JP5526317B2 - Temperature sensitive polymer compound and temperature sensitive drug release system - Google Patents

Description

  The present invention relates to a temperature-sensitive polymer compound that rapidly undergoes a phase transition from hydrophilic to hydrophobic at a specific temperature, and a temperature-sensitive drug release system comprising the temperature-sensitive polymer compound and a drug.

  The drug delivery system (hereinafter sometimes referred to as “DDS”) wraps the drug in a membrane, etc., so that it reaches the affected area without being absorbed or decomposed on the way, and releases the drug at the affected area, thereby having a therapeutic effect. In addition to enhancing the therapeutic effect of the drug, it exhibits an excellent therapeutic effect with the minimum amount of drug used, and is therefore attracting attention because it has the advantage of reducing side effects. Usually, the drug is absorbed and decomposed in the body, or diffuses to the whole body, that is, to a site other than the affected area, so that the drug that reaches the affected area is said to be a very small amount of the administered dose. This is because when the amount to be administered is determined by calculating back from the amount of drug to be reached, the dose becomes very large and the possibility of side effects increases.

  In recent years, a DDS for delivering a drug to a target site, which is composed of a liposome prepared using a phospholipid derived from a living body and a drug such as an anticancer drug or a therapeutic gene, has been developed. Liposomes are composed of lipid bilayers composed of phospholipids, etc., and are closed vesicles with a particle size of about several tens to several hundreds of nanometers and function as nanocapsules that can encapsulate various substances. Liposomes used for such applications include encapsulating a hydrophilic drug in a hydrophilic region inside the closed endoplasmic reticulum and / or carrying a hydrophobic drug on a lipid bilayer membrane. Synthesized. And it can be made to take in selectively to an affected part by adjusting the particle diameter of a liposome. However, in order to release the drug from inside the liposome that has reached the affected area, it is necessary to destroy the liposome by external stimulation, and it has been difficult to smoothly release the drug. In addition, the residence time in the blood can be extended by converting the liposome to polyethylene glycol, but since the hydrophilicity is improved when polyethylene glycol is converted, it is difficult to adsorb to the affected area. .

  On the other hand, a dendrimer is a high molecular compound having a highly branched structure and a single polymerization degree, and it is known that various substances can be encapsulated inside the compound and can be used for DDS. Development of DDS using dendrimers has been widely conducted, and dendrimers that can adjust the residence time in blood vessels, select target cells, and enhance stimulus responsiveness (especially temperature responsiveness) have been studied. ing. A temperature-responsive dendrimer can undergo a phase transition from a hydrophilic substance to a hydrophobic substance by increasing the temperature. It acts as a base for transporting the drug during transportation in the blood, and when it reaches the heated affected area, it becomes a hydrophobic substance. By making a phase transition and being easily taken into the target cell, the drug can be quickly and selectively delivered to the target cell (Non-patent Document 1). As a temperature-responsive dendrimer, for example, DDS using a polyamidoamine dendrimer has been developed (Patent Document 1). However, since polyamide amine dendrimers are questionable in biocompatibility and are expensive, there is a need for temperature-sensitive polymer compounds that are more biocompatible and can be provided at low cost. I came.

Kenji Kawano, Chie Kojima, Gene Medicine MOOK volume, “Nano DDS that can be seen in the picture-Treatment, diagnosis, prognosis and prevention from the viewpoint of materials, the latest in healthcare technology”, p. 106-112 (2007), Medical Do

JP 2006-312689 A

Accordingly, an object of the present invention is to provide a novel temperature-sensitive polymer compound that is excellent in biocompatibility and can be provided at low cost.
Another object of the present invention is to provide a temperature sensitive drug release system comprising a temperature sensitive polymer compound having the above properties and a drug.

In order to further enhance the practicality of the temperature-sensitive polymer compound encapsulating a drug in the pharmaceutical field, it is desired to have the following characteristics.
(1) The drug can be stably maintained at a temperature close to the body temperature of mammals, particularly humans, that is, around 34 to 38 ° C.
(2) The drug can be rapidly released at a temperature away from the body temperature of mammals, particularly humans, that is, 39 ° C. or higher, preferably 40 to 45 ° C.
(3) Long residence time in blood in mammals, particularly humans.

  As a result of intensive studies to solve the above problems, the present inventors have a novel temperature-sensitive polymer having a polyglycerol skeleton (particularly a highly branched polyglycerin skeleton) at the center and a temperature-sensitive group at the terminal. The compound has the characteristics (1) to (3) described above, is excellent in biocompatibility, can be provided at low cost, and can be suitably used in a drug release system. completed.

［式中、ＧＬはグリセリン残基、ＸはＧＬ側に連結基としてのカルボニル結合（−ＣＯ−）を有する炭素数１〜１０のアルキレン基、Ｒはメチルカルバモイル基、エチルカルバモイル基、プロピルカルバモイル基、イソプロピルカルバモイル基、ブチルカルバモイル基、又は、Ｘとの結合部位に連結基としてのカルボニル結合（−ＣＯ−）を有する重合度が２〜１０のポリアルキレングリコール残基（但し、前記ポリアルキレングリコール残基の末端水酸基は、炭素数１〜１０のアルキル基、３〜１０員のシクロアルキル基、炭素数６〜１０のアリール基、炭素数７〜１２のアラルキル基、炭素数１〜１１のアシル基から選択される有機基で封止されていてもよい）を示す］
で表される繰り返し単位を有する温度感受性高分子化合物を提供する。 That is, the present invention has a polyglycerin skeleton as the main chain, and at least the following formula (a):

[ Wherein, GL is a glycerin residue, X is a C 1-10 alkylene group having a carbonyl bond (—CO—) as a linking group on the GL side , R is a methylcarbamoyl group, an ethylcarbamoyl group, a propylcarbamoyl group , An isopropyl carbamoyl group, a butyl carbamoyl group, or a polyalkylene glycol residue having a carbonyl bond (—CO—) as a linking group at the bonding site with X (however, the polyalkylene glycol residue) The terminal hydroxyl group is an alkyl group having 1 to 10 carbon atoms, a 3 to 10-membered cycloalkyl group, an aryl group having 6 to 10 carbon atoms, an aralkyl group having 7 to 12 carbon atoms, or an acyl group having 1 to 11 carbon atoms. It may be sealed with an organic group selected from
The temperature sensitive high molecular compound which has a repeating unit represented by this is provided.

The temperature-sensitive polymer compound has a molecular weight of 120 to 12000 and is a monovalent organic having a functional group capable of introducing R by reaction into polyglycerol having a primary hydroxyl group of 50% or more of all hydroxyl groups. introducing a precursor X ¹ of the X is a group, then it may also be obtained by introducing said R.

In the temperature-sensitive polymer compound, the glycerin residue: the X and the precursor thereof: the R (molar ratio) is preferably 1: 0.5 to 1: 0.5-1.

（式中、Ｘ¹は、反応により前記Ｒを導入可能な官能基を有する１価の有機基である前記Ｘの前駆体を示し、Ｘ²は前記Ｒを導入した後のＸ¹に対応する２価の基を示す。ＧＬはグリセリン残基を示し、Ｒは前記に同じ。ｍ、ｎは同一又は異なって１以上の整数を示す）
で表される高分子化合物が含まれる。 The temperature sensitive polymer compound includes the following formula (1):

(In the formula, X ¹ represents a precursor of X which is a monovalent organic group having a functional group capable of introducing R by reaction, and X ² corresponds to X ¹ after introducing R. A divalent group, GL represents a glycerin residue, R is the same as above, and m and n are the same or different and represent an integer of 1 or more)
The high molecular compound represented by these is included.

As X ¹ , the following formula (2)
_{-CO- (CH 2) s- (CO} ) t-OH (2)
(In the formula, s is an integer of 1 to 10, and t represents 1 )
The monovalent organic group represented by these is mentioned.

Examples of the polyalkylene glycol residue include oligoethylene glycol residues having a degree of polymerization of oligoethylene glycol of 2 to 10 . Examples of R include an isopropylcarbamoyl group.

  The present invention also provides a temperature sensitive drug release system comprising the above temperature sensitive polymer compound and a drug.

  In this temperature sensitive drug release system, the drug amount is preferably 0.1 to 30 mol% with respect to the amount of the temperature sensitive polymer compound.

  The temperature-sensitive polymer compound of the present invention requires not only the inexpensive polyglycerin used for the skeleton but also the time and cost required to verify the safety of using it as a DDS material. Since the temperature-sensitive polymer compound according to the present invention can be formed by a fatty acid ester or the like that has been approved for use as a food additive, excellent biocompatibility and safety have already been established. And according to the temperature sensitive high molecular compound which concerns on this invention, a more practical temperature sensitive chemical | medical agent discharge | release system can be provided.

  For example, when a target cancer cell is heated to about 40 to 45 ° C., a temperature sensitive drug release system in which an anticancer drug is encapsulated as a drug is administered to the temperature sensitive polymer compound according to the present invention. The anti-cancer agent can be efficiently and specifically allowed to act on the cancer cells, and treatment can be effectively performed while reducing side effects.

1 is a diagram showing ¹ H-NMR measurement results of succinylated polyglycerin obtained in Example 1. FIG. 1 is a diagram showing ¹ H-NMR measurement results of an isopropylamide-terminated polyglycerin derivative obtained in Example 1. FIG. It is a figure which shows the temperature sensitivity and pH sensitivity of the temperature sensitive drug release system of this invention (evaluation method 1). The vertical axis represents transmittance (%), and the horizontal axis represents temperature (° C.). In addition, a graph shows the case of pH5, pH5.2, pH5.4, pH6.5 from the left. It is a figure which shows the temperature sensitivity of the temperature sensitive chemical | medical agent release system of this invention (evaluation method 2). The vertical axis represents transmittance (%), and the horizontal axis represents temperature (° C.). It is a figure which shows the RB discharge | release capability in the temperature sensitive chemical | medical agent release system of this invention (evaluation method 3). The vertical axis represents absorbance (abs), and the horizontal axis represents light wavelength (nm). FIG. 4 is a diagram showing the ¹ H-NMR measurement result of the oligoethylene glycol derivative group-terminated polyglycerin derivative obtained in Example 4. FIG. 6 shows ¹ H-NMR measurement results of the oligoethylene glycol-derived group-terminated polyglycerin derivative obtained in Example 5. FIG. 4 is a diagram showing the ¹ H-NMR measurement result of the oligoethylene glycol derivative group-terminated polyglycerin derivative obtained in Example 6. In the evaluation method 4, it is a figure which shows the result using the oligoethylene glycol derivative group terminal polyglycerin derivative obtained in Examples 4-6. The vertical axis represents transmittance (%), and the horizontal axis represents temperature (° C.). In the evaluation method 4, it is a figure which shows the result using the oligoethylene glycol derivative group terminal polyglycerin derivative obtained in Examples 5-9. The vertical axis represents transmittance (%), and the horizontal axis represents temperature (° C.).

[Temperature sensitive polymer compound]
The temperature-sensitive polymer compound according to the present invention has a polyglycerin skeleton as a main chain and has at least a repeating unit represented by the formula (a). In formula (a), GL represents a glycerin residue, X represents a linker, and R represents a thermosensitive group.

This temperature-sensitive polymer compound has, for example, a functional group capable of introducing a heat-responsive group into a polyglycerin having a molecular weight of 120 to 12000 and a primary hydroxyl group of 50% or more of all hydroxyl groups. It can be obtained by introducing a linker precursor X ¹ which is a monovalent organic group and subsequently introducing a thermosensitive group R. The linker precursor X ¹ becomes the linker X after the introduction of the thermosensitive group.

This temperature sensitive polymer compound includes the polymer compound represented by the formula (1). In formula (1), X ¹ represents a precursor of a linker which is a monovalent organic group having a functional group capable of introducing a heat-responsive group by reaction, and X ² represents a group after introducing the heat-responsive group. A linker (ie, X) which is a divalent group corresponding to X ¹ is shown. GL represents a glycerin residue, and R represents a thermosensitive group. m and n are the same or different and represent an integer of 1 or more. Each repeating unit in parentheses may be present as a block or may be present at random.

The GL (glycerin residue) has any structure represented by the following formulas (3) and (4).

X ¹ represents a monovalent organic group having a functional group capable of introducing a heat-responsive group by reaction. Examples of the “monovalent organic group” include a hydrocarbon group, a heterocyclic group, a carbonyl group, and the like. And a group in which two or more are bonded via or without a linking group. The number of carbon atoms of the monovalent organic group in X ^1, for example, 1 to 30, preferably 4 to 15. Examples of the linking group include an ether bond (—O—), a thioether bond (—S—), an amine bond (—NR′—), a carbonyl bond (—C (═O) —), and an ester bond (—C). (= O) -O-), silyl bond (-Si-), carbonate bond, amide bond, urea bond, urethane bond and the like. R ′ represents an alkyl group or a hydrogen atom.

  Examples of the functional group capable of introducing a thermosensitive group by the reaction include a halogen atom, an oxo group, a hydroxyl group, a substituted oxy group (for example, an alkoxy group, an aryloxy group, an aralkyloxy group, an acyloxy group, etc.), a carboxyl group Group, substituted oxycarbonyl group (alkoxycarbonyl group, aryloxycarbonyl group, aralkyloxycarbonyl group, etc.), substituted or unsubstituted carbamoyl group, cyano group, isocyanate group, nitro group, acyl group, substituted or unsubstituted amino group, sulfo Groups and the like.

  The hydrocarbon group in the monovalent organic group may be a group that does not inhibit this reaction (for example, a group that is non-reactive under the reaction conditions in the present method), such as an aliphatic hydrocarbon group, An alicyclic hydrocarbon group, an aromatic hydrocarbon group, and these combined groups are included. The hydrocarbon group includes a hydrocarbon group having a substituent.

  Examples of the aliphatic hydrocarbon group include 1 to 20 carbon atoms (preferably 1 to 1) such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, s-butyl, t-butyl, pentyl, hexyl, decyl, and dodecyl groups. 10, more preferably an alkyl group of about 1 to 3); an alkenyl group of about 2 to 20 carbon atoms (preferably 2 to 10, more preferably 2 to 3) such as vinyl, allyl, 1-butenyl group; Examples thereof include alkynyl groups having about 2 to 20 carbon atoms (preferably 2 to 10, more preferably 2 to 3) such as propynyl groups.

As the alicyclic hydrocarbon group, a cycloalkyl group having about 3 to 20 members (preferably 3 to 15 members, more preferably 5 to 8 members) such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cyclooctyl groups; Cycloalkenyl groups of about 3 to 20 members (preferably 3 to 15 members, more preferably 5 to 8 members) such as pentenyl and cyclohexenyl groups; perhydronaphthalen-1-yl group, norbornyl, adamantyl, tetracyclo [4 4.0.1, ^2,5 . 1 ^7,10 ] bridged cyclic hydrocarbon groups such as dodecan-3-yl groups. Examples of the monovalent aromatic hydrocarbon group include aromatic hydrocarbon groups having about 6 to 14 (preferably 6 to 10) carbon atoms such as phenyl and naphthyl groups.

The hydrocarbon group in which an aliphatic hydrocarbon group and an alicyclic hydrocarbon group are bonded includes a cycloalkyl-alkyl group such as cyclopentylmethyl, cyclohexylmethyl, 2-cyclohexylethyl group (for example, C _3-20 cycloalkyl- C _1-4 alkyl group and the like). The hydrocarbon group in which an aliphatic hydrocarbon group and an aromatic hydrocarbon group are bonded to each other includes an aralkyl group (for example, a C _7-18 aralkyl group) and an alkyl-substituted aryl group (for example, about 1 to about 4). A phenyl group substituted with a C _1-4 alkyl group or a naphthyl group).

  The hydrocarbon group includes various substituents such as halogen atoms, oxo groups, hydroxyl groups, substituted oxy groups (for example, alkoxy groups, aryloxy groups, aralkyloxy groups, acyloxy groups, etc.), carboxyl groups, substituted oxycarbonyls. Groups (alkoxycarbonyl groups, aryloxycarbonyl groups, aralkyloxycarbonyl groups, etc.), substituted or unsubstituted carbamoyl groups, cyano groups, isocyanate groups, nitro groups, acyl groups, substituted or unsubstituted amino groups, sulfo groups, heterocyclic groups It may have a group or the like. The hydroxyl group and carboxyl group may be protected with a protective group commonly used in the field of organic synthesis. In addition, an aromatic or non-aromatic heterocyclic ring may be condensed with a ring of an alicyclic hydrocarbon group or an aromatic hydrocarbon group.

The heterocyclic ring constituting the heterocyclic group in the monovalent organic group includes an aromatic heterocyclic ring and a non-aromatic heterocyclic ring. As the heterocyclic ring, for example, a heterocyclic ring containing an oxygen atom as a hetero atom (for example, 5-membered ring such as furan, tetrahydrofuran, oxazole, isoxazole, γ-butyrolactone ring, 4-oxo-4H-pyran, tetrahydropyran, morpholine 6-membered ring such as a ring, condensed ring such as benzofuran, isobenzofuran, 4-oxo-4H-chromene, chroman, isochroman ring, 3-oxatricyclo [4.3.1.1 ^4,8 ] undecan-2- An on-ring, a bridged ring such as a 3-oxatricyclo [4.2.1.0 ^4,8 ] nonan-2-one ring), a heterocycle containing a sulfur atom as a hetero atom (eg, thiophene, thiazole, iso 5-membered rings such as thiazole and thiadiazole rings, 6-membered rings such as 4-oxo-4H-thiopyran ring, and condensed benzothiophene rings Etc.), heterocycles containing nitrogen atoms as heteroatoms (for example, 5-membered rings such as pyrrole, pyrrolidine, pyrazole, imidazole and triazole rings, 6-membered rings such as pyridine, pyridazine, pyrimidine, pyrazine, piperidine and piperazine rings, indoles) , Indoline, quinoline, acridine, naphthyridine, quinazoline, condensed rings such as purine rings, etc.).

In addition to the substituents that the hydrocarbon group may have, the heterocyclic group includes an alkyl group (eg, a C _1-4 alkyl group such as a methyl or ethyl group), a cycloalkyl group, an aryl group It may have a substituent such as (for example, phenyl, naphthyl group).

X ¹ in the present invention has a hydroxyl group, an isocyanate group, an amino group, or a carboxyl group as a functional group capable of introducing a thermosensitive group by reaction, and a carbonyl bond (—C (═O) — as a linking group). ), A hydrocarbon group having a urea bond and having about 1 to 20 (preferably 1 to 10, more preferably 1 to 3) carbon atoms. X ¹ is preferably —COOH or —COCOOH.

X ¹ is preferably a monovalent organic group represented by the above formula (2). In formula (2), s and t are the same or different and represent an integer of 0 or more. In many cases, t is 0 or 1.

X ² represents a divalent group (linker X) corresponding to X ¹ after the introduction of the thermosensitive group. As X and X ² , an alkylene group having about 1 to 20 carbon atoms (preferably 1 to 10, more preferably 1 to 3) having a linking group at the terminal (GL side) is preferable. Examples of the linking group include an ether bond (—O—), a thioether bond (—S—), an amine bond (—NR′—), a carbonyl bond (—C (═O) —), an ester bond (—C (═O). ) -O-), silyl bond (-Si-), carbonate bond, amide bond, urea bond, urethane bond and the like. Preferred examples of X ² include a methylene group having an ether bond (—O—) and a carbonyl bond (—C (═O) —) at the terminal (GL side), an ethylene group, and a propylene group as a linking group. Can do. In addition, the corresponding group (for example, -CO-) after the reaction of the functional group (for example, -COOH) capable of introducing a thermosensitive group by the reaction in X ¹ is included in R.

In formula (a) and formula (1), R represents a thermosensitive group. The bonding mode of R and X (X ² ) is not particularly limited, but an amide bond, a urethane bond, an ester bond, an ether bond, or the like is preferably used.

  R preferably has a hydrophilic part and a hydrophobic part. And, it is preferably a group that undergoes a phase transition to hydrophobicity as a whole by dehydrating the hydrophilic portion at a predetermined temperature. Examples of R include a group composed of a carbamoyl group having hydrophilicity and a hydrocarbon group having hydrophobicity (N-hydrocarbon group-substituted carbamoyl group). Examples of such a group include a methyl carbamoyl group, an ethyl carbamoyl group, a propyl carbamoyl group, an isopropyl carbamoyl group, and a butyl carbamoyl group. An isopropylcarbamoyl group is preferred.

The R may be a polyalkylene glycol residue in which a terminal hydroxyl group may be sealed with an organic group and may have a linking group at the binding site with the linker (X, X ² ). preferable. In this case, since the temperature sensitive polymer compound can be composed of only carbon atoms, hydrogen atoms and oxygen atoms, biocompatibility can be further enhanced. Examples of the polyalkylene glycol in the polyalkylene glycol residue include polyethylene glycol, polypropylene glycol, polytrimethylene glycol, polybutylene glycol, polytetramethylene glycol, polyoxyethyleneoxypropylene glycol and the like. Among these, polyethylene glycol is preferable. Moreover, as this polyalkylene glycol, oligoalkylene glycol (especially oligoethylene glycol) is preferable, and the polymerization degree is 2-10, for example, Preferably it is 2-5.

  Examples of the organic group for sealing the terminal hydroxyl group include alkyl groups such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, s-butyl, t-butyl, pentyl, and hexyl groups (for example, those having 1 to 10 carbon atoms). Alkyl groups, preferably alkyl groups having 1 to 5 carbon atoms); cycloalkyl groups such as cyclopentyl and cyclohexyl groups (for example, 3 to 10-membered cycloalkyl groups); aryl groups such as phenyl groups (for example, having 6 to 6 carbon atoms) 10 aryl groups); aralkyl groups such as benzyl groups (eg aralkyl groups having 7 to 12 carbon atoms); acyl groups such as acetyl, propionyl, butyryl, pentanoyl, hexanoyl, cyclohexanecarbonyl, benzoyl groups (eg 1 carbon atoms) ˜11 acyl groups, especially aliphatic acyl groups). Among these, as an organic group which seals a terminal hydroxyl group, a C1-C10 alkyl group, especially a C1-C5 alkyl group are preferable.

Examples of the linking group at the binding site of R to X or X ² include, for example, a corresponding group after the reaction of a functional group (for example, —COOH or the like) capable of introducing a thermosensitive group by the reaction at X ¹ (for example, -CO-) and the like.

In the temperature-sensitive polymer compound of the present invention, the glycerin residue GL: linker X and its precursor: heat-responsive group R (molar ratio) can be appropriately selected depending on the type of R, but generally 1: 0.5 It is preferably ˜1: 0.5-1. When the ratio of R is less than 0.5, the temperature sensitivity decreases, and even when the target cell is reached, it tends to be difficult to release the encapsulated drug. The molar ratio of each group can be determined, for example, by ¹ H-NMR.

In the temperature sensitive polymer compound represented by the formula (1), GL: (X ¹ + X ² ): R (molar ratio) is 1: 0.01 to 0.2: 0.003. It may be 0.2.

As described above, the temperature-sensitive polymer compound according to the present invention has, for example, X ¹ added to polyglycerin (PGL) having a molecular weight of 120 to 12000 and a primary hydroxyl group of 50% or more of all hydroxyl groups. It is obtained by introducing and subsequently introducing R. As X ¹ and R, two or more of them may be used in combination.

  The polyglycerin can be synthesized using a polymerizable glycerin equivalent such as glycidol as a raw material. The molecular weight of the polyglycerin is, for example, about 120 to 12000, preferably about 240 to 10,000, and particularly preferably about 350 to 8000. It is. In the temperature-sensitive polymer compound represented by the formula (1), the molecular weight of polyglycerin is, for example, about 120 to 4000, preferably about 240 to 4000, particularly preferably 350 to 4000, depending on the type of R. Degree. When the molecular weight of polyglycerin is less than 120 or more than 12000, it tends to be difficult to adjust the drug release temperature of the temperature sensitive drug release system composed of the temperature sensitive polymer compound and the drug.

  Moreover, it is preferable that the said polyglycerol has a space | gap inside a molecule | numerator. By having the voids, the fat-soluble drug can be enclosed in the voids, so that it can also be used as a DDS of the fat-soluble drug. The porosity can be adjusted by the primary ratio of the hydroxyl groups of polyglycerin. Among the hydroxyl groups, the primary hydroxyl group is 50% or more, preferably 60% or more, particularly preferably 70% or more. . In the present invention, as the polyglycerol, highly branched poly (deca) glycerol (trade name “PGL10PS”, manufactured by Daicel Chemical Industries, Ltd.), highly branched poly (20) glycerol (trade name “PGL20P”, manufactured by Daicel Chemical Industries, Ltd.) Commercially available products such as hyperbranched poly (40) glycerin (trade name “PGLXP”, manufactured by Daicel Chemical Industries, Ltd.) and hyperbranched poly (80) glycerin (trade name “HPG80”, manufactured by Daicel Chemical Industries, Ltd.) are preferably used. can do.

As a method for introducing X ¹ into polyglycerol and subsequently introducing R, for example, X ¹ is —C (═O) CH ₂ CH ₂ —, and R is —C (═O) NH—iC ₃ H. _In the case of ₆ , there is a method in which polyglycerin is subjected to a condensation reaction of succinic anhydride, followed by a condensation reaction of isopropylamine. X ¹ is —C (═O) CH ₂ CH ₂ —, R is —C (═O) —Z (Z represents a polyalkylene glycol residue in which the terminal hydroxyl group may be sealed with an organic group. A succinic anhydride is condensed with polyglycerin, and then a polyalkylene glycol whose terminal hydroxyl group may be blocked with an organic group is subjected to a condensation reaction.

The reaction for introducing X ¹ into polyglycerol is carried out in the presence or absence of a solvent. The solvent is not particularly limited as long as it is inert under the reaction conditions. Examples thereof include organic acids such as acetic acid, propionic acid and trifluoroacetic acid; nitriles such as acetonitrile, propionitrile and benzonitrile; formamide, acetamide, N Amides such as N, dimethylformamide (DMF) and dimethylacetamide; Aliphatic hydrocarbons such as hexane, heptane, octane and cyclododecane; Aromatic hydrocarbons such as benzene, toluene and xylene; Chloroform, dichloromethane, dichloroethane, Halogenated hydrocarbons such as carbon chloride, chlorobenzene and trifluoromethylbenzene; nitro compounds such as nitrobenzene, nitromethane and nitroethane; esters such as ethyl acetate and butyl acetate; hexafluoroisopropyl alcohol, tri Fluoroalcohol such as Le Oro ethanol; etc. and mixtures of these solvents.

  The subsequent reaction for introducing R is also carried out in the presence or absence of a solvent. Any solvent may be used as long as it is inert under the reaction conditions. Examples thereof include water, DMSO, DMF, and other solvents exemplified above.

  Depending on the type of reaction, a base (an amine such as triethylamine, a nitrogen-containing aromatic heterocyclic compound such as pyridine), an acid, a condensing agent, or the like may be used for the reaction.

  The reaction temperature can be appropriately selected depending on the type of reaction, the type of compound used, the type of solvent, etc., and is not particularly limited. For example, it is about 0-250 degreeC, Preferably it is about 25-150 degreeC, More preferably, it is about 50-150 degreeC. Depending on the type of reaction, the reaction may proceed smoothly near room temperature. The reaction may be carried out under an inert gas atmosphere such as nitrogen or argon, or under an air atmosphere or an oxygen atmosphere.

[Temperature sensitive drug release system]
The temperature-sensitive drug release system according to the present invention comprises a temperature-sensitive polymer compound and a drug.

  The drug may be a hydrophilic drug or a fat-soluble drug (hydrophobic drug). When using a hydrophilic drug, it is enclosed in the hydrophilic region of the closed space inside the temperature-sensitive polymer compound, and when using a fat-soluble drug, it is supported on the hydrophobic portion of the temperature-sensitive polymer compound. Moreover, it is although it does not specifically limit as a chemical | medical agent, The anticancer agent used together with a thermotherapy, an anti-inflammatory agent, etc. are mentioned. Serious side effects caused by excessive use of anti-cancer drugs, etc. are problematic, but using the temperature-sensitive drug release system of the present application allows the anti-cancer drugs to act specifically on the target cells only. Effective treatment can be performed, and side effects caused by anticancer agents can be minimized.

  Examples of the anticancer agent include metal complexes such as cisplatin, carboplatin, tetilaplatin, and iproplatin; anticancer antibiotics such as adriamycin, mitomycin, actinomycin, ansamitocin, bleomycin, cytarabine, and baunomycin; 5-fluorouracil, methotrexate, Antimetabolites such as TAC-788; alkylating agents such as BCNU and CCNU; lymphokines such as interferon-α, β, or γ and various interleukins. Examples of the anti-inflammatory agent include prednisolone, betamethasone, betamethasone, chlorpheniramine maleate, and the like.

  The above drugs include genes for disease treatment. Examples of such genes include adenosine deaminase gene for the treatment of severe combined immunodeficiency, LDL receptor gene for the treatment of familial hypercholesterolemia, interferon-α, β for the treatment of cancer. Or γ gene, granulocyte macrophage colony stimulating factor (GM-CSF) gene, various interleukin genes, tumor necrosis factor (TNF) -α gene, lymphotoxin (LT) -β gene, granulocyte colony stimulating factor (G-CSF) ) Genes, T cell activation costimulatory genes and the like. In addition, genes for the treatment of Alzheimer's disease, spinal cord injury, Parkinson's disease, arteriosclerosis, diabetes, hypertension and the like can be mentioned.

  The temperature sensitive drug release system of the present invention preferably contains at least one pharmaceutical additive in addition to the above drugs. As the pharmaceutical additive, known and commonly used pharmaceutical additives corresponding to the dosage form of the temperature sensitive drug release system can be used. For example, when the temperature sensitive drug release system is a liquid preparation, for example, physiological saline, Carriers such as sterilized water and buffer; membrane stabilizers such as cholesterol; isotonic agents such as sodium chloride, glucose and glycerin; antioxidants such as tocopherol, ascorbic acid and glutathione; preservatives such as chlorbutanol and parabens Etc. The carrier can be used as a solvent in manufacturing a temperature sensitive drug release system.

  When the temperature sensitive drug release system is a solid formulation, for example, carriers (eg, sugars such as lactose, sucrose; starches such as corn starch; celluloses such as crystalline cellulose; gum arabic, magnesium aluminate metasilicate , Calcium phosphate, etc.), lubricants (eg, magnesium stearate, talc, polyethylene glycol, etc.), binders (eg, saccharides such as mannitol, sucrose, crystalline cellulose, polyvinyl pyrrolidone, hydroxypropyl methylcellulose, etc.), disintegrants (For example, starches such as potato starch, celluloses such as carboxymethyl cellulose, cross-linked polyvinyl pyrrolidone, etc.), coloring agents, flavoring agents and the like.

  In the production of the temperature-sensitive drug release system according to the present invention, as a method for encapsulating the drug in the temperature-sensitive polymer compound, a known method can be adopted depending on the type of drug, for example, in a solution containing the drug For example, a method in which a temperature sensitive polymer compound is immersed in a temperature sensitive polymer compound and a method in which the temperature sensitive polymer compound is synthesized in a solvent in which the drug is dissolved may be used.

  As a mechanism for delivering, for example, an anticancer drug to a target cell using a temperature-sensitive drug release system, the following may be considered. There are many holes of about 200 nm in microvessels around cancer cells. On the other hand, since such a hole does not exist in the microvessel around the normal cell, the anticancer drug-encapsulated temperature-sensitive drug release system does not leak from the blood vessel around the normal cell. Therefore, by setting the particle size of the temperature sensitive drug release system encapsulating the anticancer drug to about 200 nm, the temperature sensitive drug release system leaks from the microvessel near the target cancer cell and reaches the cancer cell. be able to. In this way, the anti-cancer drug-encapsulated temperature-sensitive drug release system can accumulate specifically and at a high concentration in the site surrounding the target cancer cell.

  In the temperature sensitive drug release system according to the present invention, the particle size is preferably about 200 nm. For example, the amount of the drug to be encapsulated is 0.1 to 30 mol% with respect to the amount of the temperature sensitive polymer compound, preferably By setting the content to 1 to 6 mol%, the particle size can be set to about 200 nm. On the other hand, when the amount of the drug to be encapsulated exceeds 30 mol%, the particle size tends to be too large, and it tends to be difficult to selectively deliver the drug to the site around the cancer cell.

  As the temperature sensitive drug release system according to the present invention, a temperature sensitive drug release system is stably present in the vicinity of 34 to 38 ° C., and the drug can be stably held in the temperature sensitive drug release system. In the vicinity of ˜45 ° C., it is preferable that the temperature sensitive drug release system rapidly collapses and the held drug can be released. Specifically, the drug release rate from the temperature sensitive drug release system after incubation at 37 ° C. for 3 minutes is 5% or less, more preferably 4.5% or less, and after incubation at 45 ° C. for 3 minutes. It is preferred that the drug release rate from the temperature sensitive drug release system is 85% or more, more preferably 87% or more. The mechanism by which a temperature sensitive drug release system collapses at a specific temperature is not clear, but it is likely to increase hydrophobicity due to dehydration of hydrophilic temperature functional groups as the temperature increases. It is thought that the temperature-sensitive drug release system collapses when the group interacts with the hyperbranched polyglycerin skeleton.

  The temperature-sensitive drug release system according to the present invention can be administered by either oral or parenteral routes. For example, when an anticancer drug is encapsulated as a drug, it is administered by the parenteral route (particularly intravenously). Administration). In addition, the temperature-sensitive drug release system according to the present invention can suppress the metabolism of the lipophilic drug in the liver by encapsulating the lipophilic drug, thereby increasing the residence time in the blood. Therefore, even if one-shot administration, for example, one-shot administration can be quickly disappeared or excreted without using special administration techniques such as continuous infusion for a long time, for example, intravenous infusion The desired therapeutic effect can be obtained. Furthermore, it is not necessary to administer a large amount in order to obtain the desired therapeutic effect, thereby making it possible to minimize the occurrence of unexpected side effects.

  In addition, the administration target of the temperature-sensitive drug release system according to the present invention is not particularly limited as long as it is an organism having a normal body temperature of 38 ° C. or lower, and for example, it is suitably used for mammals, particularly humans. be able to.

  As the administration method of the temperature sensitive drug release system according to the present invention, for example, the temperature sensitive drug release system is administered to the subject, and the target affected area is heated to a temperature of 40 ° C. or higher, preferably 40 to 45 ° C. As a result, the temperature-sensitive polymer constituting the temperature-sensitive drug release system that reaches the affected area can be phase-shifted to release the encapsulated drug. As a method of heating the affected part, a method used in normal thermotherapy can be adopted, for example, hot back, humidified air like a bubble bath, wet heat method of transferring heat via hot water, Examples thereof include a heat-sensitive method via dry air such as infrared rays and dry packs, an electromagnetic energy such as a laser and a microwave, an electric energy such as an ultrasonic wave, and a conversion heat method in which physical vibration is changed into heat in the body.

  The temperature-sensitive drug release system according to the present invention also has pH sensitivity, and if it is not heated at a high temperature of 50 ° C. or higher at pH 7 or higher, release of the encapsulated drug is not caused. By heating at 45 ° C., the encapsulated drug can be quickly released. Since normal cells have a pH of around 7.4, cancer cells tend to be acidic at about pH 6.8. Therefore, even if the temperature-sensitive drug release system reaches normal cells, 40-45. Heating to ℃ does not cause the release of the encapsulated drug, allowing the drug to act on the cancer cells more specifically, and reducing the possibility of normal cells being damaged by anticancer drugs Can do.

  As is apparent from the above, the temperature sensitive drug release system according to the present invention is formed by an inexpensive fatty acid ester recognized as a food additive, and thus has excellent biocompatibility, can reduce costs, and is inexpensive. Can be provided. In addition, since the residence time in blood is long, and it has temperature sensitivity and pH sensitivity, the drug can reach only the targeted cells more accurately, can exhibit excellent therapeutic effects, and exhibits side effects. Can be kept to a minimum.

  EXAMPLES Hereinafter, although an Example demonstrates this invention more concretely, this invention is not limited by these Examples. The succinylation rate and oligoethylene glycolation rate were calculated by the following methods.

(Calculation of succinylation rate)
From the hydroxyl value of polyglycerol used as a raw material, the average number of terminal hydroxyl groups per molecule is calculated. Next, the ratio of the linker-modified polyglycerol terminal hydroxyl group is calculated from the area of the signal derived from the ¹ H-NMR linker portion of the polyglycerol derivative obtained by condensation reaction of the linker portion, and is defined as the succinylation rate.

(Calculation of oligoethylene glycolation rate)
Next, after the oligoethylene glycolation, the ratio of the terminal carboxylic acid modified with oligoethylene glycol is calculated from the area of the signal derived from the terminal group of ¹ H-NMR, and the oligoethylene glycolation rate (modification rate) .

Example 1
(Production of succinylated polyglycerin)
5.842 g (3.9 mmol) of hyperbranched poly (20) glycerin (trade name “PGL20P”, manufactured by Daicel Chemical Industries) and 25.10 g (257.2 mmol) of succinic anhydride were dissolved in 64.27 mL of pyridine. The mixture was stirred for 7 hours under a temperature of 63 to 65 ° C., an argon atmosphere and light-shielding conditions. Thereafter, the reaction solution is cooled and the precipitate is removed by filtration, and then concentrated, washed with diethyl ether, purified by a Sephadex LH-20 column (developing solvent: methanol), and succinylated polyglycerin is added to 8. 314 g (59% yield) was obtained.

(Production of isopropylamide-terminated polyglycerol derivative)
The operation of dissolving the obtained succinylated polyglycerin in a mixed solution of acetone and ion-exchanged water and performing distillation under reduced pressure by a rotary evaporator was repeated several times.
13.20 mL of dimethylformamide was dissolved in 1.220 g (0.30 mmol) of succinylated polyglycerol after depressurizing distillation. Subsequently, 2.8 mL (20 mmol) of 99.5% triethylamine, 2.418 g of HBTU (O-benzotriazol-1-yl-N, N, N ′, N′-tetramethyluronium hexafluorophosphate) (6 .376 mmol) was added and stirred under shading conditions. Under ice-cooling, 2.058 mL (isopropylamide mole number: 4.787 mmol) of a dimethylformamide (isopropylamide concentration: 2.326 M) solution was added to the stirring reaction solution, and the mixture was stirred for 28-30 under argon atmosphere and light-shielding conditions. Stir at 0 ° C. for 3 days. Thereafter, reprecipitation and purification were performed using a Sephadex LH-20 column (developing solvent: methanol) to obtain an isopropylamide-terminated polyglycerol derivative.

Example 2
(Production of succinylated polyglycerin)
Using 5.878 g (1.974 mmol) of highly branched poly (40) glycerin (trade name “PGLXP”, manufactured by Daicel Chemical Industries) and 24.86 g (248.4 mmol) of succinic anhydride in 64.66 mL of pyridine. The product was synthesized and purified under the same conditions as in Example 1. Water was added to the purified product, and the solvent was distilled off under reduced pressure using a rotary evaporator. This operation was performed a total of 4 times and lyophilized to obtain a pale yellow viscous solid in a yield of 10.264 g and a yield of 74%.
¹ H-NMR (400 MHz, CD ₃ OD): δ 2.62 (br, —O—CO—CH ₂ —CH ₂ —CO—OH), 2.93 and 3.07 (s, —CO—N ( _{CH 3) 2), 3.4-4.07 (} br, H R PG-scaffold), 4.07-4.45 (br, -CH 2 -O-CO -), 5.0-5.3 (Br, —CH—O—CO—)

(Production of isopropylamide-terminated polyglycerol derivative)
The operation of dissolving the obtained succinylated polyglycerin in a mixed solution of acetone and ion-exchanged water and performing distillation under reduced pressure by a rotary evaporator was repeated several times.
9.34 mL of dimethylformamide was dissolved in 1.021 g (0.13 mmol) of succinylated polyglycerol after depressurizing distillation. Subsequently, 2.4 mL (17 mmol) of 99.5% triethylamine, 3.103 g of HBTU (O-benzotriazol-1-yl-N, N, N ′, N′-tetramethyluronium hexafluorophosphate) (8 182 mmol) and stirred under light shielding conditions. Dimethylformamide (1.00 mL) was added and isopropylamide (0.80 mL) was added to the stirring reaction. After temporarily cooling with ice to generate heat, the mixture was stirred for 4 days at room temperature under light-shielding conditions in an argon atmosphere, and then the solvent was distilled off under reduced pressure. Purification by reprecipitation and Sephadex LH-20 column (developing solvent: methanol) gave an isopropylamide-terminated polyglycerin derivative in a yield of 1.144 g and a yield of 93%.
¹ H-NMR (400 MHz, CD ₃ OD): δ 1.12 (d, —NH—CH (CH ₃ ) ₂ ), 2.45 (br, —O—CO—CH ₂ —CH ₂ —CO—NH) -), 2.62 (br, -O -CO-CH 2 -CH 2 -CO-OH and -O-CO-CH 2 -CH 2 -CO-NH -), 3.4-4.07 (br , HPG-scaffold), 3.94 ( m, -CO-O-NH-CH (CH 3) 2), 4.07-4.45 (br, -CH 2 -O-CO -), 5.0 -5.3 (br, -CH-O-CO-).

¹ H-NMR measurement results for the succinylated polyglycerol and the isopropylamide-terminated polyglycerol derivative obtained in Example 1 are shown in FIGS.
¹ H-NMR measurement conditions Body: JEOL Ltd., 400 MHz NMR analyzer Sample concentration: 2-10% (wt / wt)
Solvent: Heavy methanol Internal standard: TMS

Evaluation With respect to the isopropylamide-terminated polyglycerin derivatives obtained in Examples, the following evaluation methods were used to evaluate temperature sensitivity, pH sensitivity, and drug release ability using rose bengal (RB) as a test substance.

(Evaluation method 1)
UV having an isothermal unit by dissolving the isopropylamide-terminated polyglycerin derivative obtained in Example 1 in a phosphate buffer having a pH of 5.0 to 6.5 to obtain a solution having an isopropylamide-terminated polyglycerin derivative concentration of 10 mg / mL. Using a transmittance tester, turbidity was measured by UV transmittance (%) while raising the temperature in the system.
Turbidity measurement conditions Temperature rise rate: 1 ° C / min Measurement wavelength: 700nm
100% transmission standard solution: 10 mM PBS

  The evaluation results are summarized in FIG.

(Evaluation method 2)
Rose Bengal was used as a drug model, and turbidity measurement was performed using 10 mM phosphate buffered saline (PBS) (pH 7.4) and the isopropylamide-terminated polyglycerol derivative obtained in Example 2. Reference solution having a transmittance of 100%: 10 mM PBS, heating rate: 1 ° C./min, measurement wavelength: 700 nm. The measurement results when RB was not included as a control were also shown.
Turbidity measurement conditions Temperature rise rate: 1 ° C / min Measurement wavelength: 700nm
100% transmission standard solution: 10 mM PBS

  The evaluation results are shown in FIG.

(Evaluation method 3)
RB, 10 mM phosphate buffered saline (PBS) (pH 7.4), isopropylamide-terminated polyglycerol derivative (NIPAM) or hyperbranched poly (40) glycerol obtained in Example 2 (trade name “PGLXP”, Daicel Chemical) (Manufactured by Kogyo Co., Ltd.) Using HPG40, a sample was prepared so that the polymer concentration was 5 mg / ml, the RB concentration was 58.6 μM, and 10 mM PBS. The polymer was dissolved by ice cooling and ultrasonic irradiation. Thereafter, centrifugation was performed at 4 ° C. and 15000 rpm for 30 minutes using a KUBOTA micro cooling centrifuge (3700). 100 μl of the supernatant was diluted 10-fold with 10 mM PBS (pH 7.4), and spectrum measurement (at 4 ° C.) was performed using a UV-Vis spectrophotometer JASCO V630. The remaining sample was incubated for 30 minutes at 40 ° C., 300 rpm using an Eppendorf Thermomixer comfort. Thereafter, the mixture was centrifuged at 15,000 rpm for 30 minutes at 30 to 40 ° C., and the spectrum of the supernatant was measured in the same manner as described above.
As a result, in the case of using the isopropylamide-terminated polyglycerin derivative obtained in Example 2, precipitation could be confirmed after heating. As a result of the measurement, the spectrum was obtained at λmax = 560 nm and abs (absorbance) = 0.588 before heating, but was obtained at λmax = 548 nm and abs = 0.0427 after heating.
From these results, it was found that the recovery rate can be calculated as 93% as follows, and this example is effective as a temperature sensitive drug release system.
Recovery rate (%) = {1- [abs (NIPAM 40 ° C.) ÷ abs (NIPAM 4 ° C.)]} × 100

  The said evaluation result is put together in FIG.

Example 3
(Production of succinylated polyglycerin (Suc-HPG80))
Highly branched poly (80) glycerin (trade name “HPG80”, manufactured by Daicel Chemical Industries, Ltd.) was dissolved in ion-exchanged water, and the supernatant was collected and filtered to remove insoluble components. HPG80 (6.215 g) after lyophilization was dissolved in pyridine (68.37 ml), succinic anhydride (25.82 g) was added, and under an argon atmosphere, protected from light, at a temperature of 60 to 62 ° C., Heating and stirring were performed for 7 hours. After refrigeration, the precipitate was filtered and the filtrate was distilled off under reduced pressure. The concentrate was washed once with diethyl ether, a small amount of acetic acid was added to the precipitate, and the precipitate was dissolved with an aqueous sodium hydrogen carbonate solution. Thereafter, the polymer was reprecipitated with 1N hydrochloric acid, and the supernatant was removed by centrifugation. After adding high-concentration acetic acid / sodium acetate buffer (pH 4.3-4.4), mix lightly, add acetic acid and methanol to dissolve, filter, and then separate the Sephadex LH-20 column (developing solvent). : Methanol) to obtain succinylated polyglycerin (Suc-HPG80) (yield 100%).

Example 4
(Production of oligoethylene glycol-derived group-terminated polyglycerol derivative (MDEG-Suc-HPG80))
Suc-HPG80 (0.676 g, 48.1 μmol) was dissolved in DMF (9.76 ml). After adding triethylamine (≧ 99.5%) (1.7 ml, 12 mmol), HBTU (O-benzotriazol-1-yl-N, N, N ′, N′-tetramethyluronium hexafluorophosphate) ( 1.79 g, 4.72 mmol) was added and dissolved in a light-shielded state under an argon atmosphere. Methoxydiethyleneglycol (MDEG) (1.38 ml, 11.7 mmol) was added, and the mixture was stirred for 3 days at room temperature in the dark. The solvent was distilled off to some extent and then washed with diethyl ether. To suppress side reactions, a small amount of special grade acetic acid (1.50 ml) and distilled water were added, and after stirring, diluted with methanol and added dropwise to diethyl ether. The ether layer was removed, the aqueous layer was diluted with distilled water, and the supernatant was dialyzed against distilled water (MWCO = 2000). Dialysis was carried out for 1 day, but it was not purified. The solution was concentrated, diluted with methanol, filtered, and purified by Sephadex LH-20 column (developing solvent: methanol) to give a yellow viscous liquid (target compound) Got. Further, dialysis (MWCO = 2000) was performed for 3 days with distilled water. The identification was performed by ¹ H-NMR (CD ₃ OD, 400 MHz) after purification with LH-20 column (FIG. 6). After dialysis, the methanol peak only disappeared in ¹ H-NMR (D ₂ O, 400 MHz). The succinylation rate and MDEG modification rate were calculated using the integration ratio of the peak derived from the polymer skeleton as the standard (400H). The succinylation rate was 96%, the MDEG modification rate was 90%, the yield was 1.00 g, and the yield was 98%. .

Example 5
(Production of oligoethylene glycol-derived group-terminated polyglycerol derivative (MTEG-Suc-HPG80))
Suc-HPG80 (0.732 g, 52.1 μmol) was dissolved in DMF (10.32 ml). Triethylamine (≧ 99.5%) (1.8 ml, 13 mmol) was added, and then HBTU (1.93 g, 5.08 mmol) was added and dissolved in a light-shielded state under an argon atmosphere. Methoxytriethyleneglycol (MTEG) (2.03 ml, 12.7 mmol) was added, and the mixture was stirred for 4 days at room temperature in the dark. After the solvent was distilled off to some extent, a special grade acetic acid (1.50 ml) was added and stirred to prevent side reactions, and the mixture was washed with diethyl ether. A small amount of distilled water was added, and after stirring, diluted with methanol and added dropwise to diethyl ether. The ether layer was removed, the aqueous layer was diluted with distilled water, and the supernatant was dialyzed against distilled water (MWCO = 2000). The solution was concentrated, diluted with methanol and filtered, and then purified with a Sephadex LH-20 column (developing solvent: methanol) to obtain a yellow viscous liquid (target compound). Further, dialysis (MWCO = 2000) was carried out with distilled water for 2 days. The identification was performed by ¹ H-NMR (CD ₃ OD, 400 MHz) after purification of the LH-20 column (FIG. 7). The succinylation rate and MTEG modification rate were calculated using the integration ratio of the peak derived from the polymer backbone as a reference (400H), and the succinylation rate was 96%, MTEG modification rate was 89%, yield was 1.26 g, and yield was 100%. .

Example 6
(Production of oligoethylene glycol derivative group-terminated polyglycerol derivative (ETEG-Suc-HPG80))
Suc-HPG80 (0.344 g) was dissolved in DMF (3.44 ml). Triethylamine (≧ 99.5%) (0.90 ml) and HBTU (0.910 g) were added, dissolved in a light-shielded state under an argon atmosphere, and then ethoxytriethyleneglycol (ETEG) (1.05 ml). And stirred for 4 days at room temperature in a light-shielded state. After the solvent was distilled off to some extent, special grade acetic acid (0.50 ml) was added and stirred to suppress side reactions. Ion exchange water (0.50 ml) was added and stirred, and then washed several times with diethyl ether. The ether layer was removed and the mixture was purified by a Sephadex LH-20 column (developing solvent: methanol) to obtain a yellow viscous liquid. Since the purification was insufficient, the product was purified again using a Sephadex LH-20 column (developing solvent: methanol) to obtain a yellow viscous liquid (target compound). Identification was performed by ¹ H-NMR (CD ₃ OD, 400 MHz) (FIG. 8). The succinylation rate and the ETEG modification rate were calculated based on the integration ratio of the peak derived from the polymer skeleton as a reference (400H). .

Example 7
(Production of oligoethylene glycol-derived group-terminated polyglycerin derivative (MTEG / ETEG-Suc-HPG80))
Suc-HPG80 (0.422 g) was dissolved in DMF (4.22 ml). Triethylamine (≧ 99.5%) (1.1 ml) and HBTU (1.12 g) were added, dissolved in a light-shielded state under an argon atmosphere, and then ethoxytriethyleneglycol (ETEG) (0.39 ml). Then, methoxytriethyleneglycol (MTEG) (0.94 ml) was added, and the mixture was stirred for 4 days at room temperature in the dark. After the solvent was distilled off to some extent, special grade acetic acid (0.50 ml) was added and stirred to suppress side reactions. Ion exchange water (0.50 ml) was added and stirred, and then washed several times with diethyl ether. The ether layer was removed and the mixture was purified by a Sephadex LH-20 column (developing solvent: methanol) to obtain a yellow viscous liquid. Since the purification was insufficient, the product was purified again using a Sephadex LH-20 column (developing solvent: methanol) to obtain a yellow viscous liquid (target compound). ^From the measurement results of ¹ H-NMR (CD ₃ OD, 400 MHz), the succinylation rate, the MTEG modification rate, and the ETEG modification rate were calculated using the integral ratio of the peak derived from the polymer backbone as a reference (400H). %, MTEG modification rate 64%, ETEG modification rate 26%, yield 0.72 g, yield 97%.

Example 8
(Production of oligoethylene glycol-derived group-terminated polyglycerin derivative (MTEG / ETEG-Suc-HPG80))
Suc-HPG80 (0.444 g) was dissolved in DMF (4.44 ml). Triethylamine (≧ 99.5%) (1.16 ml) and HBTU (1.18 g) were added, dissolved in a light-shielded state under an argon atmosphere, and then ethoxytriethyleneglycol (ETEG) (0.68 ml). Then, methoxytriethyleneglycol (MTEG) (0.62 ml) was added, and the mixture was stirred for 4 days at room temperature in the dark. After the solvent was distilled off to some extent, special grade acetic acid (0.50 ml) was added and stirred to suppress side reactions. Ion exchange water (0.50 ml) was added and stirred, and then washed several times with diethyl ether. The ether layer was removed and the mixture was purified by a Sephadex LH-20 column (developing solvent: methanol) to obtain a yellow viscous liquid. Since the purification was insufficient, the product was purified again using a Sephadex LH-20 column (developing solvent: methanol) to obtain a yellow viscous liquid (target compound). ^From the measurement results of ¹ H-NMR (CD ₃ OD, 400 MHz), the succinylation rate, MTEG modification rate, and ETEG modification rate were calculated using the integral ratio of the peak derived from the polymer backbone as a reference (400H). %, MTEG modification rate 51%, ETEG modification rate 42%, yield 0.72 g, yield 90%.

Example 9
(Production of oligoethylene glycol-derived group-terminated polyglycerin derivative (MTEG / ETEG-Suc-HPG80))
Suc-HPG80 (0.396 g) was dissolved in DMF (3.96 ml). Triethylamine (≧ 99.5%) (1.04 ml) and HBTU (1.05 g) were added, dissolved in a light-shielded state under an argon atmosphere, and then ethoxytriethyleneglycol (ETEG) (0.97 ml). Then, methoxytriethyleneglycol (MTEG) (0.33 ml) was added, and the mixture was stirred for 4 days at room temperature in the dark. After the solvent was distilled off to some extent, special grade acetic acid (0.50 ml) was added and stirred to suppress side reactions. Ion exchange water (0.50 ml) was added and stirred, and then washed several times with diethyl ether. The ether layer was removed and the mixture was purified by a Sephadex LH-20 column (developing solvent: methanol) to obtain a yellow viscous liquid. Since the purification was insufficient, the product was purified again using a Sephadex LH-20 column (developing solvent: methanol) to obtain a yellow viscous liquid (target compound). ^From the measurement results of ¹ H-NMR (CD ₃ OD, 400 MHz), the succinylation rate, the MTEG modification rate, and the ETEG modification rate were calculated using the integral ratio of the peak derived from the polymer backbone as a reference (400H). %, MTEG modification rate 23%, ETEG modification rate 66%, yield 0.69 g, yield 97%.

Evaluation The temperature sensitivity of the oligoethylene glycol derivative group-terminated polyglycerin derivatives obtained in the examples was evaluated by the following evaluation method.

(Evaluation method 4)
The oligoethylene glycol-derived group-terminated polyglycerin derivatives obtained in Examples 4 to 9 were dissolved in 10 mM phosphate buffer (PBS) (pH 7.4), and the concentration of the oligoethylene glycol-derived group-terminated polyglycerin derivative was 10 mg / A temperature sensitivity test was performed by measuring the UV transmittance (%) while increasing the system temperature over time using a UV transmittance tester having a constant temperature unit as a mL solution. The measurement conditions are shown below.
Turbidity measurement conditions Temperature rise rate: 1 ° C / min Measurement wavelength: 700nm
100% transmission standard solution: 10 mM PBS
Solution: Polymer concentration 10 mg / mL, 10 mM phosphate buffer

FIG. 9 shows the results using the oligoethylene glycol-derived group-terminated polyglycerin derivatives obtained in Examples 4 to 6. FIG. 9 shows the results obtained using the oligoethylene glycol-derived group-terminated polyglycerin derivatives obtained in Examples 5 to 9. As shown in FIG. The symbols in the figure are as follows.
ETEG, ETEG0.85: Oligoethylene glycol derivative group-terminated polyglycerol derivative obtained in Example 6 MDEG: Oligoethylene glycol derivative group-terminated polyglycerin derivative obtained in Example 4 MTEG, MTEG0.89: In Example 5 Obtained oligoethylene glycol derivative group-terminated polyglycerol derivative MTEG0.23 / ETEG0.66: Oligoethylene glycol derivative group-terminated polyglycerin derivative obtained in Example 9 MTEG0.51 / ETEG0.42: Obtained in Example 8 Oligoethylene glycol-derived group-terminated polyglycerol derivative MTEG0.64 / ETEG0.26: oligoethyleneglycol-derived group-terminated polyglycerol derivative obtained in Example 7

  As shown in FIG. 9, the longer the ethylene oxide chain (polyethylene glycol chain), the higher the transition temperature, and the longer the terminal alkyl group (group that seals the terminal hydroxyl group), the lower the transition temperature. .

  As shown in FIG. 10, a temperature-sensitive polymer compound (temperature-responsive polymer compound) having a transition temperature near the body temperature is obtained by adjusting the ratio of two types of thermosensitive groups (MTEG and ETEG). It becomes possible.

Up to now, many researches have been made on the development of DDS targeting cancer, but there are few DDS materials in practical use. The reason is that it takes time to verify the safety of the DDS material. Therefore, there is a problem that even if the material has a high function, it is not easy to put into practical use a synthetic compound whose safety has not been established and is generally expensive. The polyglycerin used for the skeleton in the present invention is not only relatively inexpensive, but also excellent in biocompatibility because the fatty acid ester using this substrate has already been recognized as a food additive. It is.
That is, by using the material of the present invention, it is possible to design an inexpensive and safe temperature-responsive DDS as a DDS material, and contribute to the treatment of diseases such as cancer that still afflicts people.

Claims (9)

A polyglycerin skeleton as the main chain, and at least the following formula (a)

[Wherein, GL is a glycerin residue, X is a C 1-10 alkylene group having a carbonyl bond (—CO—) as a linking group on the GL side, R is a methylcarbamoyl group, an ethylcarbamoyl group, a propylcarbamoyl group , An isopropyl carbamoyl group, a butyl carbamoyl group, or a polyalkylene glycol residue having a carbonyl bond (—CO—) as a linking group at the bonding site with X (however, the polyalkylene glycol residue) The terminal hydroxyl group is an alkyl group having 1 to 10 carbon atoms, a 3 to 10-membered cycloalkyl group, an aryl group having 6 to 10 carbon atoms, an aralkyl group having 7 to 12 carbon atoms, or an acyl group having 1 to 11 carbon atoms. It may be sealed with an organic group selected from
The temperature sensitive high molecular compound which has a repeating unit represented by these. The precursor of X, which is a monovalent organic group having a functional group capable of introducing R by reaction into polyglycerol having a molecular weight of 120 to 12000 and a primary hydroxyl group of 50% or more of all hydroxyl groups The temperature-sensitive polymer compound according to claim 1, which is obtained by introducing X ¹ and subsequently introducing R.   The temperature-sensitive polymer compound according to claim 1 or 2, wherein the glycerin residue: the X and the precursor thereof: the R (molar ratio) is 1: 0.5 to 1: 0.5-1. Following formula (1)

(In the formula, X ¹ represents a precursor of X which is a monovalent organic group having a functional group capable of introducing R by reaction, and X ² corresponds to X ¹ after introducing R. A divalent group, GL represents a glycerin residue, R is the same as above, and m and n are the same or different and represent an integer of 1 or more)
The temperature-sensitive polymer compound according to any one of claims 1 to 3, which is represented by: X ¹ is the following formula (2)
_{-CO- (CH 2) s- (CO} ) t-OH (2)
(In the formula, s is an integer of 1 to 10, and t represents 1 )
The temperature-sensitive polymer compound according to claim 2, which is a monovalent organic group represented by the formula: The temperature-sensitive polymer compound according to any one of claims 1 to 5, wherein the polyalkylene glycol residue is an oligoethylene glycol residue having a degree of polymerization of oligoethylene glycol of 2 to 10 .   R is an isopropyl carbamoyl group, The temperature sensitive high molecular compound according to any one of claims 1 to 6.   A temperature-sensitive drug release system comprising the temperature-sensitive polymer compound according to any one of claims 1 to 7 and a drug.   The temperature sensitive drug release system according to claim 8, wherein the drug amount is 0.1 to 30 mol% with respect to the amount of the temperature sensitive polymer compound.

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