Polymer having upper critical solution temperature (UCST), obtained through polymerization or copolymerization of monomer mainly composed of n-acryloyl-piperidinecarboxamide, and method for producing the same and method for changing the UCST in aqueous solution

Abstract

To provide a polymer having an upper critical solution temperature (UCST) controlled over a wide temperature range.SOLUTION: The present invention relates to a polymer having an upper critical solution temperature (UCST), obtained through polymerization or copolymerization of a monomer mainly composed of N-acryloyl-piperidinecarboxamide, and to a method for producing the polymer and a method for changing the UCST in an aqueous solution.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to a polymer having an upper critical solution temperature (UCST) and obtained by polymerizing or copolymerizing a monomer mainly composed of N-acryloyl-piperidinecarboxamide, and to a method for producing the polymer and a method for changing the UCST in an aqueous solution thereof. [Background technology]

[0002] Temperature-responsive polymers that exhibit an upper critical solution temperature (UCST) in aqueous solutions have the property that the polymer aggregates and precipitates at low temperatures and dissolves at high temperatures beyond the upper critical solution temperature. The change is reversible. The temperature-dependent aggregation and dissolution of polymers is due to the formation of ionic and hydrogen bonds between polymer chains. Intelligent polymers, whose structure and physical properties change in response to temperature, pH, and light stimuli, have been widely studied and applied in regenerative medicine and tissue engineering, and the physical and structural changes of UCST-type temperature-responsive polymers are one of them that have attracted interest. In particular, temperature-responsive polymers that express UCST under physiological conditions are expected to be applied in biomaterials such as cell culture substrates, medium additives as cell growth cofactors, chromatography carriers, artificial skin for humans and robots, drug delivery systems, and wearable sensors, as well as in the fields of regenerative medicine and tissue engineering. However, the number of temperature-responsive polymers that exhibit UCST under physiological conditions is limited. Therefore, the development of temperature-responsive polymers that express UCST under physiological conditions has been actively pursued.

[0003] To date, polymers that have UCST in physiological environments (such as physiological phosphate buffer solutions) have been reported, such as poly(allylurea-allylamine) copolymer, poly(acrylamide-acetonitrile) copolymer, poly(N-acryloylglycinamide), poly(N-acryloylnipecotamide), etc. For poly(allylurea-allylamine) copolymer, a UCST-type temperature-responsive polymer that contains ionic functional groups in the polymer side chains, the effects of pH changes and counter ion species under physiological conditions have been investigated (Non-Patent Document 1).

[0004] UCST-type polymers such as poly(acrylamide-acetonitrile) copolymers, poly(N-acryloylglycinamide), and poly(N-acryloylnipecotamide), which have neutral side-chain functional groups that do not have ionic functional groups in the polymer side chains, have been developed (Non-Patent Documents 2, 3, and 4). It has been reported that these UCST-type temperature-responsive polymers can be controlled by the concentration and molecular weight of the polymer, and further, in the case of copolymer polymers such as poly(acrylamide-acetonitrile), by the composition of added monomers in the copolymer. However, conventional UCST control could only be changed by a specific molecular weight or concentration of the polymer, or the composition of the monomer components in the copolymer. In addition, there has been little research on temperature control, etc. for polymers with UCST that have N-acryloyl-piperidinecarboxamide as the main component. [Prior art documents] [Non-patent literature]

[0005] [Non-Patent Document 1] N. Shimada et al, Biomacromolecules, 2011, 12, 3418-3422 [Non-Patent Document 2] F.Liu et al.,Journal of Polymer Science Part A:Polymer Chemistry,2012,50,4920-4928 [Non-Patent Document 3] J.Seuring et al.,Macromol.Chem.Phys.2010,211,2109-2117 [Non-Patent Document 4] Y. Akiyama.,Macromol Rapid Commun.,2021,42,2100208 Summary of the Invention [Problem to be solved by the invention]

[0006] An object of the present invention is to provide a polymer having an upper critical solution temperature (UCST) that is controlled within a wide temperature range. [Means for solving the problem]

[0007] The present inventors have found that in a polymer having UCST, the main component of which is N-acryloyl-piperidinecarboxamide, the expression of UCST can be controlled over a wider temperature range by changing the end group structure of the polymer chain. Furthermore, for polymers with UCST that have N-acryloyl-piperidinecarboxamide as the main component, we synthesized new monomers by changing the number and arrangement of substituents on the carboxamide group, modifying the carboxamide group, etc., and by forming copolymers using these alone or in combination, we succeeded in synthesizing polymers with UCST over a wide temperature range.

[0008] That is, the present invention is specified by the following items. 1. Formula I [ka] (In the formula, R 11 is H or CH3, R placed in a ring structure 12 ~R 16 1 to 3 H atoms are replaced by -C(=O)NH2, (However, in the case of 1, this is limited to poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end, poly(N-acryloyl-nipecotamide) having a trithiocarboxyethyl group and a 4-cyanopentanoic acid group at the end, and poly(N-acryloyl-nipecotamide) having a trithioethyl group and a 4-cyanopentanoic acid group at the end.) m is an integer equal to or greater than 2. or a homopolymer having a structural unit represented by Formula II [ka] (In the formula, R 21 and R 31 are, independently of each other, H or CH3, R placed in a ring structure 22 ~R 26 1 to 3 H atoms are replaced independently by -C(=O)NH2 or -C(=O)-NH-CH2-C(=O)-NH2; R placed in a ring structure 32 ~R 36 1 to 3 H atoms are independently replaced by -CN or -C(=O)-NH2; m is an integer equal to or greater than 1; and n is an integer of 1 or greater.) or Formula III [ka] (In the formula, R 41 and R 51 are, independently of each other, H or CH3, R placed in a ring structure 42 ~R 46 one to three H atoms are replaced independently by -CN, -C(=O)NH2 or -C(=O)-NH-CH2-C(=O)-NH2; R 52 is -CN, -C(=O)NH2 or -C(=O)-O-C2H4OH, m is an integer equal to or greater than 1; and n is an integer of 1 or more.

[0009] 2. The constitutional unit represented by formula I is [ka] (In the formula, R 11 , m is as defined in formula (I). 2. The polymer according to claim 1,

[0010] 3. The constitutional unit represented by formula II is [ka] [ka] [ka] or [ka] (In the formula, R 21 , R 31 , m and n are as defined in formula (II). 2. The polymer according to claim 1,

[0011] 4. The constitutional unit represented by formula III is [ka] or [ka] (In the formula, R 41 , R 51 , m and n are as defined in formula (III). 2. The polymer according to claim 1,

[0012] 5. The polymer according to any one of 1 to 4 above, wherein the terminal group contains a C(═O)OH group, a maleimide group, a CN group, an SH group, an amino group or an OH group.

[0013] 6. The polymer according to any one of 1 to 5 above, which is synthesized by radical polymerization.

[0014] 7. A polymer according to any one of 1 to 6 above, synthesized by RAFT (reversible addition-fragmentation chain transfer) polymerization.

[0015] 8. The polymer according to claim 7, modified at both ends with reactant residues derived from a chain transfer agent in RAFT polymerization.

[0016] 9. Chain transfer agents for RAFT polymerization YXZ, where X is a thiocarbonylthio group (-C(=S)-S-) or a trithiocarbonate group (-SC(=S)-S-), Y and Z are each independently H, a phenyl group, or a straight-chain or branched alkyl group which may be substituted with one or more C(=O)OH groups, phenyl groups, or CN groups. [ka] (In the formula, R 11 ~R 16 and m is as defined in formula (I). and formula II is formula II-1 [ka] (In the formula, R 21 ~R 26 , R 31 ~R 36 , m and n are as defined in formula (II); or formula III is formula III-1 [ka] (In the formula, R 41 ~R 46 , R 51 ~R 52 9. The polymer according to claim 8, wherein m and n are as defined in formula (III).

[0017] 10. The polymer according to any one of 1 to 9 above, having a weight average molecular weight Mw of 1,000 to 700,000.

[0018] 11. The polymer according to any one of 1 to 10 above, wherein the ratio Mw / Mn of the number average molecular weight Mn to the weight average molecular weight Mw is 1.00 to 7.00.

[0019] 12. The polymer according to any one of 1 to 11 above, wherein UCST is 3 to 80°C.

[0020] 13. The polymer according to any one of 1 to 12 above, which has no cytotoxicity.

[0021] 14. The polymer according to any one of claims 1 to 13, which promotes cell proliferation.

[0022] 15. Formula [ka] (In the formula, R 6 is H or CH3.) A compound represented by the formula:

[0023] 16. Formula [ka] (In the formula, R 7 is H or CH3.) A compound represented by the formula:

[0024] 17. A compound according to the above 15 or 16 for producing a polymer according to the above 1 to 14.

[0025] 18. A method for producing the polymer described in 1 to 14 above, comprising the steps of: One or two types of [ka] (In the formula, R 81 is H or CH3, R 82 ~R86 1 to 3 H atoms of are replaced independently by -CN, -C(=O)NH2 or -C(=O)-NH-CH2-C(=O)-NH2. A method for radical polymerization using a monomer represented by the following formula:

[0026] 19. The method according to claim 18, wherein the radical polymerization is carried out using a radical polymerization initiator or a radical initiator and a chain transfer agent for RAFT polymerization.

[0027] 20. A method for changing the upper critical solution temperature (UCST) of a polymer in an aqueous solution obtained by polymerizing or copolymerizing a monomer mainly composed of N-acryloyl-piperidinecarboxamide, comprising the steps of: The method is Changing the terminal modification group The method according to the above.

[0028] 21. The method according to claim 20, wherein the terminal group comprises a C(=O)OH group, a maleimide group, a CN group, a SH group, an amino group or an OH group. Effect of the Invention

[0029] The polymer of the present invention, which is mainly composed of N-acryloyl-piperidinecarboxamide, can be controlled over a wider temperature range of UCST by changing the end group structure of the polymer chain, expanding its application in a wide range of fields such as biomaterials, regenerative medicine, and tissue engineering. It was suggested that the polymer of the present invention not only has no cytotoxicity but also has an effect of promoting cell proliferation.

[0030] Furthermore, when a random copolymer synthesized by RAFT polymerization instead of free radical polymerization is used for synthesizing the polymer of the present invention, the molecular weight distribution can be narrowed and better temperature control becomes possible. [Brief description of the drawings]

[0031] [Figure 1]The 1H-13C HSQC NMR chart of N-acryloyl-2-piperidinecarboxamide is shown. [Diagram 2] The 1H-13C HSQC NMR chart of N-acryloyl-4-piperidinecarbonitrile is shown. [Diagram 3] The NMR chart of N-acryloylpiperidine-3,5-dicarboxamide is shown. [Figure 4] The 1H-13C HSQC NMR chart of acryloyl-N-(2-amino-2-oxoethyl)piperidine-3-carboxamide is shown. [Diagram 5] The results of 1H-NMR measurement of poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end and poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end are shown. [Figure 6] The results of upper critical solution temperature measurements of aqueous solutions of poly(N-acryloyl-nipecotamide) (Mn 12,400) having a dodecyltrithiocarbonate group at one end and poly(N-acryloyl-nipecotamide) (Mn 13,200) having a thiomaleimide group at one end are shown. [Figure 7] The results of upper critical solution temperature measurements of aqueous solutions of poly(N-acryloyl-nipecotamide) (Mn 14,100) having a dodecyltrithiocarbonate group at one end and poly(N-acryloyl-nipecotamide) (Mn 14,800) having a thiomaleimide group at one end are shown. [Figure 8] The results of upper critical solution temperature measurements of aqueous solutions of poly(N-acryloyl-nipecotamide) (Mn 14,100) having trithiocarboxyethyl groups and 4-cyanopentanoic acid groups at the polymer end, and poly(N-acryloyl-nipecotamide) (Mn 16,400) having trithioethyl groups and 4-cyanopentanoic acid groups at the polymer end are shown. [Figure 9] The results of upper critical solution temperature measurement of poly(acryloyl-N-(2-amino-2-oxoethyl)piperidine-3-carboxamide-co-acryloyl-N-piperidine-4-carbonitrile) (Mn 55,400) in aqueous solution are shown. [Figure 10] 1 shows the results of measurement of the upper critical solution temperature of poly(N-acryloyl-3,5-piperidinedicarboxamide). [Figure 11] The results of upper critical solution temperature measurement of poly(N-acryloyl-isonipecotamide-co-acrylonitrile) copolymer in aqueous solution are shown. [Figure 12] 1 shows the results of evaluating the cytotoxicity of poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end. [Figure 13] 1 shows the results of evaluating the particle size of poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end by light scattering. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0032] The polymer used in the present invention has an upper critical solution temperature (UCST) and is obtained by polymerizing or copolymerizing a monomer containing N-acryloyl-piperidinecarboxamide as a main component. Having an upper critical solution temperature (UCST) means that the polymer has the property of agglomerating and precipitating at temperatures lower than the upper critical solution temperature and dissolving at temperatures higher than the upper critical solution temperature. The upper critical solution temperature is determined by heating a solution of a polymer dissolved in water at a constant rate to a dissolved (transparent) state, then lowering the temperature at a constant rate, and determining the temperature at which the transmittance (solubility) of visible light decreases sharply (for example, the point at which the transmittance decreases by 90%) as the upper critical solution temperature. Another method is to measure the temperature at which the absorbance increases. The rate of temperature lowering and heating of the cooling composition solution can be appropriately determined according to the magnitude of change in the transmittance of the solution containing each composition.

[0033] The method for measuring the light transmittance is not particularly limited as long as it is a method using light of a wavelength suitable for measuring the change in the light transmittance in the polymer solution, and the wavelength is preferably a wavelength in the visible light range, specifically, a wavelength of 360 to 780 nm, preferably 550 to 650 nm. The temperature inside the cell holder during the measurement of the light transmittance can be maintained using a temperature control device such as a temperature controller.

[0034] The upper critical solution temperature of the polymer of the present invention is, for example, 3 to 80°C, preferably 15 to 55°C, and more preferably 20 to 40°C.

[0035] According to one aspect of the present invention, the present invention relates to a method for changing the upper critical solution temperature (UCST) of a polymer obtained by polymerizing or copolymerizing a monomer mainly composed of N-acryloyl-piperidinecarboxamide in an aqueous solution. Specifically, the method for changing UCST is as follows: (1) Changing the polymer dissolution concentration, (2) Changing the molecular weight of the polymer; (3) This can be achieved by changing the type and composition of the copolymerized monomers.

[0036] In the present invention, the expression of UCST can be freely controlled over a wider temperature range by further changing the terminal modification group of the polymer. Specifically, the UCST can be controlled by introducing a C(=O)OH group, a maleimide group, a CN group, a SH group, an amino group, or an OH group into the terminal group. In the case of a hydrophobic terminal group, the UCST of the polymer shifts to a higher temperature, and in the case of a hydrophilic terminal group, the UCST shifts to a lower temperature. This characteristic is a finding obtained in the present invention. In the present invention, the UCST can be changed widely by combining this method with the above methods (1) to (3).

[0037] The monomer used in the method of the present invention may be any monomer having N-acryloyl-piperidinecarboxamide as the main component. The monomer having N-acryloyl-piperidinecarboxamide as the main component includes all monomers having different positions (2-6 positions) of the carboxamide group coordinated to the piperidine ring and different coordination numbers, and also includes derivatives such as those in which the carboxamide group is modified.

[0038] Specifically, radical polymerization is carried out using one or two mutually independent monomers represented by the following formulas. [ka]

[0039] During the ceremony, R 81 is H or CH3, R placed in a ring structure 82 ~R 86 One to three H atoms, preferably one or two, more preferably one H atom, are each independently replaced by -CN, -C(=O)NH2 or -C(=O)-NH-CH2-C(=O)-NH2.

[0040] for example, [ka] [ka] These include, but are not limited to:

[0041] In the present invention, the monomers to be used are selected in consideration of the solubility of the monomer when the polymer is formed. If the monomer alone has a UCST when the polymer is formed, a homopolymer can be formed using only one type. If the monomer alone is poorly soluble in an aqueous solution or is soluble (easily soluble) and does not have a UCST by itself when the polymer is formed, two or more of these monomers may be combined (for example, a poorly soluble monomer + a soluble monomer) to form a polymer having a UCST.

[0042] The monomer constituting the hardly soluble or soluble polymer is, for example, As a monomer constituting the poorly soluble polymer, N-acryloyl-piperidine-2-carboxamide (PNP2CAm) is used. When a polymer is formed by itself, N-acryloyl-nipecotamide (N-acryloyl-piperidine-3-carboxamide: NANAm) and acryloyl-(piperidine-3,5-dicarboxamide) are monomers that form a polymer having UCST. Monomers that make up soluble polymers can be classified into N-acryloyl-N'-(2-amino-2-oxoethyl)nipecotamide, N-acryloyl-isonipecotamide (NAiNAm), and the like.

[0043] Among the above compounds, acryloyl-(piperidine-3,5-dicarboxamide) and N-acryloyl-N'-(2-amino-2-oxoethyl)nipecotamide in particular were newly synthesized for use in the method of the present invention, and it was discovered in the present invention that when they form a polymer by themselves, they have UCST, and when they form a copolymer, they can form a polymer having UCST.

[0044] The monomer constituting the polymer of the present invention may be any combination of monomers other than the monomer mainly composed of N-acryloyl-piperidinecarboxamide, taking into consideration the solubility of the polymer, etc. Examples of monomers include acrylonitrile, N-(meth)acryloyl-piperidine-4-carbonitrile, (meth)acrylamide, and hydroxyethyl (meth)acrylate. (Meth)acrylamide and hydroxyethyl (meth)acrylate can be used as monomers constituting a soluble polymer, and acrylonitrile and N-(meth)acryloyl-piperidine-4-carbonitrile can be used as monomers constituting a poorly soluble polymer.

[0045] As described above, the polymer of the present invention includes both homopolymers obtained by polymerizing a monomer having N-acryloyl-piperidinecarboxamide as a main component, and copolymers obtained by copolymerizing two or more kinds of monomers.

[0046] In one aspect of the present invention, for example, Formula I [ka] The present invention relates to a polymer having an upper critical solution temperature (UCST) that is a homopolymer having a constitutional unit represented by the following formula:

[0047] In formula I, R 11 is H or CH3, R placed in a ring structure 12 ~R 16 One to three, preferably one or two H atoms are substituted with -C(=O)NH2. m is an integer of 2 or more, preferably an integer of 2-500, and more preferably an integer of 10-150. When such a polymer is constituted by the above-mentioned monomers themselves, it is a homopolymer constituted by only the monomers that constitute the polymer having UCST.

[0048] For example, the constitutional unit represented by formula I is [ka] It is a homopolymer of

[0049] In one aspect of the present invention, Formula II [ka] The present invention relates to a polymer having an upper critical solution temperature (UCST), which is a copolymer having a constitutional unit represented by the following formula:

[0050] In formula II, R 21 and R 31 are, independently of each other, H or CH3, R placed in a ring structure 22 ~R 26 in which 1 to 3 H atoms, preferably 1 or 2 H atoms, more preferably 1 H atom, are each independently replaced by -C(=O)NH2 or -C(=O)-NH-CH2-C(=O)-NH2; R placed in a ring structure 32 ~R 36 one to three H atoms, preferably one or two H atoms, more preferably one H atom, are each independently replaced by -CN or -C(=O)-NH; m is an integer of 1 or more, preferably an integer of 1 to 500, and more preferably an integer of 10 to 300; n is an integer of 1 or more, preferably 1 to 500, and more preferably 10 to 300; The ratio m:n is from 1:9 to 9:1, and preferably 8:2.

[0051] For example, the constitutional unit represented by formula II is [ka]

[0052] [ka]

[0053] [ka]

[0054] or [ka] etc. Such a polymer is a copolymer obtained by copolymerizing a combination of a monomer constituting the soluble polymer and a monomer constituting the poorly soluble polymer.

[0055] In one aspect of the present invention, Formula III [ka] The present invention relates to a copolymer having a certain upper critical solution temperature (UCST) having a constitutional unit represented by the following formula:

[0056] In formula III, R 41 and R 51 are, independently of each other, H or CH3, R placed in a ring structure 42 ~R 46 wherein 1 to 3 H atoms, preferably 1 or 2 H atoms, more preferably 1 H atom, are independently replaced by -CN, -C(=O)NH2 or -C(=O)-NH-CH2-C(=O)-NH2; R 52 is -CN, -C(=O)NH2 or -C(=O)-O-C2H4OH, m is an integer of 1 or more, preferably an integer of 1 to 500, and more preferably an integer of 10 to 300; n is an integer of 1 or more, preferably 1 to 500, and more preferably 10 to 300; The ratio m:n is from 1:9 to 9:1, and preferably 8:2.

[0057] For example, the constitutional unit represented by formula III is [ka] Such a copolymer is a copolymer obtained by copolymerizing a monomer constituting a soluble polymer and acrylonitrile, which is a monomer constituting a poorly soluble polymer, in combination. The copolymer used in the present invention may be either a random copolymer or a block copolymer, but is preferably a random copolymer.

[0058] Polymer synthesis methods include radical reactions in which polymer chains grow with radicals as the reaction center, specifically, synthesis methods using free radical polymerization that can produce polymers with a wide molecular weight distribution, synthesis methods using living radical polymerization that allows control of molecular structure while maintaining the simplicity and versatility of radical polymerization, and synthesis methods using reversible addition-fragmentation chain transfer (RAFT) polymerization that can produce copolymers with smaller molecular weight distribution (Mw / Mn). RAFT polymerization allows control of molecular weight and more precise temperature control.

[0059] The synthesis method by the free radical polymerization may be a conventionally known method using a polymerization initiator that generates sufficient radicals at the polymerization temperature and a solvent suitable for the polymerization reaction. Since the synthesized reaction solution contains impurities such as unreacted monomers, it is preferable to prepare the polymer by further carrying out a purification treatment.

[0060] The synthesis method by RAFT polymerization can be exemplified by a conventionally known method that uses a polymerization initiator that generates sufficient radicals at the polymerization temperature, a solvent suitable for the polymerization reaction, and a reversible addition-fragmentation chain transfer agent (RAFT agent) for controlling the reactivity of the terminal active radicals to cause the polymerization to proceed in a pseudo-living manner.

[0061] Examples of the polymerization initiator include, but are not limited to, azo-based polymerization initiators such as 2,2'-azobis(isobutyronitrile) (AIBN), 2,2'-azobis(4-methoxy-2,4-dimethylvaleronitrile), 4,4'-azobis(4-cyanovaleric acid), 2,2'-azobis(2-amidinopropane) dihydrochloride, 2,2'-azobis[2-(5-methyl-2-imidazolin-2-yl)propane] dihydrochloride, 2,2'-azobis(2-methylpropionamidine) disulfate, and 2,2'-azobis(N,N'-dimethyleneisobutylamidine); persulfates such as potassium persulfate and ammonium persulfate; and peroxide-based polymerization initiators such as benzoyl peroxide, t-butyl hydroperoxide, and hydrogen peroxide.

[0062] Examples of solvents suitable for the above polymerization reaction include DMSO, water, and DMF.

[0063] Examples of the RAFT agent include dithiocarbonates such as O-ethyl-S-(1-phenylethyl)dithiocarbonate, O-ethyl-S-(2-propoxyethyl)dithiocarbonate, and O-ethyl-S-(1-cyano-1-methylethyl)dithiocarbonate; dithioesters such as cyanoethyl dithiopropionate, benzyl dithiopropionate, benzyl dithiobenzoate, and acetoxyethyl dithiobenzoate; S-benzyl-N,N-dimethyldithiocarbamate, benzyl-1-pyrrolecarbamate, and benzyl-1-pyrrolecarbamate; Examples of the thiocarbamate include, but are not limited to, dithiocarbamates such as dodecylthioate, dibenzyltrithiocarbonate, 2-(2-carboxyethylsulfanylthiocarbonylsulfanyl)propionic acid, 4-cyano-4-(((ethylthio)carbonothionyl)thio)pentanoic acid, 4-((((2-carboxyethyl)thio)carbonothioyl)thio)-4-cyanopentanoic acid (CTCCA), and trithiocarbonates such as cyanomethyldodecylthiocarbonate (CMDTC).

[0064] Radical polymerization can form polymers that are end-functionalized with residues from the initiator, and RAFT polymerization in particular can form polymers that are both end-functionalized with residues from the initiator and / or reactant residues from the RAFT agent.

[0065] Specifically, the RAFT polymerization chain transfer agent YXZ, where X is a thiocarbonylthio group (-C(=S)-S-) or a trithiocarbonate group (-SC(=S)-S-), Y and Z are each independently H, a phenyl group, or a linear or branched alkyl group which may be substituted with one or more C(=O)OH groups, phenyl groups or CN groups, preferably an optionally substituted C1-C 12 When the polymer is polymerized using the alkyl group, the chain transfer agent is modified at both ends with the reactant residues derived from the chain transfer agent, for example, as shown in formula I-1. [ka] It is possible to polymerize a homopolymer represented by the following formula:

[0066] In formula I-1, R 11 ~R 16 and m is as defined in formula (I).

[0067] Similarly, Formula II-1 [ka] It is possible to polymerize a copolymer represented by the formula:

[0068] In formula II-1, R 21 ~R 26 , R 31 ~R 36 , m and n are as defined in formula (II).

[0069] Similarly, Formula III-1 [ka] It is possible to polymerize a copolymer represented by the formula: In formula III-1, R 41 ~R 46 , R 51 ~R 52 , m and n are as defined in formula (III).

[0070] When RAFT polymerization is performed, a desired end group can be introduced by selecting a RAFT agent having a desired end group, which is preferable from the viewpoint of simplification of polymer preparation.

[0071] In RAFT polymerization, the number of moles of the RAFT agent blended when the total number of moles of monomers is taken as 1 is preferably 0.0001 to 0.05, and more preferably 0.0002 to 0.01.

[0072] The weight average molecular weight (Mw) of the polymer in the present invention may be 1,000 to 700,000 in the case of free radical polymerization, preferably 25,000 to 175,000, more preferably 50,000 to 150,000, and further preferably 85,000 to 120,000, and in the case of RAFT polymerization, may be 1,000 to 200,000, preferably 3,000 to 100,000, and more preferably 5,000 to 90,000. The method for calculating the weight average molecular weight may include a method of measuring using gel permeation chromatography (GPC), preparing a calibration curve using a standard substance, and then calculating.

[0073] The ratio (Mw / Mn) of the weight average molecular weight (Mw) to the number average molecular weight (Mn) of the polymer in the present invention is preferably 1.0 to 7.0, more preferably 1.0 to 5.0, and even more preferably 1.0 to 4.5. In the case of free radical polymerization, a preferable example is 1.0 to 3.0, and in the case of RAFT polymerization, a preferable example is 1.0 to 2.0.

[0074] The polymer of the present invention has extremely low cytotoxicity and promotes cell proliferation. The polymer is evaluated by dissolving the polymer in Dulbecco's modified Eagle medium (containing 10% serum) at a concentration of 0.1 mg / mL to 6.0 mg / mL, adding the polymer to the medium of bovine vascular endothelial cells that have been cultured in advance, and culturing the cells for 24 hours. After culturing, the viability of the cells is evaluated using Cell Counting Kit-8 (Dojin Chemical).

[0075] The present invention will be described in more detail below with reference to examples, but the present invention is not limited to these examples. EXAMPLES

[0076] <Example 1: Synthesis of N-acryloyl-2-piperidinecarboxamide> [ka] 2-Piperidinecarboxamide (3.91 g) was added to dimethylformamide (80 mL), stirred at 45° C. for 30 minutes to dissolve, and then allowed to stand at room temperature. Then, triethylamine (4.88 mL) was added under an argon gas atmosphere at 4° C. A solution of acrylic acid chloride (3.27 mL) dissolved in dimethylformamide (20 mL) was slowly added dropwise to this reaction solution, and after the dropwise addition, the mixture was stirred at room temperature overnight. After stirring, triethylamine hydrochloride was removed by filtration, the solution was concentrated with an evaporator, and the solution containing N-acryloyl-2-piperidinecarboxamide was separated by silica gel chromatography (Wakogel C200, Fujifilm Wako Pure Chemical Industries, Ltd.) (elution solution: acetone:hexane = 7:3 (volume ratio)), concentrated again with an evaporator (about 30 mL), and an appropriate amount of ethyl acetate (about 50 mL) was added to the concentrated solution to precipitate a white component, which was collected by filtration and drying to obtain N-acryloyl-2-piperidinecarboxamide (1.73 g). When developed with thin-layer silica gel chromatography (developing solution: acetone:hexane = 7:3 (volume ratio)), a UV absorption spot was obtained (Rf value = 0.33 to 0.43). N-acryloyl-2-piperidinecarboxamide 1 H- 13 The C HSQC NMR chart is shown in Figure 1.

[0077] <Example 2: Synthesis of N-acryloyl-4-piperidinecarbonitrile> [ka] 4-Piperidinecarbonitrile (5.06 mL) (Sigma-Aldrich) was added to a mixed solution of acetone (80 mL) and triethylamine (6.7 mL), and the mixture was stirred at 4°C under an argon gas atmosphere. A solution of acrylic acid chloride (4.3 mL) dissolved in acetone (20 mL) was added dropwise to the reaction solution, and the mixture was stirred overnight. After stirring, triethylamine hydrochloride was removed by filtration, the solution was concentrated with an evaporator, and the solution containing N-acryloyl-4-piperidinecarbonitrile was separated by silica gel chromatography (elution solution: ethyl acetate: acetone = 5: 1 (volume ratio)), and the solution was concentrated again with an evaporator to obtain liquid N-acryloyl-4-piperidinecarbonitrile (0.27 g). When developed with thin-layer silica gel chromatography (development solution: ethyl acetate: acetone = 5: 1 (volume ratio)), a spot of UV absorption was obtained (Rf value = 0.48-0.58). N-Acryloyl-4-piperidinecarbonitrile 1 H- 13 The C HSQC NMR chart is shown in Figure 2.

[0078] <Example 3: Synthesis of N-acryloyl-3,5-piperidinedicarboxamide> [ka] Oxalyl chloride (506 mL) was added dropwise to 3,5-pyridinedicarboxylic acid (100 g) under an argon gas atmosphere and refluxed (16 hours). After refluxing, the precipitated components were collected using an evaporator, dissolved in acetonitrile (1.8 L), and ammonia gas was injected into the solution. The precipitate was collected by filtration, suspended in an appropriate amount of saturated sodium bicarbonate solution, treated (70°C, 5 minutes), and allowed to stand at room temperature to obtain a precipitate. The precipitate was washed with water, acetonitrile, and diethyl ether in that order, and the target compound intermediate 1 (pyridine-3,5-dicarboxamide) was obtained using a rotary evaporator.

[0079] Pyridine-3,5-dicarboxamide (112 g) was added dropwise to a 4 molar solution of hydrochloric acid and 1,4-dioxane under an argon atmosphere and stirred at room temperature for 16 hours. The solvent was then dried and solidified on a rotary evaporator under an inert atmosphere, and the solid was crushed with diethyl ether. This was dried under vacuum to obtain intermediate 2 (pyridine-3,5-dicarboxamide hydrochloride).

[0080] A platinum oxide methanol solution (platinum oxide (9.4 g)) was added to a methanol aqueous solution (methanol:water = 9:1 (volume ratio)) (3 L) in which pyridine-3,5-dicarboxamide hydrochloride (104.5 g) was dissolved, and the mixture was stirred in a sealed environment under hydrogen gas (150 psi) (80 °C, 24 hours). The reaction mixture was then filtered through Celite, the filtrate was evaporated in a rotavapor, and the residue was co-distilled with toluene three times and dried under vacuum to obtain intermediate 3 (piperidine-3,5-dicarboxamide hydrochloride).

[0081] Piperidine-3,5-dicarboxamide hydrochloride (48.6 g) was dissolved in dimethylformamide (1.2 L), potassium carbonate (63.9 g) was added, and the mixture was stirred at room temperature (30 min). The reaction mixture was then cooled to 0°C, acryloyl chloride (23.3 g) was added, and the mixture was stirred at room temperature (16 h). The hydrochloride was filtered and removed, and the filtrate was concentrated with Rotavapol, and purified by column chromatography using Florisil as the stationary phase and 20% methanol-dichloromethane solution as the eluent to obtain the target compound (N-acryloylpiperidine-3,5-dicarboxamide). The NMR chart of N-acryloylpiperidine-3,5-dicarboxamide is shown in Figure 3.

[0082] <Example 4: Synthesis of acryloyl-N-(2-amino-2-oxoethyl)piperidine-3-carboxamide> [ka] A mixed solution of 1-Boc-nipecotic acid (1) (163 g), triethylamine (116 mL), and THF (1.97 L) was added dropwise to a solution of isobutyl chloroformate (111 mL) in tetrahydrofuran (1.32 L) under ice cooling. After stirring for 1 hour, the reaction solution was filtered. The filtrate obtained was added dropwise to a mixed solution of glycinamide hydrochloride (196 g), sodium hydroxide (70.6 g), tetrahydrofuran (1.02 L), and water (510 mL) under ice cooling. After stirring overnight at room temperature, an aqueous NaCl solution was added to the reaction solution, and the mixture was extracted with ethyl acetate. The organic layer obtained was washed with an aqueous 1M sodium hydroxide solution, dried over magnesium sulfate, filtered, and the filtrate obtained was concentrated. Intermediate 2 (209 g) was obtained by the above operations.

[0083] To a solution of intermediate 2 in dichloromethane (1.39 L), trifluoroacetic acid (663 mL) was added dropwise. After stirring for 1 hour, the reaction solution was concentrated to obtain viscous intermediate 3 (229 g).

[0084] Sodium carbonate (243 g) was added to a solution of intermediate 3 (229 g) in methanol (1.76 L), cooled to -20°C or lower, and acryloyl chloride (185 mL) was added dropwise. After stirring for 3 hours, the reaction solution was filtered. The filtrate was concentrated, and an acetonitrile-methanol solvent (acetonitrile:methanol = 2:1 (volume ratio) (1.0 L) was added to the residue, and the mixture was filtered. The filtrate was concentrated, and the resulting crude product was purified by silica gel column chromatography to obtain the desired acryloyl-N-(2-amino-2-oxoethyl)piperidine-3-carboxamide (66.8 g). 1 H- 13 The C HSQC NMR chart is shown in Figure 4.

[0085] <Example 5: Synthesis of poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end> [ka] N-acryloyl nipecotamide (0.99 g), cyanomethyl dodecyl trithiocarbonate (21.7 mg), and 2,2'-azobis(isobutyronitrile) (2.3 mg) were added to dimethyl sulfoxide (10 mL) and degassed by the freeze-thaw method, after which a polymerization reaction was carried out at 70°C for 1 hour. The reaction solution was then dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (10 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover poly(N-acryloyl nipecotamide) having dodecyl trithiocarbonate groups at the end groups. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 7,500 and 1.32, respectively.

[0086] <Example 6: Synthesis of poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end> N-acryloyl nipecotamide (1.0 g), cyanomethyl dodecyl trithiocarbonate (12.2 mg), and 2,2'-azobis(isobutyronitrile) (1.3 mg) were added to dimethyl sulfoxide (10 mL) and degassed by the freeze-thaw method, after which a polymerization reaction was carried out at 70°C for 6 hours. The reaction solution was then dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (10 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover poly(N-acryloyl nipecotamide) having dodecyl trithiocarbonate groups at the end groups. The recovered weight was 0.86 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 12,400 and 1.35, respectively.

[0087] <Example 7: Synthesis of poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end> [ka] Poly(N-acryloyl-nipecotamide) (520 mg) having dodecyl trithiocarbonate groups at the end groups synthesized in Example 5, maleimide (2.8 g), and 2-aminoethanol (800 μL) were added to dimethyl sulfoxide (3.5 mL) and stirred for 96 hours at room temperature under a nitrogen atmosphere. After stirring, the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (3.5 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover poly(N-acryloyl-nipecotamide) having thiomaleimide groups at the end groups. The weight of the recovered polymer was 0.42 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 7,500 and 1.29, respectively.

[0088] <Example 8: Synthesis of poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end> Poly(N-acryloyl-nipecotamide) (526 mg) having dodecyl trithiocarbonate groups at the end groups synthesized in Example 6, maleimide (56.5 mg), and 2-aminoethanol (55 μL) were added to dimethyl sulfoxide (3.5 mL) and stirred for 96 hours at room temperature under a nitrogen atmosphere. After stirring, the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (6.0 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover poly(N-acryloyl-nipecotamide) having thiomaleimide groups at the end groups. The weight of the recovered polymer was 0.49 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 13,200 and 1.30, respectively.

[0089] Example 9: Poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end and poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end 1 H-NMR measurement> The polymers synthesized in Examples 5 and 7 1 H NMR measurement was performed (Figure 5). The signal of the methyl group derived from the terminal dodecyltrithiocarbonate group at around 0.8 ppm (Figure 5, top) disappeared after the terminal treatment (Figure 5, bottom). This indicated that the polymer terminal group was converted to a maleimide group.

[0090] <Example 10: Measurement of the upper critical solution temperature of aqueous solutions of poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end and poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end> The upper critical solution temperature of the aqueous solutions of the polymers synthesized in Examples 6 and 8 dissolved in phosphate buffered saline was measured (Figure 6). Poly(N-acryloyl-nipecotamide) with a dodecyltrithiocarbonate group at one end synthesized in Example 6 showed UCSTs at around 70°C at a concentration of 0.4 wt%, around 55°C at a concentration of 0.3 wt%, and around 45°C at a concentration of 0.1 wt%, and it was confirmed that the UCST was higher than 70°C at higher polymer concentrations (Figure 6, left). Poly(N-acryloyl-nipecotamide) with a thiomaleimide group at the end group synthesized in Example 8 showed UCSTs at around 55°C to 30°C at concentrations of 8.0 wt% to 2.0 wt% (Figure 6, right). It was confirmed that the UCST was lower than 1°C at a concentration of 1.0 wt%. From these comparisons, it was confirmed that the UCST could be shifted to a lower temperature by introducing a thiomaleimide group into the terminal functional group.

[0091] <Example 11: Synthesis of poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end> N-acryloyl nipecotamide (1.0 g), cyanomethyl dodecyl trithiocarbonate (10.6 mg), and 2,2'-azobis(isobutyronitrile) (1.1 mg) were added to dimethyl sulfoxide (10 mL) and degassed by the freeze-thaw method, after which a polymerization reaction was carried out at 70°C for 4 hours. The reaction solution was then dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (10 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover poly(N-acryloyl nipecotamide) with dodecyl trithiocarbonate groups at the end groups. The recovered weight was 0.97 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 14,100 and 1.36, respectively.

[0092] <Example 12: Synthesis of poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end> Poly(N-acryloyl-nipecotamide) (680 mg) having a dodecyltrithiocarbonate group at the end group synthesized in Example 11, maleimide (148 mg), and 2-aminoethanol (126 μL) were added to dimethyl sulfoxide (7 mL) and stirred for 120 hours at room temperature under a nitrogen atmosphere. After stirring, the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (7 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover poly(N-acryloyl-nipecotamide) having a thiomaleimide group at the end group. The weight of the recovered polymer was 620 mg. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 14,800 and 1.32, respectively.

[0093] <Example 13: Measurement of upper critical solution temperature of aqueous solutions of poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end and poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end> The upper critical solution temperature of the aqueous solutions of the polymers synthesized in Examples 11 and 12 dissolved in phosphate buffered saline was measured (Figure 7). Poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end in Example 11 showed UCST at around 70°C to around 20°C at a concentration of 0.6 to 0.1 wt%, and it was confirmed that the UCST was higher than 70°C at higher polymer concentrations (Figure 7, left). On the other hand, poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end (polymer synthesized in Example 12) showed UCST at around 60°C to around 10°C at a concentration of 6.0 wt% to 1.0 wt% (Figure 7, right). From these comparisons, it was confirmed that the UCST changes depending on the change in the terminal functional group, and from comparison with Figure 6, it was confirmed that it changes depending on the molecular weight.

[0094] <Example 14: Synthesis of poly(N-acryloyl-nipecotamide) having trithiocarboxyethyl and 4-cyanopentanoic acid groups at the polymer end and its behavior in aqueous solution> N-acryloylnipecotamide (0.99 g), 4-((((2-carboxyethyl)thio)carbonothioyl)thio)-4-cyanopentanoic acid (8.3 mg), and 2,2'-azobis(isobutyronitrile) (0.9 mg) were added to dimethyl sulfoxide (10 mL) and the reaction solution was degassed by the freeze-thaw method, and then polymerization reaction was carried out at 70°C for 3 hours. The reaction solution was then dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (10 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to obtain poly(N-acryloyl-nipecotamide) with trithiocarboxyethyl and 4-cyanopentanoic acid groups at the ends. The collected weight was 0.83 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 34,300 and 2.83, respectively. The obtained polymer was extremely insoluble even in phosphate buffered saline at 70°C (Table 1, Run 6), confirming that the UCST was a temperature higher than 70°C.

[0095] [Table 1]

[0096] <Example 15: Synthesis of poly(N-acryloyl-nipecotamide) having trithiocarboxyethyl and 4-cyanopentanoic acid groups at the polymer end and measurement of the upper critical solution temperature in aqueous solution> N-acryloylnipecotamide (1.0 g), 4-((((2-carboxyethyl)thio)carbonothioyl)thio)-4-cyanopentanoic acid (8.4 mg), and 4,4'-azobis(4-cyanovaleric acid) (1.54 mg) were added to dimethyl sulfoxide (10 mL) and degassed by the freeze-thaw method, after which a polymerization reaction was carried out at 65°C for 24 hours. The reaction solution was then dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (10 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to obtain poly(N-acryloyl-nipecotamide) with trithiocarboxyethyl and 4-cyanopentanoic acid groups at the ends. The collected weight was 0.94 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 14,100 and 1.36, respectively (Table 1, Run 7). The upper critical solution temperature of the obtained polymer in an aqueous solution was measured (Figure 8). It was dissolved in phosphate buffered saline at 5.0 wt% to 3.0 wt%, and showed a UCST near 20°C to 5°C (Figure 8, left). At 2.0 wt%, it was shown that the UCST was at a temperature lower than 5°C. It was revealed that the UCSTs were completely different when comparing the aqueous solutions of poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end and poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end, both of which have similar number-average molecular weights as described in Figure 7 and Example 13. It was revealed that in the case of a hydrophobic end group, the UCST of the polymer shifts to higher temperatures, and in the case of a hydrophilic end group, the UCST shifts to lower temperatures.

[0097] <Example 16: Synthesis of poly(N-acryloyl-nipecotamide) having trithiocarboxyethyl and 4-cyanopentanoic acid groups at the polymer end and measurement of the upper critical solution temperature in aqueous solution> N-acryloylnipecotamide (1.0 g), 4-((((2-carboxyethyl)thio)carbonothioyl)thio)-4-cyanopentanoic acid (14.8 mg), and 4,4'-azobis(4-cyanovaleric acid) (2.61 mg) were added to dimethyl sulfoxide (10 mL) and degassed by the freeze-thaw method, after which a polymerization reaction was carried out at 65°C for 21 hours. The reaction solution was then dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (10 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to obtain poly(N-acryloyl-nipecotamide) with trithiocarboxyethyl and 4-cyanopentanoic acid groups at the ends. The collected weight was 0.83 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 10,200 and 1.39, respectively (Table 1, Run 8). The obtained polymer was dissolved even in phosphate buffered saline at 2°C, confirming that the UCST was a temperature lower than 5°C.

[0098] <Example 17: Synthesis of poly(N-acryloyl-nipecotamide) having trithioethyl and 4-cyanopentanoic acid groups at the polymer end and measurement of the upper critical solution temperature in aqueous solution> N-acryloylnipecotamide (1.0 g), 4-cyano-4-(((ethylthio)carbonothionyl)thio)pentanoic acid (7.2 mg), and 4,4'-azobis(4-cyanovaleric acid) (1.54 mg) were added to dimethyl sulfoxide (10 mL) and degassed by the freeze-thaw method, after which a polymerization reaction was carried out at 65°C for 1 hour. The reaction solution was then dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (10 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to obtain poly(N-acryloyl-nipecotamide) with trithiocarboxyethyl and 4-cyanopentanoic acid groups at the ends. The collected weight was 0.78 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 16,400 and 1.39, respectively (Table 1, Run 9). The upper critical solution temperature of the obtained polymer in an aqueous solution was measured (Figure 8). It was dissolved in phosphate buffered saline at 5.0 wt% to 3.0 wt%, and showed a UCST from around 23°C to around 7°C (Figure 8, right). At 2.0 wt%, it was shown that the UCST was at a temperature lower than 7°C. It was revealed that the UCSTs were completely different when comparing the aqueous solutions of poly(N-acryloyl-nipecotamide) having a dodecyltrithiocarbonate group at one end and poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end, both of which have similar number average molecular weights as described in Figure 7 and Example 13. It was revealed that in the case of a hydrophobic end group, the UCST of the polymer shifts to higher temperatures, and in the case of a hydrophilic end group, the UCST shifts to lower temperatures.

[0099] <Example 18: Synthesis of poly(acryloyl-N-(2-amino-2-oxoethyl)piperidine-3-carboxamide-co-acryloyl-N-piperidine-4-carbonitrile) and measurement of its upper critical solution temperature in aqueous solution> [ka] N-acryloyl-4-piperidinecarbonitrile (0.11 g) synthesized in Example 2, acryloyl-N-(2-amino-2-oxoethyl)piperidine-3-carboxamide (0.3 g) synthesized in Example 4, and 2,2'-azobis(isobutyronitrile) (1.66 mg) were dissolved in dimethyl sulfoxide (5 mL), and the reaction solution was degassed by the freeze-thaw method, and then polymerization reaction was carried out at 70°C for 3 hours. The reaction solution was then dropped into an acetone solution to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (5 mL), and the reaction solution was dropped into acetone to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover a copolymer of poly(acryloyl-N-(2-amino-2-oxoethyl)piperidine-3-carboxamide and acryloyl-N-piperidine-4-carbonitrile). The collected weight was 0.28 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 55,400 and 5.04, respectively. The upper critical solution temperature of the obtained polymer in an aqueous solution was measured (Figure 9). The polymer was dissolved in phosphate buffered saline at 5.0 wt% to 1.0 wt%, and showed UCST from around 55°C to around 48°C (Figure 9). At a concentration of 0.5 wt%, UCST was confirmed at a temperature lower than 2°C.

[0100] <Example 19: Synthesis of poly(N-acryloyl-3,5-piperidinedicarboxamide) and measurement of upper critical solution temperature in aqueous solution> [ka] N-acryloyl-3,5-piperidinedicarboxamide (0.3 g), 2-(2-carboxyethylsulfanylthiocarbonylsulfanyl)propionic acid (8.9 mg), and 4,4'-azobis(4-cyanovaleric acid) (1.9 mg) synthesized in Example 3 were dissolved in dimethyl sulfoxide (5 mL), and the reaction solution was degassed by the freeze-thaw method, and then polymerization reaction was carried out at 65°C for 26 hours. Thereafter, the reaction solution was dropped into an acetone solution to precipitate a polymer. The precipitated polymer was filtered and collected, and dissolved again in dimethyl sulfoxide (5 mL), and the reaction solution was dropped into acetone to precipitate a polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover poly(N-acryloyl-3,5-piperidinedicarboxamide) having a trithiocarboxyethyl group and a propionic acid group at the end. The recovered weight was 0.28 g. The upper critical solution temperature of the obtained polymer in an aqueous solution was measured (Figure 10). It was dissolved in phosphate buffered saline at 4.0 wt% to 1.0 wt%, and showed a UCST at around 8°C to 5°C. At a concentration of 0.5 wt%, a UCST was confirmed at a temperature lower than 2°C.

[0101] <Example 20: Synthesis of poly(N-acryloyl-isonipecotamide-co-acrylonitrile) copolymer and measurement of upper critical solution temperature in aqueous solution> [ka] N-Acryloyl-isonipecotamide (0.3 g), acrylonitrile (46.6 mg), 2,2'-azobis(isobutyronitrile) (3.36 mg), 4-((((2-carboxyethyl)thio)carbonothioyl)thio)-4-cyanopentanoic acid (19.0 mg) were dissolved in dimethyl sulfoxide (4 mL), and the reaction solution was degassed by the freeze-thaw method, and then polymerization reaction was carried out at 65°C for 24 hours. The reaction solution was then dropped into an acetone solution to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (5 mL), and the reaction solution was dropped into acetone to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to obtain poly(N-acryloyl-isonipecotamide-co-acrylonitrile) copolymer having a trithiocarboxyethyl group and a 4-cyanopentanoic acid group at the end. The obtained polymer was dissolved in phosphate buffered saline at 3.0 wt%, and the change in turbidity of the aqueous solution due to temperature change was observed. The UCST was observed from around 15°C to around 13°C (Figure 11). At a concentration of 2.0 wt%, the polymer did not show an upper critical solution temperature and remained in a dissolved state.

[0102] Poly(N-acryloyl-nipecotamide: PNANAm), poly(N-acryloyl-isonipecotamide: PNAiNAm), and poly(N-acryloyl-piperidine-2-carboxamide: PNP2Cam) were prepared by RAFT polymerization according to the polymerization conditions in Table 2 below. [Table 2] According to the results in Table 2, poly(N-acryloyl-piperidine-2-carboxamide: PNP2CAm) is a poorly soluble polymer, PNANA has UCST, and dissolved PNAiNAm is a soluble polymer. Dissolved PNAiNAm became a polymer with UCST by copolymerization with acrylonitrile, which is believed to constitute a poorly soluble polymer.

[0103] <Example 21: Synthesis of poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end and evaluation of its cytotoxicity> N-acryloyl nipecotamide (0.99 g), cyanomethyl dodecyl trithiocarbonate (43.2 mg), and 2,2'-azobis(isobutyronitrile) (4.5 mg) were added to dimethyl sulfoxide (10 mL) and degassed by the freeze-thaw method, after which a polymerization reaction was carried out at 70°C for 24 hours. The reaction solution was then dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (10 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover poly(N-acryloyl nipecotamide) having dodecyl trithiocarbonate groups at the end groups. The recovered weight was 0.92 g. The obtained polymer (0.2 g), 2-aminoethanol (195 μL), and maleimide (0.33 g) were added to dimethyl sulfoxide (4 mL) and stirred at room temperature for 6 days under a nitrogen atmosphere. After stirring, the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer. The precipitated polymer was filtered, collected, and dissolved again in dimethyl sulfoxide (4 mL), and the reaction solution was dropped into an acetone-methanol mixed solution (acetone:methanol = 4:1 (volume ratio)) to precipitate the polymer again. The precipitated polymer was filtered, collected, and vacuum dried to recover poly(N-acryloyl-nipecotamide) having thiomaleimide groups at the terminal groups. The weight of the recovered polymer was 0.16 g. Evaluation by gel filtration chromatography (column: TSKgel SuperH4000) confirmed that the number average molecular weight (Mn) and molecular weight distribution (Mw / Mn) of the polymer were 3,900 and 1.19, respectively. The obtained polymer was dissolved in Dulbecco's modified Eagle's medium (containing 10% serum) at concentrations of 0.1 mg / mL to 6.0 mg / mL, added to the medium of previously cultured bovine vascular endothelial cells, and cultured for 24 hours. After culture, cell viability was evaluated using Cell Counting Kit-8 (Dojin Chemical), and the cell viability was equivalent to that of the control, confirming that the polymer was not toxic (Figure 12).Furthermore, at polymer concentrations of 2.0 mg / mL or higher, cell proliferation was improved (FIG. 12).

[0104] <Example 22: Evaluation of particle size of poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end by light scattering> The polymer synthesized in Example 12 was dissolved in water and the particle size was measured by light scattering at 45°C (above UCST) and 20°C (below UCST). Above UCST, the particle size was about 10 nm, and below UCST, the particle size showed aggregation of about 110 um, confirming that the aggregation of polymer chains changes with temperature (Figure 13). [Industrial Applicability]

[0105] According to the present invention, it has been demonstrated that UCST can be controlled by changing the end group of a (meth)acrylamide-based polymer having a piperidine carboxamide structure in the polymer side chain, and that these polymers have extremely low cytotoxicity and accelerate cell proliferation, thereby contributing to the proliferation effect of cultured cells that could not be confirmed with conventional UCST-type polymers.

Claims

1. Formula I 【Chemistry 1】 (In the formula, R １１ is H or CH ３ and R arranged in a ring structure １２ ~R １６ 1 to 3 H atoms are -C(=O)NH ２ is replaced by (However, in the case of 1, this is limited to poly(N-acryloyl-nipecotamide) having a thiomaleimide group at one end, poly(N-acryloyl-nipecotamide) having a trithiocarboxyethyl group and a 4-cyanopentanoic acid group at the end, and poly(N-acryloyl-nipecotamide) having a trithioethyl group and a 4-cyanopentanoic acid group at the end.) m is an integer of 2 or more. or a homopolymer having a structural unit represented by Formula II 【Chemistry 2】 (In the formula, R ２１ and R ３１ are each independently H or CH ３ and R arranged in a ring structure ２２ ~R ２６ 1 to 3 H atoms are independently -C(=O)NH ２ Or -C(=O)-NH-CH ２ -C(=O)-NH ２ is replaced by R arranged in a ring structure ３２ ~R ３６ 1 to 3 H atoms are independently -CN or -C(=O)-NH ２ is replaced by m is an integer of 1 or more, and n is an integer equal to or greater than 1.) or Formula III 【Chemistry 3】 (In the formula, R ４１ and R ５１ are each independently H or CH ３ and R arranged in a ring structure ４２ ~R ４６ 1 to 3 H atoms are independently -CN, -C(=O)NH ２ Or -C(=O)-NH-CH ２ -C(=O)-NH ２ is replaced by R ５２ is -CN, -C(=O)NH ２ Or -C(=O)-O-C ２ H ４ OH, m is an integer of 1 or more, wherein n is an integer of 1 or more.

2. The constitutional unit represented by formula I is 【Chemistry 4】 (In the formula, R １１ , m is as defined in formula (I).

2. The polymer of claim 1 ,

3. The constitutional unit represented by formula II is 【Chemistry 5】 【Chemistry 6】 【Chemistry 7】 or 【Chemistry 8】 (In the formula, R ２１ , R ３１ , m and n are as defined in formula (II).

2. The polymer of claim 1 ,

4. The constitutional unit represented by formula III is 【Chemistry 9】 or 【Chemistry 10】 (In the formula, R ４１ , R ５１ , m and n are as defined in formula (III).

2. The polymer of claim 1 ,

5. 2. The polymer of claim 1, wherein the end group comprises a C(=O)OH group, a maleimide group, a CN group, a SH group, an amino group, or an OH group.

6. The polymer of claim 1 synthesized by radical polymerization.

7. 10. The polymer of claim 1 synthesized by RAFT (reversible addition-fragmentation chain transfer) polymerization.

8. The polymer of claim 7 , modified at both ends with reactant residues derived from a chain transfer agent in RAFT polymerization.

9. A chain transfer agent for RAFT polymerization, Y-X-Z, where X is a thiocarbonylthio group (-C(=S)-S-) or a trithiocarbonate group (-S-C(=S)-S-), Y and Z are each independently H, a phenyl group, or a straight-chain or branched alkyl group which may be substituted with one or more C(=O)OH groups, phenyl groups, or CN groups. 【Chemistry 11】 (In the formula, R １１ ~R １６ and m are as defined in formula (I). and Formula II is Formula II-1 【Chemistry 12】 (In the formula, R ２１ ~R ２６ , R ３１ ~R ３６ , m and n are as defined in formula (II); or formula III is formula III-1 【Chemistry 13】 (In the formula, R ４１ ~R ４６ , R ５１ ~R ５２ 10. The polymer of claim 8, wherein m and n are as defined in formula (III).

10. 2. The polymer according to claim 1, characterized in that it has a weight average molecular weight Mw of 1,000 to 700,000.

11. 2. The polymer according to claim 1, wherein the ratio Mw / Mn of the number average molecular weight Mn to the weight average molecular weight Mw is 1.00 to 7.

00.

12. 2. The polymer of claim 1, wherein the UCST is from 3 to 80° C.

13. The polymer of claim 1 which is non-cytotoxic.

14. The polymer of claim 1 which promotes cell proliferation.

15.

14. (In the formula, R ６ is H or CH ３ It is.) A compound represented by the formula:

16.

15. (In the formula, R ７ is H or CH ３ It is.) A compound represented by the formula:

17. A compound according to claim 15 or 16 for the preparation of a polymer according to claims 1 to 14.

18. A method for producing a polymer according to any one of claims 1 to 14, comprising the steps of: One or two types of 【Chemistry 16】 (In the formula, R ８１ is H or CH ３ and R arranged in a ring structure ８２ ~R ８６ 1 to 3 H atoms are independently -CN, -C(=O)NH ２ Or -C(=O)-NH-CH ２ -C(=O)-NH ２ is replaced by.) A method for radical polymerization using a monomer represented by the following formula:

19. 19. The method according to claim 18, wherein the radical polymerization is carried out using a radical polymerization initiator or a radical initiator and a chain transfer agent for RAFT polymerization.

20. A method for changing an upper critical solution temperature (UCST) of a polymer obtained by polymerizing or copolymerizing a monomer mainly composed of N-acryloyl-piperidinecarboxamide in an aqueous solution, comprising: The method is Changing the terminal modification group The method according to the above.

21. 21. The method of claim 20, wherein the terminal group comprises a C(=O)OH group, a maleimide group, a CN group, a SH group, an amino group, or an OH group.

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