WO2016021407A1

WO2016021407A1 - Polyamino acid derivative in which nucleic acid antimetabolites are bonded

Info

Publication number: WO2016021407A1
Application number: PCT/JP2015/070785
Authority: WO
Inventors: 正行北川; 大川村; 佳奈水沼; 中村　巌; 大地長井; 千恵子瀬野; 啓一朗山本; 裕介齋藤; 綾香望月; 上村　和也
Original assignee: 日本化薬株式会社
Priority date: 2014-08-04
Filing date: 2015-07-22
Publication date: 2016-02-11
Also published as: JP2017160125A

Abstract

　The present invention addresses the problem of providing a polymerized prodrug of nucleic acid antimetabolites that improve an antitumor effect and have minimal side effects, particularly myelosuppression. Provided is a polyamino acid derivative in which polyethylene glycol segments and nucleic acid antimetabolites are bonded either directly or via linking bonds to side chain carboxyl groups, wherein the molecular mass of the polyamino acid derivative is 20 kilodaltons to 200 kilodaltons, and the mass content of the polyethylene glycol segments in the polyamino acid derivative is 30 mass% to 90%.

Description

Polyamino acid derivative bound with nucleic acid antimetabolite

The present invention relates to a polyaspartic acid derivative or polyglutamic acid derivative to which a nucleic acid metabolism antagonist is bound, and a use thereof.

Various nucleic acid metabolism antagonists have been developed for the purpose of treating malignant tumors or viral diseases. For example, as an antitumor agent (anticancer agent), there are cytarabine, gemcitabine, doxyfluridine, azacitidine, decitabine, nelarabine and the like. Moreover, as an antiviral agent, zalcitabine, lamivudine, etc. are used clinically.

These nucleic acid antimetabolites have extremely strong pharmacological activity in the in vitro evaluation. However, these drugs are susceptible to metabolism and excretion in the living body, and there is a problem that in vivo evaluation, the drug efficacy of the original drug cannot be fully exhibited. Many of these drugs require high doses for clinical treatment. For example, gemcitabine has a strong activity comparable to powerful antitumor agents such as paclitaxel and doxorubicin, in vitro cytostatic activity evaluation (IC ₅₀ value). On the other hand, the clinical use of gemcitabine requires a high dose of 1,000 mg / m ² as a use per body surface area. This is because the in-vivo utilization rate is lowered because the 4-position amino group of the nucleobase part of gemcitabine is metabolized and inactivated by cytidine deaminase, which is a metabolic enzyme of 2′-deoxycytidine. It is considered (Non-Patent Document 1).

In order to improve the bioavailability by suppressing the metabolism / inactivation of the drug, a polymerized drug in which the drug is bound to a polymer carrier has been studied. The high molecular weight drug is expected to change the pharmacokinetics based on the high molecular weight and to improve the therapeutic effect. Non-Patent Document 2 describes a polymerized derivative in which cytarabine, which is a nucleic acid antimetabolite, is bound to polyglutamic acids having an average molecular weight of about 30 kilodaltons. However, polymerized derivatives of drugs are easily recognized by foreign bodies in the living body, and there is a concern that many drugs are trapped in phagocytic tissues such as the liver. Moreover, there is a case where hypersensitivity reaction is caused by inducing immunity. In such a case, there is a concern that repeated administration as a drug cannot be performed.

As a method for avoiding foreign substance recognition of the polymerized derivative, a method using polyethylene glycol is known. For example, Patent Document 1 describes a polymerized derivative in which a cytidine derivative is bound to polyethylene glycols. Non-Patent Document 3 describes a polymerized derivative in which aspartic acid is branched at both ends of polyethylene glycols, and cytarabine is bound thereto via an appropriate linking group. Furthermore, Patent Document 2 describes a polymerized derivative having a structure in which an amino acid is branched at the end of a polyethylene glycol chain and the drug is released after each branch undergoes a benzyl elimination reaction.
These polymer derivatives have a large difference between the rate of hydrolysis in phosphate buffered saline (PBS) and the rate of hydrolysis in plasma, and the hydrolysis reaction is highly dependent on enzymes in the body. The effect may be greatly influenced by individual differences among patients.

Patent Document 3 and Patent Document 4 describe a polymerized derivative in which a nucleic acid antimetabolite and a hydrophobic substituent are bonded to a side chain carboxy group of a block copolymer of polyethylene glycols and a polyacidic amino acid. ing. Patent Document 5 describes a polymerized derivative in which a nucleic acid antimetabolite is bound to a side chain carboxy group of a block copolymer of polyethylene glycols and a polyacidic amino acid via a linker having a hydrophobic substituent. Has been.
The polymer conjugates of these nucleic acid antimetabolites are bipolar polymers having both a hydrophobic segment having a hydrophobic substituent introduced into the side chain carboxy group and polyethylene glycol, which is a hydrophilic segment. For this reason, the polymer conjugate of the nucleic acid antimetabolite is considered to form a self-aggregate having the hydrophobic segment as the inner core and the hydrophilic segment as the outer side due to intermolecular aggregation of the hydrophobic segment in an aqueous solution.

The polymer conjugate of the nucleic acid antimetabolite has a property of undergoing hydrolysis in a phosphate buffered saline (PBS) solution to dissociate the slowly bound nucleic acid antimetabolite. Therefore, these polymerized nucleic acid antimetabolites have a characteristic that they continue to exert a tumor growth inhibitory effect for a long period of time at a low dose as compared with conventional nucleic acid antimetabolites. However, such sustained-release nucleic acid antimetabolite prodrugs also cause side effects that occur due to the same action and mechanism of action over a long period of time. As for a nucleic acid antimetabolite, bone marrow suppression recognized as manifestation of leukopenia or the like is a problem as a dose limiting factor. High-molecular-weight nucleic acid antimetabolite, a sustained-release nucleic acid antimetabolite prodrug, has a tendency to prolong myelosuppression, and establishes a useful therapeutic method that achieves both improved therapeutic effects and reduced side effects. This is a big issue.

Special table 2003-524028 gazette JP-T-2004-532289 International Publication WO2006 / 120914 International Publication No. WO2008 / 056596 International Publication WO2008 / 056654

An object of the present invention is to provide a nucleic acid metabolism antagonist that improves the antitumor effect and has low side effects, particularly bone marrow suppression. Specifically, an object of the present invention is to provide a nucleic acid metabolism antagonist that exhibits a tumor growth inhibitory effect for a long period of time and does not prolong myelosuppression.

As a result of intensive studies to solve the above problems, the present inventors have bound a plurality of polyethylene glycol segments and a plurality of nucleic acid antimetabolites to a side chain carboxy group of a polyacidic amino acid such as polyaspartic acid. It has been found that polyamino acid derivatives can improve the antitumor effect and avoid prolonged myelosuppression, which is a side effect.

That is, the present invention relates to the following [1] to [11].
[1] A polyamino acid derivative containing a plurality of units of an aspartic acid derivative and / or a glutamic acid derivative, wherein a polyethylene glycol segment and a nucleic acid antimetabolite are bonded to the side chain carboxy group directly or via a linking group. The molecular weight of the polyamino acid derivative is 20 kilodaltons or more and 200 kilodaltons or less, and the mass content of the polyethylene glycol segment in the polyamino acid derivative is 30% by mass or more and 90% by mass or less. Polyamino acid derivative.

[2] The nucleic acid antimetabolite-binding polyamino acid derivative according to [1], wherein 2 to 80 units of the polyethylene glycol segment are bonded.

[3] The nucleic acid antimetabolite according to [1] or [2], wherein the nucleic acid antimetabolite is an amino acid antimetabolite having an amino group, and the nucleic acid antimetabolite is bound via an amide bond at the amino group. Antimetabolite-binding polyamino acid derivative.

[4] The nucleic acid antimetabolite-binding polyamino acid derivative according to any one of [1] to [3], wherein the mass content of the nucleic acid antimetabolite is 2% by mass to 60% by mass.

[5] The nucleic acid antimetabolite-binding polyamino acid derivative is represented by the general formula (1)

[Wherein, _{R 1} represents a hydrogen atom, a group selected from the group consisting of alkyl groups and polyethylene glycol segments of carbon atoms (C1 ~ C8), acyl _{R 2} is a hydrogen atom, the number of carbon atoms (C1 ~ C8) A group selected from the group consisting of a group and a C1-C8 alkoxycarbonyl group, R ₃ represents a polyethylene glycol segment, R ₄ represents a nucleic acid antimetabolite binding residue, and R ₅ represents an asparagine An acid bond residue and / or an aspartate imide bond residue, R ₆ represents a hydroxyl group and / or —N (R ₇ ) CONH (R ₈ ), and R ₇ and R ₈ may be the same or different. A linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with a tertiary amino group, wherein X ₁ and X ₂ are linking groups; ₃ is methylene A, b, c, d, e, f, g, h and i each independently represents an integer of 0 to 200, and the total number of polymerized polyamino acid derivatives (a + b + c + d + e + f + g + h + i) is 3 to 250, (a + b) is 1 to 95, and (c + d) is 1 to 175. The amino acid unit to which R ₃ is bonded, the amino acid unit to which R ₄ is bonded, and the R ₅ are bonded to each other Any one of the above [1] to [4], wherein the amino acid unit, the amino acid unit to which R ₆ is bonded, and the amino acid unit in which the side chain carboxy group is an intramolecular cyclization type are each independently a random sequence. The nucleic acid antimetabolite-binding polyamino acid derivative according to claim 1.

[6] X ₂ represents the following general formula (2) or general formula (3)

[Wherein R ₉ and R ₁₀ each independently represents a hydrogen atom or an alkyl group having from 1 to 8 carbon atoms (C1 to C8), and R ₁₁ represents a hydrogen atom or an optionally substituted carbon number (C1 to C8). A linear, branched or cyclic alkyl group of C20), an optionally substituted linear (C1-C20) linear, branched or cyclic aralkyl group, and a substituent. One or more groups selected from the group consisting of an amino acid-bonded residue in which an aromatic group and a carboxy group may be protected, and CX—CY represents C═C (two The nucleic acid antimetabolite-binding polyamino acid derivative according to [5] above, which exhibits a heavy bond).

[7] R ₅ represents the following general formula (4), general formula (5), and general formula (6).

[Wherein R ₉ , R ₁₀ , R ₁₁ and CX-CY have the same meaning as described above, R ₁₂ represents a hydroxyl group and / or —N (R ₁₃ ) CONH (R ₁₄ ), and R ₁₃ , R ₁₄ May be the same or different and represents a linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with a tertiary amino group]. The nucleic acid antimetabolite-binding polyamino acid derivative according to [5] or [6], which is one or more kinds of groups.

[8] The polyethylene glycol segment of R ₃ is represented by the following general formula (7)

[In the formula, R ₁₅ represents a hydrogen atom or a linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may have a substituent, and n is 5 to 2,500. The nucleic acid antimetabolite-binding polyamino acid derivative according to any one of [5] to [7], which is an integer].

[9] The nucleic acid antimetabolite is represented by the formula (8):

[Wherein, -Rf represents the formula (9):

Wherein R ₁₆ represents a hydrogen atom or an acyl group of a fatty acid ester], and is any one or more nucleic acid metabolism antagonists, [1] to [8] ] The nucleic acid antimetabolite binding polyamino acid derivative according to any one of the above.

[10] The nucleic acid antimetabolite is represented by the formula (10):

[Wherein, -Rf represents the formula (11):

The nucleic acid antimetabolite binding according to any one of [1] to [9] above, wherein R ₁₆ represents a hydrogen atom or an acyl group of a fatty acid ester] Polyamino acid derivative.

[11] A medicine containing the nucleic acid antimetabolite-binding polyamino acid derivative according to any one of [1] to [10].

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention is an aspartic acid and / or glutamic acid having a side chain carboxy group in the polymer main chain, and a plurality of polyethylene glycol segments and a plurality of nucleic acid metabolisms in the side chain carboxy group. It comprises an antagonist. The polyamino acid derivative has the property of gradually dissociating and releasing the bound nucleic acid antimetabolite, while remaining in the blood and being distributed in the body after administration in vivo. The polyamino acid derivative can be controlled to have a molecular weight within an appropriate range, and by controlling the content of the polyethylene glycol segment, side effects can be avoided while improving the efficacy of the nucleic acid antimetabolite. In particular, it is possible to provide a drug that avoids prolonged myelosuppression.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention is a polyamino acid derivative containing a plurality of units of aspartic acid derivatives and / or glutamic acid derivatives, and a polyethylene glycol segment with respect to the side chain carboxy group having a plurality And a nucleoside derivative which is a nucleic acid metabolism antagonist. The details will be described below.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention is a polymer compound having a polyamino acid derivative containing a plurality of units of aspartic acid derivatives and / or glutamic acid derivatives as a polymer main chain structure. That is, a polyamino acid derivative containing an acidic amino acid aspartic acid and / or glutamic acid as a polymer main chain and having a plurality of side chain carboxy groups is used as the main chain structure of the polymer carrier, and the side chain carboxy group is polyethylene glycol. Polymerized derivatives chemically functionalized with segments and nucleic acid antimetabolites.

The polymer main chain of the polyamino acid derivative is not particularly limited as long as it is a polymer structure containing a plurality of aspartic acids and / or glutamic acids and having a plurality of side chain carboxy groups. That is, as long as it contains a plurality of units of aspartic acid and / or glutamic acid, it may be a polyamino acid derivative that optionally contains other amino acids. The other amino acid may be a natural amino acid or a non-natural amino acid, and any of L-form and D-form can be used without particular limitation. Examples of the other amino acids include hydrocarbon amino acids such as glycine, β-alanine, alanine, leucine and phenylalanine, and basic amino acids such as lysine, arginine and histidine.
The side chain carboxy group of the polyamino acid derivative is used as a binding functional group for binding a later-described polyethylene glycol segment and a nucleic acid antimetabolite. For this reason, it is preferable that the polyamino acid derivative has a higher side chain carboxy group content. Therefore, the polymer main chain of the polyamino acid derivative is preferably a polyamino acid main chain polymer in which aspartic acid and / or glutamic acid-containing units are contained in 50 units% or more of the polyamino acid composition, and the aspartic acid and / or A main chain polymer containing 80 unit% or more of glutamic acid units is more preferable. Particularly preferred is a polyamino acid main chain composed of aspartic acid and / or glutamic acid. That is, particularly preferred polyamino acid main chain is polyaspartic acid, polyglutamic acid or poly (aspartic acid-glutamic acid) copolymer.

In the polyamino acid derivative, aspartic acid remains as a side chain so that one carboxy group can function as a binding functional group, and a polyamino acid main chain is constructed by an amide bond using the other carboxy group. The carboxy group constituting the polyamino acid main chain may be an α-amide bond polymer, a β-amide bond polymer, or a mixture thereof. Similarly, glutamic acid in the polyamino acid derivative remains as a side chain so that one carboxy group can function as a binding functional group, and a polyamino acid main chain is constructed by an amide bond using the other carboxy group. The carboxy group constituting the polyamino acid main chain may be an α-amide bond type polymer, a γ-amide bond type polymer, or a mixture thereof.

When the polymer main chain of the polyamino acid derivative is polyaspartic acid or polyglutamic acid, it is a structure in which aspartic acid or glutamic acid is polymerized in an appropriate number of units. , Β-amide bond type polymer, γ-amide bond type polymer, or a mixture thereof.
When the polymer main chain of the polyamino acid derivative is the poly (aspartic acid-glutamic acid) copolymer, it is a polyamino acid structure in which aspartic acid and glutamic acid are mixed. It may be the body. The bonding mode may be an α-amide bond polymer, a β-amide bond polymer with a side chain carboxy group or a γ-amide bond polymer, and a mixture thereof. Also good.

The number of polymerization of the amino acid main chain of the polyamino acid derivative is preferably 3 to 300 units. More preferably, the polymerization number of the amino acid main chain is 3 to 250 units, and the polymerization number is particularly preferably 10 to 200 units. Among them, the number of units containing aspartic acid and / or glutamic acid is 3 to 300, preferably 3 to 250 units, and particularly preferably 10 to 200 units.

The terminal group of the polyamino acid derivative is not particularly limited to both the N terminal group and the C terminal group, and may be an unprotected free amino group and a free carboxylic acid, and salts thereof. An appropriate modification of the C-terminal group may be used.
Examples of the modified N-terminal group of the polyamino acid derivative include acylamide-type modified products, alkoxycarbonylamide-type modified products (urethane-type modified products), alkylaminocarbonylamide-type modified products (urea-type modified products), and the like. it can.
On the other hand, examples of the modified form of the C-terminal group of the polyamino acid derivative include an ester-type modified form, an amide-type modified form, and a thioester-type modified form.
The modifying group of the N terminal group and the C terminal group of the polyamino acid derivative may be any modifying group. Both terminal groups are preferably substituents that do not interfere with the hydrophilicity of the polyamino acid derivative. Preferably, a linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may have a substituent via an appropriate bonding group bonded to the N-terminal group and the C-terminal group. And a terminal modifying group such as an aromatic group having carbon atoms (C6 to C18) which may have a substituent, an aralkyl group having carbon atoms (C7 to C20) which may have a substituent. Can do. Alternatively, it may be a polyethylene glycol substituent capable of imparting water solubility, such as a terminal modifying group bonded through an appropriate bonding group bonded to the N terminal group and the C terminal group.

That is, the N-terminal group is preferably a suitable acylamide-type modified product or alkoxycarbonylamide-type modified product (urethane-type modified product), and has the above-described substituent via a carbonyl group or a carbonyloxy group. A linear, branched or cyclic alkyl group having a carbon number (C1 to C8), an aromatic group having a carbon number (C6 to C18) which may have a substituent, and a substituent. An aralkyl group having a carbon number (C7 to C20) may be preferable.
On the other hand, the C-terminal group is preferably a suitable amide type substituent or ester type substituent, and the number of carbon atoms (C1 to C8) optionally having the above substituent via an amide group or ester group. ) Linear, branched or cyclic alkyl group, an aromatic group having a carbon number (C6 to C18) which may have a substituent, and a carbon number which has a substituent (C7 to C7) C20) is preferably an aralkyl group or a polyethylene glycol substituent.

In the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention, a polyethylene glycol segment is bonded to a side chain carboxy group directly or via a bonding group. The polyethylene glycol segment is a segment having a repeating structure of an ethyleneoxy group: (CH ₂ CH ₂ O) unit. A segment structure containing a polyethylene glycol chain having an ethyleneoxy group unit polymerization degree of 5 to 10,000 units, more preferably a polymerization degree of 5 to 5,000 units.
That is, the polyethylene glycol segment is preferably a segment part having a molecular weight corresponding to polyethylene glycol of 200 daltons to 500 kilodaltons, more preferably a structural part having a molecular weight of 200 daltons to 250 kilodaltons, and particularly preferably a molecular weight. 200 to 150 kilodaltons. Particularly preferred are polyethylene glycol segments having a molecular weight of 1,000 to 50 kilodaltons.
The molecular weight of the polyethylene glycol segment used in the present invention is determined by the GPC method based on the polyethylene glycol standard product of the polyethylene glycol segment structural compound used in preparing the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention. The average molecular weight calculated | required by the peak top molecular weight measured is employ | adopted.

One terminal group of the polyethylene glycol segment is a linking group for bonding to the carboxy group of the polyamino acid main chain directly or via a bonding group. That is, an oxygen atom of an ethyleneoxy group: (CH ₂ CH ₂ O) unit is a terminal group.

The other end group of the polyethylene glycol segment is not particularly limited, and may be a hydrogen atom, a hydroxyl group, an alkoxy group having a carbon number (C1 to C8) which may have a substituent, or a substituent. And an aralkyloxy group having a carbon number (C7 to C20) which may be included. Examples of the substituent in the alkoxy group and aralkyloxy group include a hydroxyl group, an amino group, a formyl group, and a carboxy group.

Examples of the alkoxy group having a carbon number (C1 to C8) which may have a substituent in the terminal group include a linear, branched or cyclic alkoxy group having a carbon number (C1 to C8). For example, methoxy group, ethoxy group, n-propoxy group, isopropoxy group, n-butoxy group, isobutoxy group, t-butoxy group, n-pentyloxy group, isopentyloxy group, 2-methylbutoxy group, neopentyloxy Group, 1-ethylpropoxy group, n-hexyloxy group, 4-methylpentyloxy group, 3-methylpentyloxy group, 2-methylpentyloxy group, 1-methylpentyloxy group, 3,3-dimethylbutoxy group, 2,2-dimethylbutoxy group, 1,1-dimethylbutoxy group, 1,2-dimethylbutoxy group, 1,3-dimethylbutoxy group, 2,3-dimethylbutoxy group, 2-ethylbutoxy group, cyclopropoxy group, Examples include a cyclopentyloxy group or a cyclohexyloxy group. Preferred is an alkoxy group having a carbon number (C1 to C4), such as a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, an n-butoxy group, an s-butoxy group, or a t-butoxy group. Particularly preferred is a methoxy group, an ethoxy group, an n-propoxy group or an isopropoxy group.
The aralkyloxy group having a carbon number (C7 to C20) which may have a substituent in the terminal group is a linear or branched alkyl group in which any one hydrogen atom is substituted with an aryl group. is there. For example, benzyloxy group, 2-phenylethyloxy group, 4-phenylbutyloxy group, 3-phenylbutyloxy group, 5-phenylpentyloxy group, 6-phenylhexyloxy group, 8-phenyloctyloxy group, etc. It is done. A benzyloxy group, a 4-phenylbutyloxy group, and an 8-phenyloctyloxy group are preferable.

In the polyamino acid derivative of the present invention, the polyethylene glycol segment is bonded to the side chain carboxy group directly or via a bonding group. As an aspect in which the polyethylene glycol segment is directly bonded to the side chain carboxy group, the terminal group of the ethyleneoxy group: (CH ₂ CH ₂ O) unit is an oxygen atom, and the side chain carboxy group and an ester bond It is the mode which is doing.
The present invention also includes an embodiment in which the polyethylene glycol segment is bonded to the side chain carboxy group via a bonding group. The linking group has a binding functional group in which one end group is bonded to the terminal oxygen atom of the polyethylene glycol segment in an ether bond mode, an ester bond, a urethane bond, or a carbonate bond, and the other end group is on the side. It is preferably an alkylene group having a linking functional group capable of forming an ester bond, an amide bond or a thioester bond with a chain carboxy group and having a substituent (C1 to C8).

In the linking group, as a linking group that is ether-bonded to the polyethylene glycol segment and amide bond, ester bond or thioester bond to the side chain carboxy group, for example, — (CH ₂ ) _x —NH— (x is 1-8) -(CH ₂ ) _x -O- (x represents an integer of 1 to 8) and-(CH ₂ ) _x -S- (x represents an integer of 1 to 8). As a linking group that is ester-bonded to a polyethylene glycol segment and is linked to an amide bond, ester bond or thioester bond to a side chain carboxy group, for example, —CO— (CH ₂ ) _x —NH— (x represents an integer of 1 to 8) , —CO— (CH ₂ ) _x —O— (wherein x represents an integer of 1 to 8) and —CO— (CH ₂ ) _x —S— (wherein x represents an integer of 1 to 8). —CONH— (CH ₂ ) _x —NH— (x represents an integer of 1 to 8) as a linking group that is bonded to a polyethylene glycol segment by urethane and is bonded to a side chain carboxy group by an amide bond, ester bond or thioester bond, CONH— (CH ₂ ) _x —O— (x represents an integer of 1 to 8) and —CONH— (CH ₂ ) _x —S— (x represents an integer of 1 to 8). -COO- (CH ₂ ) _x -NH- (x represents an integer of 1 to 8) as a linking group that binds to a polyethylene glycol segment and carbonate, and a side chain carboxy group to an amide bond, ester bond or thioester bond , —COO— (CH ₂ ) _x —O— (x represents an integer of 1 to 8), —COO— (CH ₂ ) _x —S— (x represents an integer of 1 to 8). it can. The linking group is particularly preferably — (CH ₂ ) _x —NH— (x represents an integer of 1 to 8), which is a linking group that has an ether bond with a polyethylene glycol segment and an amide bond with a side chain carboxy group.

Moreover, you may use an amino acid derivative as said coupling group of a polyethyleneglycol segment and a side chain carboxy group. When an amino acid derivative is used as a linking group, the N-terminal amino group of the amino acid is amide-bonded to the side chain carboxy group, and the C-terminal carboxy group is used as an ester bond with the terminal oxygen atom of the polyethylene glycol segment. .
When an amino acid derivative is used as the linking group, the amino acid used may be a natural amino acid or a non-natural amino acid, and any of L-form and D-form can be used without particular limitation. For example, hydrocarbon amino acids such as glycine, β-alanine, alanine, leucine and phenylalanine, acidic amino acids such as aspartic acid and glutamic acid, basic amino acids such as lysine, arginine and histidine can be used.

The polyethylene glycol segment preferably has 2 to 80 units bonded to a plurality of side chain carboxy groups of the polyamino acid derivative according to the present invention. That is, the nucleic acid antimetabolite-binding polyamino acid derivative preferably comprises a plurality of units of polyethylene glycol segments. More preferred are those having 2 to 70 units of polyethylene glycol segments, and particularly preferred is an embodiment in which 2 to 60 units are bound.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention binds a nucleic acid antimetabolite to the side chain carboxy group directly or via a bonding group. The nucleic acid antimetabolite is a compound having a structure of a nucleoside derivative having pharmacological activity such as antitumor activity or antiviral activity. As the nucleic acid metabolism antagonist in the present invention, it is preferable to use pyrimidine base nucleoside derivatives, purine base nucleoside derivatives, triazine base nucleoside derivatives and the like. The nucleic acid metabolism antagonist is preferably a compound having an amino group and / or a hydroxyl group in the molecule. That is, it is preferable because it can be introduced into the aforementioned polyamino acid main chain via an amide bond and / or an ester bond due to the amino group and / or hydroxyl group. In particular, it is preferable to use a nucleic acid antimetabolite having an amino group in the nucleoside nucleobase, and a pyrimidine base nucleoside derivative having an amino group, a purine base nucleoside derivative having an amino group, and a triazine base nucleoside derivative having an amino group are preferable. A nucleic acid antimetabolite having an amino group is particularly preferred because it can be introduced into a polyamino acid derivative by an amide bond with the amino group.

As the nucleic acid metabolism antagonist, a plurality of compounds having antitumor activity and antiviral activity are known. For example, cytarabine, gemcitabine, azacitidine, decitabine, nelarabine, 2′-methylidene-2′-deoxycytidine (DC), having the structure shown in the following formula (12): Troxacitabine, 3′-ethynylcytidine, 2′-cyano-2′-deoxy-1-β-D-arabinofuranosylcytosine (CNDAC), 2′-deoxy-5,6-dihydro- 5-azacytidine (DHAC), 5′-fluoro-2′-deoxycytidine (NSC-48006), 4′-thio-β-D-arabinofuranosylcytosine (OSI-7836), cladri Down (cladribine), clofarabine (Clofarabine) or fludarabine (Fludarabine), cytarabine-5'-elaidate (CP-4055), gemcitabine-5'-elaidic acid ester (CP-4126), and the like. The nucleic acid antimetabolite used in the polyamino acid derivative of the present invention is not limited to these compounds, but can be mentioned as preferred compounds to be applied.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention is obtained by binding the nucleic acid antimetabolite to a side chain carboxy group of a polyamino acid directly or via a binding group.
When the nucleic acid antimetabolite is directly bonded to the side chain carboxy group, a nucleic acid antimetabolite having an amino group and / or a hydroxyl group is used, and an amide bond with the side chain carboxy group by the amino group or the hydroxyl group What is necessary is just to couple | bond by the ester bond with this side chain carboxy group. The bonding mode may be any of an amide bond only, an ester bond only, or a mixture of an amide bond and an ester bond. Depending on the binding functional group of the nucleic acid antimetabolite used, the mode of binding to the side chain carboxy group may be appropriately selected.

The nucleic acid antimetabolite also includes an embodiment in which the side chain carboxy group is bonded via a suitable linking group. The linking group has a binding functional group in which one terminal group can bind to an amino group and / or a hydroxyl group that is a binding functional group of the nucleic acid antimetabolite, an amide bond, an ester bond, a carbonate bond, a urethane bond, or a urea bond. And the other terminal group has a linking functional group capable of forming an amide bond, an ester bond or a thioester bond with the side chain carboxy group, and optionally having a substituent (C1-C8) alkylene group It is preferable that

In the linking group that binds the nucleic acid antimetabolite to the side chain carboxy group, the amino group and / or hydroxyl group of the nucleic acid antimetabolite is linked to an amide bond or ester bond, and the side chain carboxy group is linked to an amide bond, ester bond or thioester bond. As the format, for example, —CO— (CH ₂ ) _y —NH— (y represents an integer of 1 to 8), —CO— (CH ₂ ) _y —O— (y represents an integer of 1 to 8). ), —CO— (CH ₂ ) _y —S— (y represents an integer of 1 to 8). Examples of a mode in which a urea bond or a urethane bond with a nucleic acid antimetabolite and a side chain carboxy group with an amide bond, an ester bond or a thioester bond include -CONH- (CH ₂ ) _y -NH- (where y is 1 to 8). -CONH- (CH ₂ ) _y -O- (y is an integer of 1 to 8), -CONH- (CH ₂ ) _y -S- (y is an integer of 1 to 8) Is mentioned. On the other hand, the form of carbonate bond with a nucleic acid antimetabolite and side chain carboxy group with amide bond, ester bond or thioester bond is, for example, —COO— (CH ₂ ) _y —NH— (y is an integer of 1 to 8) -COO- (CH ₂ ) _y -O- (y is an integer of 1 to 8), -COO- (CH ₂ ) _y -S- (y is an integer of 1 to 8) Can be mentioned.

The hydrogen atom of the alkylene group of the linking group may be modified with an appropriate substituent. Examples of the substituent include a hydroxyl group, an amino group, a halogen atom, an alkyl group having a carbon number (C1 to C8), an alkylcarbonylalkoxy group having a carbon number (C1 to C8), and an alkylcarbonylamide group having a carbon number (C1 to C8). An alkylcarbonylalkylamide group having a carbon number (C1 to C8), an alkylaryl group having a carbon number (C1 to C8), an alkoxy group having a carbon number (C1 to C8), an alkylamino group having a carbon number (C1 to C8), And an acylamide group having a carbon number (C1 to C8) and an alkoxycarbonylamino group having a carbon number (C1 to C8).

The linking group is a —CO— (CH ₂ ) _y —NH— (y represents an integer of 1 to 8) having a carboxy group on the binding side with a nucleic acid antimetabolite and the other having an amino group or a hydroxyl group, —CO— (CH ₂ ) _y —O— (y is an integer of 1 to 8) is preferred. Particularly preferably, —CO— (CH ₂ ) _y —NH— () having a carboxy group capable of amide bond or ester bond with the nucleic acid antimetabolite and an amino group capable of amide bond with the side chain carboxy group. y represents an integer of 1 to 8.
As the linking group, preferred —CO— (CH ₂ ) _y —NH— (y represents an integer of 1 to 8) includes —CO—CH ₂ —NH—, —CO— (CH ₂ ) ₂ —NH. —, —CO— (CH ₂ ) ₄ —NH—, —CO— (CH ₂ ) ₆ —NH—.

In the nucleic acid antimetabolite, the —CO— (CH ₂ ) _y —NH— (y represents an integer of 1 to 8) group optionally having substituent (s) as the preferred linking group, The case of 1 is synonymous with the amino acid skeleton. Therefore, an amino acid derivative may be used as the binding group. When the amino acid derivative is used as a linking group, the N-terminal amino group of the amino acid is an amide bond with the side chain carboxy group, and the C-terminal carboxy group is an amide bond or an ester bond with the amino group or hydroxyl group of the nucleic acid antimetabolite. Used as a linking group in embodiments.
When an amino acid derivative is used as the binding group, the amino acid used may be a natural amino acid or a non-natural amino acid, and any of L-form and D-form can be used without any particular limitation. For example, hydrocarbon amino acids such as glycine, β-alanine, alanine, leucine and phenylalanine, acidic amino acids such as aspartic acid and glutamic acid, basic amino acids such as lysine, arginine and histidine can be used.

When using an amino acid derivative as the binding group of the nucleic acid antimetabolite, it is preferable to use an aspartic acid derivative. The aspartic acid derivative is an aspartic acid derivative binding group in which an α-carboxy group functions as a binding group of the nucleic acid antimetabolite and the β-carboxy group is an amide. Alternatively, an aspartic acid derivative in which a β-carboxy group functions as a binding group of the nucleic acid antimetabolite and the α-carboxy group is an amide may be used. The amide in the other carboxy group that is not a binding group of the nucleic acid antimetabolite is, for example, an alkylamide having a carbon number (C1-20) which may have a substituent, or a substituent. And an aromatic amide having a good carbon number (C5 to C20), an aralkylamide having a carbon number (C7 to C20) which may have a substituent, or an amino acid residue in which a carboxy group is protected.

In the case of using an aspartic acid derivative in which one carboxy group is a linking group of a nucleic acid antimetabolite and one carboxy group is an amide derivative, the linking group that binds the nucleic acid antimetabolite to the side chain carboxy group, This is particularly preferable because reliable dissociation of the nucleic acid antimetabolite is promoted.
Examples of the alkylamide having a carbon number (C1-20) which may have a substituent of an aspartic acid derivative as a linking group include, for example, methylamide, ethylamide, isopropylamide, t-butylamide, cyclohexylamide, dodecylamide, octadecyl Examples include amides. Examples of the aromatic amide having a carbon number (C5 to C20) which may have a substituent of the aspartic acid derivative include phenylamide, 4-methoxyphenylamide, 4-dimethylaminophenylamide, 4-hydroxyphenyl. Examples include amides. Examples of the aralkyl amide having a carbon number (C7 to C20) which may have a substituent of the aspartic acid derivative include benzylamide, 2-phenylethylamide, 4-phenylbutyramide, 8-phenyloctylamide and the like. Is mentioned. Examples of the amino acid amide in which the carboxy group of the aspartic acid derivative is protected include glycinyl-methyl ester, alanyl-methyl ester, leucinyl-methyl ester, isoleucinyl-methyl ester, valinyl-methyl ester, phenylalanyl-methyl ester, Examples include alanyl-ethyl ester, leucinyl-ethyl ester, isoleucinyl-ethyl ester, alanyl-butyl ester, and leucinyl-butyl ester.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention preferably has an embodiment in which a nucleic acid antimetabolite having an amino group and / or a hydroxyl group is directly bonded to the carboxy group of the aspartic acid unit. That is, when the main chain polymer of the polyamino acid is polyaspartic acid, the nucleic acid antimetabolite may be directly bonded to the side chain carboxy group of the main chain polymer. A nucleic acid antimetabolite may be bound using an aspartic acid derivative as a binding group to be bound.
Further, when the main chain polymer of the polyamino acid is polyglutamic acid, it is preferable to bind the nucleic acid antimetabolite using an aspartic acid derivative as a binding group for binding the nucleic acid antimetabolite and the side chain carboxy group. As the aspartic acid derivative used as a linking group, an aspartic acid amide derivative is preferably used.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention comprises a polyamino acid containing a plurality of aspartic acid derivatives and / or glutamic acid derivatives as a main chain, and polyethylene glycol on the side chain carboxy group of the aspartic acid and / or glutamic acid. A polyamino acid derivative characterized by a chemical structure in which a segment and a nucleic acid antimetabolite are bonded directly or via a linking group, wherein the molecular weight of the polyamino acid derivative is 20 kilodaltons or more and 200 kilodaltons or less And More preferably, the molecular weight is 20 kilodaltons or more and 160 kilodaltons or less.

As the molecular weight of the polyamino acid derivative of the present invention, a calculated value obtained by adding the constituent molecular weights of the above-described constituent parts is adopted as the “molecular weight of the polyamino acid derivative”. That is, (1) the molecular weight of the polyamino acid main chain, (2) the total molecular weight of the polyethylene glycol segment obtained by multiplying the molecular weight of the polyethylene glycol segment by the number of bonds, and (3) the number of bonds in the molecular weight of the binding residue of the nucleic acid antimetabolite. (4) the total molecular weight of the binding group obtained by multiplying the binding group residue molecular weight of any polyethylene glycol segment by the number of bonds, and (5) the total molecular weight of the nucleic acid metabolism antagonist The calculated value obtained by adding the total molecular weight of the linking group obtained by multiplying the linking group residue molecular weight by the number of bonds is defined as the molecular weight.
The molecular weight of the polyamino acid derivative is required to be regulated by the accuracy in kilodalton units. Therefore, the analysis method for each component is not particularly limited as long as it is a sufficiently accurate analysis method for measuring the molecular weight of the polyamino acid derivative in kilodalton units, and various analysis methods are appropriately selected. Good. Below, the preferable analysis method in each component is listed.

(1) The molecular weight of the polyamino acid main chain is a calculated value obtained by multiplying the molecular weight of the polymerization monomer unit of the main chain by the number of polymerizations. When the polyethylene glycol segment is bonded to the end of the polyamino acid main chain, the calculated value is obtained by adding the molecular weight of the polyethylene glycol segment to the molecular weight of the polymerization monomer unit of the main chain multiplied by the number of polymerizations. is there. The number of polymerizations is the number of polymerizations calculated from the integral value of ¹ H-NMR, the number of polymerizations calculated by amino acid analysis, or the number of polymerizations calculated by quantifying the side chain carboxy group of polyamino acid by neutralization titration. be able to. When the polyamino acid main chain is a polymer main chain having a single amino acid structure such as polyaspartic acid, analysis of ¹ H-NMR is simple and preferable, and protons serving as a reference from the integrated value of ¹ H-NMR It is preferable to use the number of polymerizations calculated using the integral value.

(2) The total molecular weight of the polyethylene glycol segment is a calculated value obtained by multiplying the molecular weight of the polyethylene glycol segment by the binding amount. As the molecular weight of the polyethylene glycol segment, an average molecular weight determined by the peak top molecular weight of the polyethylene glycol segment structural compound to be used, which is measured by a GPC method based on a polyethylene glycol standard product, is employed.

The binding amount of the polyethylene glycol segment can be determined by cleaving the polyethylene glycol segment from the nucleic acid antimetabolite-binding polyamino acid derivative and quantitatively analyzing the released polyethylene glycol segment. Alternatively, since the constituents of the nucleic acid antimetabolite-binding polyamino acid derivative are a polyethylene glycol segment, a polyamino acid main chain, and a nucleic acid antimetabolite, the mass content of the amino acid main chain calculated by amino acid analysis and the nucleic acid antimetabolite The mass content of the nucleic acid antimetabolite calculated by hydrolyzing the bound polyamino acid derivative and quantitatively analyzing the released nucleic acid antimetabolite by high performance liquid chromatography (HPLC) is obtained, and the remaining mass content is determined as polyethylene. A method of calculating as a glycol segment may be used. Alternatively, a method of calculating from the consumption rate of the polyethylene glycol segment in the reaction of introducing the polyethylene glycol segment into the polyamino acid main chain may be used.
In addition, the total molecular weight of the bonding group of the arbitrary polyethylene glycol segment of (4) is a calculated value obtained by multiplying the bonding group residue molecular weight by the number of bonds. The number of bonds of the bonding group is the same as the number of bonds of the polyethylene glycol segment described above, and can be calculated by using the value.

The total molecular weight of the nucleic acid antimetabolite (3) is a calculated value obtained by multiplying the binding residue molecular weight of the nucleic acid antimetabolite by the number of bonds. The binding number of the nucleic acid antimetabolite is a value calculated by hydrolyzing the nucleic acid antimetabolite-binding polyamino acid derivative and quantitatively analyzing the released nucleic acid antimetabolite by high performance liquid chromatography (HPLC). Is preferably used.
The total molecular weight of the binding group of any nucleic acid antimetabolite in (5) is a calculated value obtained by multiplying the binding group residue molecular weight by the number of bonds. The number of bonds of the binding group is the same as the number of bonds of the aforementioned nucleic acid antimetabolite, and can be calculated by using the value.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention is characterized in that the mass content of the polyethylene glycol segment in the polyamino acid derivative is 30% by mass or more and 90% by mass or less. The mass content of the polyethylene glycol segment can be calculated from the content ratio of the total molecular weight of the polyethylene glycol segment of (2) above to the molecular weight of the polyamino acid derivative described above. That is, the mass content of the polyethylene glycol segment is calculated by the following formula.
(Calculation formula) Mass content of polyethylene glycol segment (%) = total molecular weight of polyethylene glycol segment / polyamino acid derivative molecular weight × 100
The polyethylene glycol segment has a mass molecular weight of 30% by mass to 90% by mass, and preferably 35% by mass to 85% by mass.

In the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention, the mass content of the nucleic acid antimetabolite in the polyamino acid derivative is preferably 2% by mass or more and 60% by mass or less.
If the content of the nucleic acid antimetabolite is less than 2% by mass, the total dose of the polyamino acid derivative is increased in order to ensure an effective amount of the nucleic acid antimetabolite, which is not preferable. On the other hand, when the content of the nucleic acid antimetabolite is more than 60% by mass, myelosuppression tends to be strongly developed. It is preferable to set the content of the nucleic acid antimetabolite in order to ensure administration convenience and achieve sufficient drug efficacy and side effect reduction.
The mass content of the nucleic acid antimetabolite in the polyamino acid derivative can be calculated from the content ratio of the total molecular weight of the nucleic acid antimetabolite (3) to the molecular weight of the polyamino acid derivative. A more preferable range of the content of the nucleic acid antimetabolite is 5% by mass or more and 50% by mass or less. It is particularly preferable that the content of the nucleic acid antimetabolite is 5% by mass or more and 40% by mass or less.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention is preferably prepared by preparing an aqueous solution of the polyamino acid derivative and administering it parenterally. The aqueous solution is prepared by dissolving in water, physiological saline, phosphate buffered saline (PBS solution), 5% glucose aqueous solution, or the like.
The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention preferably has physical properties that do not exhibit associative properties in the aqueous solution. Here, the association property indicates that the polyamino acid derivative is a physical property that forms an aggregate in which more than 10 molecules self-associate. Therefore, the “physical property not exhibiting associative property” in the present invention indicates an embodiment in which the polyamino acid derivative in the aqueous solution exists as a monomolecular substance or the self-aggregate forms an aggregate of 10 molecules or less.
It is effective to use light scattering intensity using laser light as an index of the self-association property of the nucleic acid antimetabolite-binding hyperbranched compound of the present invention. That is, the self-association property of the nucleic acid antimetabolite-binding hyperbranched compound in an aqueous solution can be confirmed using the laser light scattering intensity as an index. At that time, a method for confirming the self-association property of the nucleic acid antimetabolite-binding multibranched compound in an aqueous solution using toluene as a light scattering intensity standard sample and relative intensity with respect to toluene as an index is effective.
In the present invention, when an aqueous solution having a polyamino acid derivative concentration of 1 mg / mL is measured with a laser light scattering photometer, and the light scattering intensity is 5 times or less as the relative intensity to the light scattering intensity of toluene, It is considered that the amino acid derivative does not exhibit associative properties and is dispersed in an aqueous solution in the form of a monomolecular substance to several molecules. Preferably, it is a polyamino acid derivative whose light scattering intensity is 3 times or less as relative intensity to the light scattering intensity of toluene.

As the laser light scattering photometer, for example, a dynamic light scattering photometer DLS-8000DL (measurement temperature 25 ° C., measurement angle: 90 °, wavelength: 632.8 nm, ND filter: 5%, PH1: OPEN, manufactured by Otsuka Electronics Co., Ltd.) PH2: SLIT, sample concentration: 1 mg / mL), and measuring the light scattering intensity of an aqueous solution having a polyamino acid derivative concentration of 1 mg / mL with a laser light scattering photometer.
In addition, toluene used as a standard substance for light scattering intensity measurement is not particularly limited as long as it has a reagent level purity, and can be used. It is preferable to use toluene that has been subjected to pretreatment filtration, which is usually performed in sample preparation for light scattering analysis.
In this measurement method, the polyamino acid derivative preferably has a light scattering intensity of 5 times or less, more preferably 3 times or less as a relative intensity with respect to the light scattering intensity of toluene. In this case, the lower limit value is not particularly limited, and is a case where a clear light scattering intensity is not exhibited, indicating a state where self-association is not exhibited in an aqueous solution.

The presence or absence of self-association in the aqueous solution of the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention correlates with side effects such as bone marrow suppression. Nucleic acid metabolism antagonists have the problem that bone marrow suppression such as leukopenia occurs as a side effect, making it difficult to continue treatment using the therapeutic agent. For this reason, providing a therapeutic agent for a nucleic acid antimetabolite with low bone marrow suppression is very useful in a method for treating malignant tumors. In the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention, the molecular weight of the derivative is 20 kilodaltons or more and 200 kilodaltons or less, and the mass content of the polyethylene glycol segment in the polyamino acid derivative is 30 mass% or more and 90 mass% or less. As a result, the derivative has physical properties that do not exhibit self-association in an aqueous solution, and as a result, a medicinal product having a high therapeutic effect with little bone marrow suppression can be provided.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention is preferably a nucleic acid antimetabolite-binding polyamino acid derivative represented by the following general formula (1).

In the formula, R ₁ is a group selected from the group consisting of a hydrogen atom, an alkyl group having 1 to 8 carbon atoms and a polyethylene glycol segment, and R ₂ is a hydrogen atom and an acyl group having 1 to 8 carbon atoms (C1 to C8). And a group selected from the group consisting of C1-C8 alkoxycarbonyl groups, R ₃ represents a polyethylene glycol segment, R ₄ represents a nucleic acid antimetabolite binding residue, and R ₅ represents aspartic acid. A bonding residue and / or an aspartic imide bonding residue, R ₆ represents a hydroxyl group and / or —N (R ₇ ) CONH (R ₈ ), and R ₇ and R ₈ may be the same or different. at best, a tertiary amino group may carbon atoms optionally substituted with (C1 ~ C8) for straight, branched or cyclic alkyl group, X ₁ and X ₂ is a linking group, X ₃ Is methylene Or an ethylene group, a, b, c, d, e, f, g, h and i each independently represent an integer of 0 to 200, and the total number of polymerization of the polyamino acid derivative (a + b + c + d + e + f + g + h + i) is 3 250, (a + b) is 1 to 95, (c + d) is 1 to 175, the amino acid unit to which R ₃ is bonded, the amino acid unit to which R ₄ is bonded, and the amino acid to which R ₅ is bonded The unit, the amino acid unit to which R ₆ is bonded, and the amino acid unit in which the side chain carboxy group is an intramolecular cyclization type are each independently a random sequence of a nucleic acid antimetabolite-binding polyamino acid derivative. The molecular weight of the polyamino acid derivative is 20 kilodaltons or more and 200 kilodaltons or less, and the mass content of the polyethylene glycol segment in the polyamino acid derivative is 30 mass% or more and 90 mass% or less.

R ₁ in the general formula (1) is a substituent selected from the group consisting of a hydrogen atom, an alkyl group having from 1 to 8 carbon atoms and a polyethylene glycol segment.
The alkyl group having a carbon number (C1 to C8) is a linear, branched or cyclic alkyl group having a carbon number (C1 to C8). Examples of the linear alkyl group include a methyl group, an ethyl group, an n-propyl group, an n-butyl group, an n-hexyl group, and an n-octyl group. Examples of the branched alkyl group include isopropyl group, t-butyl group, 1-methyl-propyl group, 2-methyl-propyl group, 2,2-dimethylpropyl group and the like. Examples of the cyclic alkyl group include a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, a cyclohexyl group, a cyclohexylmethyl group, and a cyclohexylethyl group.

The polyethylene glycol segment in R ₁ of the general formula (1) is a segment having a repeating structure of ethyleneoxy group; (CH ₂ CH ₂ O) units. A segment structure containing a polyethylene glycol chain having an ethyleneoxy group unit polymerization degree of 5 to 10,000 units, more preferably a polymerization degree of 5 to 5,000 units. That is, the polyethylene glycol segment is preferably a segment portion having an average molecular weight of 200 to 500 kilodaltons equivalent to polyethylene glycol, more preferably a structural portion having an average molecular weight of 200 to 250 kilodaltons, particularly preferably. The average molecular weight is 200 to 150 kilodaltons. A polyethylene glycol segment having an average molecular weight of 1,000 to 50 kilodaltons is particularly preferred.
The average molecular weight of the polyethylene glycol segment used in the present invention is the GPC method based on the polyethylene glycol standard structure of the polyethylene glycol segment structural compound used in preparing the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention. It is an average molecular weight calculated | required by the peak top molecular weight measured by (1).

In the polyethylene glycol segment as R ₁ , the bonding side terminal group with the polyamino acid main chain is not particularly limited as long as it is a suitable linking group to the polyamino acid derivative. An alkylene group having a carbon number (C1 to C8) which may have a substituent is preferable. For example, a methylene group, ethylene group, propylene group, butylene group, hexamethylene group, octamethylene group and the like can be mentioned. That is, the polyethylene glycol segment structure is preferably a structure in which an oxygen atom of an ethyleneoxy group: (CH ₂ CH ₂ O) unit and an alkylene group having the carbon number (C1 to C8) are ether-bonded.

Further, the terminal group of the polyethylene glycol segment in R ₁ is not particularly limited, and may be a hydrogen atom, a hydroxyl group, an alkoxy group having a carbon number (C1 to C8) which may have a substituent, or a substituent. Examples thereof include an optionally substituted carbon number (C7 to C20) aralkyloxy group. Examples of the substituent in the alkoxy group, alkynyloxy group, and aralkyloxy group include a hydroxyl group, an amino group, a formyl group, and a carboxy group.
Examples of the carbon number (C1 to C8) alkoxy group which may have a substituent in the terminal group include a linear, branched or cyclic (C1 to C8) alkoxy group. For example, methoxy group, ethoxy group, n-propoxy group, isopropoxy group, n-butoxy group, isobutoxy group, t-butoxy group, n-pentyloxy group, isopentyloxy group, 2-methylbutoxy group, neopentyloxy Group, 1-ethylpropoxy group, n-hexyloxy group, 4-methylpentyloxy group, 3-methylpentyloxy group, 2-methylpentyloxy group, 1-methylpentyloxy group, 3,3-dimethylbutoxy group, 2,2-dimethylbutoxy group, 1,1-dimethylbutoxy group, 1,2-dimethylbutoxy group, 1,3-dimethylbutoxy group, 2,3-dimethylbutoxy group, 2-ethylbutoxy group, cyclopropoxy group, Examples include a cyclopentyloxy group or a cyclohexyloxy group. Preferred is a (C1 to C4) alkoxy group, such as a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, an n-butoxy group, an s-butoxy group or a t-butoxy group, and particularly preferably A methoxy group, an ethoxy group, an n-propoxy group or an isopropoxy group;
The carbon number (C7 to C20) aralkyloxy group which may have a substituent in the terminal group is a linear or branched alkyl group in which any one hydrogen atom is substituted with an aryl group . For example, benzyloxy group, 2-phenylethyloxy group, 4-phenylbutyloxy group, 3-phenylbutyloxy group, 5-phenylpentyloxy group, 6-phenylhexyloxy group, 8-phenyloctyloxy group, etc. It is done. A benzyloxy group, a 4-phenylbutyloxy group, and an 8-phenyloctyloxy group are preferable.

R ₂ in the general formula (1) represents one or more substituents selected from the group consisting of a hydrogen atom, an acyl group having a carbon number (C1 to C8) and an alkoxycarbonyl group having a carbon number (C1 to C8). Show.
The acyl group having a carbon number (C1 to C8) is a linear, branched or cyclic acyl group having a carbon number (C1 to C8). Examples include formyl group, acetyl group, propionyl group, butyroyl group, cyclopropylcarbonyl group, cyclopentanecarbonyl group and the like.
The alkoxycarbonyl group having a carbon number (C1 to C8) is a linear, branched or cyclic alkoxycarbonyl group having a carbon number (C1 to C8). For example, methoxycarbonyl group, ethoxycarbonyl group, propoxycarbonyl group, isopropoxycarbonyl group, n-butoxycarbonyl group, t-butoxycarbonyl group, pentoxycarbonyl group, hexyloxycarbonyl group, cyclopropoxycarbonyl group, cyclopentyloxycarbonyl group And a cyclohexyloxycarbonyl group.

R ₃ in the general formula (1) represents a polyethylene glycol segment. The polyethylene glycol segment is a segment having a repeating structure of an ethyleneoxy group: (CH ₂ CH ₂ O) unit. A segment structure containing a polyethylene glycol chain having an ethyleneoxy group unit polymerization degree of 5 to 10,000 units, more preferably a polymerization degree of 5 to 5,000 units. That is, the polyethylene glycol segment is preferably a segment portion having an average molecular weight of 200 to 500 kilodaltons equivalent to polyethylene glycol, more preferably a structural portion having an average molecular weight of 200 to 250 kilodaltons, particularly preferably. The average molecular weight is 200 to 150 kilodaltons. A polyethylene glycol segment having an average molecular weight of 1,000 to 50 kilodaltons is particularly preferred.
The average molecular weight of the polyethylene glycol segment used in the present invention is the GPC method based on the polyethylene glycol standard structure of the polyethylene glycol segment structural compound used in preparing the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention. It is an average molecular weight calculated | required by the peak top molecular weight measured by (1).

The terminal group of the polyethylene glycol segment in R _{3 that} is bonded to the side chain carboxy group of the polyamino acid main chain is a linking group for bonding to the side chain carboxy group directly or via a bonding group. . That is, an oxygen atom of an ethyleneoxy group: (CH ₂ CH ₂ O) unit is a terminal group.

On the other hand, the other terminal group of the polyethylene glycol segment in R ₃ is not particularly limited, and may be a hydrogen atom, a hydroxyl group, an optionally substituted alkoxy group having a carbon number (C1 to C8), a substituted group. And an aralkyloxy group having a carbon number (C7 to C20) which may have a group. Examples of the substituent in the alkoxy group, alkynyloxy group, and aralkyloxy group include a hydroxyl group, an amino group, a formyl group, and a carboxy group.
Examples of the alkoxy group having a carbon number (C1 to C8) which may have a substituent in the terminal group include a linear, branched or cyclic (C1 to C8) alkoxy group. For example, methoxy group, ethoxy group, n-propoxy group, isopropoxy group, n-butoxy group, isobutoxy group, t-butoxy group, n-pentyloxy group, isopentyloxy group, 2-methylbutoxy group, neopentyloxy Group, 1-ethylpropoxy group, n-hexyloxy group, 4-methylpentyloxy group, 3-methylpentyloxy group, 2-methylpentyloxy group, 1-methylpentyloxy group, 3,3-dimethylbutoxy group, 2,2-dimethylbutoxy group, 1,1-dimethylbutoxy group, 1,2-dimethylbutoxy group, 1,3-dimethylbutoxy group, 2,3-dimethylbutoxy group, 2-ethylbutoxy group, cyclopropoxy group, Examples include a cyclopentyloxy group or a cyclohexyloxy group. Preferred is a (C1 to C4) alkoxy group, such as a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, an n-butoxy group, an s-butoxy group or a t-butoxy group, and particularly preferably A methoxy group, an ethoxy group, an n-propoxy group or an isopropoxy group;
The aralkyloxy group having a carbon number (C7 to C20) which may have a substituent in the terminal group is a linear or branched alkyl group in which any one hydrogen atom is substituted with an aryl group. is there. For example, benzyloxy group, 2-phenylethyloxy group, 4-phenylbutyloxy group, 3-phenylbutyloxy group, 5-phenylpentyloxy group, 6-phenylhexyloxy group, 8-phenyloctyloxy group, etc. It is done. A benzyloxy group, a 4-phenylbutyloxy group, and an 8-phenyloctyloxy group are preferable.

R ₃ in the general formula (1) is more preferably a polyethylene glycol segment represented by the following general formula (7).

In the formula, R ₁₅ represents a hydrogen atom or a linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may have a substituent, and n is an integer of 5 to 2,500. Indicates.

Examples of the linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may have a substituent in R ₁₅ include, for example, a methyl group, an ethyl group, and the like. Group, n-propyl group, n-butyl group, n-hexyl group, n-decyl group and the like. Examples of the branched alkyl group include isopropyl group, t-butyl group, 1-methyl-propyl group, 2-methyl-propyl group, 2,2-dimethylpropyl group and the like. Examples of the cyclic alkyl group include a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, a cyclohexyl group, and an adamantyl group.
Examples of the substituent that may have a mercapto group, a hydroxyl group, a halogen atom, a nitro group, a cyano group, a carbocyclic or heterocyclic aryl group, an alkylthio group, an arylthio group, an alkylsulfinyl group, an arylsulfinyl group, an alkylsulfonyl group, Arylsulfonyl group, sulfamoyl group, alkoxy group, aryloxy group, acyloxy group, alkoxycarbonyloxy group, carbamoyloxy group, substituted or unsubstituted amino group, acylamino group, alkoxycarbonylamino group, ureido group, sulfonylamino group, sulfa A moylamino group, a formyl group, an acyl group, a carboxy group, an alkoxycarbonyl group, a carbamoyl group or a silyl group can be exemplified. The substitution position on the aromatic ring may be the ortho position, the meta position, or the para position.

X ₁ in the general formula (1) is a linking group that binds the polyethylene glycol segment according to R ₃ and the side chain carboxy group of the polyamino acid derivative. The linking group is not particularly limited as long as it is a linking group having functional groups at both ends capable of binding to the oxygen atom of the polyethylene glycol segment end group and the side chain carboxy group of the polyamino acid derivative. It is not something.

The linking group has a binding functional group in which one end group is bonded to the terminal oxygen atom of the polyethylene glycol segment in an ether bond mode, an ester bond, a urethane bond, or a carbonate bond, and the other end group is on the side. It is a C1-C8 alkylene group which has a linking functional group capable of forming an ester bond, an amide bond, or a thioester bond with a chain carboxy group and may have a substituent.
Examples of the linking group according to X ₁ include an ether bond with a polyethylene glycol segment, and a linking group with an amide bond, an ester bond or a thioester bond with a side chain carboxy group, such as — (CH ₂ ) _x —NH— (where x is 1 represents an integer of 1 to 8), — (CH ₂ ) _x —O— (x represents an integer of 1 to 8), — (CH ₂ ) _x —S— (x represents an integer of 1 to 8) Etc. As a linking group that is ester-bonded to a polyethylene glycol segment and is linked to an amide bond, ester bond or thioester bond to a side chain carboxy group, for example, —CO— (CH ₂ ) _x —NH— (x represents an integer of 1 to 8) , —CO— (CH ₂ ) _x —O— (x represents an integer of 1 to 8), —CO— (CH ₂ ) _x —S— (x represents an integer of 1 to 8), and the like. . As a linking group that is urethane-bonded to a polyethylene glycol segment and amide bond, ester bond or thioester bond to a side chain carboxy group, for example, —CONH— (CH ₂ ) _x —NH— (x represents an integer of 1 to 8) , —CONH— (CH ₂ ) _x —O— (x represents an integer of 1 to 8), —CONH— (CH ₂ ) _x —S— (x represents an integer of 1 to 8), and the like. . In addition, as a linking group that bonds to a polyethylene glycol segment and binds to a side chain carboxy group to an amide bond, ester bond, or thioester bond, for example, —COO— (CH ₂ ) _x —NH— (x is an integer of 1 to 8). -COO- (CH ₂ ) _x -O- (x represents an integer of 1 to 8), -COO- (CH ₂ ) _x -S- (x represents an integer of 1 to 8), etc. Can be mentioned. Preferably, it is a linking group that is ether-bonded to a polyethylene glycol segment and amide-bonded to a side chain carboxy group, and as X ₁ , — (CH ₂ ) _x —NH— (x represents an integer of 1 to 8) It is.

It is also possible to use an amino acid derivative as binding group according to the X _1. When the amino acid derivative is used as a linking group, the linking group is used in such a manner that the N-terminal amino group of the amino acid derivative is amide-bonded with the side chain carboxy group, and the C-terminal carboxy group is the terminal oxygen atom of the polyethylene glycol segment. And an ester bond.
When using an amino acid derivative as binding group according to the X _1, amino acids used may be natural amino acids or unnatural amino acids, L body, can be used without being limited particularly either D-form. For example, hydrocarbon amino acids such as glycine, β-alanine, alanine, leucine and phenylalanine, acidic amino acids such as aspartic acid and glutamic acid, basic amino acids such as lysine, arginine and histidine can be used.

The X ₁ may be a “bond”. The “bond” refers to an embodiment in which the side chain carboxy group of the polyamino acid derivative and the terminal oxygen atom of the polyethylene glycol segment are directly ester-linked without using a linking group.

R ₄ in the general formula (1) is a binding residue of a nucleic acid antimetabolite that has antitumor activity or antiviral activity and has a structure of a nucleoside derivative. The nucleic acid antimetabolite that can be used in the present invention is not particularly limited as long as it is a nucleic acid antimetabolite having an amino group and / or a hydroxyl group. Examples of the nucleic acid antimetabolite include pyrimidine antimetabolite, purine antimetabolite, triazine antimetabolite and the like. Here, the binding residue of the nucleic acid antimetabolite is a binding residue in the case of an amide bond by the amino group of the nucleic acid antimetabolite, or a nucleoside derivative in the case of an ester bond by the hydroxyl group of the nucleic acid antimetabolite Refers to the side conjugate structure.

The nucleic acid antimetabolite according to R ₄ is preferably a nucleic acid antimetabolite having an amino group at the nucleoside base. That is, R ₄ is a nucleic acid antimetabolite having an amino group, and is preferably a binding residue bonded by an amide bond.

In R ₄ , the nucleobase moiety is any one or more selected from the following formula (8), and the group (Rf) bonded thereto is selected from the following formula (9). Particularly preferred is a nucleic acid antimetabolite that is a combination of the above.

[Wherein, -Rf represents the formula (9):

And R ₁₆ represents a hydrogen atom or an acyl group residue of a fatty acid ester. ]

The acyl group of the fatty acid ester in R ₁₆ is an acyl residue in which a monocarboxylic acid having a carbon number (C4 to C30) is ester-bonded. The hydrocarbon having a carbon number (C4 to C30) may be a saturated fatty acid that is a saturated hydrocarbon, or an unsaturated fatty acid that is an unsaturated hydrocarbon containing one or more double bonds. These fatty acid esters are known as fat-soluble derivatives of the nucleic acid antimetabolite, and can be used as an active ingredient of the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention.
In the fatty acid of the acyl group residue of the fatty acid ester as R ₁₆ , the saturated fatty acid includes butanoic acid, pentanoic acid, hexanoic acid, octanoic acid, decanoic acid, dodecanoic acid, tetradecanoic acid, hexadecanoic acid, octadecanoic acid, eicosane. An acid, docosanoic acid, etc. are mentioned.
Examples of the unsaturated fatty acid include 9-hexadecenoic acid, cis-9-octadecenoic acid, trans-9-octadecenoic acid, cis, cis-9,12-octadecadienoic acid, and 9,12,15-octadecatrienoic acid. 6,9,12-octadecatrienoic acid, 5,8,11,14-eicosatetraenoic acid, and the like.

As the nucleic acid antimetabolite according to R ₄ in the general formula (1), it is particularly preferable to use a nucleic acid antimetabolite known to be effective as an antitumor agent or an antiviral agent. For example, cytarabine, gemcitabine, azacitidine, decitabine, nelarabine, 2'-methylidene-2'-deoxycytidine (DMDC), troxacitabine Cytidine (Ethynycytidine), 2′-cyano-2′-deoxy-1-β-D-arabinofuranosylcytosine (CNDAC), 2′-deoxy-5,6-dihydro-5-azacytidine (DHAC), 5′- Fluoro-2'-deoxycytidine (NSC-48006), 4'-thio-β-D-arabinofuranosylcytosine (OSI-7836), cladribine, clofala Down (Clofarabine) or fludarabine (Fludarabine), cytarabine-5'-elaidate (CP-4055), gemcitabine-5'-elaidic acid ester (CP-4126).

As the nucleic acid antimetabolite in R ₄ of the general formula (1), a cytidine antimetabolite is preferably used. The nucleobase moiety is a cytidine base represented by the following formula (10), and a group ( Rf) is particularly preferably a nucleic acid antimetabolite that is a combination of any one or more selected from the substituent group of the following formula (11). Here, R ₁₆ is a compound represented by a hydroxyl group or an acyl group of a fatty acid ester.

[Wherein, -Rf represents the formula (11):

And R ₁₆ represents a hydrogen atom or an acyl group of a fatty acid ester. ]
These cytidine antimetabolites are gemcitabine and its fatty acid ester derivatives, cytarabine and its fatty acid ester derivatives, and 3′-ethynylcytidine and its fatty acid ester derivatives. Examples of fatty acid ester derivatives include cytarabine-5′-elaidic acid ester (CP-4055), gemcitabine-5′-elaidic acid ester (CP-4126), and the like.

X ₂ in the general formula (1) is a linking group that binds the nucleic acid metabolism antagonist according to R ₄ and the side chain carbonyl group of the polyamino acid main chain. The binding group is not particularly limited as long as it is a binding group having both functional groups capable of binding to the binding functional group of the nucleic acid antimetabolite and the side chain carboxy group of the polyamino acid derivative at both ends. Is not to be done.
The terminal binding functional group of the nucleic acid metabolism antagonist side bonding groups according to the X _2, carboxy, oxy carboxy group or an amino carboxy group. Since the nucleic acid antimetabolite has an amino group and / or a hydroxyl group in the molecule, these binding functional groups include the amide bond, ester bond, urethane bond, carbonate bond and urea bond with the amino group and / or hydroxyl group. To do.
The other terminal binding functional group on the side chain carboxy group side of the binding group according to X ₂ is preferably an amino group, a hydroxyl group or a thiol group. These binding functional groups can form a side chain carboxy group and an amide bond, an ester bond, or a thioester bond.

That is, the bonding group according to X ₂ has a substituent in which one end group is a carboxy group, an oxycarboxy group or an aminocarboxy group, and the other end group is an amino group, a hydroxyl group or a thiol group. It is preferably a good (C1-C8) alkylene group.
Examples of the linking group according to X ₂ include a form of amide bond or ester bond with an amino group and / or hydroxyl group of a nucleic acid antimetabolite, and amide bond, ester bond or thioester bond with a side chain carboxy group. CO— (CH ₂ ) _y —NH— (y represents an integer of 1 to 8), —CO— (CH ₂ ) _y —O— (y represents an integer of 1 to 8), —CO— (CH ₂ ) _y -S- (y is an integer of 1 to 8). As a mode of binding a urea bond or a urethane bond with a nucleic acid antimetabolite, and a side chain carboxy group with an amide bond, an ester bond or a thioester bond, —CONH— (CH ₂ ) _y —NH— (where y is an integer of 1 to 8) -CONH- (CH ₂ ) _y -O- (y is an integer of 1 to 8), -CONH- (CH ₂ ) _y -S- (y is an integer of 1 to 8), etc. Can be mentioned. On the other hand, as a mode of forming a carbonate bond with a nucleic acid antimetabolite and binding a side chain carboxy group with an amide bond, an ester bond or a thioester bond, —COO— (CH ₂ ) _y —NH— (y represents an integer of 1 to 8) ), —COO— (CH ₂ ) _y —O— (y represents an integer of 1 to 8), —COO— (CH ₂ ) _y —S— (y represents an integer of 1 to 8). Can do.

In the alkylene group of the linking group according to X ₂ , a hydrogen atom may be modified with an appropriate substituent. Examples of the substituent include a hydroxyl group, an amino group, a halogen atom, an alkyl group having a carbon number (C1 to C8), an alkylcarbonylalkoxy group having a carbon number (C1 to C8), and an alkylcarbonylamide group having a carbon number (C1 to C8). An alkylcarbonylalkylamide group having a carbon number (C1 to C8), an alkylaryl group having a carbon number (C1 to C8), an alkoxy group having a carbon number (C1 to C8), an alkylamino group having a carbon number (C1 to C8), And an acylamide group having a carbon number (C1 to C8) and an alkoxycarbonylamino group having a carbon number (C1 to C8).

The linking group related to X ₂ is preferably a carboxy group on the binding side with the nucleic acid antimetabolite and the other is a linking group having an amino group or a hydroxyl group, and —CO— (CH ₂ ) _y —NH— (Y represents an integer of 1 to 8) and —CO— (CH ₂ ) _y —O— (y represents an integer of 1 to 8). Particularly preferably, —CO— (CH ₂ ) _y —NH— () having a carboxy group capable of amide bond or ester bond with the nucleic acid antimetabolite and an amino group capable of amide bond with the side chain carboxy group. y represents an integer of 1 to 8.
Specific examples of —CO— (CH ₂ ) _y —NH— (y represents an integer of 1 to 8), which is a preferred linking group related to X ₂ , include —CO—CH ₂ —NH—, —CO— (CH ₂ ) ₂ —NH—, —CO— (CH ₂ ) ₄ —NH—, —CO— (CH ₂ ) ₆ —NH—.

In the —CO— (CH ₂ ) _y —NH— (y represents an integer of 1 to 8) group optionally having substituent (s) listed as preferred linking groups for X ₂ , The case is synonymous with the amino acid skeleton. Therefore, a linking group according to the X _2, may be used amino acid derivatives.
When the amino acid derivative is used as a linking group, the N-terminal amino group of the amino acid is amide-bonded to the side chain carboxy group, and the C-terminal carboxy group is amide-bonded or ester-bonded to the amino group or hydroxyl group of the nucleic acid antimetabolite. Used as a linking group in embodiments.
When an amino acid derivative is used as a linking group, the amino acid used may be a natural amino acid or a non-natural amino acid, and any of L-form and D-form can be used without particular limitation. For example, hydrocarbon amino acids such as glycine, β-alanine, alanine, leucine and phenylalanine, acidic amino acids such as aspartic acid and glutamic acid, basic amino acids such as lysine, arginine and histidine can be used.

When using an amino acid derivative as binding group according to the X _2, it is preferable to use the aspartic acid derivative. The aspartic acid derivative is an aspartic acid derivative binding group in which an α-carboxy group functions as a binding group of the nucleic acid antimetabolite and the β-carboxy group is an amide. Alternatively, an aspartic acid derivative in which a β-carboxy group functions as a binding group of the nucleic acid antimetabolite and the α-carboxy group is an amide may be used. When the other carboxy group that is not the binding group of the nucleic acid antimetabolite is an amide, it may have a substituent (C1-20) alkylamide or a substituent. And an aromatic amide having a carbon number (C5 to C20), an aralkylamide having a carbon number (C7 to C20) which may have a substituent, or an amino acid residue in which a carboxy group is protected.

As coupling groups according to the X _2, is a linking group of one of the carboxy groups nucleic acid metabolism antagonist, if one of the carboxy group with aspartic acid derivative is an amide derivative, reliable dissociation of nucleic acid metabolism antagonist Is particularly preferred. Examples of the alkylamide having a carbon number (C1-20) which may have a substituent of an aspartic acid derivative as a linking group include, for example, methylamide, ethylamide, isopropylamide, t-butylamide, cyclohexylamide, dodecylamide, octadecyl Examples include amides. Examples of the aromatic amide having a carbon number (C5 to C20) which may have a substituent of the aspartic acid derivative include phenylamide, 4-methoxyphenylamide, 4-dimethylaminophenylamide, 4-hydroxyphenyl. Examples include amides. Examples of the aralkyl amide having a carbon number (C7 to C20) which may have a substituent of the aspartic acid derivative include benzylamide, 2-phenylethylamide, 4-phenylbutyramide, 8-phenyloctylamide and the like. Is mentioned. Examples of the amino acid amide in which the carboxy group of the aspartic acid derivative is protected include glycinyl-methyl ester, alanyl-methyl ester, leucinyl-methyl ester, isoleucinyl-methyl ester, valinyl-methyl ester, phenylalanyl-methyl ester, Examples include alanyl-ethyl ester, leucinyl-ethyl ester, isoleucinyl-ethyl ester, alanyl-butyl ester, and leucinyl-butyl ester.

As X ₂ , an aspartic acid derivative bonding group or a maleic acid derivative bonding group represented by the following general formula (2) or general formula (3) can be used.

Here, in the formulas (2) and (3), R ₉ and R ₁₀ each independently represent a hydrogen atom or an alkyl group having a carbon number (C1 to C8), and R ₁₁ has a hydrogen atom or a substituent. A linear, branched or cyclic alkyl group having a carbon number (C1 to C20) which may be optionally substituted, a linear or branched chain having a carbon number (C7 to C20) which may have a substituent, or One or more groups selected from the group consisting of a cyclic aralkyl group, an aromatic group having a carbon number (C5 to C20) which may have a substituent, and an amino acid bond residue in which a carboxy group is protected; CX-CY is CH-CH or Z = C (double bond) in the Z configuration.

As coupling groups of the _{X 2,} when using the formula (2) or formula aspartic acid derivative bonded group or maleic acid derivative bonded groups shown by (3), _wherein, R _9, in _{R 10,} carbon atoms (C1 ~ The alkyl group of C8) is a linear, branched or cyclic alkyl group having a carbon number (C1 to C8).
Examples of the linear alkyl group include a methyl group, an ethyl group, an n-propyl group, an n-butyl group, and an n-hexyl group.
Examples of the branched alkyl group include isopropyl group, t-butyl group, 1-methyl-propyl group, 2-methyl-propyl group, 2,2-dimethylpropyl group and the like.
Examples of the cyclic alkyl group include a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, a cyclohexyl group, and the like.

In R _{11 according} to the general formula (2) or the formula (3), the linear, branched or cyclic alkyl group having a carbon number (C1 to C20) which may have a substituent is, for example, Examples include a methyl group, an ethyl group, an isopropyl group, a t-butyl group, a cyclohexyl group, an n-octyl group, a dodecyl group, and an octadecyl group.
Examples of the linear, branched or cyclic aralkyl group having a carbon number (C7 to C20) which may have a substituent include, for example, benzyl group, 2-phenylethyl group, 4-phenylbutyl group, 8 -Phenyloctyl group and the like.
Examples of the aromatic group having a carbon number (C5 to C20) which may have a substituent include a phenyl group, a 4-methoxyphenyl group, a 4-dimethylaminophenyl group, and a 4-hydroxyphenyl group. .

R ₁₁ may be an amino acid binding residue in which a carboxy group is protected. Examples of amino acid-bonded residues in which the carboxy group is protected include glycinyl-methyl ester group, alanyl-methyl ester group, leucinyl-methyl ester group, valinyl-methyl ester group, phenylalanyl-methyl ester group, alanyl-ethyl Examples thereof include an ester group, leucinyl-ethyl ester group, alanyl-butyl ester group, and leucinyl-butyl ester group.

The X ₂ may be a “bond”. “Binding” means that the side chain carboxy group of the polyamino acid derivative and the amino group and / or hydroxyl group, which is the binding substituent of the nucleic acid antimetabolite, are not directly bound via an amide bond and / or via a binding group. Or the aspect which is ester-bonded is pointed out.

In the general formula (1), X ₃ is a methylene group or an ethylene group. That is, if the X ₃ is a methylene group, the polymer backbone of the polyamino acid derivative according to the present invention will become polyaspartic acid. On the other hand, the X ₃ is the case of an ethylene group, the polymer backbone of the polyamino acid derivative according to the present invention will become polyglutamic acid. Therefore, the polyamino acid derivative according to the general formula (1) is a polyaspartic acid derivative or a polyglutamic acid derivative.

In the present invention, it is preferable that the nucleic acid antimetabolite having an amino group and / or a hydroxyl group is directly bonded to the carboxy group of the aspartic acid unit. That is, in the general formula (1), X ₃ is a methylene group, the main case chain polymer is polyaspartic acid, nucleic acid metabolism antagonists in the side chain carboxyl group may be bonded directly, also the X ₂ As the linking group according to, a nucleic acid antimetabolite may be bound via an aspartic acid derivative. In the general formula (1), when X ₃ is an ethylene group and the main chain polymer is polyglutamic acid, a nucleic acid antimetabolite is bound via an aspartic acid derivative as the binding group related to X _2. Is preferred. As bonding groups according to the X _2, when using the aspartic acid derivative, it is preferable to use aspartic acid amide derivative.

R ₅ in the general formula (1) represents an aspartic acid residue or an aspartic imide residue. These residues comprise as a binding group according to the X _2, when using the aspartic acid derivative bonded group or maleic acid derivative bonded group, nucleic acid metabolism antagonist from the linking group is indicative of the dissociated residues. Therefore, the aspartic acid residue or aspartic imide residue according to R ₅ has the same chemical structure as the above-mentioned aspartic acid derivative-binding group or maleic acid derivative-binding group.

Examples of the aspartic acid residue or aspartic imide residue according to R ₅ include one or more selected from the substituent group consisting of the following general formula (4), general formula (5), and general formula (6). Residue.

Here, in the formula, R ₉ , R ₁₀ , R ₁₁ , CX-CY have the same meaning as described above, R ₁₂ represents a hydroxyl group and / or —N (R ₁₃ ) CONH (R ₁₄ ), R ₁₃ and R ₁₄ may be the same or different and represents a linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with a tertiary amino group.

Examples of the linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with a tertiary amino group in R ₁₃ and R ₁₄ include, for example, a methyl group, an ethyl group, n -Propyl group, isopropyl group, isobutyl group, sec-butyl group, t-butyl group, 1-methylbutyl group, 2-methylbutyl group, neopentyl group, cyclohexyl group, etc., preferably isopropyl group, cyclohexyl group Can be mentioned.
Examples of the linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with the tertiary amino group include, for example, 2-dimethylaminoethyl group, 3-dimethylaminopropyl group , 5-dimethylaminopentyl group, 6-dimethylaminohexyl group and the like. Preferred examples of R ₁₃ and R ₁₄ include an ethyl group, an isopropyl group, a cyclohexyl group, and a 3-dimethylaminopropyl group.

When R ₁₂ is a hydroxyl group, it represents a carboxylic acid embodiment. Moreover, the arbitrary salt aspect of the carboxylic acid may be sufficient.
R ₁₂ is a hydroxyl group and / or —N (R ₁₃ ) CONH (R ₁₄ ), but when it is only a hydroxyl group, a hydroxyl group and —N (R ₁₃ ) CONH (R ₁₄ ) coexist, or —N The mode in the case of only (R ₁₃ ) CONH (R ₁₄ ) can be taken. The abundance ratio of the hydroxyl group to —N (R ₁₃ ) CONH (R ₁₄ ) may be arbitrarily set.

R ₆ in the general formula (1) represents a hydroxyl group and / or —N (R ₇ ) CONH (R ₈ ). Here, R ₇ and R ₈ may be the same or different, and may be a linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with a tertiary amino group. is there.
The linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with a tertiary amino group in R ₇ and R ₈ is, for example, a methyl group, an ethyl group, n -Propyl group, isopropyl group, isobutyl group, sec-butyl group, t-butyl group, 1-methylbutyl group, 2-methylbutyl group, neopentyl group, cyclohexyl group, etc., preferably isopropyl group, cyclohexyl group Can be mentioned.
Examples of the linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with the tertiary amino group include, for example, 2-dimethylaminoethyl group, 3-dimethylaminopropyl group , 5-dimethylaminopentyl group, 6-dimethylaminohexyl group and the like. Preferred examples of R ₁₃ and R ₁₄ include an ethyl group, an isopropyl group, a cyclohexyl group, and a 3-dimethylaminopropyl group.

When R ₆ is a hydroxyl group, the side chain carboxy group of the nucleic acid antimetabolite-binding polyamino acid derivative is in the form of a carboxylic acid. Moreover, the arbitrary salt aspect of the carboxylic acid may be sufficient.
R ₆ is a hydroxyl group and / or —N (R ₇ ) CONH (R ₈ ), but when it is only a hydroxyl group, a hydroxyl group and —N (R ₇ ) CONH (R ₈ ) coexist, or —N An embodiment in which only (R ₇ ) CONH (R ₈ ) is present can be employed. The abundance ratio between the hydroxyl group and —N (R ₇ ) CONH (R ₈ ) may be arbitrarily set.

In the general formula (1), a, b, c, d, e, f, g, h, and i indicating the contents of the constituent units of the polyamino acid derivative each independently represent an integer of 0 to 200. The polyamino acid derivative includes an amino acid structural unit to which R ₃ is bonded, an amino acid structural unit to which R ₄ is bonded, an amino acid structural unit to which R ₅ is bonded, an amino acid structural unit to which R ₆ is bonded, and a side chain carboxy group. However, there are intramolecular cyclized amino acid structural units, and each of these amino acid structural units has a structure polymerized in a random sequence in an arbitrary order. That is, the amino acid structural unit in which R ₃ , R ₄ , R ₅ , and R ₆ are bonded to the side chain carboxy group and the amino acid structural unit in which the side chain carboxylic acid has an intramolecular cyclized structure are localized in the form of a sequence. It may be a polymer structure composed of a random sequence having no regularity in each constituent unit, that is, a sequence having no particular regularity in the sequence of the side chain modifications.
Further, (a + b + c + d + e + f + g + h + i), which is the total number of polymerized constituent units of the polyamino acid, is an integer of 3 to 250. Preferably, the total polymerization number is 5 to 200. The average value of the total number of polymerizations is 3 to 250, preferably 5 to 200.

Here, (a + b), which is the total number of structural units to which R ₃ which is a polyethylene glycol segment is bonded, is an integer of 1 to 95. The average value of (a + b) is also 1 to 95. That is, the polyamino acid constituent unit to which R _{3 of the} polyethylene glycol segment is bonded is an essential constituent, and the polyamino acid derivative has at least one unit of polyethylene glycol segment. In the polyamino acid derivative, the polyethylene glycol segment as R ₃ is preferably bonded to 2 units or more, and preferably 80 units or less. That is, (a + b) is preferably an integer of 2 to 80, and the average value of (a + b) is also preferably 2 to 80.

Further, the total number of structural units (c + d) to which R ₄ which is a binding residue of the nucleic acid antimetabolite is bonded is an integer of 1 to 175. The average value of (c + d) is also 1 to 175. That is, the polyamino acid constituent unit to which R ₄ which is a nucleic acid antimetabolite binding residue is bound is an essential constituent, and the polyamino acid derivative has at least one unit of nucleic acid antimetabolite binding residue. In the polyamino acid derivative, the nucleic acid antimetabolite binding residue that is R ₄ is preferably 5 units or more, and preferably 120 units or less. That is, the (c + d) is preferably an integer of 5 to 120, and the average value of the (c + d) is also preferably 5 to 120.

In the general formula (1), the amino acid structural unit to which R ₅ and R ₆ are bonded and the amino acid structural unit in which the side chain carboxy group is cyclized in the molecule are arbitrary. In particular, since the amino acid structural unit R ₅ and R ₆ are bonded, when the X ₁ and X ₂ are "binding", which usually amino acid building units R ₅ and R ₆ is attached is not present, the e , F, and g are each 0.

The total molecular weight of the nucleic acid antimetabolite-binding polyamino acid derivative represented by the general formula (1) is 20 kilodaltons or more and 200 kilodaltons or less. It is preferably 20 kilodaltons or more and 180 kilodaltons or less, more preferably 20 kilodaltons or more and 160 kilodaltons or less.

The method for calculating the total molecular weight is the same as described above, and a calculated value obtained by adding the constituent molecular weights of the constituent parts is adopted as the “molecular weight of the polyamino acid derivative”. That is, (1) the molecular weight of the polyamino acid main chain, (2) the total molecular weight of the polyethylene glycol segment obtained by multiplying the molecular weight of the polyethylene glycol segment by the number of bonds, and (3) the number of bonds in the molecular weight of the binding residue of the nucleic acid antimetabolite. (4) the total molecular weight of the binding group obtained by multiplying the binding group residue molecular weight of any polyethylene glycol segment by the number of bonds, and (5) the total molecular weight of the nucleic acid metabolism antagonist The calculated value obtained by adding the total molecular weight of the linking group obtained by multiplying the linking group residue molecular weight by the number of bonds is defined as the molecular weight.
The molecular weight of the polyamino acid derivative is required to be regulated by the accuracy in kilodalton units. Therefore, the analysis method for each component is not particularly limited as long as it is a sufficiently accurate analysis method for measuring the molecular weight of the polyamino acid derivative in kilodalton units, and various analysis methods are appropriately selected. Good.

Further, the polyethylene glycol segment according to R ₃ in the general formula (1) has an average molecular weight per segment of 200 daltons to 500 kilodaltons. Preferably, it is 500 to 100 kilodaltons, more preferably 1 to 50 kilodaltons.
The molecular weight of the polyethylene glycol segment is a peak top molecular weight measured by a GPC method based on a polyethylene glycol standard product. In the general formula (1), since the number of amino acid structural units (a + b) having R ₃ is 1 to 95, the total molecular weight of the polyethylene glycol segment according to R ₃ is 400 to 180 kilodaltons, preferably Is 1 to 150 kilodaltons, more preferably 2 to 130 kilodaltons.

The nucleic acid antimetabolite-binding polyamino acid derivative of the general formula (1) is characterized in that the mass content of the polyethylene glycol segment in the polyamino acid derivative is 30% by mass or more and 90% by mass or less. The mass content of the polyethylene glycol segment is as defined above, and can be calculated from the content ratio of the total molecular weight of the polyethylene glycol segment to the total molecular weight of the polyamino acid derivative.
The polyethylene glycol segment has a mass molecular weight of 30% by mass to 90% by mass, and preferably 35% by mass to 85% by mass.

The nucleic acid antimetabolite binding residue of R ₄ in the general formula (1) is preferably 2 to 60% by mass in terms of mass content based on the binding molar equivalent of the nucleic acid antimetabolite. More preferably, it is 5 to 50% by mass, and most preferably 5 to 40% by mass.
If the content of the nucleic acid antimetabolite is less than 2% by mass, the total dose of the polyamino acid derivative is increased in order to ensure an effective amount of the nucleic acid antimetabolite, which is not preferable. On the other hand, when the content of the nucleic acid antimetabolite is more than 60% by mass, myelosuppression tends to be strongly developed. It is preferable to set the content of the nucleic acid antimetabolite in order to ensure administration convenience and achieve sufficient drug efficacy and side effect reduction.
The mass content of the nucleic acid antimetabolite in the polyamino acid derivative can be calculated from the content ratio of the total molecular weight of the nucleic acid antimetabolite (3) to the molecular weight of the polyamino acid derivative. A more preferable range of the content of the nucleic acid antimetabolite is 5% by mass or more and 50% by mass or less. It is particularly preferable that the content of the nucleic acid antimetabolite is 5% by mass or more and 40% by mass or less.

The nucleic acid antimetabolite-binding polyamino acid derivative of the general formula (1) preferably has a physical property that does not show associative properties in an aqueous solution of the polyamino acid derivative. Whether or not the polyamino acid derivative is associated in an aqueous solution has the same meaning as described above, and 1 mg / mL of the aqueous solution can be measured with a laser light scattering photometer and defined by the light scattering intensity. That is, the light scattering intensity may be 5 times or less as a relative intensity with respect to the light scattering intensity of toluene, and a polyamino acid derivative that is 3 times or less is more preferable.
The method of measuring the light scattering intensity with a laser light scattering photometer is as defined above.

Next, a method for producing the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention will be described.
The polyamino acid derivative of the present invention is a polyamino acid comprising an aspartic acid derivative constituent unit and / or a glutamic acid derivative constituent unit in which a polyethylene glycol segment and a nucleic acid antimetabolite are bound to a side chain carboxy group of aspartic acid and / or glutamic acid. Is a derivative.
As a method for producing the polyamino acid derivative, the polyamino acid derivative may be produced by sequentially polymerizing each amino acid structural unit. As another method, a polymer main chain of a polyamino acid containing aspartic acid and / or glutamic acid in which a plurality of side chains are free carboxylic acids is constructed, and then a polyethylene glycol segment and a nucleic acid are added to the free carboxylic acid side chains. Examples thereof include a method of chemically binding an antimetabolite. As the production method, the latter method is preferred because the amount of polyethylene glycol segment introduced and the amount of binding of the nucleic acid antimetabolite are easily controlled.

For example, L-aspartic acid-N-carboxylic acid anhydride is subjected to ring-opening polymerization to obtain polyaspartic acid. The polyamino acid derivative according to the present invention can be produced by reacting this with a compound containing a polyethylene glycol segment and a nucleic acid antimetabolite. When an arbitrary linking group is used in the binding of the polyethylene glycol segment and / or the nucleic acid antimetabolite, the polyethylene glycol segment and / or the nucleic acid antimetabolite having the linking group is prepared and reacted with polyaspartic acid. Thus, the polyamino acid derivative according to the present invention using a linking group can be produced. After completion of the reaction, a purification step may be optionally performed, and a polyaspartic acid derivative in which a polyethylene glycol segment and a nucleic acid antimetabolite that can be applied as pharmaceuticals are introduced into a side chain carboxy group can be produced.

The nucleic acid antimetabolite-binding polyamino acid derivative of the present invention has a property of gradually releasing a nucleic acid antimetabolite after administration into a living body, and has a use as a pharmaceutical comprising the nucleic acid antimetabolite as an active ingredient.

The use of the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention as a pharmaceutical is not particularly limited as long as it is a disease having a therapeutic effect by the nucleic acid antimetabolite. For example, it is suitable for pharmaceuticals used for the treatment of malignant tumors, viral diseases and the like. Particularly preferred is a medicament for the treatment of malignant tumors. Examples of malignant tumors include non-small cell lung cancer, pancreatic cancer, gastric cancer, colon cancer, rectal cancer, breast cancer, ovarian cancer, bladder cancer, AIDS-related Kaposi's sarcoma and the like.

The medicament containing the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention may have other additives usually accepted as pharmaceuticals. Such additives include excipients, extenders, fillers, binders, wetting agents, disintegrants, lubricants, surfactants, dispersants, buffers, preservatives, solubilizers, preservatives, flavoring agents. Agents, soothing agents, stabilizers, tonicity agents and the like.

The medicament containing the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention may be prepared as a therapeutic pharmaceutical preparation. The preparation can be administered by any method such as oral, injection, intrarectal administration, intraportal administration, mixing with organ perfusate, and local administration to the affected organ, preferably parenteral administration. More preferably, intravenous administration by injection, intraarterial administration, or local administration to an affected organ.

The dosage of the pharmaceutical containing the nucleic acid antimetabolite-binding polyamino acid derivative of the present invention varies depending on the disease state, administration method, patient state, age, weight, etc., but is usually 1 mg per 1 m ² of body surface area in terms of nucleic acid antimetabolite. 5,000 mg, preferably 10 mg to 2,000 mg, which may be administered once or divided into several times a day. Moreover, although this administration can be performed every day, repeated administration may be performed after several days to several months. As needed, administration methods, dosages, and administration schedules other than those described above can be used.

The present invention will be further described below with reference to examples. However, the present invention is not limited to these examples.
The “molecular weight of the polyamino acid derivative” in Examples and Comparative Examples was calculated by the following calculation formula.
[Polyamino acid derivative molecular weight] = [molecular weight of main chain polyamino acid unit × polymerization number, or molecular weight of polyethylene glycol segment bonded to terminal of main chain polyamino acid + molecular weight of main chain polyamino acid unit × polymerization number] + [(polyethylene glycol) Segment + polyethylene glycol binding group residue) molecular weight x number of bonds] + [nucleic acid antimetabolite residue molecular weight x number of bonds] + [nucleic acid antimetabolite binding group residue molecular weight x number of bonds]
In this example, “polyethylene glycol segment” uses a polyethylene glycol segment compound in which a propyleneamino group as a bonding group is integrated with a polyethylene glycol segment, and these are combined to obtain a polyethylene glycol segment molecular weight.
Therefore, the “molecular weight of the polyamino acid derivative” in this example is the main component, that is, the molecular weight of the main chain polyamino acid segment, the total molecular weight of (polyethylene glycol segment + the binding group residue), and the total molecular weight of the nucleic acid antimetabolite. , As well as the calculated total value of the total molecular weight of the binding group residues of any nucleic acid antimetabolite.

The number of polymerizations of the main chain polyamino acid was calculated from its integral value by ¹ H-NMR analysis of the polyamino acid derivative precursor compound before the introduction of the polyethylene glycol segment and the nucleic acid antimetabolite.

As the molecular weight of the polyethylene glycol segment, the peak top molecular weight in the GPC analysis based on the polyethylene glycol standard substance in the polyethylene glycol segment compound before the introduction reaction was adopted.

The number of bonds of the polyethylene glycol segment was calculated from the consumption rate in the reaction relative to the charged amount of the polyethylene glycol segment compound in the coupling reaction of the polyamino acid derivative precursor compound and the polyethylene glycol segment compound.

The consumption amount of the polyethylene glycol compound was calculated using numerical values obtained by the following analysis.
10 μL of the reaction solution of the polyamino acid derivative precursor compound, the polyethylene glycol segment compound and the nucleic acid antimetabolite before the start of the reaction (before addition of diisopropylcarbodiimide) was diluted with 90 μL of 1% phosphoric acid, and HPLC (use column: Superdex 75 10 / 300 GL, manufactured by GE Healthcare, detector: RI). The peak area corresponding to the polyethylene glycol segment compound at this time is As, and 10 μL of the reaction solution at the end of the reaction is diluted with 90 μL of 1% phosphoric acid, and the peak area corresponding to the polyethylene glycol compound when analyzed by HPLC is At. It was. And the consumption rate of the polyethylene glycol segment was computed with the following formula | equation.
[Consumption rate of polyethylene glycol segment compound] = 1-At / As

The number of binding of the nucleic acid antimetabolite is 10 mg of the polyamino acid derivatives of the obtained Examples and Comparative Examples, precisely dissolved in 1 mL of acetonitrile, mixed with 1 mL of 1 mol / L sodium hydroxide aqueous solution, It hydrolyzed by stirring for 30 minutes. To this hydrolyzed solution, 1 mL of 1 mol / L hydrochloric acid was added, and a water / acetonitrile mixture (1: 1) was added to make exactly 10 mL. This solution was calculated by quantitative analysis of the released nucleic acid antimetabolite using HPLC.

“Polyethylene glycol segment content” in Examples and Comparative Examples was calculated by the following calculation formula.
“Polyethylene glycol segment content” (%) = “polyethylene glycol segment total molecular weight” / “polyamino acid derivative molecular weight” × 100
As the “total molecular weight of the polyethylene glycol segment”, a value calculated by multiplying the molecular weight of the polyethylene glycol segment by the number of bonds of the polyethylene glycol segment was used.
As the “polyamino acid derivative molecular weight”, a numerical value calculated from the sum of the molecular weights of the respective constituent parts was used.

The scattering intensity of the polyamino acid derivatives of Examples and Comparative Examples was measured by a dynamic light scattering photometer DLS-8000DL (measurement temperature 25 ° C., measurement angle: 90 °, wavelength: 632.8 nm, ND filter: 5%, manufactured by Otsuka Electronics Co., Ltd.) , PH1: OPEN, PH2: SLIT).
As a measurement sample for the measurement of scattering intensity, a solution prepared by adding 5% glucose injection so as to have a polyamino acid derivative concentration of 1 mg / mL and irradiating with ultrasound for 3 minutes under ice cooling was used.
Toluene (manufactured by Junsei Co., Ltd., special grade) used for measurement of light scattering intensity was used after being filtered three times with a 0.2 μm membrane filter.
The light scattering intensity of the toluene standard solution measured by the light scattering intensity meter was 12,934 cps.

The number of associated molecules in the measurement sample solution of the polyamino acid derivatives of Examples and Comparative Examples was calculated by the following calculation formula.
[Number of associated molecules] = [SEC-MALS measured molecular weight] / [Polyamino acid derivative molecular weight]
The molecular weight measured by SEC-MALS was measured using DAWN EOS (light scattering detector) and Optilab rEX (RI detector) manufactured by Wyatt Technology, and dn / dc was calculated using the value of polyethylene glycol (0.135). .
Column used: Superdex 200 Increase 10/300 GL, manufactured by GE Healthcare Inc. For the measurement sample, 5% glucose injection solution was added so that the polyamino acid derivative concentration was 1 mg / mL, and ultrasound was irradiated for 3 minutes under ice cooling. The prepared solution was used.

For the polyamino acid derivatives of Examples and Comparative Examples, the peak top molecular weight in GPC analysis based on a polyethylene glycol standard was measured. The peak top molecular weight in the GPC analysis was calculated by the following analysis.
25 mg of polyamino acid derivatives of Examples and Comparative Examples were dissolved in 0.05 mmol / L citrate buffer (pH 3.5) / acetonitrile mixture (9: 1), and HPLC (column used: shodex GF510A-4E, Showa Denko Manufactured, detector: RI). By comparing the retention time of the peak apex at this time with the retention time when the polyethylene glycol standard was analyzed, the peak top in the GPC analysis based on the polyethylene glycol standard of the polyamino acid derivatives of the examples and comparative examples Molecular weight was calculated.

[Synthesis Example 1] Synthesis of polyaspartic acid having an average polymerization number of 89 (Compound 1)
n-Butylamine (manufactured by Tokyo Chemical Industry, 99.2 mg) was dissolved in DMSO (202 mL) and then γ-benzyl-L-aspartic acid-N-carboxylic acid anhydride (BLA-NCA, 33.8 g, 99.9 equivalents). And stirred at 30 ° C. overnight. The reaction solution was dropped into a mixed solvent of ethanol (750 mL) and diisopropyl ether (3200 mL) over 1 hour and stirred at room temperature for 3 hours. The precipitate was collected by filtration and dried in vacuo to obtain a solid (22.6 g).
Dimethylimidazolidinone (DMI) (400 mL) was added to the solid (21.8 g), dissolved at 80 ° C., and 1 mL of acetic anhydride was added at 65 ° C. After stirring for 3 hours, the reaction solution was dropped into a mixed solution of ethyl acetate (1.1 L) and diisopropyl ether (10 L) over 1 hour and stirred at room temperature for 1 hour. The precipitate was collected by filtration and dried in vacuo to obtain a solid (23.2 g).
Acetonitrile (200 mL) and 0.2 N aqueous sodium hydroxide solution (700 mL) were added to the solid (23.0 g), and the mixture was stirred overnight at room temperature. After the reaction, acetonitrile was removed by concentration under reduced pressure, and the concentrated solution was washed three times with ethyl acetate (700 mL). The aqueous layer was concentrated under reduced pressure, passed through a column of Dowex 50W (Dow Chemical Co., proton type, 40 mL), eluted and freeze-dried to obtain Compound 1 (13.0 g).
¹ H-NMR (400 MHz, D ₂ O, NaOD, ppm): 0.7 (n-butylamine-terminated CH ₃ , 3H, integrated value 3.00), 4.2-4.6 (aspartic acid αCH, 1H, Integration value 89.2)
From the molar ratio calculated from the integral value of n-butylamine and aspartic acid, the polymerization number was calculated to be 89. Therefore, the molecular weight of Compound 1 was 10 kilodaltons.

[Synthesis Example 2] Synthesis of polyaspartic acid having an average polymerization number of 22 (Compound 2)
The title compound 2 was obtained by using 22.5 equivalents of BLA-NCA to n-butylamine according to the method described in Synthesis Example 1.
¹ H-NMR (400 MHz, D ₂ O, NaOD, ppm): 0.7 (n-butylamine-terminated CH ₃ , 3H, integrated value 3.00), 4.2-4.6 (aspartic acid αCH, 1H, Integration value 22.4)
From the molar ratio calculated from the integrated value of n-butylamine and aspartic acid, the polymerization number was calculated to be 22. Therefore, the molecular weight of Compound 2 was 2.6 kilodaltons.

[Synthesis Example 3] Synthesis of polyaspartic acid having an average polymerization number of 60 (compound 3)
The title compound 3 was obtained by using 67.4 equivalents of BLA-NCA based on n-butylamine according to the method described in Synthesis Example 1.
¹ H-NMR (400 MHz, D ₂ O, NaOD, ppm): 0.7 (n-butylamine-terminated CH ₃ , 3H, integrated value 3.00), 4.2-4.6 (aspartic acid αCH, 1H, Integration value 60.4)
From the molar ratio calculated from the integrated value of n-butylamine and aspartic acid, the polymerization number was calculated to be 60. Therefore, the molecular weight of Compound 3 was 7.0 kilodalton.

Synthesis Example 4 Synthesis of polyaspartic acid having an average polymerization number of 110 (Compound 4)
According to the method described in Synthesis Example 1, 135 equivalents of BLA-NCA was used with respect to n-butylamine to obtain the title compound 4.
¹ H-NMR (400 MHz, D ₂ O, NaOD, ppm): 0.7 (n-butylamine-terminated CH ₃ , 3H, integrated value 3.00), 4.2-4.6 (aspartic acid αCH, 1H, Integration value 109.8)
From the molar ratio calculated from the integrated value of n-butylamine and aspartic acid, the polymerization number was calculated to be 110. Therefore, the molecular weight of Compound 4 was 13 kilodaltons.

[Synthesis Example 5] Synthesis of aspartic acid-1-alanine methyl ester-4-gemcitabine amide (Compound 5)
After dissolving N- (t-butoxycarbonyl) aspartic acid-4-benzyl ester (15.0 g) and L-alanine methyl ester (6.5 g) in DMF (160 mL), 1-ethyl-3- [3- (Dimethylamino) propyl] carbodiimide (WSCD) hydrochloride (13.3 g), 1-hydroxybenzotriazole (HOBt) (8.53 g), triethylamine (6.5 mL) were added, and the mixture was stirred for 4 hours in an ice bath. . Water was added to the reaction solution, extracted with ethyl acetate, and washed with 5% aqueous citric acid solution, saturated aqueous sodium hydrogen carbonate solution and saturated brine. After drying over magnesium sulfate, ethyl acetate was distilled off under reduced pressure, followed by vacuum drying to obtain an oil (13.9 g).
This oil (13.9 g) was dissolved in methanol (350 mL), 10% palladium carbon (moisture content 50%) (1.39 g) was added, and the system was replaced with hydrogen, followed by 3 hours at room temperature. Stir. 10% Palladium carbon was filtered and washed with methanol (50 mL), and then the methanol was distilled off under reduced pressure, followed by vacuum drying to obtain an oil (10.6 g).
This oil and gemcitabine (3.0 g, manufactured by SCINO PHARM) were dissolved in DMF (56 mL), then HOBt (2.1 g) and WSCD hydrochloride (3.3 g) were added, and the temperature was raised from 0 ° C. to room temperature. And stirred overnight. Purified water was added to the reaction mixture, and the mixture was extracted with ethyl acetate (170 mL). The organic layer was washed twice with saturated brine and dried over sodium sulfate. After removing ethyl acetate by concentration under reduced pressure, purification by silica gel column chromatography was performed, followed by vacuum drying to obtain an oily substance (3.9 g).
This oil was dissolved in ethyl acetate (41 mL), 4N hydrochloric acid-ethyl acetate solution (31 mL) was added, and the mixture was stirred at room temperature for 2 hr. A mixed solvent of ethyl acetate (80 mL) and n-hexane (20 mL) was added to the reaction mixture, and the precipitate was collected by filtration and dried in vacuo to give compound 5 (2.99 g).
¹ H-NMR (400 MHz, DMSO-d6, ppm): 1.20 (d, 3H), 2.95-3.10 (m, 2H), 3.63 (s, 3H), 3.67 (dd , 1H), 3.90-4.02 (m, 5H), 4.18-4.31 (m, 3H), 6.18 (dd, 1H), 7.22 (d, 1H), 8. 25 (s, 2H), 8.92 (d, 1H), 11.3 (br, 1H)

[Synthesis Example 6] Synthesis of aspartic acid-1-leucine methyl ester-4-gemcitabine amide (Compound 6)
N- (t-butoxycarbonyl) -L-aspartate-4-benzyl (4.9 g manufactured by Watanabe Chemical Co., Ltd.) and L-leucine-methyl ester hydrochloride (2.7 g manufactured by Kokusan Chemical Co., Ltd.) were combined with DMF ( 75 mL), HOBt (2.8 g), diisopropylethylamine (2.6 mL), and WSCD hydrochloride (4.3 g) were added, and the mixture was stirred at 0 ° C. for 2 hours. Purified water was added to the reaction mixture, and the mixture was extracted with ethyl acetate (250 mL). The organic layer was washed with saturated aqueous sodium hydrogen carbonate and saturated brine, and dried over sodium sulfate. After removing ethyl acetate by concentration under reduced pressure, purification by silica gel column chromatography was performed, followed by vacuum drying to obtain an oily substance (6.8 g).
This oil was dissolved in methanol (150 mL), 10% (w / w) palladium on carbon was added, and the mixture was stirred under a hydrogen atmosphere for 2 hr. The reaction solution was filtered, and the filtrate was vacuum-dried to obtain an oil (5.37 g).
This oil and gemcitabine (3.0 g, manufactured by SCINO PHARM) were dissolved in DMF (57 mL), HOBt (2.1 g) and WSCD hydrochloride (3.3 g) were added, and the temperature was raised from 0 ° C. to room temperature. And stirred overnight. Purified water was added to the reaction mixture, and the mixture was extracted with ethyl acetate (170 mL). The organic layer was washed twice with saturated brine and dried over sodium sulfate. After removing ethyl acetate by concentration under reduced pressure, purification by silica gel column chromatography was performed, followed by vacuum drying to obtain an oily substance (4.9 g).
This oil was dissolved in ethyl acetate (44 mL), 4N ethyl acetate solution (33 mL) was added, and the mixture was stirred at room temperature for 2 hr. A mixed solvent of ethyl acetate (80 mL) and n-hexane (20 mL) was added to the reaction mixture, and the precipitate was collected by filtration and dried in vacuo to give compound 6 (3.62 g).
¹ H-NMR (400 MHz, DMSO-d6, ppm): 0.89 (dd, 6H), 1.49-1.71 (m, 3H), 2.97-3.17 (m, 4H), 3 .63 (s, 3H), 3.66 (dd, 1H), 3.80-3.93 (m, 2H), 4.10-4.28 (m, 4H), 6.18 (dd, 1H) ), 7.22 (d, 1H), 8.32 (s, 2H), 8.91 (d, 1H), 11.3 (br, 1H)

[Synthesis Example 7] Synthesis of polyethylene glycol-polyaspartic acid block copolymer (polyethylene glycol molecular weight 12 kilodalton, polyaspartic acid polymerization number 67) (compound 7)
After dissolving polyethylene glycol having one end methoxy group and one end 3-aminopropyl group (SUNBRIGHT MEPA-12T, NOF Corporation, average molecular weight 12 kilodalton, 10.0 g) in DMSO (200 mL), γ-benzyl-L -Aspartic acid-N-carboxylic anhydride (BLA-NCA, 22.3 g, 107.7 equivalents) was added and stirred at 32.5 ° C. overnight. The reaction solution was dropped into a mixed solvent of ethanol (400 mL) and diisopropyl ether (1600 mL) over 1 hour and stirred at room temperature for 30 minutes. The precipitate was collected by filtration and dried in vacuo to obtain a solid (24.1 g).
Acetonitrile (460 mL) was added to the solid (23.0 g), dissolved at 35 ° C., and then 0.8 mL of acetic anhydride was added. After stirring for 3 hours, the temperature was lowered to 23 ° C., 0.2N aqueous sodium hydroxide solution (540 mL) was added, and the mixture was stirred for 3 hours. After the reaction, acetonitrile was removed by concentration under reduced pressure, and the concentrated solution was washed three times with ethyl acetate (690 mL). The aqueous layer was concentrated under reduced pressure, the pH of the solution was adjusted to about 10 with 1N aqueous sodium hydroxide solution, and sodium chloride (57.5 g) was added. It was passed through a partition adsorption resin column followed by a column of Dowex 50W (manufactured by Dow Chemical Co., proton type, 40 mL), eluted and freeze-dried to obtain Compound 7 (13.2 g).
In Compound 7, the polymerization number of aspartic acid was calculated to be 66.9 by a titration method using 0.1N potassium hydroxide. Therefore, the molecular weight of Compound 7 was 20 kilodaltons.

Example 1 In the general formula (1), the average value of a + b + c + d + e + f + g + h + i = 89, the average value of a + b = 3.6, the average value of c + d = 41, e + f = 0, g + h + i = 44.4, R ₁ = n-butyl Group, R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₁₅ = methyl group, average value of n = 272, X ₁ = — (CH ₂ ) ₃ —NH— group, X ₂ = Synthesis of Compound with X ₃ = Methylene Group Compound 1 obtained in Synthesis Example 1 (molecular weight: 10 kilodalton, 3.8 g) and polyethylene glycol compound having one end methoxy group and one end 3-aminopropyl group ( SUNBRIGHT MEPA-12T, manufactured by NOF Corporation, average molecular weight 12 kilodalton, 15.7 g) and gemcitabine (6.9 g, manufactured by SCINO PHARM) were combined with DMF (3 And dissolved at 35 ° C. in 5 mL), 1-hydroxybenzotriazole at 25 ℃ (HOBt) (4.9g), was added diisopropylcarbodiimide (DIPCI) (10.3mL), and stirred overnight. The reaction solution was dropped into a mixed solvent of ethanol (1.5 L) and diisopropyl ether (6 L) over 30 minutes and stirred at room temperature for 30 minutes. The precipitate was collected by filtration and washed with ethanol / diisopropyl ether (1/4 (v / v), 1.5 L). The obtained precipitate was dissolved in acetonitrile (720 mL), and purified water (128 mL) and an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 160 mL) were added. After stirring for 30 minutes, the mixture was filtered, and the filtrate was concentrated under reduced pressure and lyophilized to obtain the title compound of Example 1 (22.2 g).

After alkaline hydrolysis of the compound of Example 1, the released gemcitabine was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content in this compound. As a result, the gemcitabine content in Example 1 was 17.1% (w / w). Therefore, the binding rate to the aspartic acid polymerization number 89, which is the polyamino acid main chain, was calculated to be 46.0%.
As a result, the total molecular weight of gemcitabine in Example 1 was 11 kilodaltons.

The binding amount of the polyethylene glycol compound of Example 1 was 3.6 molecules from the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. Therefore, the total polyethylene glycol segment molecular weight was 43 kilodaltons.
In addition, the feed equivalent of the polyethylene glycol segment compound with respect to the main-chain aspartic acid derivative in this reaction = 3.6 equivalent, and the consumption rate of the polyethylene glycol segment compound = 1.

From the main chain polyaspartic acid molecular weight, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Example 1 was calculated to be 64 kilodaltons.
The polyethylene glycol segment content was calculated to be 67% by mass.
Further, the degree of association of the polyamino acid derivative of Example 1 was measured by the laser light scattering intensity. As a result, the light scattering intensity was 23,505 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 1.82. The molecular weight measured by SEC-MALS was 104,300, and the number of associated molecules was 1.6.
Moreover, the peak top molecular weight of the polyamino acid derivative of Example 1 in GPC analysis based on polyethylene glycol standard was 40,955.

[Example 2] In the general formula (1), the average value of a + b + c + d + e + f + g + h + i = 89, the average value of a + b = 8.3, the average value of c + d = 39.6, e + f = 0, g + h + i = 41.1, R ₁ = n-butyl group, R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₁₅ = methyl group, average value of n = 114, X ₁ = — (CH ₂ ) ₃ —NH— group , X ₂ = bond, X ₃ = synthesis of compound of methylene group Compound 1 obtained in Synthesis Example 1 (molecular weight 10 kilodalton, 0.47 g) and polyethylene glycol having one end methoxy group and one end 3-aminopropyl group (SUNBRIGHT MEPA-50H, manufactured by NOF Corporation, average molecular weight 5 kilodalton, 2.04 g) and gemcitabine (0.86 g, manufactured by SCINO PHARM) were combined with DMF (48 And dissolved at 35 ° C. in L), HOBt (0.61g) at 20 ° C., adding DIPCI (1.3 mL), and stirred overnight. The reaction solution was dialyzed against purified outer solution water, then dialyzed against outer solution acetonitrile, and the inner solution was lyophilized to obtain the title compound (2.62 g) according to Example 2.

After alkaline hydrolysis of the compound of Example 2, the released gemcitabine was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content of this compound. As a result, the gemcitabine content in Example 2 was 17.0% (w / w). Therefore, the binding rate to the polyamino acid main chain aspartic acid polymerization number 89 was 44.5%.
As a result, the total molecular weight of gemcitabine in Example 2 was 10 kilodaltons.

The binding amount of the polyethylene glycol compound of Example 2 was 8.3 molecules from the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. Therefore, the total polyethylene glycol segment molecular weight was calculated to be 42 kilodaltons.
In Example 2, the feed equivalent of the polyethylene glycol segment compound to the main chain aspartic acid derivative was 8.9 equivalents, and the consumption rate of the polyethylene glycol segment compound was 0.93.

From the main chain polyaspartic acid molecular weight, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Example 2 was calculated to be 62 kilodaltons.
The polyethylene glycol segment content was calculated to be 67% by mass.
Further, the degree of association of the polyamino acid derivative of Example 2 was measured by the laser light scattering intensity. As a result, the light scattering intensity was 7,430 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 0.57 times. Further, the molecular weight measured by SEC-MALS was 74,150, and the number of associated molecules was 1.2.

Example 3 In the general formula (1), the average value of a + b + c + d + e + f + g + h + i = 60, the average value of a + b = 3.0, the average value of c + d = 23.0, e + f = 0, g + h + i = 34.0, R ₁ = n -Butyl group, R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₁₅ = methyl group, average value of n = 272, X ₁ =-(CH ₂ ) ₃ -NH- group, Synthesis of Compound with X ₂ = Bond, X ₃ = Methylene Group Compound 3 obtained in Synthesis Example 3 (molecular weight 7.0 kilodalton, 0.37 g), polyethylene having one end methoxy group and one end 3-aminopropyl group Glycol (SUNBRIGHT MEPA-12T, NOF Corporation, average molecular weight 12 kilodalton, 1.92 g) and gemcitabine (0.67 g, SCINO PHARM) DMF (3 And dissolved at 35 ° C. in mL), HOBt (0.48g) at 20 ° C., adding DIPCI (1.0 mL), and stirred overnight. The reaction solution was dropped into a mixed solvent of ethyl acetate (105 mL) and diisopropyl ether (0.9 L) over 30 minutes, and stirred at room temperature for 1 hour. The precipitate was collected by filtration and washed with ethyl acetate / diisopropyl ether (1/9 (v / v), 0.5 L). The obtained precipitate was dissolved in purified water (50 mL), and then an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 12 mL) was added. After stirring for 20 minutes, the mixture was filtered and freeze-dried to obtain the title compound (2.38 g) according to Example 3.

After alkaline hydrolysis of the compound of Example 3, the released gemcitabine was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content of this compound. As a result, the gemcitabine content in Example 3 was 12.5% (w / w). Therefore, the binding rate to the polyamino acid main chain aspartic acid polymerization number 60 was 38.3%.
As a result, the total molecular weight of gemcitabine in Example 3 was 6.0 kilodaltons.

The binding amount of the polyethylene glycol compound of Example 3 was 3.0 molecules from the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. Therefore, the total polyethylene glycol segment molecular weight was calculated to be 36 kilodaltons.
In addition, the feed equivalent of the polyethylene glycol compound with respect to the main chain aspartic acid derivative = 3.0 equivalent, and the consumption rate of the polyethylene glycol compound = 1.

From the main chain polyaspartic acid molecular weight, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Example 3 was calculated to be 49 kilodaltons.
The polyethylene glycol segment content was calculated to be 73% by mass.
Further, the degree of association of the polyamino acid derivative of Example 3 was measured by the laser light scattering intensity, whereby the light scattering intensity was 10,270 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 0.79 times. Further, the molecular weight measured by SEC-MALS was 102,500, and the number of associated molecules was 2.1.
Moreover, the peak top molecular weight in GPC analysis of the polyamino acid derivative of Example 3 on the basis of the polyethylene glycol standard substance was 42,642.

Example 4 In the general formula (1), the average value of a + b + c + d + e + f + g + h + i = 110, the average value of a + b = 5.5, the average value of c + d = 44, e + f = 0, g + h + i = 60.5, R ₁ = n-butyl Group, R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₁₅ = methyl group, average value of n = 272, X ₁ = — (CH ₂ ) ₃ —NH— group, X ₂ = Synthesis of Compound with X ₃ = Methylene Group Compound 4 obtained in Synthesis Example 4 (molecular weight: 13 kilodalton, 0.50 g) and polyethylene glycol having one terminal methoxy group and one terminal 3-aminopropyl group (SUNBRIGHT MEPA) -12T, NOF Corporation, average molecular weight 12 kilodalton, 2.6 g) and gemcitabine (0.92 g, SCINO PHARM) DMF (50 mL) And dissolved at 35 ° C. in, HOBt (0.65 g) at 20 ° C., adding DIPCI (2.0 mL), and stirred overnight. The reaction solution was dropped into a mixed solvent of ethanol (0.2 L) and diisopropyl ether (0.8 L) over 30 minutes and stirred at room temperature for 1 hour. The precipitate was collected by filtration and washed with ethanol / diisopropyl ether (1/4 (v / v), 0.2 L). The obtained precipitate was dissolved in acetonitrile (90 mL), and purified water (15 mL) and an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 30 mL) were added. After stirring for 1 hour, the mixture was filtered, and the filtrate was concentrated under reduced pressure and lyophilized to obtain the title compound (3.01 g) according to Example 4.

After alkaline hydrolysis of the compound of Example 4, the released gemcitabine was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content of this compound. As a result, the gemcitabine content in Example 4 was 12.9% (w / w). Therefore, the binding rate to the aspartic acid polymerization number 110 which is the polyamino acid main chain was 44.0%.
As a result, the total molecular weight of gemcitabine in Example 4 was 13 kilodaltons.

The binding amount of the polyethylene glycol compound was 5.5 molecules from the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. Therefore, the total polyethylene glycol segment molecular weight was calculated to be 66 kilodaltons.
In addition, the feed equivalent of the polyethylene glycol compound with respect to the main chain aspartic acid derivative = 5.5 equivalent, and the consumption rate of the polyethylene glycol compound = 1.

From the main chain polyaspartic acid molecular weight, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Example 4 was calculated to be 91 kilodaltons.
The polyethylene glycol segment content was calculated to be 72% by mass.
Further, when the degree of association of the polyamino acid derivative of Example 4 was measured by the laser light scattering intensity, the light scattering intensity was 11,215 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 0.87 times. The molecular weight measured by SEC-MALS was 112,200, and the number of associated molecules was 1.2.
Moreover, the peak top molecular weight of the polyamino acid derivative of Example 4 in the GPC analysis based on the polyethylene glycol standard substance was 58,443.

Example 5 In the general formula (1), the average value of a + b + c + d + e + f + g + h + i = 89, the average value of a + b = 3.6, the average value of c + d = 45, e + f = 0, g + h + i = 40.4, R ₁ = n-butyl Group, R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₁₅ = methyl group, average value of n = 113, X ₁ = — (CH ₂ ) ₃ —NH— group, X ₂ = Synthesis of Compound with X ₃ = Methylene Group Compound 1 obtained in Synthesis Example 1 (molecular weight 10 kilodalton, 0.5 g) and polyethylene glycol having one terminal methoxy group and one terminal 3-aminopropyl group (SUNBRIGHT MEPA) -50H, NOF Corporation, average molecular weight 5 kilodalton, 0.870 g) and gemcitabine (0.915 g, SCINO PHARM) DMF (50 mL) And dissolved at 35 ° C. in, HOBt (0.646 g) at 20 ° C., adding DIPCI (2.04 mL), and stirred overnight. The reaction solution was dropped into a mixed solvent of ethanol (0.2 L) and diisopropyl ether (0.8 L) over 15 minutes, and stirred at room temperature for 30 minutes. The precipitate was collected by filtration and washed with ethanol / diisopropyl ether (1/4 (v / v), 0.2 L). The obtained precipitate was dissolved in acetonitrile (90 mL), and purified water (16 mL) and an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 20 mL) were added. After stirring for 1 hour, the mixture was filtered, and the filtrate was concentrated under reduced pressure and lyophilized to obtain the compound according to Example 5 (1.38 g).

After alkaline hydrolysis of the compound of Example 5, the released gemcitabine was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content of this compound. As a result, the gemcitabine content in Example 5 was 27.9% (w / w). Therefore, the binding rate with respect to 89 aspartic acid polymerization number which is a polyamino acid main chain was 50.6%.
As a result, the total molecular weight of gemcitabine in Example 5 was 12 kilodaltons.

The binding amount of the polyethylene glycol compound was 3.6 molecules from the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. Therefore, the total polyethylene glycol segment molecular weight was calculated to be 18 kilodaltons.
The feed equivalent of the polyethylene glycol compound to the main chain aspartic acid derivative was 3.6 equivalents, and the consumption rate of the polyethylene glycol compound was 1.

From the main chain polyaspartate molecular weight, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Example 5 was calculated to be 42 kilodaltons.
The polyethylene glycol segment content was calculated to be 43% by mass.
Further, the degree of association of the polyamino acid derivative of Example 5 was measured by the laser light scattering intensity. As a result, the light scattering intensity was 35,280 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 2.73 times. Further, the molecular weight measured by SEC-MALS was 347,200, and the number of associated molecules was 8.4.

[Example 6] In general formula (1), the average value of a + b + c + d + e + f + g + h + i = 89, the average value of a + b = 24.0, the average value of c + d = 27.5, e + f = 0, g + h + i = 37.5, R ₁ = n -Butyl group, R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₉ and R ₁₀ = hydrogen atom, R ₁₁ = alanine methyl ester group, R ₁₂ = hydroxyl group, R ₁₅ = methyl group , Average value of n = 45, synthesis of compound of X ₁ = — (CH ₂ ) ₃ —NH— group, X ₃ = methylene group Compound 1 obtained in Synthesis Example 1 (molecular weight 10 kilodalton, 0.53 g) And polyethylene glycol having one end methoxy group and one end 3-aminopropyl group (SUNBRIGHT MEPA-20H, manufactured by NOF Corporation, average molecular weight 2 kilodaltons, 2.47 g) It melt | dissolved in MF (50 mL) at 35 degreeC, HOBt (0.68g) and DIPCI (1.4mL) were added at 20 degreeC, and it stirred for 3.5 hours. After 3.5 hours, Compound 5 (1.1 g) obtained in Synthesis Example 5 and diisopropylethylamine (0.4 mL) were added and stirred overnight. The reaction solution was added dropwise to diisopropyl ether (1.5 L) over 30 minutes and stirred at room temperature for 1 hour. The precipitate was collected by filtration and washed with diisopropyl ether (0.5 L). The resulting precipitate was dissolved in purified water (50 mL), and then an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 20 mL) was added. After stirring for 20 minutes, the mixture was filtered and freeze-dried to obtain the title compound (3.55 g) according to Example 6.

After alkaline hydrolysis of the compound of Example 6, the gemcitabine released was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content of this compound. As a result, the gemcitabine content in Example 6 was 10.1% (w / w). Therefore, the binding ratio to the polyamino acid main chain aspartic acid polymerization number 89 was 30.1%.
As a result, the total molecular weight of gemcitabine in Example 6 was 7.1 kilodalton.

The binding amount of the polyethylene glycol compound was 24 molecules based on the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. Therefore, the total polyethylene glycol segment molecular weight was calculated to be 48 kilodaltons.
The feed equivalent of the polyethylene glycol compound to the main chain aspartic acid derivative was 24 equivalents, and the consumption rate of the polyethylene glycol compound was 1.

From the main chain polyaspartic acid molecular weight, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Example 6 was calculated to be 70 kilodaltons.
The polyethylene glycol segment content was calculated to be 68% by mass.
Further, when the association degree of the polyamino acid derivative of Example 5 was measured by the laser light scattering intensity, the light scattering intensity was 6,025 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 0.47 times. The molecular weight measured by SEC-MALS was 64,790, and the number of associated molecules was 0.9.

[Example 7] In general formula (1), the average value of a + b + c + d + e + f + g + h + i = 89, the average value of a + b = 24.0, the average value of c + d = 18.7, e + f = 0, g + h + i = 46.3, R ₁ = n -Butyl group, R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₉ and R ₁₀ = hydrogen atom, R ₁₁ = leucine methyl ester group, R ₁₂ = hydroxyl group, R ₁₅ = methyl group , Average value of n = 45, synthesis of compound of X ₁ = — (CH ₂ ) ₃ —NH— group, X ₃ = methylene group Compound 1 obtained in Synthesis Example 1 (molecular weight 10 kilodalton, 0.56 g) And polyethylene glycol having one end methoxy group and one end 3-aminopropyl group (SUNBRIGHT MEPA-20H, manufactured by NOF Corporation, average molecular weight 2 kilodaltons, 2.64 g) It melt | dissolved in MF (50 mL) at 35 degreeC, HOBt (0.73 g) and DIPCI (1.5 mL) were added at 20 degreeC, and it stirred for 4 hours. After 4 hours, Compound 6 (1.3 g) obtained in Synthesis Example 6 and diisopropylethylamine (0.4 mL) were added to the reaction solution, and the mixture was stirred overnight. The reaction solution was dropped into a mixed solvent of ethyl acetate (150 mL) and diisopropyl ether (1.4 L) over 30 minutes, and stirred at room temperature for 1 hour. The precipitate was collected by filtration and washed with ethyl acetate / diisopropyl ether (1/9 (v / v), 0.5 L). The resulting precipitate was dissolved in purified water (50 mL), and then an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 20 mL) was added. After stirring for 20 minutes, the mixture was filtered and freeze-dried to obtain the title compound according to Example 7 (3.73 g).

After alkaline hydrolysis of the compound of Example 7, the released gemcitabine was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content of this compound. As a result, the gemcitabine content in Example 7 was 7.3% (w / w). Therefore, the binding rate with respect to 89 aspartic acid polymerization number which is a polyamino acid main chain was 21.0%.
As a result, the total molecular weight of gemcitabine in Example 7 was 4.9 kilodaltons.

From the main chain polyaspartic acid molecular weight, the combined gemcitabine total molecular weight, the combined polyethylene glycol segment molecular weight and the total molecular weight of the aspartic acid unit residue, the molecular weight of the polyamino acid derivative of Example 7 was calculated to be 69 kilodaltons.
The polyethylene glycol segment content was calculated to be 70% by mass.
Further, the degree of association of the polyamino acid derivative of Example 7 was measured by the laser light scattering intensity. As a result, the light scattering intensity was 5,740 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 0.44 times. The molecular weight measured by SEC-MALS was 64,630 and the number of associated molecules was 0.9.

[Example 8] In the general formula (1), the average value of a + b + c + d + e + f + g + h + i = 89, the average value of a + b = 34.0, the average value of c + d = 10.9, e + f = 0, g + h + i = 44.4, R ₁ = n -Butyl group, R ₂ = acetyl group, R ₄ = CP-4126 binding residue, R ₆ = hydroxyl group, R ₁₅ = methyl group, average value of n = 45, X ₁ =-(CH ₂ ) ₃ -NH- Synthesis of a compound having a group, X ₂ = bond, X ₃ = methylene group Compound 1 obtained in Synthesis Example 1 (molecular weight: 10 kilodalton, 0.58 g) and polyethylene having one end methoxy group and one end 3-aminopropyl group Glycol (SUNBRIGHT MEPA-20H, manufactured by NOF Corporation, average molecular weight 2 kilodaltons, 1.97 g) and Gemcitabine 5′-elaidate (CP-4126, 0.45 g (manufactured by MedKoo Biosciences) was dissolved in DMF (53 mL) at 35 ° C., HOBt (0.845 g) and DIPCI (0.75 mL) were added at 25 ° C., and the mixture was stirred overnight. The reaction solution was dropped into a mixed solvent of diisopropyl ether (1060 mL) over 30 minutes and stirred at room temperature for 30 minutes. The precipitate was collected by filtration and washed with diisopropyl ether (50 mL). The obtained precipitate was dissolved in purified water (53 mL), and an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 25 mL) was added. After stirring for 30 minutes, the mixture was filtered, and the filtrate was concentrated under reduced pressure and lyophilized to obtain the title compound (4.79 g) according to Example 8.

After alkaline hydrolysis of the compound of Example 8, the released gemcitabine was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content of this compound. As a result, the gemcitabine content in Example 8 was 3.5% (w / w). Therefore, the binding rate to the polyamino acid main chain aspartic acid polymerization number 89 was 12.3%.
As a result, the total molecular weight of gemcitabine in Example 8 was 2.9 kilodaltons.

The binding amount of the polyethylene glycol compound was 34.0 molecules from the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. As a result, the total polyethylene glycol segment molecular weight was calculated to be 68 kilodaltons.
The charged equivalent of the polyethylene glycol compound to the main chain aspartic acid derivative was 37 equivalents, and the consumption rate of the polyethylene glycol compound was 0.92.

From the main chain polyaspartic acid molecular weight, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Example 8 was calculated to be 83 kilodaltons.
The polyethylene glycol segment content was calculated to be 82% by mass.
Further, the association degree of the polyamino acid derivative of Example 8 was measured by the laser light scattering intensity, whereby the light scattering intensity was 20,545 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 1.59 times. Further, the molecular weight measured by SEC-MALS was 252,800, and the number of associated molecules was 3.1.

[Example 9] In general formula (1), average value of a + b + c + d + e + f + g + h + i = 67, average value of a + b = 3.5, average value of c + d = 36.5, e + f = 0, g + h + i = 27, R ₁ = polyethylene glycol segment R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₁₅ = methyl group, average value of n = 272, X ₁ = — (CH ₂ ) ₃ —NH— group, X ₂ = Bond, Synthesis of Compound with X ₃ = Methylene Group Compound 7 obtained in Synthesis Example 7 (molecular weight 20 kilodalton, 11.0 g) and polyethylene glycol compound (SUNBRIGHT with one end methoxy group and one end 3-aminopropyl group) MEPA-12T, manufactured by NOF Corporation, average molecular weight 12 kilodalton, 23.4 g) and gemcitabine hydrochloride (8.9 g, SCINO P ARM) was dissolved in DMF (317 mL) at 35 ° C., and 1-hydroxybenzotriazole (HOBt) (6.3 g) and diisopropylcarbodiimide (DIPCI) (11.5 mL) were added at 20 ° C. Stir overnight. The reaction solution was dropped into a mixed solvent of ethanol (1.3 L) and diisopropyl ether (5.1 L) over 40 minutes and stirred at room temperature for 30 minutes. The precipitate was collected by filtration and washed with ethyl acetate (486 mL). The resulting precipitate was dissolved in purified water (395 mL), and purified water (9 mL) and an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 113 mL) were added. After stirring for 40 minutes, the mixture was filtered and lyophilized to obtain the title compound of Example 9 (37.8 g).

After alkaline hydrolysis of the compound of Example 9, the gemcitabine released was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content in this compound. As a result, the gemcitabine content in Example 9 was 13.6% (w / w). Therefore, the binding rate of compound 7 to the polyaspartic acid polymerization number 67 was calculated to be 54.4%.
As a result, the total molecular weight of gemcitabine in Example 9 was 9.6 kilodaltons.

The binding amount of the polyethylene glycol compound of Example 9 was 3.5 molecules from the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. Therefore, the total polyethylene glycol segment molecular weight was 42 kilodaltons.
In addition, the preparation equivalent of the polyethylene glycol segment compound with respect to the compound 7 in this reaction = 3.5 equivalent, and the consumption rate of the polyethylene glycol segment compound = 1.

From the molecular weight of Compound 9, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Example 9 was calculated to be 71 kilodaltons.
The polyethylene glycol segment content was calculated as 77% by mass.
Further, the degree of association of the polyamino acid derivative of Example 9 was measured by the laser light scattering intensity, whereby the light scattering intensity was 17,715 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 1.37 times. Further, the molecular weight measured by SEC-MALS was 83,470, and the number of associated molecules was 1.2.
Moreover, the peak top molecular weight in GPC analysis of the polyamino acid derivative of Example 9 based on the polyethylene glycol standard substance was 52,341.

[Comparative Example 1] In the general formula (1), the average value of a + b + c + d + e + f + g + h + i = 22, the average value of a + b = 4.4, the average value of c + d = 9.2, e + f = 0, g + h + i = 8.4, R ₁ = n -Butyl group, R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₁₅ = methyl group, average value of n = 45, X ₁ =-(CH ₂ ) ₃ -NH- group, Synthesis of compound of X ₂ = bond, X ₃ = methylene group Compound 2 obtained in Synthesis Example 2 (molecular weight 2.6 kilodalton, 0.56 g) and polyethylene having one end methoxy group and one end 3-aminopropyl group Glycol (SUNBRIGHT MEPA-20H, manufactured by NOF Corporation, average molecular weight 2 kilodalton, 1.94 g) and gemcitabine (1.0 g, manufactured by SCINO PHARM) in DMF (50 mL) Was dissolved at 5 ℃, HOBt (0.72g) at 20 ° C., adding DIPCI (1.5 mL), and stirred overnight. The reaction solution was dropped into a mixed solvent of ethyl acetate (150 mL) and diisopropyl ether (1.4 L) over 30 minutes, and stirred at room temperature for 1 hour. The precipitate was collected by filtration and washed with ethyl acetate / diisopropyl ether (1/9 (v / v), 0.5 L). The obtained precipitate was dissolved in purified water (50 mL), and then an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 12 mL) was added. After stirring for 20 minutes, the mixture was filtered and freeze-dried to obtain the title compound (2.65 g) according to Comparative Example 1.

After alkaline hydrolysis of the compound of Comparative Example 1, the released gemcitabine was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content of this compound. As a result, the gemcitabine content in Comparative Example 1 was 17.6% (w / w). Therefore, the binding rate with respect to the aspartic acid polymerization number 22 as the polyamino acid main chain was 38.2%.
As a result, the total molecular weight of gemcitabine in Comparative Example 1 was 2.2 kilodalton.

The binding amount of the polyethylene glycol compound was 4.4 molecules from the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. As a result, the total polyethylene glycol segment molecular weight was calculated to be 8.8 kilodaltons.
In addition, the feed equivalent of the polyethylene glycol compound to the main chain aspartic acid derivative = 4.4 equivalent, and the consumption rate of the polyethylene glycol compound = 1.

From the main chain polyaspartic acid molecular weight, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Comparative Example 1 was calculated to be 14 kilodaltons.
The polyethylene glycol segment content was calculated to be 64% by mass.
Further, the degree of association of the polyamino acid derivative of Comparative Example 1 was measured by the laser light scattering intensity. The light scattering intensity was 3,030 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 0.23 times. The SEC-MALS measurement molecular weight was 17,810, and the number of associated molecules was 1.3.
Moreover, the peak top molecular weight of the polyamino acid derivative of Comparative Example 1 in the GPC analysis based on the polyethylene glycol standard substance was 8,162.

Comparative Example 2 In general formula (1), the average value of a + b + c + d + e + f + g + h + i = 89, the average value of a + b = 3.6, the average value of c + d = 49, e + f = 0, g + h + i = 36.4, R ₁ = n-butyl Group, R ₂ = acetyl group, R ₄ = gemcitabine binding residue, R ₆ = hydroxyl group, R ₁₅ = methyl group, average value of n = 45, X ₁ = — (CH ₂ ) ₃ —NH— group, X ₂ = Synthesis of compound of X ₃ = methylene group Compound 1 obtained in Synthesis Example 1 (molecular weight: 10 kilodalton, 0.50 g) and polyethylene glycol having one terminal methoxy group and one terminal 3-aminopropyl group (SUNBRIGHT MEPA) -20H, NOF Corporation, average molecular weight 2 kilodaltons, 0.35 g) and gemcitabine (0.915 g, SCINO PHARM) DMF (50 mL) Was dissolved at 35 ℃, HOBt (0.65g) at 20 ° C., adding DIPCI (2.0 mL), and stirred overnight. The reaction solution was dropped into a mixed solvent of ethanol (0.2 L) and diisopropyl ether (0.8 L) over 15 minutes, and stirred at room temperature for 30 minutes. The precipitate was collected by filtration and washed with ethanol / diisopropyl ether (1/4 (v / v), 0.2 L). The obtained precipitate was dissolved in acetonitrile (90 mL), and purified water (16 mL) and an ion exchange resin (Dow Chemical Dowex 50 (H ⁺ ), 20 mL) were added. After stirring for 30 minutes, the mixture was filtered, and the filtrate was concentrated under reduced pressure and lyophilized to obtain the title compound (0.89 g) according to Comparative Example 2.

After alkaline hydrolysis of the compound of Comparative Example 2, the released gemcitabine was quantified by high performance liquid chromatography (HPLC) to determine the gemcitabine content of this compound. As a result, the gemcitabine content in Comparative Example 1 was 41.5% (w / w). Accordingly, the binding rate to the polyamino acid main chain aspartic acid polymerization number 89 was 55.1%.
As a result, the total molecular weight of gemcitabine in Comparative Example 2 was 13 kilodaltons.

The binding amount of the polyethylene glycol compound was 3.6 molecules from the charged equivalent of the polyethylene glycol compound and the consumption rate of the polyethylene glycol compound. Therefore, the total polyethylene glycol segment molecular weight was calculated to be 7.2 kilodaltons.
The feed equivalent of the polyethylene glycol compound to the main chain aspartic acid derivative was 3.6 equivalents, and the consumption rate of the polyethylene glycol compound was 1.

From the main chain polyaspartic acid molecular weight, the combined gemcitabine total molecular weight, and the combined polyethylene glycol segment molecular weight, the molecular weight of the polyamino acid derivative of Comparative Example 2 was calculated to be 30 kilodaltons.
The polyethylene glycol segment content was calculated to be 24% by mass.
Further, the degree of association of the polyamino acid derivative of Comparative Example 2 was measured by laser light scattering intensity. As a result, the light scattering intensity was 205,765 cps. Therefore, the relative ratio with the light scattering intensity of toluene was 15.91 times. Further, the molecular weight measured by SEC-MALS was 1,520,000, and the number of associated molecules was 50.1.

[Test Example 1] Hematological toxicity evaluation for non-cancer-bearing mice Using the compounds of Examples 1 to 6 and Comparative Examples 1 and 2 and gemcitabine hydrochloride as a control drug, non-tumor-bearing mice (ICR mice (Crlj: CD- A blood toxicity evaluation test for 1)) was conducted.
The compounds of Examples 1 to 6 and Comparative Examples 1 and 2 were dissolved in a 5% glucose injection solution, and gemcitabine hydrochloride as a control drug was used as a physiological saline solution, and the compounds of Examples 1 to 6 and Comparative Examples 1 and 2 and Gemcitabine hydrochloride was 40 mg / kg in terms of gemcitabine, and Comparative Example 2 was administered once intravenously at a dose of 20 mg / kg in terms of gemcitabine. In addition, a group to which a solvent (5% glucose injection solution or physiological saline, 10 mL / kg) was administered was set, and 5% glucose injection solution was administered to the compounds of Examples 1 to 6 and Comparative Examples 1 and 2. For the group, and for gemcitabine hydrochloride, the physiological saline administration group was the control group.
Seven days after administration, blood was collected, and the reticulocyte count was measured with a blood cell analyzer (XT-2000iV). On the 7th day after administration, the relative value of the reticulocyte count of each compound administration group relative to the solvent control group was calculated. The results are shown in Table 1.

[Table 1]

As a result, although the dose of the compound of Comparative Example 2 was low, the reticulocyte count was remarkably reduced as compared with gemcitabine hydrochloride as a control drug, and hematologic toxicity was observed. This phenomenon indicates that the recovery of the reticulocyte count is delayed even 7 days after administration, and is considered to be a prolongation of blood toxicity. In contrast, in the compounds according to Examples 1 to 6 of the present invention, no decrease in the number of reticulocytes was confirmed, indicating that no hematological toxicity was observed at 7 days after administration. Gemcitabine hydrochloride is a major side effect of hematological toxicity in clinical use. Therefore, it is considered that the compounds according to Examples 1 to 6 did not prolong hematologic toxicity and showed rapid recovery.

[Test Example 2] Antitumor effect test on nude mice transplanted with human pancreatic cancer Antitumor effect tests of the compounds of Examples 1 to 6 and 9 and Comparative Examples 1 and 2 were performed.
A tumor mass of human pancreatic cancer AsPC-1 subcultured subcutaneously in nude mice was made into blocks of about 3 mm square and transplanted subcutaneously on the dorsal side of nude mice using a trocar. When the average tumor volume after tumor transplantation reached 300 mm ³ or more, the compounds of Examples 1 to 6 and 9 were dissolved in 5% glucose injection and administered at a dose of 40 mg / kg in terms of gemcitabine. Further, Comparative Examples 1 and 2 were dissolved in a 5% glucose injection solution, and Comparative Example 1 was administered at 40 mg / kg in terms of gemcitabine, and Comparative Example 2 was administered at 20 mg / kg in terms of gemcitabine. As a control drug, gemcitabine hydrochloride was dissolved in physiological saline and administered at 40 mg / kg. Each compound and control drug was administered into the tail vein 4 times at 3 day intervals.
The relative tumor volume was determined from the tumor volume on the administration start date and the evaluation date (16th day or 14th day after the start of administration) and used as an index of the antitumor effect. The tumor volume was calculated by the formula (L × W ² ) / 2 by measuring the major axis (L: mm) and minor axis (W: mm) of the tumor. The test was divided into four parts. The results are shown in Tables 2, 3, 4 and 5.

[Table 2]

[Table 3]

[Table 4]

[Table 5]

As a result, the compound of Comparative Example 1 was not significantly different in antitumor effect compared to the control drug. On the other hand, the compound of the present invention showed a strong antitumor effect and sustained its effect as compared with the control drug. This is probably because the compound of Comparative Example 1 was poor in blood retention compared to the compound of the present invention, and could not maintain the drug concentration required to exhibit a stronger antitumor effect than the control drug. Low molecular weight compounds are known to be excreted by the kidney and the like. The compound of Comparative Example 1 has a molecular weight of 14 kilodaltons and is lower in molecular weight than the compound of the present invention. The compounds according to Examples 1 to 6 and 9 of the present invention are more potent than the control drug because the retention in the blood is increased by controlling the molecular weight within an appropriate range, and the drug concentration sufficient to show the drug effect is maintained. It is considered that it showed an antitumor effect.

Claims

A polyamino acid derivative containing a plurality of units of an aspartic acid derivative and / or a glutamic acid derivative, wherein a polyethylene glycol segment and a nucleic acid antimetabolite are bound to the side chain carboxy group directly or via a binding group; The nucleic acid antimetabolite-binding polyamino acid, wherein the polyamino acid derivative has a molecular weight of 20 kilodaltons or more and 200 kilodaltons or less, and a mass content of polyethylene glycol segment in the polyamino acid derivative is 30 mass% or more and 90 mass% or less Derivative.
2. The nucleic acid antimetabolite-binding polyamino acid derivative according to claim 1, wherein the polyethylene glycol segment is bound by 2 to 80 units.
The nucleic acid antimetabolite-binding polyamino acid according to claim 1 or 2, wherein the nucleic acid antimetabolite is a nucleic acid antimetabolite having an amino group, and the nucleic acid antimetabolite is bound via an amide bond at the amino group. Derivative.
The nucleic acid antimetabolite-binding polyamino acid derivative according to any one of claims 1 to 3, wherein the nucleic acid antimetabolite has a mass content of 2% by mass or more and 60% by mass or less.
The nucleic acid antimetabolite-binding polyamino acid derivative has the general formula (1)

[Wherein, R 1 represents a hydrogen atom, a group selected from the group consisting of alkyl groups and polyethylene glycol segments of carbon atoms (C1 ~ C8), acyl R 2 is a hydrogen atom, the number of carbon atoms (C1 ~ C8) A group selected from the group consisting of a group and a C1-C8 alkoxycarbonyl group, R 3 represents a polyethylene glycol segment, R 4 represents a nucleic acid antimetabolite binding residue, and R 5 represents an asparagine An acid bond residue and / or an aspartate imide bond residue, R 6 represents a hydroxyl group and / or —N (R 7 ) CONH (R 8 ), and R 7 and R 8 may be the same or different. A linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with a tertiary amino group, wherein X 1 and X 2 are linking groups; 3 is methylene A, b, c, d, e, f, g, h and i each independently represents an integer of 0 to 200, and the total polymerization number of the polyamino acid derivative (a + b + c + d + e + f + g + h + i) is 3 to 250, (a + b) is 1 to 95, and (c + d) is 1 to 175. The amino acid unit to which R 3 is bonded, the amino acid unit to which R 4 is bonded, and the R 5 are bonded to each other The amino acid unit, the amino acid unit to which R 6 is bonded, and the amino acid unit in which the side-chain carboxy group is an intramolecular cyclization type are each independently a random sequence]. A nucleic acid antimetabolite-binding polyamino acid derivative according to 1.
X 2 represents the following general formula (2) or general formula (3)

[Wherein R 9 and R 10 each independently represents a hydrogen atom or an alkyl group having from 1 to 8 carbon atoms (C1 to C8), and R 11 represents a hydrogen atom or an optionally substituted carbon number (C1 to C8). A linear, branched or cyclic alkyl group of C20), an optionally substituted linear (C1-C20) linear, branched or cyclic aralkyl group, and a substituent. One or more groups selected from the group consisting of an amino acid-bonded residue in which an aromatic group and a carboxy group may be protected, and CX—CY represents C═C (two The nucleic acid antimetabolite-binding polyamino acid derivative according to claim 5, wherein
R 5 represents the following general formula (4), general formula (5), and general formula (6).

[Wherein R 9 , R 10 , R 11 and CX-CY have the same meaning as described above, R 12 represents a hydroxyl group and / or —N (R 13 ) CONH (R 14 ), and R 13 , R 14 May be the same or different and represents a linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may be substituted with a tertiary amino group]. The nucleic acid antimetabolite-binding polyamino acid derivative according to claim 5 or 6, wherein the polyamino acid derivative is one or more kinds of groups.
The polyethylene glycol segment of R 3 is represented by the following general formula (7)

[In the formula, R 15 represents a hydrogen atom or a linear, branched or cyclic alkyl group having a carbon number (C1 to C8) which may have a substituent, and n is 5 to 2,500. The nucleic acid antimetabolite-binding polyamino acid derivative according to any one of claims 5 to 7, which represents an integer.
The nucleic acid metabolism antagonist is represented by the formula (8):

[Wherein, -Rf represents the formula (9):

Any one or more nucleic acid metabolism antagonists represented by: R 16 represents a hydrogen atom or an acyl group of a fatty acid ester]. The nucleic acid antimetabolite-binding polyamino acid derivative according to claim 1.
The nucleic acid metabolism antagonist is represented by the formula (10):

[Wherein, -Rf represents the formula (11):

A nucleic acid antimetabolite-binding polyamino acid derivative according to any one of claims 1 to 9, wherein R 16 represents a hydrogen atom or an acyl group of a fatty acid ester]. .
A medicament comprising the nucleic acid antimetabolite-binding polyamino acid derivative according to any one of claims 1 to 10.