mg, 0.02mmo) and DMAP 24.4 mg (0.2mmol) were added to the reaction flask, dissolved in a mixed solvent of methylene chloride and N,N-dimethylformamide, cooled in an ice bath under nitrogen protection, and then dropped into DCC. 36.1 mg (0.175 mmol) of a solution dissolved in dichloromethane, which was naturally warmed up overnight after the completion of the dropwise addition. The reaction solution was concentrated the next day, and the residue was recrystallized from isopropanol to give Y-type polyethylene glycol glutamic acid dipeptide proline (40K) - dasatinib conjugate (DSR-5) 0.39 g. 1H -NMR (DMSO-d 6 ): 0.91 (s, 18H), 2.12 (s, 9H), 2.13 (m, 2H), 2.17 (m, 2H), 2.40, (t, 4H), 2.44 (s, 9H) ), 2.67 (m, 3H), 2.97 (t, 6H), 3.30 (s, 6H), 3.37 (t, 2H), 3.44 (t, 12H), 3.46 (d, 3H), 3.54 (m, 1800H) , 3.62 (t, 12H), 3.76 (t, 2H), 4.09 (s, 2H), 4.18 (t, 6H), 4.26 (s, 2H), 4.45 (t, 1H), 4.53 (t, 1H), 5.28 (s, 3H), 7.01 (m, 3H), 7.26 (m, 3H), 7.44 (m, 3H), 8.03 (s, 2H), 8.05 (s, 3H), 8.10 (s, 3H), 9.15 (s, 3H), 11.44 (s, 3H).
[00158】 实施例 6: 单甲氧基聚乙二醇谷氨酸三肽缬氨酸 (数均分子量 20000) 一达沙替尼结合物 （DSR-6) 的制备
Figure imgf000040_0001
[00159】 N-叔丁氧羰基谷氨酸苄酯二肽（化合物 2) 6.47g (O.Olmol) 溶解在 15 mL二氯甲垸中， 加入三氟乙酸 6mL， 室温反应 2h。 除去溶剂， 加入 lOOmL 二氯甲垸， 用 5%碳酸氢钠溶液调节 pH=7-8。 萃取分液， 有机相用 5%碳酸氢 钠溶液洗两次， 无水硫酸钠干燥。 过滤， 滤液直接加到反应瓶中， 氮气保护下， 加入叔丁氧羰基 -L-谷氨酸 -5-苄酯 3.37g (O.Olmol) ， DMAP 1.22g ( O.Olmol) 、 HOBt l.35g (O.Olmol) ， 完全溶解后滴加 DCC 2.39g (O.Ol lmol) 的二氯甲垸 溶液。 滴毕， 密闭体系反应过夜。 TLC监测反应完毕。 过滤， 依次用 10%柠檬 酸30mL*3)、 5%碳酸氢钠 C30mL*3)、 5%饱和氯化钠水溶液 C30mL*3)洗涤， 无 水硫酸钠干燥。过滤， 除去溶剂后， 浓缩液中加入 25 mL乙酸乙酯， 滤除固体， 母液加入石油醚 400 mL沉淀， 过滤得产品 N-叔丁氧羰基谷氨酸苄酯三肽 （化 合物 14) 6.8g。 Example 6: Preparation of monomethoxypolyethylene glycol glutamic acid tripeptide valine (number average molecular weight 20000) - Dasatinib conjugate (DSR-6)
Figure imgf000040_0001
[00159] N-tert-Butoxycarbonyl glutamic acid benzyl ester dipeptide (Compound 2) 6.47 g (0.1 mol) was dissolved in 15 mL of dichloromethane, 6 mL of trifluoroacetic acid was added, and reacted at room temperature for 2 h. The solvent was removed, 100 mL of dichloromethane was added, and the pH was adjusted to 7-8 with a 5% sodium hydrogen carbonate solution. The extract was separated and the organic phase was washed twice with 5% sodium hydrogen carbonate solution and dried over anhydrous sodium sulfate. Filtration, the filtrate was directly added to the reaction flask, under the protection of nitrogen, tert-butoxycarbonyl-L-glutamic acid-5-benzyl ester 3.37g (O.Olmol), DMAP 1.22g (O.Olmol), HOBt l. 35 g (O.Olmol), after completely dissolved, a solution of 2.39 g (O.Ol.lmol) of dichloromethane was added dropwise. After the dropwise addition, the closed system was reacted overnight. The reaction was monitored by TLC. Filtration, washing with 10% citric acid 30 mL*3), 5% sodium hydrogencarbonate C30 mL*3), 5% saturated sodium chloride aqueous solution C30 mL*3), dried over anhydrous sodium sulfate. After filtration, the solvent was removed, 25 mL of ethyl acetate was added to the concentrate, and the solid was filtered off. The mother liquid was added to petroleum ether 400 mL to precipitate and filtered to obtain the product N-tert-butoxycarbonyl glutamic acid benzyl ester tripeptide (Compound 14) 6.8 g .
[00160】 N-叔丁氧羰基谷氨酸苄酯三肽 （化合物 14) 1.04g (1 mmol) 溶解在 7 mL二氯甲垸中， 加入三氟乙酸 3mL， 室温反应 2h。 除去溶剂， 加入 200mL二 氯甲垸， 用 5%碳酸氢钠溶液调节 pH=7-8。 萃取分液， 有机相用 5%碳酸氢钠 溶液洗两次， 无水硫酸钠干燥。 过滤， 滤液直接加入到反应瓶中， 氮气保护下， 加入单甲氧基聚乙二醇乙酸 (20K) 20.0g(l mmol) DMAP 245 mg(2 mmol)、HOBt 135 mg(l mmol)o全部溶解后， 加入 DCC 412mg(2 mmol)。 室温搅拌反应过夜。 过滤， 旋转蒸发除去多余溶剂， 残余物中添加 500mL异丙醇， 过滤， 产物真空 干燥。将该产物溶于 200mL无水甲醇中， 加入钯炭 1.0g， 通入氢气室温反应过 夜。 过滤除去钯炭， 旋转蒸发除去多余溶剂， 残余物中添加 500mL异丙醇， 过 滤， 产物真空干燥。 得到单甲氧基聚乙二醇谷氨酸三肽酸 (20K ) (化合物 17) 15.4g。  [00160] N-tert-Butoxycarbonyl glutamic acid benzyl ester tripeptide (Compound 14) 1.04 g (1 mmol) was dissolved in 7 mL of dichloromethane, 3 mL of trifluoroacetic acid was added, and reacted at room temperature for 2 h. The solvent was removed, 200 mL of dichloromethane was added, and pH = 7-8 was adjusted with a 5% sodium hydrogen carbonate solution. The extract was separated, and the organic phase was washed twice with a 5% sodium hydrogen carbonate solution and dried over anhydrous sodium sulfate. Filtration, the filtrate was directly added to the reaction flask, and under nitrogen protection, monomethoxypolyethylene glycol acetic acid (20K) 20.0 g (1 mmol) DMAP 245 mg (2 mmol), HOBt 135 mg (1 mmol) o were added. After dissolution, DCC 412 mg (2 mmol) was added. The reaction was stirred at room temperature overnight. After filtration, the excess solvent was removed by rotary evaporation, and 500 mL of isopropyl alcohol was added to the residue, which was filtered and dried. This product was dissolved in 200 mL of anhydrous methanol, and 1.0 g of palladium on carbon was added thereto, and the mixture was reacted with hydrogen at room temperature overnight. The palladium on charcoal was removed by filtration, and excess solvent was removed by rotary evaporation. 500 mL of isopropyl alcohol was added to the residue, and the product was filtered and dried. Monomethoxypolyethylene glycol glutamic acid tripeptide acid (20K) (Compound 17) 15.4 g was obtained.
[00161】 根据实施例 4所述的方法制备缬氨酸达沙替尼酯三氟乙酸盐 (化合物 11 ) 。 将单甲氧基聚乙二醇谷氨酸三肽酸 (化合物 17) (20K, 0.6 g， 0.03mmol)、 缬氨酸达沙替尼酯三氟乙酸盐 （化合物 ll) 224 mg(0.24mmol)、 HOBt (4 mg, 0.03mmol) 禾卩 DMAP 29.3 mg(0.24mmol) 加到反应瓶中， 用二氯甲垸和 N，N-二 甲基甲酰胺混合溶剂溶解， 氮气保护下冰浴冷却， 再滴入 DCC 43.3 mg [00161] The valine dasatinib trifluoroacetate (Compound 11) was prepared according to the method described in Example 4. Monomethoxy polyethylene glycol glutamic acid tripeptide (Compound 17) (20K, 0.6 g, 0.03 mmol), Valine dasatinib trifluoroacetate (compound ll) 224 mg (0.24 mmol), HOBt (4 mg, 0.03 mmol) and hydrazine DMAP 29.3 mg (0.24 mmol) were added to the reaction flask with dichloro Methyl hydrazine and N,N-dimethylformamide were dissolved in a mixed solvent, cooled in an ice bath under nitrogen protection, and then dropped into DCC 43.3 mg.
(0.21mmol) 溶于二氯甲垸的溶液， 滴完后自然升温反应过夜。 次日将反应液浓 缩， 残分用异丙醇重结晶， 得到单甲氧基聚乙二醇谷氨酸三肽缬氨酸 （20K) 一达沙替尼结合物 （DSR-6) 0.45g。 1H-NMR (DMSO-d6): 0.91 (s， 24H)， 1.93 (m， 2H), 2.05 (t， 2H), 2.12(s, 12H), 2.13 (m， 2H), 2.17 (m， 2H), 2.40, (t， 4H)， 2.44 (s， 12H), 2.67 (m， 4H)， 2.97 (t， 8H)， 3.30 (s， 3H), 3.44 (t， 16H), 3.46 (d， 4H),3.50 (s， 2H), 3.54(m,1800H), 3.56 (t， 2H), 3.62 (t， 16H), 4.18 (t， 8H)， 4.45 (t， 1H), 4.53 (t， 1H), 5.28 (s， 4H)， 7.01 (m， 4H)， 7.26 (m， 4H)， 7.44 (m， 4H)， 8.03 (s， 3H), 8.05 (s， 4H), 8.10 (s, 4H), 9.15 (s, 4H), 11.44 (s, 4H). (0.21 mmol) A solution dissolved in dichloromethane was naturally warmed up overnight after the completion of the dropwise addition. The reaction solution was concentrated the next day, and the residue was recrystallized from isopropanol to give monomethoxy polyethylene glycol glutamic acid tripeptide valine (20K)-dasatinib conjugate (DSR-6) 0.45 g . 1H-NMR (DMSO-d 6 ): 0.91 (s, 24H), 1.93 (m, 2H), 2.05 (t, 2H), 2.12 (s, 12H), 2.13 (m, 2H), 2.17 (m, 2H) ), 2.40, (t, 4H), 2.44 (s, 12H), 2.67 (m, 4H), 2.97 (t, 8H), 3.30 (s, 3H), 3.44 (t, 16H), 3.46 (d, 4H) ), 3.50 (s, 2H), 3.54 (m, 1800H), 3.56 (t, 2H), 3.62 (t, 16H), 4.18 (t, 8H), 4.45 (t, 1H), 4.53 (t, 1H) , 5.28 (s, 4H), 7.01 (m, 4H), 7.26 (m, 4H), 7.44 (m, 4H), 8.03 (s, 3H), 8.05 (s, 4H), 8.10 (s, 4H), 9.15 (s, 4H), 11.44 (s, 4H).
[00162】 实施例 7: 化合物 DSR1〜6的体外降解试验  Example 7: In vitro degradation test of compound DSR1~6
[00163] 将化合物 DSR1〜6溶解在 0.01 M的 PBS缓冲液中，考察化合物在 PBS 缓冲液中的降解情况， 每隔 30min 取样分析， 降解情况如图 1所示。  [00163] Compounds DSR1~6 were dissolved in 0.01 M PBS buffer, and the degradation of the compound in PBS buffer was investigated and sampled and analyzed every 30 minutes. The degradation is shown in Fig. 1.
[00164】 从图中可以看出， 几种化合物在 PBS缓冲液中随时间延长都有不同 程度的降解。化合物 DSR— 1降解最快， 60min时即有 60%的降解；化合物 DSR-2 和 DSR-3降解稍缓， 但分别在 90min时和 150min时， 也有 50%的降解； 化合 物 DSR-4, DSR-5和 DSR-6则不同于前三个化合物， 降解较慢， 在 240min时， 仍只有约 20%的降解， 稳定性大大高于 DSR-l，DSR-2和 DSR-3。 从而可见， 通过两种氨基酸连接的药物结合物（即 DSR-2, DSR-3, DSR-4, DSR-5和 DSR-6) 的稳定性要高于仅通过一个氨基酸连接的药物化合物 （即 DSR-1 ) 的稳定性。 当使用缬氨酸作为第二氨基酸连接药物时， 结合物的稳定更是极大的提高了。 [00164] As can be seen from the figure, several compounds have different degrees of degradation over time in PBS buffer. Compound DSR-1 degraded the fastest, 60% degradation at 60 min; compounds DSR-2 and DSR-3 degraded slightly, but also 50% degradation at 90 min and 150 min, respectively; Compound DSR-4, DSR -5 and DSR-6 are different from the first three compounds, and the degradation is slower. At 240 min, there is still only about 20% degradation, and the stability is much higher than DSR-1, DSR-2 and DSR-3. Thus, it can be seen that drug conjugates linked by two amino acids (ie, DSR-2, DSR-3, DSR-4, DSR-5, and DSR-6) are more stable than drug compounds linked by only one amino acid (ie, Stability of DSR-1). When valine is used as the second amino acid to link the drug, the stability of the conjugate is greatly enhanced.
[00165】 实施例 8: 不同聚乙二醇达沙替尼结合物在 K562人慢性髓系白血病 皮下瘤模型中的抗肿瘤作用。 [00166】 实验方法： Example 8: Antitumor effects of different polyethylene glycol dasatinib conjugates in a K562 human chronic myeloid leukemia subcutaneous tumor model. [00166] Experimental method:
[00167】 于 NOD/SCID小鼠右侧背部皮下接种 K562细胞，建立人慢性髓系白 血病异种移植动物皮下模型。 当平均肿瘤体积达到 130mm3时， 将实验小鼠分 组， 每组 8只， 静脉注射给药， 一周两次。 根据相对肿瘤增殖率 （T/C%)进行 疗效评价。  [00167] K562 cells were subcutaneously inoculated into the right side of NOD/SCID mice to establish a subcutaneous model of human chronic myeloid leukemia xenograft animals. When the average tumor volume reached 130 mm3, the experimental mice were divided into groups of 8 and administered intravenously twice a week. The efficacy was evaluated based on the relative tumor growth rate (T/C%).
[00168】 实验步骤：  [00168] Experimental steps:
[00169】 （1 ) 细胞培养  [00169] (1) Cell culture
[00170】 K562细胞系在补充了 10%胎牛血清， L-谷氨酰胺 (；2 mM)的  [00170] K562 cell line is supplemented with 10% fetal bovine serum, L-glutamine (2 mM)
RPMI-1640培养基中， 在 37°C， 空气中含 5% C02的条件下体外培养。 肿瘤细 胞每周常规传代 2次。 收集处于指数生长期的肿瘤细胞， 悬于等体积的 PBS : 基质胶混合物中， 置于冰上， 用于肿瘤接种。  In RPMI-1640 medium, culture in vitro at 37 ° C in the presence of 5% CO 2 in air. Tumor cells were routinely passaged twice a week. Tumor cells in the exponential growth phase were collected and suspended in an equal volume of PBS: Matrigel mixture and placed on ice for tumor inoculation.
[00171】 （2) 动物分组 [00171] (2) Animal grouping
[00172】 实验小鼠于右侧背部皮下接种 5x 106 K562细胞， 定期观察肿瘤生长 情况， 待肿瘤生长至平均 130mm3时根据肿瘤大小和小鼠体重随机分组并开始 给药治疗。  [00172] Experimental mice were subcutaneously inoculated with 5 x 106 K562 cells on the right side of the right side, and tumor growth was periodically observed. When the tumors were grown to an average of 130 mm3, the tumor size and the body weight of the mice were randomly grouped and treatment was started.
[00173】 （3 ) 实验观察  [00173] (3) Experimental observation
[00174] 本研究中根据 Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC)批准的指南进行与操作、 护理、 治疗相关的 所有操作。 在接种后， 每天观察动物的发病与死亡。 在日常观察时， 注意肿瘤 生长对动物正常行为 （如运动、 进食、 饮水、 体重增减、 眼睛、 被毛） 的影响 以及其他任何异常。 记录每组内动物的死亡和临床症状。  [00174] All operations related to operation, care, and treatment were performed in this study in accordance with guidelines approved by the Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC). After vaccination, the morbidity and mortality of the animals were observed daily. During daily observation, pay attention to the effects of tumor growth on normal animal behavior (such as exercise, eating, drinking, weight gain, eyes, coat) and any other abnormalities. The death and clinical symptoms of the animals in each group were recorded.
[00175】 （4) 结果判断 [00175] (4) Result judgment
[00176】 在整个实验过程中，每周测量两次小鼠的体重和肿瘤的大小。肿瘤大 小计算公式： 肿瘤体积 (mm3) =0.5 x (肿瘤长径 x 肿瘤短径 2)。根据实验组和 对照组肿瘤体积大小， 计算 RTV和 T/C比值。 RTV是指相对肿瘤体积。 T/C 比值是指在结束治疗后某一时间点，治疗组和对照组相对肿瘤体积的百分比值， 反映不同治疗组的抗瘤药效作用。 [00177】 （5 ) 统计分析 [00176] The body weight and tumor size of the mice were measured twice a week throughout the experiment. Tumor size calculation formula: Tumor volume (mm3) = 0.5 x (tumor long diameter x tumor short diameter 2). According to the experimental group and The tumor volume of the control group was calculated, and the RTV and T/C ratios were calculated. RTV refers to the relative tumor volume. The T/C ratio is the percentage of the tumor volume relative to the treatment group and the control group at some point after the end of treatment, reflecting the antitumor efficacy of the different treatment groups. [00177] (5) Statistical analysis
[00178】 所有实验结果以平均瘤体积 ± SE (标准误差） 表示， 统计分析时， 相对肿瘤体积数据在统计时方差不齐，使用单因素方差分析（one-way ANOVA) 及 Dunnett T3多重比较方法对各组间进行相对肿瘤体积显著性差异的比较， ρ < 0.05为差异显著。 [00178] All experimental results are expressed as mean tumor volume ± SE (standard error). For statistical analysis, relative tumor volume data are statistically uneven, using one-way ANOVA and Dunnett T3 multiple comparison method. A comparison of the significant differences in tumor volume between the groups was significant, with ρ < 0.05 being significant.
[00179】 实验结果： 各治疗组和溶剂对照组肿瘤生长情况见表 1和表 2。 [00180】 表 1. 各治疗组 （5mg/kg) 肿瘤体积 17〔％值 （与溶剂对照组比较） 分组后第 14天 [00179] Experimental results: The tumor growth of each treatment group and solvent control group is shown in Table 1 and Table 2. [00180] Table 1. Tumor volume of each treatment group (5 mg/kg) 17 [% value (compared with solvent control group) Day 14 after grouping
实验组 肿瘤体积 相对肿瘤体积 T/C  Experimental group Tumor volume Relative tumor volume T/C
P值 P value
( SE) ( SE) (%) (SE) (SE) (%)
第 1组溶剂组 2447±329 1819.4±120.2 ― ― 第 2组达沙替尼 311±55 226.9±30.0 12.4 <0.001 第 3组 DSR-4 39.2±6 29.1±4.8 1.6 <0.001 第 4组 DSR-5 178.6±58 132.8±35.9 7.3 <0.001 第 5组 DSR-6 137±41 101.9±19.3 5.8 <0.001 Group 1 solvent group 2447±329 1819.4±120.2 ― ― Group 2 dasatinib 311±55 226.9±30.0 12.4 <0.001 Group 3 DSR-4 39.2±6 29.1±4.8 1.6 <0.001 Group 4 DSR-5 178.6±58 132.8±35.9 7.3 <0.001 Group 5 DSR-6 137±41 101.9±19.3 5.8 <0.001
[00181】 表 2. 各治疗组 （5mg/kg) 肿瘤体积 17〔％值 （与达沙替尼组比较） 分组后第 21天 [00181] Table 2. Tumor volume (5 mg/kg) in each treatment group 17 [% value (compared to dasatinib group) 21st day after grouping
实验组 肿瘤体积 相对肿瘤体积 T/C  Experimental group Tumor volume Relative tumor volume T/C
P值 P value
( 士 SE) ( 士 SE) (%) 第 2组达沙替尼 391±79 289.1±46.9 ― ― 第 3组 DSR-4 176±59 130±28 45 <0.001 第 4组 DSR-5 203±78 150.3±32 52 <0.001 第 5组 DSR-6 258±89 191±55 66 <0.001 (士SE) (士SE) (%) Group 2 dasatinib 391±79 289.1±46.9 ― ― Group 3 DSR-4 176±59 130±28 45 <0.001 Group 4 DSR-5 203±78 150.3±32 52 <0.001 Group 5 DSR- 6 258±89 191±55 66 <0.001
[00182】 K562人慢性髓系白血病皮下瘤模型由于肿瘤生长迅速， 溶剂对照组 在分组治疗后第 14天因平均肿瘤体积 （2447mm3 ) 超过 2000mm3而结束。 在 分组治疗后第 14天， 阳性药达沙替尼 （5mg/kg) 药效与溶剂对照组在统计学 上有显著性差异（p<0.001 )，平均肿瘤体积为 311mm3，相对肿瘤增殖率（T/C%) 为 12.4%; 受试药物 DSR-4, DSR-5和 DSR-6 ( 5mg/kg)药效与溶剂对照组在统 计学上均有显著性差异（p <0.001 )，平均肿瘤体积分别为 39.2 mm3, 178.6 mm3, 和 137mm3， 相对肿瘤增殖率 （T/C%) 分别为 1.6%，7.3%和 5.8%。 [00182] K562 human chronic myeloid leukemia subcutaneous tumor model in the rapid growth of the tumor, the solvent control group by the mean tumor volume of treatment group (2447mm 3) on day 14 than 2000mm 3 ends. On the 14th day after group treatment, the positive drug dasatinib (5mg/kg) was statistically significantly different from the solvent control group (p<0.001), and the average tumor volume was 311mm 3 , the relative tumor proliferation rate. (T/C%) was 12.4%; the drug effects of the test drugs DSR-4, DSR-5 and DSR-6 (5mg/kg) were statistically significantly different from the solvent control group (p < 0.001). mean tumor volume were 39.2 mm 3, 178.6 mm 3, and 137mm 3, the relative tumor proliferation rate (T / C%) was 1.6%, 7.3% and 5.8%.
[00183】 在分组治疗后第 21天， 达沙替尼 （5mg/kg) 组的平均肿瘤体积为 391mm3, 受试药物 DSR-4, DSR-5和 DSR-6 ( 5mg/kg) 药效与之相比在统计学 上均有显著性差异 （p<0.001 ) ， 平均肿瘤体积分别为 176 mm3， 203mm3和 258mm3, 相对肿瘤增殖率 （T/C%) 分别为 45%， 52%和 66%。 [00183] On day 21 after group therapy, the average tumor volume of the dasatinib (5 mg/kg) group was 391 mm 3 , and the test drugs DSR-4, DSR-5 and DSR-6 (5 mg/kg) were effective. in contrast there were significant differences (p <0.001) statistically, the mean tumor volume were 176 mm 3, 203mm 3 and 258mm 3, the relative tumor proliferation rate (T / C%) were 45%, 52 % and 66%.
[00184】 与溶剂对照组相比， 阳性药物达沙替尼（5mg/kg)、受试药物 DSR-4, DSR-5禾卩 DSR-6 ( 5mg/kg) 的相对肿瘤增殖率 （T/C%) 分别为 12.4%， 1.6%, 7.3%和 5.8%， 这显示了所有化合物均有显著的抗 K562肿瘤生长作用 （p均 <0.001 ) 。 与达沙替尼 （5mg/kg) 相比， 相同剂量的 DSR-4，DSR-5和 DSR-6 ( 5mg/kg) 的抗肿瘤作用更加显著 均<0.001 ) 。 [00184] Relative tumor proliferation rate of the positive drug dasatinib (5 mg/kg), test drug DSR-4, DSR-5 and DSR-6 (5 mg/kg) compared with the solvent control group (T/ C%) were 12.4%, 1.6%, 7.3%, and 5.8%, respectively, indicating that all compounds had significant anti-K562 tumor growth effects (p<0.001). Compared with dasatinib (5 mg/kg), the antitumor effects of the same doses of DSR-4, DSR-5 and DSR-6 (5 mg/kg) were all significantly more <0.001).
[00185】 实施例 9: 不同聚乙二醇达沙替尼结合物对 PC-3人前列腺癌皮下瘤 模型的药效学研究。  [00185] Example 9: Pharmacodynamic study of different polyethylene glycol dasatinib conjugates on a PC-3 human prostate cancer subcutaneous tumor model.
[00186】 实验方法： [00187】 于 Balb/c nude小鼠右侧背部皮下接种 PC-3细胞，建立人前列腺癌异 种移植动物皮下模型。 当平均肿瘤体积达到 160mm3时， 将实验小鼠分组， 每 组 8只， 静脉注射给药， 一周两次。 根据相对肿瘤增殖率 （T/C%)进行疗效评 价。 [00186] Experimental method: [00187] PC-3 cells were subcutaneously inoculated into the right side of Balb/c nude mice to establish a subcutaneous model of human prostate cancer xenograft animals. When the average tumor volume reached 160 mm 3 , the experimental mice were grouped into groups of 8 and administered intravenously twice a week. Efficacy evaluation was performed based on the relative tumor growth rate (T/C%).
[00188】 实验步骤：  [00188] Experimental steps:
[00189】 （1 ) 细胞培养 [00189] (1) Cell culture
[00190】 PC-3细胞系在补充了 10%胎牛血清， L-谷氨酰胺 (2 mM)的 Ham's F12K培养基中，在 37°C，空气中含 5% C02的条件下体外培养肿瘤细胞每周常 规传代 2次。 收集处于指数生长期的肿瘤细胞， 悬于等体积的 PBS : 基质胶混 合物中， 置于冰上， 用于肿瘤接种。 [00190] The PC-3 cell line was cultured in vitro in Ham's F12K medium supplemented with 10% fetal calf serum, L-glutamine (2 mM) at 37 ° C in air containing 5% CO 2 . Tumor cells were routinely passaged twice a week. Tumor cells in the exponential growth phase were collected, suspended in an equal volume of PBS: Matrigel mixture, placed on ice for tumor inoculation.
[00191】 （2) 动物分组 [00191] (2) Animal grouping
[00192】 实验小鼠于右侧背部皮下接种 5x l06 PC-3细胞， 定期观察肿瘤生长 情况， 待肿瘤生长至平均 160mm3时根据肿瘤大小和小鼠体重随机分组并开始 给药治疗。 [00192] Experimental mice were subcutaneously inoculated with 5×10 6 PC-3 cells on the right side of the right side, and tumor growth was observed periodically, and when the tumors were grown to an average of 160 mm 3 , the tumor size and the body weight of the mice were randomly grouped and treatment was started.
[00193】 （3 ) 实验观察  [00193] (3) Experimental observation
[00194] 本研究中根据 Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC)批准的指南进行与操作、 护理、 治疗相关的 所有操作。 在接种后， 每天观察动物的发病与死亡。 在日常观察时， 注意肿瘤 生长对动物正常行为 （如运动、 进食、 饮水、 体重增减、 眼睛、 被毛） 的影响 以及其他任何异常。 记录每组内动物的死亡和临床症状。  [00194] All operations related to operation, care, and treatment were performed in this study in accordance with guidelines approved by the Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC). After vaccination, the morbidity and mortality of the animals were observed daily. During daily observation, pay attention to the effects of tumor growth on normal animal behavior (such as exercise, eating, drinking, weight gain, eyes, coat) and any other abnormalities. The death and clinical symptoms of the animals in each group were recorded.
[00195】 （4) 结果判断 [00195] (4) Result judgment
[00196】 在整个实验过程中，每周测量两次小鼠的体重和肿瘤的大小。肿瘤大 小计算公式： 肿瘤体积 (mm3) =0.5 x (肿瘤长径 X 肿瘤短径 2)。 根据实验组和 对照组肿瘤体积大小， 计算 RTV和 T/C比值。 RTV是指相对肿瘤体积。 T/C 比值是指在结束治疗后某一时间点，治疗组和对照组相对肿瘤体积的百分比值， 反映不同治疗组的抗瘤药效作用。 实验结束时， 按以下两种方式对肿瘤进行照 片记录： 1.每组动物经 co2安乐死后， 荷瘤一面朝上， 按组别分别拍照记录； 2. 肿瘤取下后， 先称量肿瘤重量， 计算 T/C (治疗组和对照组肿瘤重量的百 分比值） ， 然后各组肿瘤按顺序同时摆放并拍照记录。 [00196] The body weight and tumor size of the mice were measured twice a week throughout the experiment. Tumor size calculation formula: tumor volume (mm 3 ) = 0.5 x (tumor long diameter X tumor short diameter 2 ). The RTV and T/C ratios were calculated based on the tumor volume of the experimental and control groups. RTV refers to the relative tumor volume. T/C The ratio refers to the percentage of tumor volume in the treatment group and the control group at a certain point after the end of treatment, reflecting the antitumor efficacy of different treatment groups. At the end of the experiment, the tumors were recorded in the following two ways: 1. After each group of animals was euthanized by co 2 , the tumor-bearing side was facing upwards and photographed separately according to the group; 2. After the tumor was removed, weighed first. Tumor weight, T/C (% of the tumor weight of the treatment group and the control group) was calculated, and then the tumors of each group were simultaneously placed and photographed in order.
[00197】 （5 ) 统计分析 [00197] (5) Statistical analysis
[00198】 所有实验结果以平均瘤体积 ± SE (标准误差） 表示， 统计分析时， 相对肿瘤体积数据使用单因素方差分析 （one-way ANOVA) 及 LSD多重比较 方法对各组间进行相对肿瘤体积显著性差异的比较， p < 0.05为差异显著。  [00198] All experimental results were expressed as mean tumor volume ± SE (standard error). For statistical analysis, relative tumor volume data were analyzed by one-way ANOVA and LSD multiple comparison methods for relative tumor volume between groups. A significant difference was compared, p < 0.05 for the difference.
[00199】 实验结果： [00199] Experimental results:
[00200】 各治疗组和溶剂对照组肿瘤生长情况见表 3和表 4。 [00200] The tumor growth of each treatment group and solvent control group is shown in Tables 3 and 4.
[00201】 表 3. 各治疗组 （10mg/kg) 肿瘤体积 17〔％值 （与溶剂对照组比较） 分组后第 22天 实验组 肿瘤体积 T/C  [00201] Table 3. Tumor volume of each treatment group (10 mg/kg) 17 [% value (compared with solvent control group) Day 22 after grouping Experimental group Tumor volume T/C
相对肿瘤体积（ 士 SE ) P值 ( 士 SE ) (%) 第 1组溶剂组 2462±158 1572.6±133.0 ― ― 第 2组达沙替尼 1752±76 1148.9±112.7 73 0.002 第 3组 DSR-4 1187±67 745.9±54.4 47 <0.001 第 4组 DSR-5 1380±112 894.2±77.8 57 <0.001 第 5组 DSR-6 1439±136 885.8±66.9 56 <0.001  Relative tumor volume (士SE) P value (士SE) (%) Group 1 solvent group 2462±158 1572.6±133.0 ― ― Group 2 dasatinib 1752±76 1148.9±112.7 73 0.002 Group 3 DSR-4 1187±67 745.9±54.4 47 <0.001 Group 4 DSR-5 1380±112 894.2±77.8 57 <0.001 Group 5 DSR-6 1439±136 885.8±66.9 56 <0.001
[00202] 表 4. 各治疗组 （10mg/kg) 肿瘤体积 17〔％值 （与达沙替尼组比较） 实验组 分组后第 25天 肿瘤体积 T/C [00202] Table 4. Tumor volume 17 (% value (compared to dasatinib group) on each of the treatment groups (10 mg/kg) 25 days after the experimental group Tumor volume T/C
相对肿瘤体积 ^士 SE) P值 ( 士 SE) (%) 第 2组达沙替尼 1868±76 1235.3±119.6 - - 第 3组 DSR-4 1200±113 751.9±78.9 61 0.030 第 4组 DSR-5 1406±116 928.6±106.6 75 0.015 第 5组 DSR-6 1449±164 887.4±79.1 72 0.001  Relative tumor volume ^ SE) P value (Shi SE) (%) Group 2 dasatinib 1868 ± 76 1235.3 ± 119.6 - - Group 3 DSR-4 1200 ± 113 751.9 ± 78.9 61 0.030 Group 4 DSR- 5 1406±116 928.6±106.6 75 0.015 Group 5 DSR-6 1449±164 887.4±79.1 72 0.001
[00203】 PC-3人前列腺癌皮下瘤模型肿瘤生长迅速并且导致小鼠体重随肿瘤 负荷增加而降低，溶剂对照组在分组治疗后第 22天因平均肿瘤体积（2462mm3 ) 超过 2000mm3而结束。 在分组治疗后第 22天， 阳性药达沙替尼 （10mg/kg)药 效与溶剂对照组在统计学上有显著性差异 (p<0.01 )，平均肿瘤体积为 1752mm3， 相对肿瘤增殖率（T/C%)为 73%;受试药物 DSR-4, DSR-5和 DSR-6 ( 10mg/kg) 药效与溶剂对照组在统计学上均有显著性差异 （p <0.001 ) ， 平均肿瘤体积分 别为 1187 mm3, 1380 mm3禾口 1439mm3，相对肿瘤增殖率 ( T/C%)分别为 47%， 57%和 56%。 [00203] PC-3 human prostate carcinoma subcutaneous tumor model in mice led to rapid tumor growth, and weight decrease with increasing tumor burden, solvent control group after day 22 due to the treatment group mean tumor volume (2462mm 3) ends than 2000mm 3 . On the 22nd day after group treatment, the drug efficacy of the positive drug dasatinib (10mg/kg) was statistically significantly different from the solvent control group (p<0.01), and the average tumor volume was 1752mm 3 , the relative tumor proliferation rate. (T/C%) was 73%; the drug effects of the test drugs DSR-4, DSR-5 and DSR-6 (10 mg/kg) were statistically significantly different from the solvent control group (p < 0.001). The average tumor volumes were 1187 mm 3, 1380 mm 3 Hekou 1439mm 3, the relative tumor proliferation rate (T / C%) were 47%, 57% and 56%.
[00204】 在分组治疗后第 25天， 达沙替尼 （10mg/kg) 组的平均肿瘤体积为 1868mm3, 受试药物 DSR-4, DSR-5和 DSR-6 ( 10mg/kg) 药效与之相比在统 计学上均有显著性差异（p<0.05，p<0.05和 p<0.01 )，平均肿瘤体积分别为 1200 mm3, 1406mm3和 1449mm3，相对肿瘤增殖率（T/C%)分别为 61%， 75%和 72%。 [00204] On day 25 after group therapy, the average tumor volume of the dasatinib (10 mg/kg) group was 1868 mm 3 , and the test drugs DSR-4, DSR-5 and DSR-6 (10 mg/kg) were effective. were compared statistically significant difference (p <0.05, p <0.05 and p <0.01), mean tumor volumes were 1200 mm 3 1406mm 3 and 1449mm 3, relative tumor proliferation rate, (T / C %) are 61%, 75% and 72% respectively.
[00205】 与溶剂对照相比， 阳性药物达沙替尼 （10mg/kg) 、 受试药物 DSR-4, DSR-5和 DSR-6 ( 10mg/kg)的相对肿瘤增殖率（T/C%)分别为 73%， 47%,57% 和 56%， 这显示了所有化合物均有统计学显著的抗 PC-3肿瘤生长作用 （p均 <0.01 ) 。 与达沙替尼 ( 10mg/kg) 相比， 相同剂量的 DSR-4，DSR-5和 DSR-6 ( 10mg/kg) 的抗肿瘤作用更加显著 （p<0.05 ) 。 [00206】 实施例 -有关雷帕霉素的结合物 [00205] Relative tumor growth rate (T/C%) of the positive drug dasatinib (10 mg/kg), test drugs DSR-4, DSR-5 and DSR-6 (10 mg/kg) compared to the solvent control ) were 73%, 47%, 57%, and 56%, respectively, indicating that all compounds had statistically significant anti-PC-3 tumor growth effects (p<0.01). The antitumor effects of the same doses of DSR-4, DSR-5 and DSR-6 (10 mg/kg) were more significant (p < 0.05) compared with dasatinib (10 mg/kg). [00206] Example - conjugate of rapamycin
[00207】 实施例中所用的雷帕霉素从武汉远成共创科技有限公司购得，溴乙酸 叔丁酯、 三苯基磷从国药集团化学试剂有限公司购得。  [00207] The rapamycin used in the examples was purchased from Wuhan Yuancheng Co-creation Technology Co., Ltd., and t-butyl bromoacetate and triphenylphosphine were purchased from Sinopharm Chemical Reagent Co., Ltd.
[00208】 实施例 10: 单甲氧基聚乙二醇乙酸甘氨酸（数均分子量 20000)—雷 帕霉素结合物 （LPR-1 ) 的制备  Example 10: Preparation of monomethoxypolyethylene glycol glycine acetate (number average molecular weight 20000) - rapamycin conjugate (LPR-1)
Figure imgf000049_0001
Figure imgf000049_0001
LPR-1  LPR-1
[00209】 将溴乙酸叔丁酯 （5.82g，30 mmol)加到反应瓶中， 用丙酮 （80 mL) 溶解， 再加入叠氮化钠 （4.55g，70 mmol) 溶于水 （40 mL) 的溶液， 加热回流 过夜。 反应液蒸去丙酮， 残分用乙醚提取， 提取液用饱和盐水洗涤， 干燥， 减 压浓缩后得油状液体。 将此液体用甲醇 （90 mL) 溶解， 再加入 1 N氢氧化钠 溶液（90 mL)， 搅拌， 加热回流 3 h。 冷却后减压蒸去甲醇， 残夜用冰浴冷却， 加 6 N 盐酸将 PH调至 2， 再用乙醚提取， 提取液加水洗涤， 干燥， 浓缩后得 叠氮乙酸 （化合物 18)， MS m/z: 124 [M+Na]+。 [00209] tert-Butyl bromoacetate (5.82 g, 30 mmol) was added to a reaction flask, dissolved in acetone (80 mL), then sodium azide (4.55 g, 70 mmol) dissolved in water (40 mL) The solution was heated to reflux overnight. The reaction mixture was evaporated to dryness. EtOAc was evaporated. This liquid was dissolved in methanol (90 mL), then 1N sodium hydroxide solution (90 mL) was added, stirred and heated to reflux for 3 h. After cooling, the methanol was evaporated under reduced pressure, and the mixture was cooled with ice-cooled overnight. The pH was adjusted to 2 with 6 N hydrochloric acid, and then extracted with diethyl ether. The extract was washed with water, dried and concentrated to give azide acetic acid (compound 18), MS m /z: 124 [M+Na] + .
[00210】 将叠氮乙酸 （化合物 18) (253 mg， 2.5 mmoL)和雷帕霉素（2.28g， 2.5 mmoL)加到反应瓶中， 用二氯甲垸溶解， 冰浴冷却， 再加入 4一二甲氨基吡啶 (DMAP, 611 mg, 5 mmoL) 禾卩 N，N-二环己基碳酰亚胺 （DCC, 1.03g, 5 mmoL) 加到反应瓶中， 加完后继续在室温下搅拌过夜。 反应液浓缩后残分用柱层析纯 化， 得叠氮乙酸雷帕霉素酯 （化合物 19)1.42g， 收率 57%， MS m/z: 1020 [00210] Azidoacetic acid (Compound 18) (253 mg, 2.5 mmoL) and rapamycin (2.28 g, 2.5) mmoL) was added to the reaction flask, dissolved in dichloromethane, cooled in an ice bath, and then added with 4-dimethylaminopyridine (DMAP, 611 mg, 5 mmoL) and N,N-dicyclohexylcarbimide ( DCC, 1.03 g, 5 mmoL) was added to the reaction flask and stirring was continued at room temperature overnight. After the reaction mixture was concentrated, the residue was purified by column chromatography to give the titled product of the the the the the the the the the the the the the the the the the the the the
[M+Na]+。 [M+Na]+.
[00211】 将叠氮乙酸雷帕霉素酯 （化合物 19) (0.7 g, 0.7 mmoL) 和三苯基磷 (0.37g, 1.4 mmoL) 加到反应瓶中， 再加入四氢呋喃和水的混合液 （5:1, 180 mL) ， 加热到 50°C反应过夜， 反应液浓缩后残分用乙酸乙酯提取， 提取液用 饱和盐水洗涤，干燥。减压浓缩后残分用柱层析纯化，得甘氨酸雷帕霉素酯 （化 合物 20) 0 .48 g， 收率 70%， MS m/z: 994 [M+Na]+。  [00211] Azoxyacetic acid rapamycin ester (Compound 19) (0.7 g, 0.7 mmoL) and triphenylphosphine (0.37 g, 1.4 mmoL) were added to the reaction flask, followed by a mixture of tetrahydrofuran and water ( 5:1, 180 mL), heated to 50 ° C overnight, and the residue was concentrated with ethyl acetate. The extract was washed with saturated brine and dried. After concentration under reduced pressure, the residue was purified by column chromatography to yield EtOAc (yield: 20), yield: 70%, MS m/z: 994 [M+Na]+.
[00212】 将单氧基聚乙二醇乙酸（20K， 1 g， 0.05mmoL)，甘氨酸雷帕霉素酯 (化 合物 20) (97mg, 0.1 mmoL)， 1一羟基苯并三氮唑 (HOBt, 6.8 mg, 0.05 mmoL) 禾口 DMAP ( 12.2 mg, 0.1 mmoL) 加到反应瓶中， 用二氯甲垸溶解， 冰浴冷却， 再滴入 DCC ( 15.5 mg, 0.075 mmoL)溶于二氯甲垸的溶液， 滴完后自然升到室 温， 反应过夜， 次日将反应液浓缩， 残分用异丙醇结晶得单甲氧基聚乙二醇乙 酸甘氨酸 （数均分子量 20000) —雷帕霉素结合物 （LPR-1 ) 0.82 go  [00212] Monooxypolyethylene glycol acetic acid (20K, 1 g, 0.05 mmoL), glycine rapamycin ester (Compound 20) (97 mg, 0.1 mmoL), 1-hydroxybenzotriazole (HOBt, 6.8 mg, 0.05 mmoL) and DMAP (12.20 mg, 0.1 mmoL) were added to the reaction flask, dissolved in dichloromethane, cooled in an ice bath, and added to DCC (15. 5 mg, 0.075 mmoL) in dichloromethane. The solution is naturally raised to room temperature after the completion of the dropwise addition, and the reaction is allowed to stand overnight. The reaction solution is concentrated the next day, and the residue is crystallized from isopropanol to obtain monomethoxypolyethylene glycol glycine acetate (number average molecular weight 20000) - rapamycin. Conjugate (LPR-1) 0.82 go
[00213] 1H-NMR (300MHz, CDC13): 0.90 (Me, 3H), 0.92 (Me, 3H), 0.94 (Me, 3H), 0.96 (Me, 3H), 0.97 (Me, 3H), 1.10 (CH2， 2H), 1.11 (CH2， 2H), 1.20 (CH2， 2H) 1.33 (CH2， 2H), 1.37 (CH， 1H)， 1.45 (CH2， 2H), 1.47 (CH2， 2H), 1.60 (CH2, 2H), 1.61 (CH2， 2H), 1.65 (CH2， 2H), 1.65 (CH2， 2H), 1.74 (Me, 3H), 1.75 (CH， 1H)， 2.07 (CH， 4H)， 2.08 (CH2， 2H), 3.14 (Me, 3H), 3.33 (CH， 1H), 3.36 (Me, 3H), 3.37 (CH2， 2H), 3.42 (CH， 1H)， 3.44 (Me, 3H), 3.56 (CH， 1H)， 3.64 (CH2， 1800H), 3.71 (CH， 1H)， 3.72 (CH， 1H)， 3.86 (CH， 1H)， 4.17 (CH2， 2H), 4.19 (CH， 1H)， 5.16 (CH， 1H), 5.17 (CH， 1H), 5.29(=CH, 1H), 5.39(=CH, 1H), 5.95(=CH, 1H), 6.13(=CH, 1H), 6.31(=CH, 1H)， 6.38(=CH, 1H)， 8.34 (CH， 1H). [00213] 1H-NMR (300MHz, CDC1 3 ): 0.90 (Me, 3H), 0.92 (Me, 3H), 0.94 (Me, 3H), 0.96 (Me, 3H), 0.97 (Me, 3H), 1.10 ( CH 2 , 2H), 1.11 (CH 2 , 2H), 1.20 (CH 2 , 2H) 1.33 (CH 2 , 2H), 1.37 (CH, 1H), 1.45 (CH 2 , 2H), 1.47 (CH 2 , 2H ), 1.60 (CH 2 , 2H), 1.61 (CH 2 , 2H), 1.65 (CH 2 , 2H), 1.65 (CH 2 , 2H), 1.74 (Me, 3H), 1.75 (CH, 1H), 2.07 ( CH, 4H), 2.08 (CH 2 , 2H), 3.14 (Me, 3H), 3.33 (CH, 1H), 3.36 (Me, 3H), 3.37 (CH 2 , 2H), 3.42 (CH, 1H), 3.44 (Me, 3H), 3.56 (CH, 1H), 3.64 (CH 2 , 1800H), 3.71 (CH, 1H), 3.72 (CH, 1H), 3.86 (CH, 1H), 4.17 (CH 2 , 2H), 4.19 (CH, 1H), 5.16 (CH, 1H), 5.17 (CH, 1H), 5.29 (=CH, 1H), 5.39 (=CH, 1H), 5.95 (=CH, 1H), 6.13 (=CH, 1H), 6.31 (=CH, 1H), 6.38 (=CH, 1H), 8.34 (CH, 1H).
[00214】 实施例 1 1 : 单甲氧基聚乙二醇谷氨酸二肽甘氨酸 （数均分子量  Example 1 1 : Monomethoxypolyethylene glycol glutamic acid dipeptide glycine (number average molecular weight)
一雷帕霉素结合物 （LPR-2) 的制备  Preparation of a rapamycin conjugate (LPR-2)
Figure imgf000051_0001
Figure imgf000051_0001
[00215】 根据实施例 1所述的方法制备单甲氧基聚乙二醇谷氨酸二肽酸 （化 合物 5)。 Monomethoxypolyethylene glycol glutamic acid dipeptide acid (Compound 5) was prepared according to the method described in Example 1.
[00216】 将单甲氧基聚乙二醇谷氨酸二肽酸 （化合物 5) (20K，0.5g，  [00216] Monomethoxy polyethylene glycol glutamic acid dipeptide acid (Compound 5) (20K, 0.5g,
0.025mmol)，甘氨酸雷帕霉素酯 （化合物 20) 48.6 mg (0.05mmol)， HOBt (3.4 mg: 0.025mmoL)禾 B DMAP 6.1 mgC0.05mmoL) 加到反应瓶中，用二氯甲垸溶解，冰 浴冷却， 再滴入 DCC 15.5 mg (0.075mmol) 溶于二氯甲垸的溶液， 滴完后自然 升到室温， 反应过夜。 次日将反应液浓缩， 残分用异丙醇结晶， 得单甲氧基聚 乙二醇 （20K) 谷氨酸二肽甘氨酸-雷帕霉素结合物 (LPR-2) 0.41g。 0.025 mmol), glycine rapamycin ester (Compound 20) 48.6 mg (0.05 mmol), HOBt (3.4 mg : 0.025 mmoL) and B DMAP 6.1 mgC 0.05 mmoL) were added to the reaction flask and dissolved with dichloromethane. After cooling in an ice bath, a solution of 15.5 mg (0.075 mmol) of DCC in DCC was added dropwise, and after the completion of the dropwise addition, the mixture was allowed to warm to room temperature overnight. The reaction solution was concentrated the next day, and the residue was crystallized from isopropanol to obtain monomethoxypoly. Ethylene glycol (20K) glutamic acid dipeptide glycine-rapamycin conjugate (LPR-2) 0.41 g.
[00217] 1H-NMR (300MHz, CDC13): 0.90 (Me, 9H)， 0.92 (Me, 9H)， 0.94 (Me, 9H )， 0.96 (Me, 9H)， 0.97 (Me, 9H)， 1.10 (CH2， 6H)， 1.11 (CH2， 6H)， 1.20 (CH2， 6H)， 1.33 (CH2， 6H)， 1.37 (CH， 3H), 1.45 (CH2， 6H)， 1.47 (CH2， 6H)， 1.60 (CH2， 6H)， 1.61 (CH2， 6H)， 1.65 (CH2， 6H)， 1.65 (CH2， 6H)， 1.74 (Me, 9H)， 1.75 (CH， 3H) 2.07 (CH， 12H), 2.08 (CH2， 6H)， 3.14 (Me, 9H)， 3.33 (CH， 3H), 3.36 (Me, 9H)， 3.37 (CH2， 6H)， 3.42 (CH， 3H), 3.44 (Me, 9H)， 3.56 (CH， 3H), 3.64 (CH2， 1800H), 3.71 (CH， 3H), 3.72 (CH， 3H), 3.86 (CH， 3H), 4.17 (CH2， 6H), 4.19 (CH， 3H), 5.16 (CH， 3H), 5.17 (CH， 3H), 5.29(=CH, 3H), 5.39(=CH, 3H), 5.95(=CH, 3H), 6.13(=CH, 3H), 6.31(=CH, 3H), 6.38(=CH, 3H), 8.34 (CH， 3H). [00217] 1H-NMR (300MHz, CDC1 3 ): 0.90 (Me, 9H), 0.92 (Me, 9H), 0.94 (Me, 9H), 0.96 (Me, 9H), 0.97 (Me, 9H), 1.10 ( CH 2 , 6H), 1.11 (CH 2 , 6H), 1.20 (CH 2 , 6H), 1.33 (CH 2 , 6H), 1.37 (CH, 3H), 1.45 (CH 2 , 6H), 1.47 (CH 2 , 6H), 1.60 (CH 2 , 6H), 1.61 (CH 2 , 6H), 1.65 (CH 2 , 6H), 1.65 (CH 2 , 6H), 1.74 (Me, 9H), 1.75 (CH, 3H) 2.07 ( CH, 12H), 2.08 (CH 2 , 6H), 3.14 (Me, 9H), 3.33 (CH, 3H), 3.36 (Me, 9H), 3.37 (CH 2 , 6H), 3.42 (CH, 3H), 3.44 (Me, 9H), 3.56 (CH, 3H), 3.64 (CH 2 , 1800H), 3.71 (CH, 3H), 3.72 (CH, 3H), 3.86 (CH, 3H), 4.17 (CH 2 , 6H), 4.19 (CH, 3H), 5.16 (CH, 3H), 5.17 (CH, 3H), 5.29 (=CH, 3H), 5.39 (=CH, 3H), 5.95 (=CH, 3H), 6.13 (=CH, 3H), 6.31 (=CH, 3H), 6.38 (=CH, 3H), 8.34 (CH, 3H).
[00218】 实施例 12: Y型聚乙二醇谷氨酸二肽甘氨酸 （数均分子量 40000) 一雷帕霉素结合物 （LPR-3) 的制备 Example 12: Y-type polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight 40000) Preparation of a rapamycin conjugate (LPR-3)
Figure imgf000053_0001
Figure imgf000053_0001
[00219】 根据上述实施例所述制备 Y型聚乙二醇谷氨酸二肽酸 （化合物 13)。 Y-type polyethylene glycol glutamic acid dipeptide acid (Compound 13) was prepared as described in the above examples.
[00220】 将 Y型聚乙二醇谷氨酸二肽酸 （化合物 13)(40K，0.5g，0.0125mmol)， 甘氨酸雷帕霉素酯 （化合物 20) 24.3 mg (0.025mmol)， HOBt (1.7 mg， 0.0125mmo) 和 DMAP 3 mg(0.025mmol) 加到反应瓶中，用二氯甲垸溶解，冰浴冷却，再滴入 DCC 4.1 mg (0.02mmol) 溶于二氯甲垸的溶液， 滴完后自然升到室温， 反应过 夜。 次日将反应液浓缩， 残分用异丙醇结晶， 得 Y型聚乙二醇谷氨酸二肽甘氨 酸 （数均分子量 40000) —雷帕霉素结合物 (LPR-3) 0.44g。 Y-type polyethylene glycol glutamic acid dipeptide acid (Compound 13) (40K, 0.5g, 0.0125mmol), glycine rapamycin ester (Compound 20) 24.3 mg (0.025mmol), HOBt (1.7 Mg, 0.0125mmo) and DMAP 3 mg (0.025mmol) were added to the reaction flask, dissolved in dichloromethane, cooled in ice bath, and then added dropwise to DCC 4.1 mg (0.02 mmol) in dichloromethane solution. After that, it naturally rose to room temperature and reacted overnight. The reaction solution was concentrated the next day, and the residue was crystallized from isopropanol to obtain Y-type polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight: 40000) - rapamycin conjugate (LPR-3) 0.44 g.
[00221] 1H-NMR (300MHz, CDC13): 0.90 (Me, 9H)， 0.92 (Me, 9H)， 0.94 (Me, 9H)， 0.96 (Me, 9H)， 0.97 (Me, 9H)， 1.10 (CH2， 6H)， 1.11 (CH2， 6H)， 1.20 (CH2， 6H)， 1.33 (CH2， 6H)， 1.37 (CH， 3H), 1.45 (CH2， 6H)， 1.47 (CH2， 6H)， 1.60 (CH2， 6H)， 1.61 (CH2， 6H)， 1.65 (CH2， 6H)， 1.65 (CH2， 6H)， 1.74 (Me, 9H)， 1.75 (CH， 3H), 2.07 (CH， 12H), 2.08 (CH2， 6H)， 3.14 (Me, 9H)， 3.33 (CH， 3H), 3.36 (Me, 9H)， 3.37 (CH2， 6H)， 3.42 (CH， 3H), 3.44 (Me, 9H)， 3.56 (CH， 3H), 3.64 (CH2， 1800H), 3.71 (CH， 3H), 3.72 (CH， 3H), 3.86 (CH， 3H), 4.17 (CH2， 6H), 4.19 (CH， 3H), 5.16 (CH， 3H), 5.17 (CH， 3H), 5.29(=CH, 3H), 5.39(=CH, 3H), 5.95(=CH, 3H), 6.13(=CH, 3H), 6.31(=CH, 3H), 6.38(=CH, 3H), 8.34 (CH， 3H). [00221] 1H-NMR (300MHz, CDC1 3 ): 0.90 (Me, 9H), 0.92 (Me, 9H), 0.94 (Me, 9H), 0.96 (Me, 9H), 0.97 (Me, 9H), 1.10 ( CH 2 , 6H), 1.11 (CH 2 , 6H), 1.20 (CH 2 , 6H), 1.33 (CH 2 , 6H), 1.37 (CH, 3H), 1.45 (CH 2 , 6H), 1.47 (CH 2 , 6H), 1.60 (CH 2 , 6H), 1.61 (CH 2 , 6H), 1.65 (CH 2 , 6H), 1.65 (CH 2 , 6H), 1.74 (Me, 9H), 1.75 (CH, 3H), 2.07 (CH, 12H), 2.08 (CH 2 , 6H), 3.14 (Me, 9H), 3.33 (CH, 3H), 3.36 (Me, 9H), 3.37 (CH 2 , 6H), 3.42 (CH, 3H) , 3.44 (Me, 9H), 3.56 (CH, 3H), 3.64 (CH 2 , 1800H), 3.71 (CH, 3H), 3.72 (CH, 3H), 3.86 (CH, 3H), 4.17 (CH 2 , 6H ), 4.19 (CH, 3H), 5.16 (CH, 3H), 5.17 (CH, 3H), 5.29 (=CH, 3H), 5.39 (=CH, 3H), 5.95 (=CH, 3H), 6.13 (= CH, 3H), 6.31 (=CH, 3H), 6.38 (=CH, 3H), 8.34 (CH, 3H).
[00222】 实施例 13 不同聚乙二醇雷帕霉素结合物对肿瘤细胞的抑制活性 [00223】 （1 ) 实验方法与步骤 [00224】 （a) 细胞培养 Example 13 Inhibitory Activity of Different Polyethylene Glycol Rapamycin Conjugates on Tumor Cells [00223] (1) Experimental Methods and Procedures [00224] (a) Cell Culture
[00225】 Plc/prf/5细胞体外单层培养， 培养条件为 MEM培养基中加 10%热 灭活胎牛血清， 于 37°C、 含 5%CO2空气的培养箱中培养。 一周两次用胰酶 -EDTA进行消化处理传代。 当细胞呈指数生长期时， 收取细胞， 计数， 接种。 [00225] Plc/prf/5 cells were cultured in vitro in a single layer in a MEM medium supplemented with 10% heat-inactivated fetal bovine serum and cultured in an incubator containing 5% CO2 air at 37 °C. Digestion was performed twice with trypsin-EDTA twice a week. When cells are exponentially growing, cells are harvested, counted, and vaccinated.
[00226】 （b) 肿瘤细胞接种、 分组及给药 [00226] (b) Tumor cell inoculation, grouping and administration
[00227】 将 1 χ107 plc/prf/5肿瘤细胞悬浮于 0.1 ml 混合液（ PBS: Matrigel 为 4:1 )， 接种于每只 NOD/SCID小鼠背部右侧肩胛处。接种 24天后肿瘤平均体 积达到约 350mm3时， 剔除瘤体积过小或过大的小鼠， 余下小鼠根据瘤体积随 机分组并开始给药。 [00228】 （C) 实验方案 表 5实验动物分组及给药方案 [00227] 1 χ10 7 plc/prf/5 tumor cells were suspended in 0.1 ml of a mixture (PBS: Matrigel 4:1) and inoculated into the right scapula of each NOD/SCID mouse. When the average tumor volume reached about 350 mm 3 24 days after the inoculation, the mice whose tumor volume was too small or too large were knocked out, and the remaining mice were randomly grouped according to the tumor volume and started to be administered. [00228] (C) Experimental protocol Table 5 Experimental animal grouping and dosing schedule
剂量 a 给药容积 给药途  Dosage a dose volume
组别 N 化合物治疗 给药方案  Group N compound therapy dosing regimen
(mg/kg) (μΐ/s) 径  (mg/kg) (μΐ/s) diameter
1 5 溶媒对照 ― 10 i.v. 02W 2W 1 5 Solvent Control ― 10 i.v. 02W 2W
2 5 LPR-1 10mg/kg 10 i.v. Q2W 2W2 5 LPR-1 10mg/kg 10 i.v. Q2W 2W
3 5 LPR-2 10mg/kg 10 i.v. Q2W 2W3 5 LPR-2 10mg/kg 10 i.v. Q2W 2W
4 5 LPR-3 10mg/kg 10 i.v. Q2W 2W 4 5 LPR-3 10mg/kg 10 i.v. Q2W 2W
[00229] (2) 实验结果 [00230】 （a) 体重 [00229] (2) Experimental results [00230] (a) Weight
[00231】 各处理组荷瘤鼠的体重变化如表 6和图 2所示。 表 6.各处理组不同时间点的体重  [00231] The body weight changes of the tumor-bearing mice in each treatment group are shown in Table 6 and Figure 2. Table 6. Body weight at different time points in each treatment group
动物体重 (g)  Animal weight (g)
接种天数 LPR-1 LPR-2 LPR-3  Inoculation days LPR-1 LPR-2 LPR-3
溶媒对照  Solvent control
10 mg/kg 10 mg/kg 10 mg/kg 10 mg/kg 10 mg/kg 10 mg/kg
24 17-6 0-5 19-2 0-7 19-6 1 -5 19-2 0-724 17-6 0-5 19-2 0-7 19-6 1 -5 19-2 0-7
27 18.3 0.3 19.5 ± 0.8 18.6 1.2 18.0 0.427 18.3 0.3 19.5 ± 0.8 18.6 1.2 18.0 0.4
31 17.4 ± 0.1 18.4 0.6 18.0 1.2 17.7 ± 0.431 17.4 ± 0.1 18.4 0.6 18.0 1.2 17.7 ± 0.4
34 18.0 0.2 19.1 ± 0.8 18.5 1.2 18.2 0.734 18.0 0.2 19.1 ± 0.8 18.5 1.2 18.2 0.7
38 18.1 0.2 18.8 0.5 18.1 1.2 17.8 ± 0.7 注: a. 平均值 ±标准误差 [00232】 （b) 瘤体积 38 18.1 0.2 18.8 0.5 18.1 1.2 17.8 ± 0.7 Note: a. Average ± standard error [00232] (b) Tumor volume
[00233】 各处理组肿瘤体积变化如表 7和图 3所示。 表 7.各处理组不同时间点的瘤体积  [00233] The tumor volume changes of each treatment group are shown in Table 7 and Figure 3. Table 7. Tumor volume at different time points in each treatment group
肿瘤体积 (mm3)1 Tumor volume (mm 3 ) 1
接种天数  Inoculation days
LPR-1 LPR-2 LPR-3 溶媒对照  LPR-1 LPR-2 LPR-3 vehicle control
10 mg/kg 10 mg/kg 10 mg/kg 10 mg/kg 10 mg/kg 10 mg/kg
24 355 64 352 49 353 ± 54 358 ± 6224 355 64 352 49 353 ± 54 358 ± 62
27 763 ± 102 598 80 404 89 455 8527 763 ± 102 598 80 404 89 455 85
31 1048 ± 104 670 ± 74 391 ± 72 569 ± 10031 1048 ± 104 670 ± 74 391 ± 72 569 ± 100
34 1439 ± 130 738 74 456 77 754 ± 15134 1439 ± 130 738 74 456 77 754 ± 151
38 1801 162 919 78 536 77 848 ± 178 注: a. 平均值 ±标准误差 38 1801 162 919 78 536 77 848 ± 178 Note: a. Average ± standard error
[00234】 （C ) 抗肿瘤药效评价指标 [00234] (C) Antitumor efficacy evaluation index
[00235】 LPR1 ,2,3对 plc/prf/5皮下移植瘤模型的抑瘤药效评价指标如表 8 各处理组的抑瘤药效评价  [00235] LPR1, 2, 3 anti-tumor efficacy evaluation indexes of plc/prf/5 subcutaneous xenograft model are shown in Table 8. Evaluation of anti-tumor effect of each treatment group
肿瘤抑制率 τ/c 肿瘤生长延迟天数  Tumor inhibition rate τ/c tumor growth delay days
组别 瘤体积（mm3)a P值 Group tumor volume (mm 3 ) a P value
(%) (至 1000 mm3) 溶剂对照 1801 ± 162 (%) (to 1000 mm 3 ) Solvent control 1801 ± 162
LPR-1 919 ± 78 51 7 0.000 LPR-1 919 ± 78 51 7 0.000
LPR-2 536 ± 77 >7 0.000LPR-2 536 ± 77 >7 0.000
LPR-3 848 ± 178 >7 0.000 注: a. 平均值 ±标准误差 LPR-3 848 ± 178 >7 0.000 Note: a. Average ± standard error
[00236】 （3) 实验结果总结及讨论  [00236] (3) Summary and discussion of experimental results
[00237】 在本实验中，我们评价了 LPR-1， LPR-2和 LPR-3在人肝癌 plc/prf/5 皮下移植瘤模型中的体内药效。 各处理组在不同时间的瘤体积如表 2， 图 2所 示。 Plc/prf/5肿瘤细胞接种 NOD/SCID小鼠后 38天， 溶媒对照组瘤体积达到 1 801 mm3 o 受试物 LPR-1， LPR-2和 LPR-3都表现出一定的抑瘤作用， 其中 LPR-2抑瘤作用最为明显， 与溶媒组相比 T/C小于 40%， p值为 0.000， 具有 显著性差异。  [00237] In this experiment, we evaluated the in vivo efficacy of LPR-1, LPR-2 and LPR-3 in a human liver cancer plc/prf/5 subcutaneous xenograft model. The tumor volume of each treatment group at different times is shown in Table 2 and Figure 2. 38 days after Plc/prf/5 tumor cells were inoculated with NOD/SCID mice, the tumor control volume reached 1 801 mm3 o. The test substances LPR-1, LPR-2 and LPR-3 all showed some anti-tumor effects. Among them, LPR-2 has the most obvious anti-tumor effect. Compared with the vehicle group, T/C is less than 40%, and the p value is 0.000, which has significant difference.
[00238】 各组荷瘤鼠的体重变化影响如表 6、 图 2所示。各给药组在实验过程 中未观察到明显的毒性反应。  [00238] The effects of body weight change of each group of tumor-bearing mice are shown in Table 6 and Figure 2. No significant toxicity was observed in each of the drug-administered groups during the experiment.
[00239】 综上所述， 在本实验中， 受试药 LPR-1 , LPR-2和 LPR-3对人肝癌 plc/prf/5皮下移植瘤模型表现出抗肿瘤效果，其中以 LPR-2抑瘤作用最为明显， 各给药组实验过程中未观察到明显的毒性反应。 LPR-1和 LPR-2使用相同结构 和相同数均分子量的聚乙二醇， 所不同的是， LPR-1 的聚乙二醇仅通过甘氨酸 与雷帕霉素键合，聚乙二醇分子的一个端基只键合一个雷帕霉素分子;而 LPR-2 的聚乙二醇通过谷氨酸二肽和甘氨酸与雷帕霉素键合， 聚乙二醇分子的一个端 基可键合三个雷帕霉素分子。 LPR-2的药物负载率是 LPR-1 的 3倍，其抗肿瘤 活性也明显高于 LPR-1。  [00239] In summary, in this experiment, the test drugs LPR-1, LPR-2 and LPR-3 showed anti-tumor effects on the human liver cancer plc/prf/5 subcutaneous xenograft model, among which LPR-2 The anti-tumor effect was the most obvious, and no obvious toxicity was observed during the experiment. LPR-1 and LPR-2 use polyethylene glycol of the same structure and the same number average molecular weight, except that polyethylene glycol of LPR-1 is only bonded to rapamycin via glycine, polyethylene glycol molecule One end group is only bonded to one rapamycin molecule; and the polyethylene glycol of LPR-2 is bonded to rapamycin through glutamic acid dipeptide and glycine, and one terminal group of the polyethylene glycol molecule can be bonded Combine three rapamycin molecules. The drug loading rate of LPR-2 is three times that of LPR-1, and its antitumor activity is also significantly higher than that of LPR-1.
[00240】 实施例 -有关伊诺替康的结合物 Example - Combination of Inocomcan
[00241】 实施例中所用的盐酸伊诺替康从上海龙翔生物医药开发有限公司购 得。 [00242] 实施例 14:单甲氧基聚乙二醇谷氨酸二肽甘氨酸 (数均分子量 20000)[00241] The irinotecan hydrochloride used in the examples was purchased from Shanghai Longxiang Biomedical Development Co., Ltd. Example 14: monomethoxy polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight 20000)
-伊诺替康结合物 （YNR-1) 的制备 - Preparation of INOT for the conjugate (YNR-1)
Figure imgf000057_0001
Figure imgf000057_0001
[00243】 盐酸依诺替康 13.3g和 N-叔丁氧羰基甘氨酸 20.1g， 溶解在 120mL 无水二氯甲垸中， 加入二环己基碳二亚胺 （DCC) 18.8g和 4-二甲基氨基吡啶 (DMAP) 7.4g， 室温下搅拌过夜。 过滤除去反应产生的固体， 减压浓缩溶液， 加入 lOOmL石油醚。 过滤收集沉淀真空干燥， 制得 N-叔丁氧羰基甘氨酸伊诺 替康酯 （化合物 21) 23g。 [00243] 13.3g of irinotecan hydrochloride and 20.1g of N-tert-butoxycarbonylglycine were dissolved in 120mL of anhydrous dichloromethane, and added with dicyclohexylcarbodiimide (DCC) 18.8g and 4-dimethyl The aminopyridine (DMAP) 7.4 g was stirred at room temperature overnight. The solid produced by the reaction was removed by filtration, and the solution was concentrated under reduced pressure, and then 100 mL of petroleum ether was added. The precipitate was collected by filtration and dried under vacuum to give 23 g of N-tert-butoxycarbonylglycine inocate (Compound 21).
[00244】 N-叔丁氧羰基甘氨酸伊立替康酯 （化合物 21) 23g溶解在 lOOmL二氯 甲垸中，添加 30mL三氟乙酸，室温下搅拌 5小时。减压浓缩溶液，加入 500mL 乙醚。 过滤， 收集沉淀， 真空干燥， 制得甘氨酸依诺替康酯 （化合物 22) 20g。  [00244] 23 g of N-tert-butoxycarbonylglycine irinotecan (Compound 21) was dissolved in 100 mL of dichloromethane, and 30 mL of trifluoroacetic acid was added thereto, followed by stirring at room temperature for 5 hours. The solution was concentrated under reduced pressure and 500 mL diethyl ether was added. Filtration, collection of the precipitate, and drying in vacuo gave 20 g of enonotene glycine (Compound 22).
[00245】 根据实施例 1所述方法制备单甲氧基聚乙二醇谷氨酸二肽酸 (化合物 5)。 单甲氧基聚乙二醇谷氨酸二肽酸 (化合物 5) (分子量为 20000) 5.0g、 甘氨酸 依诺替康酯 （化合物 22) 1.02g、 N-羟基琥珀酰亚胺 （NHS) 115mg、 4-二甲基氨 基吡啶 （DMAP) 153mg溶于 50mL无水二氯甲垸中， 氮气保护下， 再添加二 环己基碳二亚胺 （DCC ) 309mg。 室温过夜搅拌反应。 过滤除去固体物质， 多 余溶剂旋转蒸发除去， 残余物添加 lOOmL异丙醇 （IPA) ， 过滤， 产品经真空 干燥， 得到单甲氧基聚乙二醇谷氨酸二肽甘氨酸 （数均分子量 20000) —伊诺 替康结合物 （YNR-l) 4.4g。 iH-NMR MSO-c^): 0.84-0.89(m, 8H)， 1.24(m, 9H)， 1.40-1.50(m, 21H), 1.79(m, 8H)， 2.12(m, 10H), 2.89(m, 4H)， 3.50(m, 1800H), 4.05(m, 9H)， 4.26(m, 5H)， 5.26(m, 5H)， 5.48(m，5H)， 7.07(m, 2H), 7.55-7.60(m, 3H), 7.89(m, 3H), 8.1 l(m, 4H)， 8.20(m, 3H)。 [00245] Monomethoxypolyethylene glycol glutamic acid dipeptide acid (Compound 5) was prepared according to the method described in Example 1. Monomethoxypolyethylene glycol glutamic acid dipeptide acid (Compound 5) (molecular weight: 20,000) 5.0 g, enonotecan glycine (Compound 22) 1.02 g, N-hydroxysuccinimide (NHS) 115 mg , 4-dimethylaminopyridine (DMAP) 153mg dissolved in 50mL anhydrous dichloromethane, under the protection of nitrogen, add two Cyclohexylcarbodiimide (DCC) 309 mg. The reaction was stirred at room temperature overnight. The solid matter was removed by filtration, and the excess solvent was removed by rotary evaporation. The residue was added with 100 mL of isopropyl alcohol (IPA), filtered, and the product was dried in vacuo to give monomethoxy polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight 20000) - Inotropin conjugate (YNR-l) 4.4g. iH-NMR MSO-c^): 0.84-0.89 (m, 8H), 1.24 (m, 9H), 1.40-1.50 (m, 21H), 1.79 (m, 8H), 2.12 (m, 10H), 2.89 ( m, 4H), 3.50 (m, 1800H), 4.05 (m, 9H), 4.26 (m, 5H), 5.26 (m, 5H), 5.48 (m, 5H), 7.07 (m, 2H), 7.55-7.60 (m, 3H), 7.89 (m, 3H), 8.1 l (m, 4H), 8.20 (m, 3H).
[00246】 实施例 15 : Y型聚乙二醇谷氨酸二肽甘氨酸（数均分子量 40000) - 伊诺替康结合物 （YNR-2) 的制备  Example 15: Y-type polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight 40000) - Preparation of INOTECK conjugate (YNR-2)
Figure imgf000058_0001
Figure imgf000058_0001
[00247】 根据上述实施例所述方法制备 Y型聚乙二醇谷氨酸二肽酸 （化合物 13) (分子量为 40000) 。 Y型聚乙二醇谷氨酸二肽酸（化合物 13) (分子量为 40000) 10g、 甘氨酸依诺替康酯 （化合物 22)1.02g、 N-羟基琥珀酰亚胺 （NHS) 115mg、 4-二甲基氨基吡啶（DMAP) 153mg溶于 50mL无水二氯甲垸中， 氮气 保护下， 再添加二环己基碳二亚胺 （DCC ) 309mg。 室温过夜搅拌反应。 过滤 除去固体物质， 多余溶剂旋转蒸发除去， 残余物添加 lOOmL异丙醇 （IPA) ， 过滤，产物真空干燥。得到 Y型聚乙二醇谷氨酸二肽甘氨酸 (数均分子量 40000) 一伊诺替康结合物 （YNR-2) 9.4g。 1H-NMR (DMSO-d6): 0.82(m, 9H), 1.23(m, 11H), 1.40-1.50(m, 20H), 1.79(m, 8H)， 2.12(m, 10H), 3.50(m, 3600H), 3.98-4.22(m, 20H), 5.26-5.48(m， 8H)， 7.07(m, 2H), 7.80(m, 2H), 7.90(m, 3H), 8.1 l(m, 4H), 8.20(m, 2H [00247] Y-type polyethylene glycol glutamic acid dipeptide acid (compound 13) (molecular weight: 40,000) was prepared according to the method described in the above examples. Y-type polyethylene glycol glutamic acid dipeptide acid (compound 13) (molecular weight 40,000) 10 g, quinolidine glycine (compound 22) 1.02 g, N-hydroxysuccinimide (NHS) 115 mg, 4- Dimethylaminopyridine (DMAP) 153 mg was dissolved in 50 mL of anhydrous dichloromethane, and 309 mg of dicyclohexylcarbodiimide (DCC) was added under nitrogen. The reaction was stirred at room temperature overnight. The solid matter was removed by filtration, and the excess solvent was removed by rotary evaporation. &lt;RTI ID=0.0&gt;&gt; A Y-type polyethylene glycol glutamic acid dipeptide glycine (number average molecular weight: 40000) - an innotecan conjugate (YNR-2) of 9.4 g was obtained. 1H-NMR (DMSO-d 6 ): 0.82 (m, 9H), 1.23 (m, 11H), 1.40-1.50 (m, 20H), 1.79 (m, 8H), 2.12 (m, 10H), 3.50 (m, 3600H), 3.98-4.22 (m, 20H), 5.26-5.48 (m, 8H) , 7.07(m, 2H), 7.80(m, 2H), 7.90(m, 3H), 8.1 l(m, 4H), 8.20(m, 2H
[00248】 实施例 16:单甲氧基聚乙二醇谷氨酸三肽甘氨酸 (数均分子量 20000)  Example 16: Monomethoxypolyethylene glycol glutamic acid tripeptide glycine (number average molecular weight 20000)
Figure imgf000059_0001
Figure imgf000059_0001
[00249】 根据上述实施例所述方法制备单甲氧基聚乙二醇谷氨酸三肽酸 （化 合物 17) (分子量为 20000)。单甲氧基聚乙二醇谷氨酸三肽酸（化合物 17) (分子 量为 20000) 5.0g、 依诺替康甘氨酸酯 （化合物 22) 1.36g、 N-羟  Monomethoxypolyethylene glycol glutamic acid tripeptide acid (Compound 17) (molecular weight: 20,000) was prepared according to the method described in the above examples. Monomethoxy polyethylene glycol glutamic acid tripeptide acid (compound 17) (molecular weight 20000) 5.0 g, ennotecan glycine ester (compound 22) 1.36 g, N-hydroxyl
(NHS) 115mg、 4-二甲基氨基吡啶（DMAP) 153mg溶于 50mL无水二氯甲垸中， 氮气保护下， 再添加二环己基碳二亚胺 （DCC) 309mg。 室温过夜搅拌反应。 过滤除去固体物质，多余溶剂旋转蒸发除去，残余物添加 lOOmL异丙醇（IPA)， 过滤， 产物真空干燥。 得到单甲氧基聚乙二醇谷氨酸三肽甘氨酸 （数均分子量 —伊诺替康结合物 （YNR-3) 4 -NMR (DMSO-d6): .84 .89(m, 8H)， 1.24(m, 9H)，， 1.71-1.74(m, 23H), 2.12(m, 16H), 2.95-2.99(m, 19H), 3.50(m, 1800H), 4.22(m, 18H), 5.45(m， 5H)， 5.49(m，5H)， 7.07(m, 2H), 7.55-7.60(m, 3H), 7.89(m, 3H), 8.1 l(m, 4H)， 8.20(m, 3H)。 (NHS) 115 mg, 4-dimethylaminopyridine (DMAP) 153 mg was dissolved in 50 mL of anhydrous dichloromethane, and 309 mg of dicyclohexylcarbodiimide (DCC) was added under nitrogen. The reaction was stirred at room temperature overnight. The solid matter was removed by filtration, and the excess solvent was removed by rotary evaporation. &lt;RTI ID=0.0&gt;&gt; Monomethoxypolyethylene glycol glutamic acid tripeptide glycine (number average molecular weight - INOTECO conjugate (YNR-3) 4 -NMR (DMSO-d6): .84.89 (m, 8H), 1.24(m, 9H),, 1.71-1.74(m, 23H), 2.12(m, 16H), 2.95-2.99(m, 19H), 3.50(m, 1800H), 4.22(m, 18H), 5.45(m 5H), 5.49 (m, 5H), 7.07 (m, 2H), 7.55-7.60 (m, 3H), 7.89 (m, 3H), 8.1 l (m, 4H), 8.20 (m, 3H).
[00250】 实施例 17:聚乙二醇伊诺替康结合物对人肠癌 HCT-1 16裸小鼠移植 瘤的生长抑制作用  Example 17: Growth Inhibition of Human Intestinal Carcinoma HCT-1 16 Nude Mice Transplanted with Polyethylene Glycosin Inocomb Conjugate
[00251】 受试物 YNR-1为单甲氧基聚乙二醇乙酸甘氨酸一依诺替康结合物。  [00251] The test substance YNR-1 is a monomethoxypolyethylene glycol acetate glycine-inonotecan conjugate.
[00252】 阳性对照药物盐酸依诺替康注射液（CPT-1 1 ) ， 40mg/2ml， 批号为 8UL002-B, 由 Aventis Pharma (Dagenham)公司制造。 临用时用生理盐水稀 释至所需浓度。 [00252] Positive control drug enonotecan hydrochloride injection (CPT-1 1 ), 40mg/2ml, batch number is 8UL002-B, manufactured by Aventis Pharma (Dagenham). Dilute with physiological saline to the desired concentration at the time of use.
[00253】 剂量设置 [00253] Dose setting
[00254】 YNR-1剂量设为 45mg/kg (按所含依诺替康的量计算） ， 静脉注射 给药， 每周一次， 连续三周； CPT-1 1剂量为 45mg/kg， 每周静脉注射给药一 次； 及 15 mg/kg， 每周静脉注射给药三次， 连续三周。  [00254] YNR-1 dose was set to 45 mg / kg (calculated according to the amount of INOTECK), administered intravenously, once a week for three weeks; CPT-1 1 dose was 45 mg / kg, weekly Administered once intravenously; and 15 mg/kg, administered intravenously three times a week for three consecutive weeks.
[00255】 动物： BALB/cA裸小鼠， 雄性， 5-6周龄， 体重 19±2 g， 由中国科学 院上海药物研究所提供， 生产合格证编号： SCXK (沪） 2008-0017。 每组动 物数： 阴性对照组 12只， 给药组 6只。  [00255] Animals: BALB/cA nude mice, male, 5-6 weeks old, weighing 19±2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017. The number of animals in each group: 12 in the negative control group and 6 in the drug-administered group.
[00256】 细胞株 [00256] Cell line
[00257】 人肠癌 HCT-1 16细胞株购自 ATCC。 用该细胞株接种裸小鼠右侧腋 窝皮下， 细胞接种量为 5 X 106/只， 形成移植瘤后再在裸小鼠体内传 2代后使 用。  [00257] Human intestinal cancer HCT-1 16 cell line was purchased from ATCC. The cell strain was inoculated subcutaneously into the right axillary fossa of the nude mice, and the inoculation amount was 5×106/head, and the transplanted tumor was formed and then used in nude mice for 2 generations.
[00258】 实验方法  [00258] Experimental method
[00259】 取生长旺盛期的瘤组织剪切成 1 .5mm3左右， 在无菌条件下， 接种于 裸小鼠右侧腋窝皮下。 裸小鼠皮下移植瘤用游标卡尺测量移植瘤直径， 待肿瘤 生长至 100-200 mm3后将动物随机分组。 YNR-1按 45mg/kg剂量组、 对照组 给等量生理盐水， 每周静脉注射一次， 连续三周。 CPT-1 1 (15 mg/kg)作为阳性 对照药， 每周静脉注射三次， 连续给药三周。 给药结束后， 继续观察一周。 整 个实验过程中，每周 2次测量移植瘤直径，同时称量小鼠体重。肿瘤体积 (tumor volume, TV)的计算公式为： TV = 1 /2 X a X b2， 其中 a、 b分别表示长、 宽。 根据测量的结果计算出相对肿瘤体积 （relative tumor volume, RTV) ， 计算 公式为： RTV = Vt/V0。 其中 V0为分笼给药时 (即 d0)测量所得肿瘤体积， Vt 为每一次测量时的肿瘤体积。 抗肿瘤活性的评价指标为相对肿瘤增殖率 T/C (%) ， 计算公式如下： T/C (%) = (TRTV/CRTV) X100 %, TRTV: 治疗组 RTV CRTV: 阴性对照组 RTV [00260】 结果与讨论 [00259] The tumor tissue in the vigorous growth period was cut into about 1.5 mm 3 and inoculated subcutaneously in the right axilla of the nude mice under aseptic conditions. The diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm 3 . YNR-1 was administered in the 45 mg/kg dose group and the control group was given the same amount of normal saline once a week for three consecutive weeks. CPT-1 1 (15 mg/kg) was used as a positive control drug, administered intravenously three times a week for three weeks. After the end of the administration, continue to observe for one week. The diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed. The tumor volume (TV) is calculated as: TV = 1 /2 X a X b 2 , where a and b represent length and width, respectively. According to the measured results, the relative tumor volume (RTV) is calculated, and the formula is: RTV = Vt/V0. Where V0 is the measured tumor volume at the time of sub-cage administration (i.e., d0), and Vt is the tumor volume at each measurement. The anti-tumor activity evaluation index is the relative tumor growth rate T/C. (%), the formula is as follows: T/C (%) = (TRTV/CRTV) X100 %, TRTV: treatment group RTV CRTV: negative control group RTV [00260] Results and discussion
[00261】 实验结果如表 9所示。 YNR-1 (45 mg/kg)每周静脉注射给药一次， 连续三周，对人肠癌 HCT-116裸小鼠皮下移植瘤的生长有明显抑制， T/C百分 数为 27.60%，抑瘤效果优于同样剂量同样给药方案的 CPT-1145mg/kg， 同样 给药方案实验治疗的 CPT-11也能一定程度抑制 HCT-116皮下移植瘤的生长， 但 T/C百分数仅为 63.56%。 阳性对照 CPT-11 (15mg/kg)每周静脉给药三次， 连续三周， 同样能显著抑制 HCT-116皮下移植瘤的生长， T/C值为 39.84% 整个实验过程中， 各组裸小鼠体重均有下降， 与溶剂对照组相比， 仅 CPT-11 45mg/kg组裸小鼠体重减轻稍强于溶剂对照组。 The experimental results are shown in Table 9. YNR-1 (45 mg/kg) was administered intravenously once a week for three weeks, and the growth of subcutaneous xenografts of human intestinal cancer HCT-116 nude mice was significantly inhibited. The percentage of T/C was 27.60%. The effect is better than the same dose of CPT-1145mg/kg of the same dosage regimen. The same treatment scheme CPT-11 can also inhibit the growth of HCT-116 subcutaneous xenografts to a certain extent, but the percentage of T/C is only 63.56%. The positive control CPT-11 (15mg/kg) was administered intravenously three times a week for three weeks, which also significantly inhibited the growth of HCT-116 subcutaneous xenografts. The T/C value was 39.84%. The body weight of the mice decreased. Compared with the solvent control group, the weight loss of the nude mice in the CPT-11 45 mg/kg group was slightly stronger than that in the solvent control group.
[00262】 表 9. PEG化依诺替康对人肠癌 HCT-116裸小鼠移植瘤的实验治疗作用 Table 9. Experimental therapeutic effect of PEGylated ennotecan on human colon cancer HCT-116 nude mice xenografts
TV (mm3, TV (mm 3 ,
动物数 体重 (g) RTV T/C 组别 剂量、 给药方式 mean SD)  Number of animals Weight (g) RTV T/C group Dose, mode of administration mean SD)
- (mean SD) (%) 开始 最后 开始 最后 do d28 - (mean SD) (%) Start last and finally do d 2 8
溶剂对照 0.4mV只 qwx3w i v 12 12 19.7 17.3 132±32 804±82 6.45±1.83Solvent control 0.4mV only qwx3w i v 12 12 19.7 17.3 132±32 804±82 6.45±1.83
CPT-11 15mg/kg, q3w<3w i v 6 6 19.1 16.6 129±26 322±37 2.57±0.66** 39.84CPT-11 15mg/kg, q3w<3w i v 6 6 19.1 16.6 129±26 322±37 2.57±0.66** 39.84
CPT-11 45mg/kg, qw<3w i v 6 6 19.2 15.1 126±25 512±99 4.10±0.73* 63.56CPT-11 45mg/kg, qw<3w i v 6 6 19.2 15.1 126±25 512±99 4.10±0.73* 63.56
YNR-1 45mg/kg, qw<3w i v 6 6 18.8 16.8 132±30 234±54 1.78±0.23**' 27.60 注： t检验， vs翻对照组， * p<0.01, **p<0.001; vs CPT-1145mg/kg组， #ρ<0·05 ## p<0.001 YNR-1 45mg/kg, qw<3w iv 6 6 18.8 16.8 132±30 234±54 1.78±0.23**' 27.60 Note: t test, vs turn control group, * p<0.01, **p<0.001; vs CPT-1145mg/kg group, #ρ<0·05 ## p<0.001
[00263】 实施例 18: 聚乙二醇伊诺替康结合物对人肠癌 HT-29裸小鼠移植瘤 的生长抑制作用 Example 18: Growth Inhibition of Human Intestinal Carcinoma HT-29 Nude Mice Transplanted Tumor by Polyethylene Glycosin Integate Conjugate
[00264】 受试物 YNR-1为单甲氧基聚乙二醇谷氨酸二肽甘氨酸一依诺替康结 合物。 [00264] The test substance YNR-1 is a monomethoxy polyethylene glycol glutamic acid dipeptide glycine-inonotecan complex.
[00265】 阳性对照药物盐酸依诺替康注射液（CPT-11) 40mg/2ml， 批号为 8UL002-B, 由 Aventis Pharma (Dagenham)公司制造。 临用时用生理盐水稀 释至所需浓度。 [00266】 剂量设置 [00265] The positive control drug inonotecan hydrochloride (CPT-11) 40 mg/2 ml, lot number 8UL002-B, manufactured by Aventis Pharma (Dagenham). Dilute with physiological saline to the desired concentration at the time of use. [00266] Dose setting
[00267】 YNR-1剂量设为 45mg/kg (按所含依诺替康的量计算） ， 静脉注射 给药， 每周一次， 连续三周； CPT-11剂量为 45mg/kg， 每周静脉注射给药一 次； 及 15mg/kg， 每周静脉注射给药三次， 连续三周。  [00267] YNR-1 dose is set to 45mg / kg (calculated according to the amount of INOTEC), intravenous administration, once a week for three weeks; CPT-11 dose is 45mg / kg, weekly vein The drug was administered once; and 15 mg/kg, administered intravenously three times a week for three consecutive weeks.
[00268】 动物  [00268] animals
[00269】 BALB/cA裸小鼠， 雄性， 5-6周龄， 体重 18±2g， 由中国科学院上 海药物研究所提供， 生产合格证编号： SCXK (沪） 2008-0017。 每组动物数： 阴性对照组 12只， 给药组 6只。  [00269] BALB/cA nude mice, male, 5-6 weeks old, weighing 18±2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017. The number of animals in each group: 12 in the negative control group and 6 in the drug-administered group.
[00270】 细胞株  [00270] cell line
[00271】 人肠癌 HT-29细胞株购自 ATCC。 用该细胞株接种裸小鼠右侧腋窝 皮下， 细胞接种量为 5X106/只， 形成移植瘤后再在裸小鼠体内传 2代后使用。  [00271] Human intestinal cancer HT-29 cell line was purchased from ATCC. The cell strain was used to inoculate the right axilla of the nude mice subcutaneously, and the inoculation amount of the cells was 5×106/head, and the transplanted tumor was formed and then used in nude mice for 2 generations.
[00272】 实验方法 [00272] Experimental method
[00273】 取生长旺盛期的瘤组织剪切成 1.5mm3左右， 在无菌条件下， 接种于 裸小鼠右侧腋窝皮下。 裸小鼠皮下移植瘤用游标卡尺测量移植瘤直径， 待肿瘤 生长至 100-200 mm3后将动物随机分组。 YNR-1及 CPT-11按 45mg/kg剂量 组、对照组给等量生理盐水，每周静脉注射一次，连续三周。 CPT-11 (15mg/kg) 作为阳性对照药， 每周静脉注射三次， 连续给药三周。 给药结束后， 继续观察 一周。 整个实验过程中， 每周 2次测量移植瘤直径， 同时称量小鼠体重。 肿瘤 体积 (tumor volume, TV)的计算公式为： TV =1/2XaXb2， 其中 a、 b分别表 示长、宽。根据测量的结果计算出相对肿瘤体积（relative tumor volume, RTV)， 计算公式为： RTV = Vt/V0。其中 V0为分笼给药时 (即 d0)测量所得肿瘤体积， vt为每一次测量时的肿瘤体积。 抗肿瘤活性的评价指标为相对肿瘤增殖率 T/C (%) ， 计算公式如下： T/C (%) = (TRTV/CRTV) X100%， TRTV: 治疗组 RTV ； CRTV: 阴性对照组 RTV。 [00274】 结果与讨论 [00273] The tumor tissue in the vigorous growth period was cut into 1.5 mm 3 and inoculated subcutaneously in the right axilla of nude mice under aseptic conditions. The diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm 3 . YNR-1 and CPT-11 were given the same amount of normal saline in the 45 mg/kg dose group and the control group, intravenously once a week for three weeks. CPT-11 (15 mg/kg) was administered as a positive control drug three times a week for three weeks. After the end of the administration, continue to observe for one week. The diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed. The formula for calculating tumor volume (TV) is: TV = 1/2XaXb 2 , where a and b represent length and width, respectively. According to the measured results, the relative tumor volume (RTV) is calculated, and the formula is: RTV = Vt/V0. Where V0 is the measured tumor volume at the time of sub-cage administration (ie, d0), and vt is the tumor volume at each measurement. The anti-tumor activity was evaluated as the relative tumor growth rate T/C (%), and the formula was as follows: T/C (%) = (TRTV/CRTV) X100%, TRTV: treatment group RTV; CRTV: negative control group RTV. [00274] Results and Discussion
[00275】 实验结果如表 10所示。 YNR-1 (45 mg/kg)每周静脉注射给药一次， 连续三周，对人肠癌 HT-29裸小鼠皮下移植瘤的生长有明显抑制，给药一周后， 实验治疗组小鼠所带肿瘤体积增长即有减缓。 至实验结束， T/C百分数为 41 .08%。 阳性对照 CPT-1 1 (15mg/kg)每周静脉给药三次， 连续三周， 同样能 显著抑制 HT-29皮下移植瘤的生长， T/C值为 27.27%。 整个实验过程中， 各 组裸小鼠生长状态良好， 仅 CPT-1 1两个不同剂量组裸小鼠体重有下降。 [00275] The experimental results are shown in Table 10. YNR-1 (45 mg/kg) was administered intravenously once a week for three weeks, and the growth of subcutaneous xenografts of human intestinal cancer HT-29 nude mice was significantly inhibited. One week after administration, mice in the experimental treatment group The increase in tumor volume is slowed down. By the end of the experiment, the percentage of T/C was 41.08%. The positive control CPT-1 1 (15 mg/kg) was administered intravenously three times a week for three consecutive weeks, which also significantly inhibited the growth of HT-29 subcutaneous xenografts with a T/C value of 27.27%. During the whole experiment, the nude mice in each group grew well, and only the two different dose groups of CPT-1 1 had a decrease in body weight.
[00276]表 10. PEG化依诺替康对人肠癌 HT-29裸小鼠移植瘤的实验治疗作用  [00276] Table 10. Experimental treatment of PEGylated ennotecan on human colon cancer HT-29 nude mice xenografts
TV (mm3, TV (mm 3 ,
动物数 体重 RTV T/C 组别 剂量、 给药方式 ω mean SD)  Number of animals Weight RTV T/C group Dose, mode of administration ω mean SD)
- (mean SD) (%) 开始 最后 开始 最后 d0 d28 - (mean SD) (%) Start last and last d 0 d 28
溶剂对照 0.4mV只 qwx3w iv 12 12 19.0 19.2 119±29 1588 Solvent control 0.4mV only qwx3w iv 12 12 19.0 19.2 119±29 1588
13.90± 5.38 13.90± 5.38
578 578
CPT-11 15mg/kg, q3wx 3w iv 6 6 18.2 16.0 121±14 460± 152 3.79 1.21*' 27.27 CPT-11 15mg/kg, q3wx 3w iv 6 6 18.2 16.0 121±14 460± 152 3.79 1.21*' 27.27
CPT-11 45mg/kg, qw <3w iv 6 6 18.6 16.2 116±13 995± 368 8.69± 3.43*** 62.52CPT-11 45mg/kg, qw <3w iv 6 6 18.6 16.2 116±13 995± 368 8.69± 3.43*** 62.52
YNR-1 45mg/kg, qw <3w iv 6 6 18.7 18.4 115±25 642 105 5.71 1.21 ** 41.08 注： t 检验， vs 溶剂对照组， * ρ<0.05, **ρ<0.01 , ***ρ<0.001 vs CPT-1 1 45mg/kg组， # p<0.05 YNR-1 45mg/kg, qw <3w iv 6 6 18.7 18.4 115±25 642 105 5.71 1.21 ** 41.08 Note: t test, vs solvent control group, * ρ<0.05, **ρ<0.01, ***ρ <0.001 vs CPT-1 1 45mg/kg group, # p<0.05
[00277】 实施例 19: 聚乙二醇伊诺替康结合物对人肺癌 A549裸小鼠移植瘤 的生长抑制作用 Example 19: Growth Inhibition of Human Lung Cancer A549 Nude Mice Transplanted with Polyethylene Glycosin Inocomb Conjugate
[00278】 受试物 YNR-1为单甲氧基聚乙二醇谷氨酸二肽甘氨酸一依诺替康结 合物。 [00278] The test substance YNR-1 is a monomethoxy polyethylene glycol glutamic acid dipeptide glycine-inonotecan complex.
[00279】 阳性对照药物盐酸依诺替康注射液（CPT-1 1 ) 40mg/2ml， 批号为 8UL002-B, 由 Aventis Pharma (Dagenham)公司制造。 临用时用生理盐水稀 释至所需浓度。 [00280】 剂量设置 [00279] The positive control drug inonotecan hydrochloride (CPT-1 1 ) 40 mg/2 ml, lot number 8UL002-B, was manufactured by Aventis Pharma (Dagenham). When used, it is diluted with physiological saline to the desired concentration. [00280] Dose setting
[00281】 YNR-1剂量设为 45mg/kg (按所含依诺替康的量计算） ， 静脉注射 给药， 每周一次， 连续三周； CPT-1 1剂量为 45mg/kg， 每周静脉注射给药一 次； 及 15mg/kg， 每周静脉注射给药三次， 连续三周。 [00281] YNR-1 dose was set to 45 mg / kg (calculated according to the amount of INOTECK), administered intravenously, once a week for three weeks; CPT-1 1 dose was 45 mg / kg, weekly Intravenous administration And 15 mg/kg, administered intravenously three times a week for three consecutive weeks.
[00282】 动物 [00282] Animals
[00283】 BALB/cA裸小鼠， 雌性， 5-6周龄， 体重 18±2g， 由中国科学院上 海药物研究所提供， 生产合格证编号： SCXK (沪） 2008-0017。 每组动物数： 阴性对照组 12只， 给药组 6只。  [00283] BALB/cA nude mice, female, 5-6 weeks old, weighing 18±2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017. The number of animals in each group: 12 in the negative control group and 6 in the drug-administered group.
[00284】 细胞株  [00284] Cell line
[00285】 人肺癌 A549细胞株购自 ATCC。用该细胞株接种裸小鼠右侧腋窝皮 下， 细胞接种量为 5X106/只， 形成移植瘤后再在裸小鼠体内传 2代后使用。  [00285] Human lung cancer A549 cell line was purchased from ATCC. The cell strain was used to inoculate the right axilla of the nude mice, and the inoculation amount was 5×106/head. After the transplanted tumor was formed, it was used in nude mice for 2 generations.
[00286】 实验方法 [00286] Experimental method
[00287】 取生长旺盛期的瘤组织剪切成 1.5mm3左右， 在无菌条件下， 接种于 裸小鼠右侧腋窝皮下。 裸小鼠皮下移植瘤用游标卡尺测量移植瘤直径， 待肿瘤 生长至 100-200 mm3后将动物随机分组。 YNR-1及 CPT-11按 45mg/kg剂量 组、对照组给等量生理盐水，每周静脉注射一次，连续三周。 CPT-11 (15mg/kg) 作为阳性对照药， 每周静脉注射三次， 连续给药三周。 给药结束后， 继续观察 一周。 整个实验过程中， 每周 2次测量移植瘤直径， 同时称量小鼠体重。 肿瘤 体积 (tumor volume, TV)的计算公式为： TV = 1/2XaXb2， 其中 a、 b分别表 示长、宽。根据测量的结果计算出相对肿瘤体积（relative tumor volume, RTV)， 计算公式为： RTV = Vt/V0。其中 V0为分笼给药时 (即 d0)测量所得肿瘤体积， vt为每一次测量时的肿瘤体积。 抗肿瘤活性的评价指标为相对肿瘤增殖率 T/C (%) ， 计算公式如下： T/C (%) = (TRTV/CRTV) X100%, TRTV: 治疗组 RTV ； CRTV: 阴性对照组 RTV。 [00287] The tumor tissue in the vigorous growth period was cut into 1.5 mm 3 and inoculated subcutaneously in the right axilla of the nude mice under aseptic conditions. The diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm 3 . YNR-1 and CPT-11 were given the same amount of normal saline in the 45 mg/kg dose group and the control group, intravenously once a week for three weeks. CPT-11 (15 mg/kg) was administered as a positive control drug three times a week for three weeks. After the end of the administration, continue to observe for one week. The diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed. The tumor volume (TV) is calculated as: TV = 1/2XaXb 2 , where a and b represent length and width, respectively. According to the measured results, the relative tumor volume (RTV) is calculated, and the formula is: RTV = Vt/V0. Where V0 is the measured tumor volume at the time of sub-cage administration (ie, d0), and vt is the tumor volume at each measurement. The anti-tumor activity was evaluated as the relative tumor growth rate T/C (%), and the formula was as follows: T/C (%) = (TRTV/CRTV) X100%, TRTV: treatment group RTV; CRTV: negative control group RTV.
[00288】 结果与讨论 [00288] Results and Discussion
[00289】 实验结果如表 11所示。 YNR-1和 CPT-11 (45 mg/kg)每周静脉注射 给药一次， 连续三周， 对人肺癌 A549裸小鼠皮下移植瘤的生长有明显抑制， T/C百分数为 20.62% YNR-1的抑瘤效果远优于同样剂量同样给药方案的 CPT-11。 与溶剂对照组相比， YNR-1实验治疗一周后， 荷瘤鼠皮下移植瘤的 生长就有减缓。 阳性对照 CPT-11(15mg/kg)每周静脉给药三次， 连续三周， 同 样能显著抑制 A549皮下移植瘤的生长， T/C值为 53.26%。 整个实验过程中， 各治疗组裸小鼠生长状况良好， 体重增加稍缓于溶剂对照组。 [00289] The experimental results are shown in Table 11. YNR-1 and CPT-11 (45 mg/kg) were administered intravenously once a week for three weeks, which significantly inhibited the growth of subcutaneous xenografts in human lung cancer A549 nude mice. The T/C percentage is 20.62%. The anti-tumor effect of YNR-1 is much better than that of CPT-11 of the same dosage regimen. Compared with the vehicle control group, the growth of subcutaneous xenografts in tumor-bearing mice was slowed after one week of YNR-1 treatment. The positive control CPT-11 (15 mg/kg) was administered intravenously three times a week for three consecutive weeks, which also significantly inhibited the growth of A549 subcutaneous xenografts with a T/C value of 53.26%. During the whole experiment, the nude mice in each treatment group grew well, and the weight gain was slightly slower than the solvent control group.
[00290]表 11. PEG化依诺替康对人肺癌 A549裸小鼠移植瘤的实验治疗作用 动物数 体重 (g) TV (mm3, mean±SD) RTV T/C 组别 剂量、 给药方式 [00290] Table 11. Experimental therapeutic effect of PEGylated irinotecan on human lung cancer A549 nude mice xenografts Animal body weight (g) TV (mm 3 , mean ± SD) RTV T/C group dose, administration the way
开始 最后 开始 最后 d d2S (mean±SD) (%) 溶剂对照 0.4mV只 qwx3w iv 12 12 18.4 23.1 108±21 1546 496 14.40±4.38 CPT-11 15mg/kg, q3w<3w iv 6 6 18.1 21.2 112±21 831 ±305 7.67±2.88**' 53.26 CPT-11 45mg/kg, qw<3w iv 6 6 18.4 21.7 110±15 1055±432 9.87±4.87 68.54 YNR-1 45mg/kg, qw<3w iv 6 6 19.1 22.0 105±21 320 154 2.97±1.04*' 20.62 注： t检验， vs 溶剂对照组， *p<0.001; vsCPT-1145mg/kg组， # p<0.01Start last and finally dd 2S (mean±SD) (%) Solvent control 0.4mV only qwx3w iv 12 12 18.4 23.1 108±21 1546 496 14.40±4.38 CPT-11 15mg/kg, q3w<3w iv 6 6 18.1 21.2 112± 21 831 ±305 7.67±2.88**' 53.26 CPT-11 45mg/kg, qw<3w iv 6 6 18.4 21.7 110±15 1055±432 9.87±4.87 68.54 YNR-1 45mg/kg, qw<3w iv 6 6 19.1 22.0 105±21 320 154 2.97±1.04*' 20.62 Note: t test, vs solvent control group, *p<0.001; vsCPT-1145mg/kg group, #p<0.01
[00291】 实施例 20: 聚乙二醇伊诺替康结合物对人卵巢癌 SKOV-3裸小鼠移 植瘤的生长抑制作用 Example 20: Growth inhibition of ectopic conjugates of polyethylene glycol on human ovarian cancer SKOV-3 nude mice
[00292】 受试物 YNR-1为单甲氧基聚乙二醇谷氨酸二肽甘氨酸一依诺替康结 合物。  [00292] The test substance YNR-1 is a monomethoxy polyethylene glycol glutamic acid dipeptide glycine-inonotecan complex.
[00293】 阳性对照药物盐酸依诺替康注射液（CPT-11) 40mg/2ml， 批号为 8UL002-B, 由 AventisPharma(Dagenham)公司制造。 临用时用生理盐水稀 释至所需浓度。  [00293] The positive control drug inonotecan hydrochloride (CPT-11) 40 mg/2 ml, batch number 8UL002-B, was manufactured by Aventis Pharma (Dagenham). When used, it is diluted with physiological saline to the desired concentration.
[00294】 剂量设置 [00294] Dose setting
[00295】 YNR-1剂量设为 45mg/kg (按所含依诺替康的量计算） ， 静脉注射 给药， 每周一次， 连续三周； CPT-11剂量为 45mg/kg， 每周静脉注射给药一 次； 和 15mg/kg， 每周静脉注射给药三次， 连续三周。 [00296】 动物 [00295] YNR-1 dose was set to 45 mg / kg (calculated according to the amount of INOTECK), administered intravenously, once a week for three weeks; CPT-11 dose was 45 mg / kg, weekly vein The drug was administered once; and 15 mg/kg, administered intravenously three times a week for three consecutive weeks. [00296] animals
[00297】 BALB/cA裸小鼠， 雌性， 5-6周龄， 体重 19±2g， 由中国科学院上 海药物研究所提供， 生产合格证编号： SCXK (沪） 2008-0017。 每组动物数： 阴性对照组 12只， 给药组 6只。  [00297] BALB/cA nude mice, female, 5-6 weeks old, weighing 19±2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017. The number of animals in each group: 12 in the negative control group and 6 in the drug-administered group.
[00298】 细胞株  [00298] Cell line
[00299】 人卵巢癌 SKOV-3细胞株购自 ATCC。 用该细胞株接种裸小鼠右侧 腋窝皮下， 细胞接种量为 5X 106/只， 形成移植瘤后再在裸小鼠体内传 2代后 使用。  [00299] Human ovarian cancer SKOV-3 cell strain was purchased from ATCC. The cell strain was inoculated subcutaneously into the right axilla of the nude mice, and the inoculation amount was 5×106/piece, and the transplanted tumor was formed and then used in nude mice for 2 generations.
[00300】 实验方法  [00300] Experimental method
[00301】 取生长旺盛期的瘤组织剪切成 1.5mm3左右， 在无菌条件下， 接种于 裸小鼠右侧腋窝皮下。 裸小鼠皮下移植瘤用游标卡尺测量移植瘤直径， 待肿瘤 生长至 100-200 mm3后将动物随机分组。 YNR-1及 CPT-11按 45mg/kg剂量 组、对照组给等量生理盐水，每周静脉注射一次，连续三周。 CPT-11 (15mg/kg) 作为阳性对照药， 每周静脉注射三次， 连续给药三周。 给药结束后， 继续观察 一周。 整个实验过程中， 每周 2次测量移植瘤直径， 同时称量小鼠体重。 肿瘤 体积 (tumor volume, TV)的计算公式为： TV = 1/2XaXb2， 其中 a、 b分别表 示长、宽。根据测量的结果计算出相对肿瘤体积（relative tumor volume, RTV)， 计算公式为： RTV = Vt/V0。其中 V0为分笼给药时 (即 d0)测量所得肿瘤体积， vt为每一次测量时的肿瘤体积。 抗肿瘤活性的评价指标为相对肿瘤增殖率 T/C (%) ， 计算公式如下： T/C (%) = (TRTV/CRTV) X100%, TRTV: 治疗组 RTV ； CRTV: 阴性对照组 RTV。 [00301] The tumor tissue in the vigorous growth period was cut into 1.5 mm 3 and inoculated subcutaneously in the right axilla of nude mice under aseptic conditions. The diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm3. YNR-1 and CPT-11 were given the same amount of normal saline in the 45 mg/kg dose group and the control group, intravenously once a week for three weeks. CPT-11 (15 mg/kg) was administered as a positive control drug three times a week for three weeks. After the end of the administration, continue to observe for one week. The diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed. The tumor volume (TV) is calculated as: TV = 1/2XaXb 2 , where a and b represent length and width, respectively. According to the measured results, the relative tumor volume (RTV) is calculated, and the formula is: RTV = Vt/V0. Where V0 is the measured tumor volume at the time of sub-cage administration (ie, d0), and vt is the tumor volume at each measurement. The anti-tumor activity was evaluated as the relative tumor growth rate T/C (%), and the formula was as follows: T/C (%) = (TRTV/CRTV) X100%, TRTV: treatment group RTV; CRTV: negative control group RTV.
[00302】 结果与讨论 [00302] Results and Discussion
[00303】 实验结果如表 12所示。 YNR-1每周静脉注射给药一次，连续三周， 对人卵巢癌 SKOV-3裸小鼠皮下移植瘤的生长有明显抑制， T/C百分数为 0.53%, 抑瘤效果远优于同样剂量同样给药方案的 CPT-1 1 ,该条件下， CPT-1 1 的 T/C百分数为 96.46% YNR— 1给药一周后， 各小鼠荷瘤体积就有缩小， 至试验结束时， YNR— 1组有 4只荷瘤鼠肿瘤完全消退。 阳性对照 [00303] The experimental results are shown in Table 12. YNR-1 was administered intravenously once a week for three weeks, and the growth of subcutaneous xenografts of human ovarian cancer SKOV-3 nude mice was significantly inhibited. The percentage of T/C was 0.53%, the tumor inhibition effect is much better than the same dose of CPT-1 1 in the same dosage regimen. Under this condition, the T/C percentage of CPT-1 1 is 96.46%. One week after administration, each mouse has tumor The volume was reduced, and by the end of the experiment, 4 tumor-bearing mice in the YNR-1 group had complete tumor regression. Positive control
CPT-1 1 (15mg/kg)每周静脉给药三次， 连续三周， 同样能显著抑制 SKOV-3皮 下移植瘤的生长， T/C值为 55.69%。 整个实验过程中， 溶剂对照组和 CPT-1 1 两种不同治疗组小鼠体重则有轻微下降； 而 YNR— 1实验治疗组裸小鼠生长状 况良好， 体重增加。 CPT-1 1 (15 mg/kg) was administered intravenously three times a week for three consecutive weeks, which also significantly inhibited the growth of SKOV-3 subcutaneous xenografts with a T/C value of 55.69%. During the whole experiment, the weight of mice in the solvent control group and CPT-1 1 treatment group decreased slightly; while the YNR-1 experimental treatment group had good growth and weight gain.
[00304]表 12. PEG化依诺替康对人卵巢癌 SKOV-3裸小鼠移植瘤的实验治疗 作用 动物数 体重 ω TV (mm3, mean SD) RTV T/C 组别 剂量、 给药方式 [00304] Table 12. Experimental therapeutic effect of PEGylated irinotecan on human ovarian cancer SKOV-3 nude mice xenografts Animal body weight ω TV (mm 3 , mean SD) RTV T/C group dose, administration the way
开始 最后 开始 最后 (mean SD) (%) 溶剂对照 0.4mV只 qwx3w iv 12 12 19.8 18.7 136±28 1770± 588(0) 13.27± 4.75  Start Last Start Finally (mean SD) (%) Solvent control 0.4mV only qwx3w iv 12 12 19.8 18.7 136±28 1770± 588(0) 13.27± 4.75
CPT-11 15mg/kg, q3wx 3w iv 6 6 18.9 18.0 137±33 924± 443(0) 7.39± 5.22**' 55.69 CPT-11 15mg/kg, q3wx 3w iv 6 6 18.9 18.0 137±33 924± 443(0) 7.39± 5.22**' 55.69
CPT-11 45mg/kg, qw <3w iv 6 6 19.7 18.4 136±44 1753± 878(0) 12.80± 3.91 96.46 CPT-11 45mg/kg, qw <3w iv 6 6 19.7 18.4 136±44 1753± 878(0) 12.80± 3.91 96.46
YNR-1 45mg/kg, qw <3w iv 6 6 20.7 22.7 135±35 1.27± 1.98(4) 0.07±0.01 *' 0.53 注： t检验， vs溶剂对照组， * p<0.001 ; vs CPT-1 1 45mg/kg组， # p<0.001 为肿瘤消退动物数 YNR-1 45mg/kg, qw <3w iv 6 6 20.7 22.7 135±35 1.27± 1.98(4) 0.07±0.01 *' 0.53 Note: t test, vs solvent control group, * p<0.001; vs CPT-1 1 45mg/kg group, #p<0.001 is the number of tumor regression animals
[00305】 实施例 21： 依诺替康不同制剂对人肠癌 SW-620裸小鼠移植瘤的生 长抑制作用 [00305] Example 21: Inhibition of growth of human intestinal cancer SW-620 nude mice xenografts by different formulations of irinotecan
[00306】 受试物 YNR-1为单甲氧基聚乙二醇谷氨酸二肽甘氨酸一依诺替康结 合物。 [00306] The test substance YNR-1 is a monomethoxy polyethylene glycol glutamic acid dipeptide glycine-inonotecan complex.
[00307】 阳性对照药物盐酸依诺替康注射液（CPT-1 1 ) 40mg/2ml， 批号为 8UL002-B, 由 Aventis Pharma (Dagenham)公司制造。 临用时用生理盐水稀 释至所需浓度。 [00308】 剂量设置 [00307] The positive control drug inonotecan hydrochloride (CPT-1 1 ) 40 mg / 2 ml, batch number 8UL002-B, manufactured by Aventis Pharma (Dagenham). Dilute with physiological saline to the desired concentration at the time of use. [00308] Dose setting
[00309】 YNR-1剂量设为 45mg/kg (按所含伊诺替康的量计算） ， 静脉注射 给药， 每周一次， 连续三周； CPT-11剂量为 45mg/kg， 每周静脉注射给药一 次； 及 15mg/kg， 每周静脉注射给药三次， 连续三周。  [00309] YNR-1 dose was set to 45 mg / kg (calculated according to the amount of INOTECK), administered intravenously, once a week for three weeks; CPT-11 dose was 45 mg / kg, weekly vein The drug was administered once; and 15 mg/kg, administered intravenously three times a week for three consecutive weeks.
[00310】 动物  [00310] animals
[00311】 BALB/cA裸小鼠， 雄性， 4-6周龄， 体重 19±2g， 由中国科学院上 海药物研究所提供， 生产合格证编号： SCXK (沪） 2008-0017。 每组动物数： 阴性对照组 12只， 给药组 6只。  [00311] BALB/cA nude mice, male, 4-6 weeks old, weighing 19±2 g, provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences, production certificate number: SCXK (Shanghai) 2008-0017. The number of animals in each group: 12 in the negative control group and 6 in the drug-administered group.
[00312】 细胞株  [00312] cell line
[00313】 人肠癌 SW-620细胞株购自 ATCC。 用该细胞株接种裸小鼠右侧腋 窝皮下， 细胞接种量为 5X106/只， 形成移植瘤后再在裸小鼠体内传 2代后使 用。  [00313] Human intestinal cancer SW-620 cell line was purchased from ATCC. The cell strain was inoculated subcutaneously into the right axillary fossa of the nude mice, and the inoculation amount was 5×106/head, and the transplanted tumor was formed and then used in nude mice for 2 generations.
[00314】 实验方法  Experimental method
[00315】 取生长旺盛期的瘤组织剪切成 1.5mm3左右， 在无菌条件下， 接种于 裸小鼠右侧腋窝皮下。 裸小鼠皮下移植瘤用游标卡尺测量移植瘤直径， 待肿瘤 生长至 100-200 mm3后将动物随机分组。 YNR-1及 CPT-11按 45mg/kg剂量 组、对照组给等量生理盐水，每周静脉注射一次，连续三周。 CPT-11 (15mg/kg) 作为阳性对照药， 每周静脉注射三次， 连续给药三周。 给药结束后， 继续观察 一周。 整个实验过程中， 每周 2次测量移植瘤直径， 同时称量小鼠体重。 肿瘤 体积 (tumor volume, TV)的计算公式为： TV= 1/2XaXb2， 其中 a、 b分别表示 长、宽。根据测量的结果计算出相对肿瘤体积（relative tumor volume, RTV)， 计算公式为： RTV = Vt/V0。其中 V0为分笼给药时 (即 d0)测量所得肿瘤体积， vt为每一次测量时的肿瘤体积。 抗肿瘤活性的评价指标为相对肿瘤增殖率 T/C (%) ， 计算公式如下： T/C (%) = (TRTV/CRTV) X100%， TRTV: 治疗组 RTV ； CRTV: 阴性对照组 RTV。 [00315] The tumor tissue in the vigorous growth period was cut into 1.5 mm 3 and inoculated subcutaneously in the right axilla of the nude mice under aseptic conditions. The diameter of the transplanted tumor was measured with a vernier caliper in a nude mouse subcutaneous xenograft, and the animals were randomly divided into groups after the tumor was grown to 100-200 mm 3 . YNR-1 and CPT-11 were given the same amount of normal saline in the 45 mg/kg dose group and the control group, intravenously once a week for three weeks. CPT-11 (15 mg/kg) was administered as a positive control drug three times a week for three weeks. After the end of the administration, continue to observe for one week. The diameter of the transplanted tumor was measured twice a week during the entire experiment, and the body weight of the mice was weighed. The tumor volume (TV) is calculated as: TV = 1/2XaXb 2 , where a and b represent length and width, respectively. According to the measured results, the relative tumor volume (RTV) is calculated, and the formula is: RTV = Vt/V0. Where V0 is the measured tumor volume at the time of sub-cage administration (ie, d0), and vt is the tumor volume at each measurement. The anti-tumor activity evaluation index is the relative tumor proliferation rate T/C (%), and the calculation formula is as follows: T/C (%) = (TRTV/CRTV) X100%, TRTV: treatment group RTV; CRTV: negative control RTV.
[00316】 结果与讨论  [00316] Results and Discussion
[00317】 实验结果如表 13所示。 YNR-1和 CPT-1 1 (45 mg/kg)每周静脉注射 给药一次， 连续三周， 对人肠癌 SW-620裸小鼠皮下移植瘤的生长有明显抑制 作用， 而且 YNR-1的抑瘤效果均优于同样剂量同样给药方案的 CPT-1 1 , 每组 6只实验治疗荷瘤小鼠实验性治疗三周后， YNR-1治疗组有 2只荷瘤小鼠肿 瘤完全消退， 停药一周后也未见反弹。 阳性对照 CPT-1 1 (15mg/kg)每周静脉给 药三次， 连续三周， 同样能显著抑制 SW-620皮下移植瘤的生长， T/C值为 [00317] The experimental results are shown in Table 13. YNR-1 and CPT-1 1 (45 mg/kg) were administered intravenously once a week for three weeks, which significantly inhibited the growth of subcutaneous xenografts in human intestinal cancer SW-620 nude mice, and YNR-1. The anti-tumor effect was better than the same dose of CPT-1 1 in the same dosage regimen. After three weeks of experimental treatment of tumor-bearing mice in each group, two tumor-bearing mice in the YNR-1 treatment group had complete tumors. Regressed, there was no rebound after one week of withdrawal. The positive control CPT-1 1 (15 mg/kg) was administered intravenously three times a week for three consecutive weeks, which also significantly inhibited the growth of SW-620 subcutaneous xenografts. The T/C value was
0.13%。 整个实验过程中， 各治疗组裸小鼠生长状况良好， 体重增加超过溶剂 对照组。  0.13%. During the whole experiment, the nude mice in each treatment group grew well and the weight gain exceeded the solvent control group.
[00318]表 13. PEG化依诺替康对人肠癌 SW-620裸小鼠移植瘤的实验治疗作 用 动物数 体重 ω TV (mm3, mean SD) RTV T/C 组别 剂量、 给药方式 [00318] Table 13. Experimental therapeutic effect of PEGylated irinotecan on human colon cancer SW-620 nude mice xenografts Animal body weight ω TV (mm 3 , mean SD) RTV T/C group dose, administration the way
开始 最后 开始 最后 do (mean SD) (%) 溶剂对照 0.4mV只 qwx3w i ' 12 12 19.4 19.9 132±29 1920± 590(0) 15.40±6.4O  Start Last Start Last do (mean SD) (%) Solvent control 0.4mV only qwx3w i ' 12 12 19.4 19.9 132±29 1920± 590(0) 15.40±6.4O
CPT-11 15mg/kg, q3w <3w i ' 6 6 19.7 24.2 133±33 2.紐 1.51(0) 0.13 CPT-11 15mg/kg, q3w <3w i ' 6 6 19.7 24.2 133±33 2. New 1.51(0) 0.13
CPT-11 45mg/kg, qw <3w i ' 6 6 18.8 21.8 132±25 877± 324(0) 6艇 1.56* 42.86 CPT-11 45mg/kg, qw <3w i ' 6 6 18.8 21.8 132±25 877± 324(0) 6 boats 1.56* 42.86
YNR-1 45mg/kg, qw <3w i ' 6 6 19.2 24.6 130±43 037±0.29(2) 0^3+0.002*^ 0.02 注： t检验， vs 溶剂对照组， * p<0.01， **p<0.001 ; vs CPT-1 1 45mg/kg组， # p<0.001， "()"为肿瘤消退动物数。 YNR-1 45mg/kg, qw <3w i ' 6 6 19.2 24.6 130±43 037±0.29(2) 0^3+0.002*^ 0.02 Note: t test, vs solvent control group, * p<0.01, ** p<0.001; vs CPT-1 1 45 mg/kg group, #p<0.001, "()" is the number of tumor regression animals.

Claims

O 2014/114262 权 I」 要 求 书 PCT/CN2014/071395 O 2014/114262 Right I" Request PCT/CN2014/071395
1. 一种水溶性聚合物-氨基酸寡肽-药物的结合物， 具有如下式 （I) 所示的结 构， A water-soluble polymer-amino acid oligopeptide-drug combination having a structure represented by the following formula (I),
Figure imgf000070_0001
式 （I) 其中， P为水溶性聚合物；
Figure imgf000070_0001
Formula (I) wherein P is a water soluble polymer;
X为连接基团， 所述连接基团连接 P和 A1 ; X is a linking group, and the linking group is bonded to P and A 1 ;
、 A2和 A3各自独立的为相同或不同的氨基酸或氨基酸类似物的残基； , A 2 and A 3 are each independently a residue of the same or different amino acid or amino acid analog;
Di和 D2各自独立的为相同或不同的药物分子残基； Di and D 2 are each independently the same or different drug molecule residues;
a为 0或 1 ; a is 0 or 1;
b为 2-12的整数； b is an integer from 2-12;
c为 0-7的整数； c is an integer from 0 to 7;
d为 0或 1。 d is 0 or 1.
2. 如权利要求 1所述的结合物， 其中所述 P选自下组： 聚乙二醇、 聚丙二醇， 聚谷氨酸， 聚天门冬氨酸， 聚乙烯吡咯垸酮， 聚乙烯醇， 聚丙烯吗啉， 葡聚糖， 羧甲基纤维素及其类似物或共聚物。  2. The conjugate according to claim 1, wherein the P is selected from the group consisting of polyethylene glycol, polypropylene glycol, polyglutamic acid, polyaspartic acid, polyvinylpyrrolidone, polyvinyl alcohol, Polypropylene morpholine, dextran, carboxymethylcellulose and the like or copolymers thereof.
3. 如权利要求 2所述的结合物， 其中所述 P为聚乙二醇， 且所述聚乙二醇的 分子量为 300-60,000。  The conjugate according to claim 2, wherein the P is polyethylene glycol, and the polyethylene glycol has a molecular weight of 300 to 60,000.
4. 如权利要求 3所述的结合物，其中所述聚乙二醇的分子量为 20，000-40，000。 4. The conjugate according to claim 3, wherein the polyethylene glycol has a molecular weight of from 20,000 to 40,000.
5. 如权利要求 2-4任一项所述的结合物， 其中所述聚乙二醇为直链、 Y型支 链或多分支链聚乙二醇。 The conjugate according to any one of claims 2 to 4, wherein the polyethylene glycol is a linear, Y-branched or multi-branched polyethylene glycol.
6. 如权利要求 2所述的结合物， 其中所述聚乙二醇具有如下式（Π)所示的结 构， R1-0-^CH2CH20^ ~ The conjugate according to claim 2, wherein the polyethylene glycol has a structure represented by the following formula (Π), R 1 -0-^CH 2 CH 2 0^ ~
e 式 （Π)  e type (Π)
其中所述 为 _12垸基、 d_12杂垸基、 氢或芳基垸基， e为 10-1,500的整数。Wherein the base is _ 12 embankment, embankment D_ 12 heteroaryl group, or aryl alkyl with hydrogen, e is an integer of 10-1,500.
7. 如权利要求 1或 2任一项所述的结合物， 其中所述 P具有如下式 （III) 所 示的结构，
Figure imgf000071_0001
（m) 其中 R2、 R3各自独立的为 d_12垸基、 d_12杂垸基、 氢、 芳基垸基； The conjugate according to any one of claims 1 to 2, wherein the P has a structure represented by the following formula (III),
Figure imgf000071_0001
(m) wherein each of R 2 and R 3 is independently d— 12 fluorenyl, d— 12 heterofluorenyl, hydrogen, aryl fluorenyl;
、 R5各自独立的为 d_12垸基、 d_12垸基羰基； And R 5 is independently d_ 12 fluorenyl, d_ 12 fluorenylcarbonyl;
f、 g各自独立的为 10-1,500的整数。 f and g are each independently an integer of 10-1,500.
8. 如权利要求 7所述的结合物， 其中 R2、 R3为甲基， 为乙基， R5为亚甲基 8. The conjugate according to claim 7, wherein R 2 and R 3 are methyl, ethyl, and R 5 is methylene.
9. 如权利要求 1或 2任一项所述的结合物， 其中所述 P具有如下式 （IV) 所 示的结构， The conjugate according to any one of claims 1 to 2, wherein the P has a structure represented by the following formula (IV),
R6—- 0-^CH2CH20- h R 6 —- 0-^CH 2 CH 2 0- h
式 （IV) 其中， R6为季戊四醇， 甲基葡萄糖苷， 蔗糖， 二甘醇， 丙二醇， 甘油或聚甘油 的羟基去除氢的残基； i为 3， 4， 6或 8; h为 10-1,500的整数。 Wherein R 6 is a residue at which a hydroxyl group of pentaerythritol, methyl glucoside, sucrose, diethylene glycol, propylene glycol, glycerin or polyglycerol removes hydrogen; i is 3, 4, 6 or 8; h is 10- An integer of 1,500.
10. 如权利要求 1-9任一项所述的结合物， 其中 X为 CCH2)n， (CH2)nCO, (CH2)nOCO， （CH2)nNHCO， -S -， -S02-， -S04-; n为 1-12的整数。 The conjugate according to any one of claims 1 to 9, wherein X is CCH 2 ) n , (CH 2 ) n CO, (CH 2 ) n OCO, (CH 2 ) n NHCO, -S -, -S0 2 -, -S0 4 -; n is an integer from 1-12.
11. 如权利要求 10所述的结合物， 其中 X为 CH2CO。 11. The conjugate according to claim 10, wherein X is CH 2 CO.
12. 如权利要求 1-11任一项所述的结合物， 其中 为具有至少两个羧酸基和 一个氨基的氨基酸或氨基酸类似物的残基。 The conjugate according to any one of claims 1 to 11, which is a residue of an amino acid or an amino acid analog having at least two carboxylic acid groups and one amino group.
13. 如权利要求 12所述的结合物， 其中 A具有如下式 （V) 所示的结构，
Figure imgf000072_0001
（V) 其中， R7为 d_2。垸基、 Cwo杂垸基。 The conjugate according to claim 12, wherein A has a structure represented by the following formula (V),
Figure imgf000072_0001
(V) where R 7 is d_ 2 .垸 base, Cwo 垸 base.
14. 如权利要求 12所述的结合物， 其中 具有如下式 （VI) 所示的结构,
Figure imgf000072_0002
（vi) The conjugate according to claim 12, which has a structure represented by the following formula (VI),
Figure imgf000072_0002
(vi)
其中， R8、 R9独立的为氢、 d_12垸基、 d_12杂垸基、 芳基、 杂芳基、 芳垸基、 杂芳垸基、 且在每个重复单元中的 R8、 R9可以相同或不同； j为 1-10的整数。Wherein R 8 and R 9 are independently hydrogen, d_ 12 fluorenyl, d- 12 heterofluorenyl, aryl, heteroaryl, aryl fluorenyl, heteroaryl fluorenyl, and R 8 in each repeating unit, R 9 may be the same or different; j is an integer from 1 to 10.
15. 如权利要求 12所述的结合物， 其中 ^为天冬氨酸或谷氨酸的残基。 15. The conjugate according to claim 12, wherein ^ is a residue of aspartic acid or glutamic acid.
16. 如权利要求 1-15任一项所述的结合物，其中 A2、A3独立的具有如下式 (VII) 所示的结构，
Figure imgf000072_0003
（VII) 其中， R1()、 Ru独立的为氢、 d_6垸基、 d_6杂垸基且在每个重复单元中的 0、 可以相同或不同； h为 1-10的整数。 The conjugate according to any one of claims 1 to 15, wherein A 2 and A 3 independently have a structure represented by the following formula (VII),
Figure imgf000072_0003
(VII) wherein, R 1 (), R u is independently hydrogen, d_ 6 alkyl with, d_ 6 0 heteroaryl and alkyl with in each repeating unit may be the same or different; H is an integer of 1-10.
17. 如权利要求 16所述的结合物， 其中 A2、 A3独立的为甘氨酸、 丙氨酸、 亮 氨酸、 异亮氨酸、 缬氨酸、 脯氨酸、 苯丙氨酸、 蛋氨酸、 丝氨酸、 苏氨酸、 半 胱氨酸和酪氨酸的残基。 The conjugate according to claim 16, wherein A 2 and A 3 are independently glycine, alanine, leucine, isoleucine, valine, valine, phenylalanine, methionine , residues of serine, threonine, cysteine and tyrosine.
18. 如权利要求 17所述的结合物， 其中 A2、 A3为缬氨酸。 18. The conjugate according to claim 17, wherein A 2, A 3 is valine.
19. 如权利要求 1-18任一项所述的结合物， 其中 D B D2各自独立的为抗肿瘤 药物的残基。 19. The conjugate according to any one of claims 1 to 18, wherein DBD 2 is independently anti-tumor The residue of the drug.
20. 如权利要求 19所述的结合物， 其中所述抗肿瘤药物的残基能够与八2或八3 形成肽键或酯键。 20. The conjugate according to claim 19, wherein said antineoplastic residue capable of eight or eight 2 3 forming a peptide bond or an ester bond.
21. 如权利要求 19所述的结合物，其中所述抗肿瘤药物为达沙替尼、雷帕霉素、 依诺替康、 伊马替尼、 厄洛替尼、 吉非替尼、 拉帕提尼、 索拉非尼、 舒尼替尼、 紫杉醇、 喜树碱、 华蟾酥毒基、 甘草次酸、 东莨菪内酯。  21. The conjugate of claim 19, wherein the anti-tumor drug is dasatinib, rapamycin, inonotecan, imatinib, erlotinib, gefitinib, pull Patini, sorafenib, sunitinib, paclitaxel, camptothecin, scutellaria, glycyrrhetinic acid, and terpene lactone.
22. 如权利要求 21所述的结合物， 其中所述抗肿瘤药物为达沙替尼。  22. The conjugate of claim 21, wherein the anti-tumor drug is dasatinib.
23. 如权利要求 1-22任一项所述的结合物， 其中所述结合物具有如下式（VIII) 所示  The conjugate according to any one of claims 1 to 22, wherein the conjugate has the following formula (VIII)
Figure imgf000073_0001
式 （vm) 其中， R7为 Cwo垸基、 Cwo杂垸基， R1Q、 Ru独立的为氢、 d_6垸基且在每个 重复单元中的 R1Q、 Ru可以相同或不同； h为 1-10的整数。
Figure imgf000073_0001
Wherein R 7 is Cwo fluorenyl, Cwo heteroindolyl, R 1Q , R u are independently hydrogen, d 6 fluorenyl and R 1Q , R u in each repeating unit may be the same or different; h is an integer from 1 to 10.
24. 如权利要求 23所述的结合物， 其中所述结合物具有如下式（IX) 、 (X) (XI) 所示的结构， The conjugate according to claim 23, wherein the conjugate has a structure represented by the following formulas (IX), (X) (XI),
式 （X) Formula (X)
Figure imgf000074_0001
式 （XI) 其中， e、 f、 g各自独立的为 10-1,500的整数。
Figure imgf000074_0001
Wherein e, f, and g are each independently an integer of from 10 to 1,500.
25. 如权利要求 24所述的结合物， 其中 为达沙替尼。  25. The conjugate of claim 24, wherein is dasatinib.
26. 包含如权利要求 1-25任一项所述的结合物以及药学上可接受的载体或赋形 剂的药物组合物。  26. A pharmaceutical composition comprising a conjugate according to any of claims 1-25 and a pharmaceutically acceptable carrier or excipient.
27. 如权利要求 26所述的药物组合物， 其中所述药物组合物为片剂、 胶囊剂、 丸剂、 颗粒剂、 散剂、 栓剂、 注射剂、 溶液剂、 混悬剂、 膏剂、 贴剂、 洗剂、 滴剂、 擦剂、 喷雾剂。  The pharmaceutical composition according to claim 26, wherein the pharmaceutical composition is a tablet, a capsule, a pill, a granule, a powder, a suppository, an injection, a solution, a suspension, a paste, a patch, and a wash. Agents, drops, liniments, sprays.
28. 如权利要求 1-27任一项所述的结合物和 /或药物组合物在制备抗肿瘤、真菌 感染、 类风湿性关节炎、 多发性硬化症、 心瓣再狭窄或肺炎的药物中的应用。 28. The conjugate and/or pharmaceutical composition according to any one of claims 1 to 27 for the preparation of a medicament for antitumor, fungal infection, rheumatoid arthritis, multiple sclerosis, heart stenosis or pneumonia Applications.
29. 如权利要求 28所述的应用，其特征在于，所述抗肿瘤药物应用于以下病症： 白血病、 急性粒细胞性白血病、 慢性粒细胞性白血病、 慢性淋巴性白血病、 急 性淋巴性白血病、脊髓发育不良、多发性骨髓瘤、何杰金氏病或非何杰金氏病、 小细胞或非小细胞性肺癌、 胃癌、 肠癌、 食道癌、 结肠直肠癌、 前列腺癌、 卵 巢癌、 乳腺癌、 脑癌、 泌尿道癌、 肾癌、 膀胱癌、 恶性黑素瘤、 肝癌、 子宫癌、 胰腺癌、 骨髓瘤癌、 子宫内膜癌、 头颈癌、 小儿肿瘤、 肉瘤。 29. The use according to claim 28, wherein the antitumor drug is applied to the following conditions: leukemia, acute myeloid leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia, acute lymphocytic leukemia, spinal cord Dysplasia, multiple myeloma, Hodgkin's disease or non-Hodgkin's disease, small cell or non-small cell lung cancer, gastric cancer, colon cancer, esophageal cancer, colorectal cancer, prostate cancer, egg Nest cancer, breast cancer, brain cancer, urinary tract cancer, kidney cancer, bladder cancer, malignant melanoma, liver cancer, uterine cancer, pancreatic cancer, myeloma, endometrial cancer, head and neck cancer, pediatric tumor, sarcoma.
30. 一种治疗主体中肿瘤、 真菌感染、 类风湿性关节炎、 多发性硬化症、 心瓣 再狭窄或肺炎的方法，包括向所述主体给与有效量的权利要求 1-26任一项所述 的结合物或药物组合物。  30. A method of treating a tumor, a fungal infection, rheumatoid arthritis, multiple sclerosis, heart valve restenosis or pneumonia in a subject, comprising administering to the subject an effective amount of any of claims 1-26 Said conjugate or pharmaceutical composition.
31. 如权利要求 30所述的方法， 其中所述主体是哺乳动物。  31. The method of claim 30, wherein the subject is a mammal.
32. 如权利要求 31所述的方法， 其中所述主体是人。  32. The method of claim 31, wherein the subject is a person.
33. 如权利要求 30所述的方法， 其中给药方式包括口腔、 粘膜、 舌下、 眼部、 局部、 肠道外、 直肠、 脑池、 阴道、 腹膜、 膀胱、 鼻部给药。  33. The method of claim 30, wherein the mode of administration comprises oral, mucosal, sublingual, ocular, topical, parenteral, rectal, cerebral, vaginal, peritoneal, bladder, nasal administration.
34. 一种制备如权利要求 1所述的结合物， 包括：  34. A conjugate according to claim 1 comprising:
使药物化合物与氨基酸发生酯化或酰胺化反应形成结合物 Di-As和 /或 D2-A3 ; 使结合物 Di-As和 /或 D2-A3
Figure imgf000075_0001
Esterification or amidation of a drug compound with an amino acid to form a conjugate Di-As and/or D 2 -A 3 ; a conjugate of Di-As and/or D 2 -A 3
Figure imgf000075_0001
所示化合物。 The compound shown.
PCT/CN2014/071395 2013-01-28 2014-01-24 Water soluble polymer-amino acid oligopeptide-medicine combination, preparation method therefore, and use thereof WO2014114262A1 (en)

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Citations (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1283643A (en) * 2000-07-05 2001-02-14 天津大学 Precursor of polyglycol carried taxusol or polyene taxusol
US6566506B2 (en) * 1993-10-27 2003-05-20 Enzon, Inc. Non-antigenic branched polymer conjugates
CN1442440A (en) * 2002-03-05 2003-09-17 北京键凯科技有限公司 Binding agent of hydrophilic polymer-glutamic acid oligopeptide and medicinal molecular, composition containing said binding agent and use
CN1556828A (en) * 2002-03-13 2004-12-22 ƽ Hydrophili polymer derirative having Y shaped branch, its preparation method, combined object with medicine molecule and medical compositon containing combined object
CN1611524A (en) * 2003-10-28 2005-05-04 北京键凯科技有限公司 Polyethylene glycol amino acid N-internal ring carbonyl anhydride active derivatives, and medicinal bonding compound and gel thereof
CN1706865A (en) * 2004-06-11 2005-12-14 北京键凯科技有限公司 Branched polyglycol-amino acid oligopeptide and its active derivative and medicinal composition
CN1756758A (en) * 2003-12-02 2006-04-05 北京键凯科技有限公司 Novel cantharidimide and go first cantharidimide derivative and the application in medicine thereof
CN1840201A (en) * 2006-01-20 2006-10-04 中国药科大学 Polyethylene glycol prodrug of gambogicacid, its preparation method, formulation and use
CN101104078A (en) * 2006-07-11 2008-01-16 北京美倍他药物研究有限公司 Polypeptide and protein medicine polyethylene coupling compound
CN102145178A (en) * 2011-04-15 2011-08-10 北京凯因科技股份有限公司 PEG interleukin 15
CN102766258A (en) * 2011-05-03 2012-11-07 山东靶点药物研究有限公司 Resveratrol hydrophilic conjugate, its preparation method and application

Patent Citations (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6566506B2 (en) * 1993-10-27 2003-05-20 Enzon, Inc. Non-antigenic branched polymer conjugates
CN1283643A (en) * 2000-07-05 2001-02-14 天津大学 Precursor of polyglycol carried taxusol or polyene taxusol
CN1442440A (en) * 2002-03-05 2003-09-17 北京键凯科技有限公司 Binding agent of hydrophilic polymer-glutamic acid oligopeptide and medicinal molecular, composition containing said binding agent and use
CN1556828A (en) * 2002-03-13 2004-12-22 ƽ Hydrophili polymer derirative having Y shaped branch, its preparation method, combined object with medicine molecule and medical compositon containing combined object
CN1611524A (en) * 2003-10-28 2005-05-04 北京键凯科技有限公司 Polyethylene glycol amino acid N-internal ring carbonyl anhydride active derivatives, and medicinal bonding compound and gel thereof
CN1756758A (en) * 2003-12-02 2006-04-05 北京键凯科技有限公司 Novel cantharidimide and go first cantharidimide derivative and the application in medicine thereof
CN1706865A (en) * 2004-06-11 2005-12-14 北京键凯科技有限公司 Branched polyglycol-amino acid oligopeptide and its active derivative and medicinal composition
CN1840201A (en) * 2006-01-20 2006-10-04 中国药科大学 Polyethylene glycol prodrug of gambogicacid, its preparation method, formulation and use
CN101104078A (en) * 2006-07-11 2008-01-16 北京美倍他药物研究有限公司 Polypeptide and protein medicine polyethylene coupling compound
CN102145178A (en) * 2011-04-15 2011-08-10 北京凯因科技股份有限公司 PEG interleukin 15
CN102766258A (en) * 2011-05-03 2012-11-07 山东靶点药物研究有限公司 Resveratrol hydrophilic conjugate, its preparation method and application

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