JPH04213308A - Copolymer of propenamide derivative with anionic monomer and its use - Google Patents

Abstract

PURPOSE:To obtain a copolymer which is water-soluble and is useful as a metastasis inhibitor, a wound healing agent, an animal cell adhesion inhibitor, a platelet aggregation inhibitor or a substrate for cell culture by copolymerizing a specified propenamide derivative with a specified anionic monomer. CONSTITUTION:A copolymer of a propenamide derivative having an adhesive peptide of formula II [wherein X, Y and Z are each a residue of an amino acid such as Ser, Gly or Val or a peptide residue; Z is -O- or -NH-; one of R<4> and R<5> is H, and the other is 1-11C (substituted) alkylene or 6-11C (substituted) arylene; and n is 1-5] at the side chain of formula I (wherein R<1> and R<2> are each H or COOH; R<3> is H, CH3, C2H5, halogen or -CH2COOH) with an anionic monomer of formula III [wherein R<6> is H or 1-3C (substituted) alkyl; W is -O- or -NH-; R<7> is H, 1-12C alkyl or 6-11C aryl which may be substituted with COOH, sulfo or phosphoro and may be further substituted] or its salt.

Description

Detailed Description of the Invention [0001] [Industrial Application Field] The present invention relates to Arg-Gly-As
A novel anionic propenamide derivative copolymer having p tripeptide as an essential unit and its salt, and an animal cell adhesion inhibitor, platelet aggregation/adhesion inhibitor, and Arg-Gly-Asp containing the same as an active ingredient. The present invention relates to an anionic propenamide derivative crosslinked copolymer having a tripeptide as an essential unit, a salt thereof, and an animal cell culture substrate containing the same as an active ingredient. [0002] Fibronectin is a protein involved in cell-extracellular matrix adhesion, and is also thought to be involved in platelet aggregation and cancer metastasis. These interactions are mediated by a series of cell surface receptors, and although fibronectin is a large molecule with a molecular weight of approximately 250,000, these receptors
It was revealed that it specifically recognizes the Asp sequence,
It has been reported that it is important for interaction with receptors (Nature, No. 309).
Vol. 30, 1984). Since then, research has been carried out using oligos or polypeptides having the Arg-Gly-Asp sequence. For example, methods for inhibiting platelet aggregation using various linear and cyclic oligopeptides having the Arg-Gly-Asp sequence (Polymer Science Society of Japan Proceedings)
mer Preprints, Japan), No. 38
Volume, 3149 pages, 1989, JP-A-2-174797
No.), a method of using a peptide having the Arg-Gly-Asp sequence as a cell migration inhibitor (Japanese Patent Laid-Open No. 2-47
No. 16), P with immobilized Arg-Gly-Asp
Method of using MMA membrane as a cell adhesion membrane (Polymer Preprints, Jap
an), Vol. 37, p. 705, 1988). Furthermore, Arg-Gly-As in the polymer
A method of covalently bonding peptides having p as an essential constituent unit and using them as animal cell culture substrates and biocomposite artificial organ substrates (JP-A-1-309682, JP-A-1-30596)
0), a method of using a polypeptide having the Arg-Gly-Asp-Ser sequence as a platelet protective agent for extracorporeal blood has been disclosed (Japanese Patent Application Laid-Open No. 64-6217).
Furthermore, a method of suppressing cancer metastasis using an oligopeptide having an Arg-Gly-Asp sequence or a polypeptide having a repeating structure thereof is known ((Int. J. Biol. Macromol.
), vol. 11, p. 23, 1989, same magazine, vol. 11,
226 pages, 1989, (Jpn. J. Cance
rRes. ) Vol. 60, p. 722, 1989)
. [0004] In general, polymeric substances have diverse properties and functions, and their interactions with living organisms are very different from those of low-molecular substances. Therefore, research is actively being conducted to suppress the interaction of these compounds with cells and their behavior in vivo by binding small-molecule drugs, bioactive peptides, etc. to polymers. The application of polymers to the life sciences has been widely attempted in such areas as medical polymer materials, polymer drugs, diagnostic materials, bioreactors, and bioaffinity materials. is co-edited by Kiichi Takemoto, Junzo Sunamoto, Mitsuru Akashi “Polymer and Medical” (Mita Publishing), “Biotechnology Series Immobilized Enzyme” (edited by Ichiro Chibata)
It is described in detail in "Affinity Chromatography" (Kodansha), edited by Makoto Yamazaki, Shinichi Ishii, and Koichi Iwai. Propenamide derivatives synthesized from propenoic acid derivatives have vinyl groups and can easily form copolymers with various anionic vinyl monomers through vinyl polymerization, and can be used in various materials. I can do it. There are examples in which oligopeptides having Arg-Gly-Asp as an essential unit are introduced into water-insoluble vinyl polymers through polymer reactions; Water-soluble anionic copolymers and hydrogels of polymerizable propenamide derivatives are not known, and these can be expected to enhance binding ability with receptors and stabilize in blood. [0006] An object of the present invention is to solve the problem of Ar
A novel anionic propenamide derivative copolymer having g-Gly-Asp tripeptide as an essential unit and its salt, and an animal cell adhesion inhibitor, platelet aggregation/adhesion inhibitor containing the same as an active ingredient, and Arg -Gl
The object of the present invention is to provide a novel anionic propenamide derivative crosslinked copolymer having a tripeptide of y-Asp as an essential unit, a salt thereof, and an animal cell culture substrate containing the same as an active ingredient. [Means for Solving the Problems] The present invention provides the following general formula [
A copolymer of a propenamide derivative having an adhesive peptide represented by the following general formula [II] as an essential unit in the side chain of [I] and an anionic monomer represented by the following general formula [III], or a copolymer thereof It provides salt. General formula [I] R1R2C =CR3 -CO-[NH]- In the formula, R1
, R2 represents a hydrogen atom or a carboxyl group, R3
represents a hydrogen atom, a methyl group, an ethyl group, a halogen atom or a carboxymethyl group. General formula [II] -[R4]-[CO]-([X]-Arg-Gly-A
sp-[Y])n-[Z]-[R5]- where X, Y are Ser, Gly, Val, Asn
, Pro or a peptide residue consisting of these amino acids, and Z represents -O- or -NH-. Either one of R4 and R5 represents a hydrogen atom, and the other represents a linear or branched alkylene group having 1 to 11 carbon atoms, or an arylene group having 6 to 11 carbon atoms, even if it has a substituent. good. n represents an integer of 1 to 5. Examples of substituents include carbonyl groups, carboxyl groups, amino groups, hydroxyl groups, sulfo groups, halogen atoms, aryl groups, nitro groups, cyano groups, and double bonds and triple bonds of unsaturated groups. , two or more may be present in the same chain. General formula [III] H2C=CR6 -[CO]-[W]-R7 In the formula, R6
may be a hydrogen atom or an alkyl group having 1 to 3 carbon atoms, and may have a substituent. W represents -O- or -NH-. R7 is a hydrogen atom, a linear or branched alkyl group having 1 to 12 carbon atoms, or an aryl group having 6 to 11 carbon atoms, and at least one of a carboxyl group, a sulfo group, and a phosphoro group is used as a substituent. and may further have a substituent. Examples of substituents include carbonyl groups, carboxyl groups, amino groups, hydroxyl groups, sulfo groups, halogen atoms, aryl groups, nitro groups, cyano groups, and double bonds and triple bonds of unsaturated groups. , two or more may be present in the same chain. Further, the same chain may have two or more amide bonds, ester bonds, ether bonds, urea bonds, carbamate ester bonds, carbonate ester bonds, etc., and may further have a heterocycle. [0013] In the general formulas [I], [II], and [III], [ ] indicates that the group within [ ] may or may not exist. The present invention further provides an animal cell adhesion inhibitor and a platelet aggregation/adhesion inhibitor containing the above-mentioned anionic propenamide derivative copolymer and its salt as an active ingredient. The content of the structural unit derived from the propenamide derivative in the copolymer is preferably 0.1 to 90 mol%, more preferably 0.5 to 60 mol%. The molecular weight of the anionic propenamide derivative copolymer and its salt is preferably 300,000 or less, particularly 300,000 or less.
000-200,000 and is preferably water-soluble at room temperature. The present invention also provides Arg-Gly-Asp
The object of the present invention is to provide an anionic crosslinked copolymer of a propenamide derivative formed by covalently bonding a peptide sequence as an essential unit, and a salt thereof. The crosslinked copolymer is preferably in the form of a hydrogel in an aqueous solution. When synthesizing a crosslinked copolymer, a polyfunctional monomer is added in addition to the propenamide derivative and anionic monomer. Examples of polyfunctional monomers include dimethacrylates such as triethylene glycol dimethacrylate and methylene bisacrylamide, diacrylates, dimethacrylamide, polyfunctional monomers having an aromatic ring such as diacrylamide, and divinylbenzene, and cell-adhesive monomers. A polyfunctional monomer having fragments, etc. can be used. In the crosslinked copolymer, the content of structural units derived from propenamide derivatives is preferably 0.1 to 9.
The content of the structural unit derived from the polyfunctional monomer is preferably 0.1 to 30 mol%, more preferably 0.5 to 60 mol%.
It is 20 mol%. The present invention further provides an animal cell culture substrate containing this crosslinked copolymer or a salt thereof as an active ingredient. [0017] The amino acids used in the adhesive peptide according to the present invention may be either L-form or D-form, but L-form is preferable. The anionic monomer that can be used in the present invention is not particularly limited as long as it is a conventional polymerizable vinyl monomer. For example, acrylic acid, methacrylic acid, itaconic acid, maleic acid, fumaric acid, crotonic acid, 2-acrylamidoglycolic acid, 2-methacrylamidoglycolic acid, styrenesulfonic acid, vinylsulfonic acid, 2-methacrylamidoglycolic acid,
Acrylamide-2-methylpropanesulfonic acid amide, vinyl monomers with carboxyl groups and sulfonic acid groups such as 4-vinylbenzoic acid, N-methacryloyl of 5-aminovaleric acid, 6-aminocaproic acid, and 12-aminolauric acid derivatives, N-acryloyl derivatives, and the like. Furthermore, N-methacryloylglycine, N
-methacryloyl-β-alanine, N-methacryloylalanine, N-methacryloyl-γ-aminobutyric acid, N-
Methacryloylvaline, N-methacryloylleucine,
N-methacryloylisoleucine, N-methacryloylnorvaline, N-methacryloylnorleucine, N-methacryloylserine, N-methacryloylthreonine,
N-methacryloylmethionine, N-methacryloylphenylalanine, N-methacryloyltyrosine, N-α
-methacryloyltryptophan, N-methacryloylproline, N-methacryloylhydroxyproline, N
-methacryloyl aspartic acid, N-methacryloyl asparagine, N-methacryloyl glutamic acid, N-
Methacryloylglutamine, N-α-methacryloylarginine, N-α-methacryloylcitrulline, N-acryloylglycine, N-acryloyl-β-alanine, N-acryloylalanine, N-acryloyl-γ-
Aminobutyric acid, N-acryloylvaline, N-acryloylleucine, N-acryloylisoleucine, N-acryloylnorvaline, N-acryloylnorleucine,
N-acryloylserine, N-acryloylthreonine, N-acryloylmethionine, N-acryloylphenylalanine, N-acryloyltyrosine, N-α-acryloyltryptophan, N-acryloylproline, N-acryloylhydroxyproline, N-acryloylaspartic acid, N -acryloyl asparagine,
Examples include vinyl monomers having amino acid residues such as N-acryloylglutamic acid, N-acryloylglutamine, N-α-acryloyl arginine, and N-α-acryloyl citrulline. [0021] Vinyl monomers having a phosphoric acid group in the side chain, such as methacryloyloxyethyl phosphoric acid, methacryloyloxyethyl phenyl phosphoric acid, and methacryloyloxydecanoyl phosphoric acid, can also be used. Preferably glycine, β-alanine, 4
-Aminobutyric acid, 5-aminovaleric acid, 6-aminocaproic acid, 12-aminolauric acid, 4-aminobenzoic acid, leucine, N-methacryloyl derivatives of glutamic acid, and N-acryloyl derivatives, leucine, N-methacryloyl derivatives of glutamic acid , and N-acryloyl derivatives, as well as acrylic acid, methacrylic acid, itaconic acid, and maleic acid. Examples of the salts of the anionic propenamide derivative copolymer of the present invention include hydrochloride, sulfate, nitrate,
Examples include salts with inorganic acids such as phosphates and borates, and salts with organic acids such as acetates, trifluoroacetates, trifluoromethanesulfonates, lactates, and tartrates. The conversion to can be performed by conventional means. [0024] The peptide synthesis method is not particularly limited, and examples thereof include liquid phase method, solid phase method, and synthesis method using an automatic synthesizer. For details on these synthesis methods, please refer to the Biochemistry Experiment Course “Protein Chemistry IV” p.207.
-495 (edited by the Japanese Biochemical Society, Tokyo Kagaku Doujin), “Protein Chemistry Experiment Course (Part 2)” (edited by the Japanese Biochemical Society, Tokyo Kagaku Doujin), “Basics and Experiments of Peptide Synthesis” (edited by Izumiya et al.) (Maruzen )It is described in. It is also possible to use commercially available synthetic peptides. [0025] Examples of methods for bonding propenoic acid derivatives, aminoalkyl carboxylic acids, and cell adhesive peptides include active ester method, mixed acid anhydride method, azide method, acid chloride method, symmetric acid anhydride method, DCC method, and DCC method. -Amide bond synthesis methods using additive methods, carbonyldiimidazole methods, etc. can be mentioned. Copolymers and crosslinked copolymers of propene derivatives and anionic monomers can be obtained by general radical polymerization methods and ionic polymerization methods. A water-soluble polymer can be subjected to specific molecular weight fractionation by gel filtration, dialysis, or the like. Crosslinked copolymers can be obtained as hypergels with different physical properties such as water content and gel strength by changing the composition of the polyfunctional monomer. The anionic propenamide derivative copolymer and its salt of the present invention have the core sequence Arg-Gly-Asp of a cell adhesion protein, and through this core sequence, a similar mechanism as that of the cell adhesion protein is produced. to adhere to cells. Therefore, it exhibits various biological activities as an agonist or antagonist of cell adhesion proteins, and has a wide range of biological activities such as immunomodulation, wound healing, inhibition of platelet aggregation by cancer cells that occur in capillaries, and healing of neurological diseases. It recognized. [0027] Therefore, the anionic propenamide derivative copolymer and its salt of the present invention can be used as a cancer metastasis inhibitor, a wound healing agent, or an immunomodulator, in combination with at least one of them, optionally with a conventional carrier or pharmaceutical auxiliary agent. , can be administered to patients as a platelet aggregation and adhesion inhibitor. especially,
Use as an animal cell adhesion inhibitor or platelet aggregation adhesion inhibitor is preferred. The dose is 0.2 μg/kg to 4
00 mg/kg, determined based on symptoms, age, body weight, etc. The anionic propenamide derivative copolymer and its salt of the present invention can be administered by the administration method generally used for peptide drugs, that is, by parenteral administration, such as intravenous administration, intramuscular administration, subcutaneous administration, etc. Preferably, it is administered. When producing such an injectable preparation, the anionic propenamide derivative copolymer of the present invention and its salt may be dissolved in PBS or physiological saline as shown in the examples below, and the injectable preparation may be prepared. Alternatively, it may be dissolved in approximately 0.1N acetic acid water, etc., and then lyophilized. Conventional stabilizers such as glycine and albumin may be added to such formulations. Furthermore, if the anionic propenamide derivative copolymer and its salt of the present invention are formed into microcapsules, microspheres, or hydrogels encapsulated in liposomes, for example,
It can also be administered orally, and if it is in the form of suppositories, sublingual tablets, nasal sprays, etc., it can be absorbed through mucous membranes other than the gastrointestinal tract. [0029] The method of using the cell culture substrate in culturing animal cells is not particularly limited and may be carried out by a conventional method. For example, a method in which animal cells are suspended in a bead culture solution treated with covalently bonded adhesive peptides and stirred at low speed to allow animal cells to adhere to the surface of microcarriers and cultured, and a Petri dish roller coated with adhesive peptides that are covalently bonded. A method of culturing animal cells on a bottle, etc., a method of circulating a culture medium through a hollow fiber that has been covalently bonded with an adhesive peptide, and culturing the animal cells by adhering to the inner surface of the hollow fiber, and a method of culturing animal cells that have been covalently bonded with an adhesive peptide. Examples include a method using a column filled with a carrier. [0030] The cell culture substrate of the present invention can be used for culturing various types of cells, and the types of cells are not particularly limited, and include cells derived from living organisms, hybridomas, and the like. [Examples] The present invention will be further explained below with reference to Examples, but the present invention is not limited thereto. Production Example 1 Carboxyethyl methacrylamide was synthesized by Schotten-Baumann reaction. That is, 17.8 g of β-alanine
20.9 g (0.2 mol) of methacrylic acid chloride was added dropwise to (0.2 mol) of an aqueous sodium hydroxide solution under ice cooling, and after stirring for 4 hours, the mixture was neutralized with hydrochloric acid. Concentrate under reduced pressure,
The precipitated sodium chloride was filtered off. The concentrated solution was extracted with chloroform, dried, and concentrated under reduced pressure to remove chloroform. The concentrate was washed with ether to obtain 17.6 g of Product 1 as a white solid. (Yield 56%) Product 1 CH2=CCH3-CO-NH-C2H4-COOH [
Production Examples 2 to 8 Acrylic acid chloride, methacrylic acid chloride, ethacrylic acid chloride and 4-aminobutyric acid, 5-aminovaleric acid, 6-aminocaproic acid, 12-aminobutyric acid were prepared by the same method as in Production Example 1.
The following propenoic acid derivatives were produced by reaction with aminolauric acid, leucine, glutamine, p-aminobenzoic acid, glycine, and glutamic acid. Product 2 CH2=CH-CO-NH-(CH2)3-COOH yield 52% Product 3 CH2=CC2H5 -CO-NH-(CH2)4-C
OOH yield 61% Product 4 CH2=CCH3-CO-NH-(CH2)5-COO
H yield 69% Product 5 CH2=CH-CO-NH-(CH2)11 -COO
H yield 71% Product 6 CH2=CH-CO-NH-CH(CH2CH(CH3
)2) -COOH yield 64% Product 7 CH2=CCH3-CO-NH-CH(C2H4CON
H2) -COOH Yield 59% Product 8 CH2=CH-CO-NH-p -C6H4-COOH
Yield 68% Product 9 CH2=CCH3-CO-NH-CH2 -COOHYield 63% Product 10 CH2=CCH3-CO-NH-CH(C2H4COO
H)-COOH yield: 57% Production Example 11 Aminoethyl methacrylamide was synthesized by Schotten-Baumann reaction. That is, 120 g of ethylenediamine (
Chloroform solution (400ml) containing 2mol)
20.9g (0.2mol) of methacrylic acid chloride
was added dropwise under ice-cooling, and after stirring for 4 hours, the mixture was concentrated under reduced pressure to remove chloroform. 50 ml of a 5% aqueous sodium hydrogen carbonate solution was added to the concentrate, and the mixture was extracted with chloroform. After drying over sodium sulfate, the residue was concentrated and purified by alumina column chromatography using chloroform/methanol=7/3 as an eluent. (Product 11) CH2=CCH3-CO-NH-C2H4-NH2 Yield 14.8g (57.8%, 0.116mol) 00
35 Production Example 12 Carboxyethyl methacrylamide synthesized in Production Example 1 was polymerized by radical polymerization. 2 g of carboxyethyl methacrylamide was dissolved in 20 ml of DMF, and radical initiator V65 (2,2-azobis(2,
10 mg of 4-dimethylvaleronitrile) was added and polymerized at 65° C. for 4 hours under a nitrogen stream. After precipitating the polymer with ethyl acetate, it was precipitated using Spectrapore 7 (molecular fraction 3000).
After removing the low molecular weight fraction by dialysis against pure water using
Lyophilized. Yield 1.24g (product 12) 003
6] Molecular weight is TSKgel G300 manufactured by Tosoh Corporation.
Measurement was carried out using an 0SW column with a mobile phase of 0.2M phosphate buffer (pH 7.4) at a flow rate of 1.0 ml/min. The molecular weight in terms of PEG was about 30,000. Production Example 13 Carboxyethyl methacrylamide synthesized in Production Example 1 was polymerized by radical polymerization to create a hydrogel. [0038] Silane-treated glass plate (5 cm x 6 cm
×1cm) and a gasket were prepared. Dissolve 2 g of carboxyethyl methacrylamide and 100 mg (5 wt%) of methylene bisacrylamide in 12 ml of distilled water.
The pH was adjusted to 7.4 with N NaOH. After adding 10 mg of ammonium persulfate to the solution and thoroughly purging with nitrogen, the solution was injected between glass plates. Hold the glass plate in a vise at 60℃.
Polymerization was carried out for 20 hours. The generated hydrogel was washed with distilled water to remove unreacted monomers. (Sterilized using an ultraviolet lamp and then subjected to cell culture experiments) (Product 13) Synthesis Examples 1 to 14 Solid-phase synthesis of adhesive peptides Merrifiel
Synthesis was performed using a peptide synthesizer using the d method. For the protection of α-amino acids, Arg-G
An oligopeptide containing ly-Asp as an essential unit was synthesized, and the propenoic acid derivative shown in Production Example 1-8 and acrylic acid, methacrylic acid, and ethacrylic acid were condensed to the terminal thereof. Trifluoromethanesulfonic acid was used to cleave the resin and remove the side chain protecting groups, resulting in preparative H
Purification was performed by PLC (high performance liquid chromatography) to obtain a propenamide derivative exhibiting a single peak. This was applied to an anion exchange resin column (Amberlite IRA-400
; Cl form) to give the hydrochloride. Hereinafter, Arg will be referred to as R, Gly as G, Asp as D, Ser as S, and Pro as P. Furthermore, the abbreviations of protecting groups and reagents are as follows. Boc: t-butoxycarbonyl OB
zl: benzyl ester HOBt
:Hydroxybenzotriazole OSu :
N-Hydroxysuccinimide ONb: Nitrobenzyl ester TFA: Trifluoroacetic acid DCC: Dicyclohexylcarbodiimide DC urea: Cyclohexylurea M
ts: mesitylenesulfonyl DMF
:dimethylformamide [0042] [0043] [0044] [0045] [0046] [0047] [0048] [0049] [0050] Synthesis 9 CH2=CH-CO-NH-CH(
CH2CH(CH3)2) -CO-RGDS
(SEQ ID NO: 9) Yield 31% Amino acid analysis (nmol/50 μl)
R:0.9814
G:1.0519
D:0.9731S
:0.8989 Leucine:0.985
3 Mass spectrum M+: 601 005
1] Compound 10 CH2=CCH3-CO-NH-C
H(C2H4CONH2) -CO-RGDS
(SEQ ID NO: 10) Yield 33% Amino acid analysis (nmol/50 μl)
R:0.9771
G:1.0501
D:0.9651S
: 0.8969 Glutamic acid: 0.9587 Mass spectrum M+ : 630 0052
] Synthesis Example 15 Synthesis 15 CH2=CCH3-CO-NH-C2H4-CO-RG
DS (SEQ ID NO: 15) Compound 15 was synthesized by a liquid phase method using a sequential extension method. (1) Synthesis Bo of BocSer(Bzl)OBzl
cSer(Bzl) 60g (0.2mol) 400
ml of ethyl acetate plus 21 ml of triethylamine
g (0.2 mol), benzyl bromide 35.4 g (0.2
mol) was added and reacted under reflux for 4 hours. After cooling, the salt was filtered off and washed with an aqueous NaHCO3 solution and an aqueous NaCl solution. This was dried over Na2SO4 and then dried under reduced pressure to obtain 54 g of white powder (yield 68%). (2) BocAsp(OBzl)Ser
Synthesis of (Bzl)OBzlBocSer(Bzl)OB
zl 30g (78mmol) of TFA/CH2Cl2
= 1/1 After adding 200 ml and stirring at room temperature for 1 hour, TFA and CH2Cl2 were concentrated under reduced pressure. This was dissolved in ethyl acetate, neutralized with NaHCO3 aqueous solution, and then NaCl
Washed with aqueous solution. After drying with Na2SO4, ethyl acetate was distilled off under reduced pressure. This compound and BocAsp(OBzl)
32.8g (78mmol) of OSu in CH2Cl25
00ml and stirred overnight. After CH2Cl2 was distilled off under reduced pressure, the residue was dissolved in ethyl acetate. NaHCO3 aqueous solution, 1M citric acid aqueous solution, NaC
1 aqueous solution, dried over Na2SO4, and evaporated to dryness to obtain 41 g of white powder (yield: 89%). (3) BocGlyAsp(OBzl)
Synthesis of Ser(Bzl)OBzl BocAsp(OB
zl)Ser(Bzl)OBzl 35g(59mm
After adding TFA:CH2Cl2=1:1200 ml to the mixture and stirring at room temperature for 1 hour, TFA and CH2Cl2 were concentrated under reduced pressure. This was dissolved in ethyl acetate, neutralized with an aqueous NaHCO3 solution, and then washed with an aqueous NaCl solution. Na2S
After drying with O4, ethyl acetate was removed under reduced pressure. [0061] This compound and 9.8 g of BocGly (
59 mmol) was dissolved in CH2Cl2 and DCC12.
After adding 2 g (59 mmol) under ice-cooling and stirring for 3 hours, the mixture was further stirred at room temperature overnight. DCurea was filtered off, concentrated under reduced pressure, and dissolved in ethyl acetate. It was washed with an aqueous NaHCO3 solution, a 1M aqueous citric acid solution, and an aqueous NaCl solution, dried over Na2SO4, and then dried under reduced pressure to obtain 30.5 g of a white powder (yield 75%). (4) BocArg(Mts)GlyA
Synthesis of sp(OBzl)Ser(Bzl)OBzlBocGlyAsp(OBzl)Ser(Bzl)OB
zl 25g (39mmol) of TFA:CH2Cl2
= 1: After adding 1200 ml and stirring at room temperature for 1 hour, T
FA and CH2Cl2 were concentrated under reduced pressure. This was dissolved in ethyl acetate, neutralized with an aqueous NaHCO3 solution, and then washed with an aqueous NaCl solution. After drying with Na2SO4, ethyl acetate was distilled off under reduced pressure. This compound and BocArg(Mts) 1
7.8g (39mmol) was dissolved in DMF400ml, DCC8.0g (39mmol), HOBt6.
8 g (45 mmol) was added under ice cooling, stirred for 3 hours, and then further stirred at room temperature overnight. DCurea was filtered off, concentrated under reduced pressure, and dissolved in ethyl acetate. It was washed with an aqueous NaHCO3 solution, a 1M aqueous citric acid solution, and an aqueous NaCl solution, dried over Na2SO4, and then dried under reduced pressure to obtain 19.5 g of a white powder (yield 50%). (5) Synthesis of compound 15 BocArg(
Mts)GlyAsp(OBzl)Ser(Bzl)O
TFA:CH2C to 15.0g (15mmol) of Bzl
After adding 100 ml of l2=1:1 and stirring at room temperature for 1 hour, TFA and CH2Cl2 were concentrated under reduced pressure. This was dissolved in ethyl acetate, neutralized with an aqueous NaHCO3 solution, and then washed with an aqueous NaCl solution. After drying with Na2SO4, ethyl acetate was distilled off under reduced pressure. This compound and 2.4 g (15 mmol) of carboxyethylmethacrylamide were mixed in CH2Cl2200
3.1 g (15 mmol) of DCC was added under ice-cooling, stirred for 3 hours, and further stirred overnight at room temperature. After concentrating under reduced pressure and adding acetone, the resulting DCurea
was filtered out. After concentration under reduced pressure, it was washed with ethyl acetate and then ether, and dried under reduced pressure to obtain 10.0 g of white powder (yield: 6
5%) was obtained. T of 10 g (9.8 mmol) of this compound
A TFA solution of 1M trifluoromethanesulfonic acid-thioanisole-m-cresol was added to the FA solution under ice cooling, and the mixture was reacted for 1 hour to remove the peptide side chain and terminal protecting group. The reaction solution was poured into ether, and the oily precipitate was dissolved in distilled water and washed with ethyl acetate, followed by anion exchange resin column (Amberlite IRA-4
00; Cl type) to form a hydrochloride and freeze-dry it. 4.8 g (80% yield) of white solid was obtained. Synthesis Example 16 Synthesis 16 CH2=CCH3-CO-NH-C2H4-CO-RG
DSGNH2 (SEQ ID NO: 16) The peptide chain was extended in the same manner as in Synthesis Example 15. The outline is shown below. (1) Synthesis of BocSer(Bzl)GlyNH2 BocSer(Bzl): 59g (
0.2mol)GlyNH2・HCl
:22.1g (0.2mol) N-methylmorpholine :20.2g (0.2mol) CH2Cl2
:800mlDCC
:41.2g (0.2mol) (1
) yield 58.3g (yield 83%) 006
9] (2) BocAsp(OBzl)Ser(Bzl)G
Product of synthesis (1) of lyNH2:
56.2g (0.16mol) TFA/CH2Cl2
:200ml/200mlBocA
sp(OBzl): 51.7g (0.
16 mol) CH2Cl2
:800mlDCC :
Yield of 33g (0.16mol) (2) 71
．． 2g (yield 80%) (3) BocGl
yAsp(OBzl)Ser(Bzl)GlyNH2
Synthesis (2) Product: 66.7g (0.
12mol) TFA/CH2Cl2
:200ml/200mlBocGly
:51.7g (0.12mol) CH
2Cl2: 700mlD
CC: 24.7 (0.1
2mol) (3) yield 61.8g (yield 8
(4%) BocArg(Mts)GlyAsp(OBz
l) Synthesis of Ser(Bzl)GlyNH2 (3) Product: 61.3g (0.1mol)
TFA/CH2Cl2: 200m
l/200mlBocArg(Mts)
:45.6g (0.1mol) DMF
:800mlDCC
:22.5 (0.1 mol) HOBt
:14g (0.1mol) (4)
Yield: 42.8g (yield 45%) 0072
] (5) Synthesis of compound 16 (4) Product
:5.0g (5.3mmol) TFA/CH2Cl2
: 50ml/50ml carboxyethyl methacrylamide: 0.83g (5.3mmol) DMF: 50ml DCC

:1.1g (5.3mmol) HOBt
:0.72g (5.3mmol) TFA solution of 1M-trifluoromethanesulfonic acid-thioanisole-m-cresol
:250ml Synthesis Example 17 Synthesis 17 RGDG-NH-C2H4-NH-CO-CCH3=C
H2 (SEQ ID NO: 17) Compound 17 was synthesized by a liquid phase method using a sequential extension method. (1) Synthesis of BocGlyONb 35 g (0.2 mol) of BocGly, 28 ml (0.2 mol) of triethylamine, and 43.2 g (0.2 mol) of p-nitrobenzyl bromide were added to 400 ml of ethyl acetate and refluxed for 5 hours, then at room temperature. It was left overnight. The precipitate was filtered off, and the filtrate was washed with an aqueous NaHCO3 solution, then water, and dried over Na2SO4.The filtrate was concentrated under reduced pressure and recrystallized from ethyl acetate-hexane. 52.7g (yield 85%
). The peptide chain was then extended in the same manner as in Synthesis Example 15. The outline is shown below. (2) Synthesis of BocAsp(OBzl)GlyONb Product of (1): 46.5g (0.
15mol) TFA/CH2Cl2
:200ml/200mlBocAsp(OBzl)
:48.5g (0.15mol) CH
2Cl2: 750mlD
CC: 30.9g (0.
15 mol) (2) Yield 64.0 g (yield 80%) (3) BocGlyAsp(OBzl)GlyONb
Synthesis (2) Product: 64.0
g (0.12 mol) TFA/CH2Cl2
:200ml/200mlBocGly
:21g (0.12mol)
CH2Cl2: 750m
lDCC: 24.7g (
0.12 mol) (3) yield 58.8 g (
(Yield 83%) (4) BocArg(Mts)GlyAsp(OBz
l) Synthesis of GlyONb (3) Product
:53.7g (91mmol) TFA/CH2C
l2: 200ml/200mlB
ocArg(Mts): 41.5g
(91 mmol)DMF
:800mlDCC :1
8.7g (91mmol) HOBt
: Yield of 13.5g (0.1mol) (4)
46.5g (yield 55%) (5) BocArg(Mts)GlyAsp(OBz
l) Synthesis of Gly (4) 9.29 g (10 m
mol) in 300 ml of 90% acetic acid, and dissolve Zn powder 3
2.7 g (0.5 mol) was added and stirred at 0°C for 3 hours. The Zn powder was filtered off, and the filtrate was concentrated under reduced pressure, acidified by adding citric acid, and extracted with ethyl acetate. After drying over Na2SO4 and concentrating under reduced pressure, ether was added to obtain 6.26 g of white powder (yield 79%). (6) Synthesis of Compound 17 Product of (5):
5.31g (6.7mmol) aminoethyl methacrylamide: 0.86g (6.7mmol) DMF
:60ml DC
C:1
．． 4g (6.7mmol) HOBt
:0.95g (7mmol)1
M-trifluoromethanesulfonic acid-thioanisole-
TFA solution of m-cresol
:250ml Synthesis Example 18 Synthesis 18 CH2=CCH3-CO-NH-(CH2)3-RGD
G-NH-C2H4-NH-CO-CCH3=CH2 (SEQ ID NO: 18) After carrying out the same synthesis as in (1) to (5) of Synthesis Example 17, compound 18 was obtained by the following synthesis. Ta. (1) BocArg(Mts)GlyAsp(OBz
l) Gly-NH-C2H4-NH-CO-CCH3
=CH2 BocArg(Mts)GlyAsp(OBzl)Gl
y: 5.19g (6.7mmol) Aminoethyl methacrylamide: 0.86g (6.7mmol) DMF
:60
mlDCC
:1.4g (6.7mmol) HOBt
:0.95g (7mm
ol) (1) Yield 4.7 g (yield 80%) Mass spectrum M+: 885 (2) Synthesis of compound 18 Product of (1)
:4.4g (5mmol) TFA/CH2Cl2
:50ml/5
0ml carboxyethyl methacrylamide: 0.79g
(5 mmol) DMF
:50ml DCC
:1.03g (5mmol
) HOBt
:0.68g (5mmol) 1M trifluoromethanesulfonic acid-thioanisole-
TFA solution of m-cresol
:250ml Amberlight IRA-400 :CI type treatment [
Synthesis Example 19 A radical copolymer of Compound 15 and Product 1 was synthesized. 500 mg (0.82 mmol) of Synthesis 15 and 677 mg (3.28 mmol) of the monomer of Production Example 1 were added to 10 mg of water.
After adjusting the pH to 7.4 with 1N NaOH, 5.1 mg of potassium persulfate and 2.0 mg of sodium bisulfite were added as initiators, and the mixture was heated at 20°C under a nitrogen stream for 2 hours.
Polymerization was carried out for 0 hours. Spectrapore 7 (molecular fraction 3000)
After dialysis against pure water to remove low molecular weight components,
Freeze-dried. Yield 490mg (synthetic product 19) 00
[84] The composition of the copolymer was determined from the values of amino acid analysis, and it was found that 19% of Compound 15 was introduced. (Calculated from β-alanine and Gly values) 0085
] Compound 19 was subjected to molecular weight fractionation by gel chromatography. The molecular weight was measured in the same manner as in Production Example 12. Fraction 1 Molecular weight approximately 53,000 (Synthetic product 19-
1) Fraction 2 Molecular weight approximately 21,000 (Synthetic product 1
9-2) Fraction 3 Molecular weight: about 8000 (Synthesis 19-3) Synthesis Example 20 Copolymerization of Synthesis 15 and the monomer of Production Example 1 was carried out in the same manner as in Synthesis Example 19, with the amount of initiator I changed it and did it. Synthesis Examples 21 and 22 Copolymerization was carried out in the same manner as in Synthesis Example 19 by changing the monomer composition. Synthesis Example 21 Composition of Synthesis 15 48% (β-alanine and Gl
Synthesis Examples 23 to 38 Synthesis Examples 23 to 38 Synthesis Examples 23 to 38 Synthesis Examples 1 to 14, 16 and 1
7 and various anionic monomers were copolymerized. [0091] [0092] [0094] [0095] [0096] [0097] [0098] Synthesis Example 30 Monomer Synthesis 8
500mg (0.69mmol)
Acrylic acid (manufactured by Tokyo Kasei) 202mg (2
．． 76 mmol) Initiator potassium persulfate
3.5mg Sodium bisulfite 1.4mg yield
233mg (Synthetic product 30) Molecular weight 100
00 Composition of Compound 8 14% Elemental analysis N value 9.62% Water/methanol = 1/1 was used as the polymerization solvent and dialysate. [0099] Molecular weight: 12000 [0100] [0101] [0102] [0103] [0104] [0105] [0106] [0107] Synthesis example 39 Hydrogels of compound 15, compound 18 and product 1 were subjected to radical polymerization It was synthesized by Two silane-treated glass plates (5 cm x 6 cm x 1 cm) and a gasket were prepared. Synthesis 15 0.23g (0.4mmol),
0.28 g (0.4 mmol) of Synthetic Product 18 and 1.49 g (9.4 mmol) of Product 1 were dissolved in distilled water to obtain a 10 wt % monomer solution. Adjust the pH to 7.4 with 1N NaOH, then add 10% ammonium persulfate.
After adding 4 mg and thoroughly purging with nitrogen, the solution was injected between glass plates. The glass plate was held in a vise and polymerized at 60° C. for 40 hours. The generated hydrogel was washed with distilled water to remove unreacted monomers. The monomer ratio of Compound 15 and Compound 18 in Compound 39 was about 8%. (Elemental analysis N value
After sterilizing the gel (12.07%) using an ultraviolet lamp, it was subjected to a cell culture experiment. Test Example 1 Measurement of Cell Adhesion Inhibitory Activity The propenamide derivative copolymer salt of the present invention inhibits cell adhesion to fibronectin and vitronectin. The method for measuring the activity is shown below. The competition method used in this experimental example is basically one that is widely used in the biochemical field, such as "Methods in Enzymo".
82, 803-831 (1981), and JP-A-1-309682 and JP-A-2-174797. Experimental method 1. Preparation of fibronectin and vitronectin adsorption plates Commercially available fibronectin (human-derived) : Purchased from Seikagaku Corporation) or vitronectin (human origin, purchased from Funakoshi Pharmaceutical Co., Ltd.) was diluted with PBS to 1.0 μl/ml and 2.0 μl/ml, respectively, and 0.5 ml of the diluted solution was added to 24 wells. Put it in the plastic plate of 3
It was kept warm at 7°C overnight and coated. Next, bovine serum albumin (BSA 1%) was added to prevent non-specific adsorption.
Incubate at 7℃ for 1 hour, then wash as usual (PBS)
was added and drained thoroughly to prepare a fibronectin adsorption plate. A vitronectin adsorption plate was prepared using the same procedure. 2. Adhesion inhibition experiment Salt of copolymer of propenamide derivative obtained by freeze-drying was added to Dulbecco's Modified Eag.
Dissolved in les Medium (hereinafter abbreviated as DMEM), 0, 0.25, 0.5, 1.0, 1.5 mg/m
1 of each concentrated solution. Put 0.25 ml of this solution into the plate prepared by the above method, and add it to the vascular endothelial cells (
4×106 cells/ml) suspension at 0.25
ml was added and kept warm at 37°C for 1 hour to allow cells to adhere. D
Wash 3 times with MEM medium, detach non-adherent cells, and add 2%
The number of cells was counted by staining with trypan blue. The results are shown in Tables 1 and 2. Table 1 Cell adhesion inhibition to fibronectin (cells/well)

Concentration (mg/ml)
0 0.25 0
．． 5 1.0 1.5
Additive compound Compound 19-1 × △
○ ○
○ Compound 19-2 ×
△ ○ ○
○ Compound 19-3 ×
△ ○ ○
○ Compound 20 ×
△ ○
○ ○ Compound 21
× △ ○
○ ○ Compound 22
× △ △
△ ○ Compound 23 × ×
△ △ ○
Composite 24 × △
○ ○
○ Compound 25 ×
△ ○ ○
○ Compound 26 ×
△ ○ ○
○ Compound 27
× △ ○
○ ○ Compound 28
× △ △
○ ○ Composite 2
9 × △
○ ○ ○
Composite 30 × △
△ ○ ○
Compound 31 ×
△ △ ○
○ Compound 32 ×
△ ○ ○
○ Compound 33
× △ ○
○ ○ Compound 34
× △ ○
○ ○ Compound 35
× △
○ ○ ○ Compound 36 × △
○ ○ ○
Compound 37 ×
△ ○ ○
○ Compound 38 ×
△ ○ ○
○ Manufactured product 12 ×
× ×
× × RGD
× × ×
× △ RGDS
× ×
× △ △
(cells/well); ○: 50 or less, △
: 51-99, ×: 100 or more Table 2 Cell adhesion inhibition to vitronectin (cells/well)

Concentration (mg/ml)
0 10
50 100 300
Additive compound Compound 19-1 × ○
○ ○
○ Compound 19-2 ×
○ ○ ○
○ Compound 19-3 ×
○ ○ ○
○ Compound 20 ×
○ ○
○ ○ Compound 21
× ○ ○
○ ○ Compound 22
× △ △
○ ○ Compound 23 × ×
△ △ ○
Compound 24 × ○
○ ○
○ Compound 25 ×
○ ○ ○
○ Compound 26 ×
○ ○ ○
○ Compound 27
× ○ ○
○ ○ Compound 28
× △ △
○ ○ Compound 2
9 × ○
○ ○ ○
Composite 30 × △
○ ○ ○
Compound 31 ×
△ △ ○
○ Compound 32 ×
○ ○ ○
○ Compound 33
× ○ ○
○ ○ Compound 34
× ○ ○
○ ○ Compound 35
× ○
○ ○ ○ Compound 36 × ○
○ ○ ○
Compound 37 ×
○ ○ ○
○ Compound 38 ×
○ ○ ○
○ Manufactured product 12 ×
× ×
× × RGD
× × ×
△ △ RGDS
× ×
△ △ ○
(cells/well); ○: 100 or less, △:
101-199, ×: 200 or more Test Example 2 Measurement of platelet aggregation inhibitory activity In of the salt of the copolymer of propenamide derivatives synthesized in the present invention
The platelet aggregation inhibitory effect in vitro was investigated using human platelet-rich plasma. The experimental method is shown below. Experimental Method Fresh human blood was added with 1/9 volume of 3.8% sodium citrate and centrifuged (1000 rpm, 10 minutes), and the upper layer was collected as platelet-rich plasma. 0-1.5 mg/salt of copolymer of propenamide derivative obtained by freeze-drying
It was dissolved in physiological saline to various concentrations of ml. Add 25 μl of this salt solution to 200 μl of plasma, incubate at 37°C for 3 minutes, then add 25 μl of 50 μM adenone siniphosphate (ADP) solution or 200 μg/ml collagen solution and check the permeability using an aggregometer. It was verified by measuring. The results are shown in Table 3. Aggregation inhibition rate (1-T/T0) x 100% T0 = Permeability T when salt of copolymer of propenamide derivative is not added = Permeability when salt of copolymer of propenamide derivative is added Table 3 Platelet aggregation inhibition Additive compounds
inhibitory activity
ADP stimulation Collagen stimulation
Compound 19-1 ○
○ Compound 19-2
○ ○
Compound 19-3 ○
○ Compound 20
○
○ Compound 21
○ ○ Compound 22 ○
△ Composite 23
△
○ Compound 24
○ ○ Compound 25 ○
○ Compound 26
○ ○
Compound 27 ○
○ Compound 28
○
△ Composite 29
○ ○
Composite 30 △
○ Compound 31
○
○ Compound 32
○ ○
Compound 33 ○
○ Compound 34
○
○ Compound 35
○ ○ Compound 36 ○
○ Compound 37
○
○ Compound 38 ○
○ Manufactured product 1
2 ×
× RGD
△〜○ △〜○
RGDS △〜○
△〜○ IC5
0 (μg/ml); ○: 40 or less, △: 40-80, ×
: 80 or more [0117] Test Example 3 Evaluation of proliferation of animal cells Cell culture was carried out using the animal cell culture substrates prepared in Examples and Comparative Examples. The cells used are vascular endothelial cells,
Culture medium is DMEM and 10% fetal calf serum (FCS)
DMEM containing . Add 1 x 10 cells to this culture medium.
The cross-linked copolymer was suspended at a ratio of 4 cells/ml in a plastic petri dish in which the cross-linked copolymer was placed in advance.
They were added at a rate of x104 pieces/cm2. This is 3
The cells were cultured at 7°C under a 5% carbon dioxide atmosphere. After culturing,
Adhesion and proliferation were observed using a phase contrast microscope. The results are shown in Table 4. Table 4 Evaluation of animal cell proliferation
DMEM medium DMEM/FCS medium
Adhesion Proliferation Adhesion Proliferation Example (Synthesis 39) ○
○ ○ ○
Comparative example (Product 13) ×～△ ×～△
△〜○ △〜○
○: Good, △: Slightly poor, ×: Poor [0119] Product 13 used for comparison had cell adhesion and proliferation properties in DMEM medium not containing FCS compared to the substrate of Synthesis 39 of Example. It was inferior. [Sequence Listing] SEQ ID NO: 1 Sequence length: 4 Sequence type: Amino acid topology: Linear sequence type: Peptide fragment type: C-terminal fragment Sequence characteristics: Symbols representing characteristics: modified site Location: 1 Method for determining characteristics: E Other information: Xaa is replaced with 4Abu Sequence Xaa Arg Gly Asp 1 SEQ ID NO: 2 Sequence length: 7 Sequence type: Amino acid topology : Type of linear sequence: Peptide fragment type: Characteristics of C-terminal fragment sequence: Symbol representing characteristics: modified site Location: 1 Method for determining characteristics: E Other information: Ala is a sequence substituted with bAla Ala Arg Gly Asp Arg Gly A
sp 1 5 012
3] Sequence number: 3 Sequence length: 10 Sequence type: Amino acid topology: Type of linear sequence: Peptide fragment type: C-terminal fragment Sequence characteristics: Symbol representing characteristics: modified site Location: 1 Characteristics Determined method: E Other information: Ala is replaced with bAla Sequence Ala Arg Gly Asp Arg Gly A
sp Arg GlyAsp 1
5 1
SEQ ID NO: 4 Sequence length: 16 Sequence type: Amino acid topology: Linear sequence type: Peptide fragment type: C-terminal fragment Characteristics of the sequence: Characteristic symbol: modified site Location: 1 How the characteristics were determined: E Other information: Sequence Ala Arg Gly Asp Arg Gly A where Ala is replaced by bAla
sp Arg GlyAsp Arg GlyAsp
Arg Gly Asp 1 5
10
15 SEQ ID NO: 5 Sequence length: 5 Sequence type: Amino acid topology: Linear sequence type: Peptide fragment type: C-terminal fragment Sequence characteristics: Characteristic symbol: modified site Location: 1 Method of characterization: E Other information: Sequence Xaa Arg Gly Asp Ser 1 where Xaa is replaced with 4Abu SEQ ID NO: 6 Sequence length: 5 Sequence type: Amino acids Topology: Linear sequence Type: Peptide fragment type: C-terminal fragment Sequence characteristics: Symbol representing characteristics: modified site Location: 1 Method for determining characteristics: E Other information: Xaa is replaced with Nva Sequence Xaa Arg Gly Asp Ser 1
5 SEQ ID NO: 7 Sequence length: 5 Sequence type: Amino acid topology: Linear sequence type: Peptide fragment type: C-terminal fragment Sequence characteristics: Characteristic symbol: modified site Location: 1 How the characteristics were determined: E Other information: Sequence Xaa Arg Gly Asp Ser 1 where Xaa is replaced with Acp
5 SEQ ID NO: 8 Sequence length: 5 Sequence type: Amino acid topology: Type of linear sequence: Peptide fragment type: C-terminal fragment Sequence characteristics: Characteristic symbol: modified site Location: 1 Method of characterization: E Other information: Xaa is replaced with 12-aminolauric acid Sequence Xaa Arg Gly Asp Ser 1
5 SEQ ID NO: 9 Sequence length: 5 Sequence type: Amino acid Topology: Linear Sequence type: Peptide fragment type: C-terminal fragment sequence Leu Arg Gly Asp Ser 1
5 SEQ ID NO: 10 Sequence length: 5 Sequence type: Amino acid Topology: Linear Sequence type: Peptide fragment type: C-terminal fragment sequence Gln Arg Gly Asp Ser 1
5 SEQ ID NO: 11 Sequence length: 5 Sequence type: Amino acid topology: Linear sequence type: Peptide fragment type: C-terminal fragment Sequence characteristics: Characteristic symbol: modified site Location: 1 Method of characterization: E Other information: Xaa is replaced with p-aminobenzoic acid Sequence Xaa Arg Gly Asp Ser 1
5 SEQ ID NO: 12 Sequence length: 6 Sequence type: Amino acid topology: Linear sequence type: Peptide fragment type: C-terminal fragment Sequence characteristics: Characteristic symbol: modified site Location: 1 How the characteristics were determined: E Other information: Sequence Xaa Gly Arg Gly Asp Ser 1 where Xaa is replaced by Nva
5 SEQ ID NO: 13 Sequence length: 6 Sequence type: Amino acid Topology: Linear Sequence type: Peptide fragment type: C-terminal fragment sequence Gly Arg Gly Asp Ser Pro 1
5 SEQ ID NO: 14 Sequence length: 7 Sequence type: Amino acid Topology: Linear Sequence type: Peptide fragment type: C-terminal fragment sequence Gly Gly Gly Arg Gly Asp S
er 1 5 0
135] SEQ ID NO: 15 Sequence length: 5 Sequence type: Amino acid topology: Linear sequence type: Peptide fragment type: C-terminal fragment Sequence characteristics: Symbol representing characteristics: modified site Location: 1 Characteristics Determined method: E Other information: Ala is replaced with bAla Sequence Ala Arg Gly Asp Ser 1
5 SEQ ID NO: 16 Sequence length: 6 Sequence type: Amino acid topology: Linear sequence type: Peptide fragment type: C-terminal fragment Characteristics of the sequence: Symbol representing characteristics: modified site Location: 1 How the characteristics were determined: E Other information: Sequence Ala Arg Gly Asp Ser Gly 1 where Ala is replaced by bAla
5 013
7] SEQ ID NO: 17 Sequence length: 4 Sequence type: Amino acid topology: Linear sequence type: Peptide fragment type: N-terminal fragment sequence Arg Gly Asp Gly 1 0138] SEQ ID NO: 18 Sequence length :5 Sequence type: Amino acid topology: Type of linear sequence: Peptide fragment type: Features of intermediate fragment sequence: Symbol representing feature: modified site Location: 1 Method for determining feature: E Other information: Ala is the sequence replaced with bAla

Claims (8)

[Claims] Claim 1: A propenamide derivative having an adhesive peptide represented by the following general formula [II] as an essential unit in the side chain of the following general formula [I], and the following general formula [III]
A copolymer with an anionic monomer represented by or a salt thereof. General formula [I] R1R2C=CR3 -CO-[NH]- in the formula, R1
, R2 represents a hydrogen atom or a carboxyl group, R3
represents a hydrogen atom, a methyl group, an ethyl group, a halogen atom or a carboxymethyl group. General formula [II] -[R4]-[CO]-([X]-Arg-Gly-A
sp-[Y])n-[Z]-[R5]- where X, Y are Ser, Gly, Val, Asn
, Pro, and Z represents -O- or -NH-. R4,
Either one of R5 has a hydrogen atom, and the other has 1 carbon number.
~11 linear or branched alkylene groups, or 6 carbon atoms
-11 represents an arylene group, which may have a substituent. n represents an integer of 1 to 5. General formula [III] H2C=CR6 -[CO]-[W]-R7 In the formula, R6
may have a substituent with a hydrogen atom or an alkyl group having 1 to 3 carbon atoms. W represents -O- or -NH-. R
7 is a hydrogen atom, a linear or branched alkyl group having 1 to 12 carbon atoms, or an aryl group having 6 to 11 carbon atoms, and at least a carboxyl group, a sulfo group,
It contains any one of the phosphoro groups and may further have a substituent. General formula [I], [II], [III]
In [ ] indicates that the group within [ ] may be present or absent. Claim 2: Molecular weight of about 3,000 to 200,0
00. The propenamide derivative copolymer or salt thereof according to claim 1, wherein the propenamide derivative copolymer or a salt thereof is in the range of 0. 3. The copolymer or salt thereof according to claim 1, wherein the content of the structural unit derived from the propenamide derivative is 0.1 to 90 mol%. 4. A crosslinked copolymer of the propenamide derivative according to claim 1 and an anionic monomer, or a salt thereof. 5. The content of the structural unit derived from the propenamide derivative and the structural unit derived from the polyfunctional monomer is 0.1 to 90 mol% and 0.1 to 30 mol%, respectively. A crosslinked copolymer of or a salt thereof. 6. An animal cell adhesion inhibitor comprising the propenamide derivative copolymer or its salt according to claim 1, 2 or 3 as an active ingredient. 7. A platelet aggregation/adhesion inhibitor comprising the propenamide derivative copolymer or its salt according to claim 1, 2 or 3 as an active ingredient. 8. An animal cell culture substrate comprising a crosslinked copolymer of the propenamide derivative according to claim 4 or 5 and an anionic monomer or a salt thereof as an active ingredient.