Hyaluronic acid derivative composition

Abstract

To provide a hyaluronic acid derivative composition which can be used as a hair growing agent or a hair restoring agent and has an excellent hair growth or hair restoration effect.SOLUTION: There is provided a hyaluronic acid derivative composition which comprises (A) a hyaluronic acid derivative into which a steryl group is introduced and (B) a component for promoting hair growth and hair restoration. The content of the hyaluronic acid derivative into which a steryl group is introduced (A) may be 0.01 pt. mass or more and 10 pts.mass or less based on 100 pts.mass of the hyaluronic acid derivative composition and the content of the component for hair growth and hair restoration (B) may be 0.01 pt.mass or more and 40 pts.mass or less based on the 100 pts.mass of the hyaluronic acid derivative composition.SELECTED DRAWING: None

Description

[Technical Field]

[0001] This invention relates to hyaluronic acid derivative compositions. [Background technology]

[0002] Androgenetic alopecia (male pattern hair loss) is a type of hair loss that begins after puberty and progresses gradually. In women, however, it differs from men in that it is observed as thinning of hair over a relatively wide area on the crown of the head. The timing of onset also differs from men, with it becoming more common during menopause. In recent years, this condition has been called "female pattern hair loss." In both men and women, hair loss is a physiological phenomenon, but because it greatly affects one's appearance, it has a significant impact on quality of life (QOL).

[0003] Various products have been developed over time for the purpose of preventing, reducing, promoting hair growth, or stimulating hair regeneration in alopecia. For example, since minoxidil has excellent hair growth effects, technologies for incorporating it into hair growth agents and the like are being investigated. Patent Document 1 discloses a scalp and hair cosmetic containing specific ingredients such as polyhydric alcohols, organic acids, menthol, and minoxidil. [Prior art documents] [Patent Documents]

[0004] [Patent Document 1] Japanese Patent Publication No. 2007-45712 [Overview of the project] [Problems that the invention aims to solve]

[0005] The present invention provides a hyaluronic acid derivative composition that has excellent hair growth or hair restoration effects and can be used as a hair growth agent or hair restoration agent. [Means for solving the problem]

[0006] That is, the present invention includes the following aspects. (1) (A) A hyaluronic acid derivative into which a steric group has been introduced, and (B) a component that promotes hair growth or hair regrowth, contain fruit , The aforementioned (B) ingredient that promotes hair growth or hair development is minoxidil, finasteride, or ethinylestradiol. The hyaluronic acid derivative to which the steryl group (A) is introduced has a repeating unit represented by the following general formula (I), A hyaluronic acid derivative composition for direct application to a site on a target animal that is the target of hair growth or hair regrowth 。

[0007]

Chemical formula

[0008] (In the formula, R , R 2 , R 3 , and R 4 are each independently selected from the group consisting of a hydrogen atom, C 1-6 alkyl, formyl, and C 1-6 alkylcarbonyl; Z represents a direct bond or a peptide linker consisting of 2 or more and 30 or less arbitrary amino acid residues; X 1 is the following formula: -NR b -R, -NR b -COO-R, -NR b -CO-R, -NR <000001l>-CO-NR c -R, -COO-R, -O-COO-R, -S-R, -CO-Y a -S-R, -O-CO-Y b -S-R, -NR b -CO-Y b -S-R, and -S-S-R, A group selected from the group consisting of groups represented by; R a , R b and R c These are, independently, hydrogen atoms and C 1-20 Alkyl, amino C 2-20 Alkyl and hydroxy C 2-20 Selected from the group consisting of alkyl groups, where the alkyl portion of the group is -O- and -NR f A group selected from the group consisting of - may be inserted; R f C is a hydrogen atom. 1-12 Alkyl, amino C 2-12 Alkyl and hydroxy C 2-12 A group selected from the group consisting of alkyl groups may have a group selected from the group consisting of -O- and -NH- inserted into the alkyl portion of the group; R is a steryl group; Y is C 2-30 Alkylene, or -(CH2CH2O) m The alkylene is -CH2CH2-, where the alkylene is -O-, -NR g A group selected from the group consisting of - and -SS- may be inserted; R g C is a hydrogen atom. 1-20 Alkyl, amino C 2-20 Alkyl and hydroxy C 2-20 A group selected from the group consisting of alkyl groups may have a group selected from the group consisting of -O- and -NH- inserted into the alkyl portion of the group; Y a C 1-5 It is alkylene; Y b C 2-8 Alkylene or C 2-8 It is an alkenylene; m is an integer between 1 and 100 (inclusive).

[0009] (2) The hyaluronic acid derivative composition according to (1), wherein the content of the hyaluronic acid derivative into which the steryl group (A) is introduced is 0.01 parts by mass or more and 10 parts by mass or less per 100 parts by mass of the hyaluronic acid derivative composition. (3) The hyaluronic acid derivative composition according to (1) or (2), wherein the content of the ingredient that promotes hair growth or hair development (B) is 0.01 parts by mass or more and 40 parts by mass or less per 100 parts by mass of the hyaluronic acid derivative composition. (4) A hyaluronic acid derivative composition according to any one of (1) to (3), wherein the ratio of the mass of the hair growth or hair development component (B) to the mass of the hyaluronic acid derivative into which the steryl group (A) has been introduced is 0.01% or more and 10000% or less. (5) The hyaluronic acid derivative composition according to any one of (1) to (4), wherein the rate of introduction of the steryl group to the hyaluronic acid derivative into which the steryl group of (A) has been introduced is 1% or more and less than 60%. (6) A hyaluronic acid derivative composition according to any one of (1) to (5), wherein the molecular weight of the hyaluronic acid derivative into which the (A) steryl group is introduced is 1,000 or more and less than 1,000,000. (7) The R is a cholesteryl group. (1) to (6) The hyaluronic acid derivative composition described above. ( 8 ) A hair growth stimulant composition or a hair tonic composition, (1)~( 7 A hyaluronic acid derivative composition as described in any one of the following. [Effects of the Invention]

[0010] The hyaluronic acid derivative composition according to the above embodiment has excellent hair growth promoting or hair nourishment promoting effects and can be used as a hair growth agent or hair nourishment agent. [Brief explanation of the drawing]

[0011] [Figure 1] This is a fluorescence image of a mouse skin section in Test Example 2. [Modes for carrying out the invention]

[0012] The following describes in detail embodiments of the present invention (hereinafter referred to as "this embodiment"), but the present invention is not limited thereto, and various modifications are possible without departing from its spirit.

[0013] The following is a definition of terms used in this specification.

[0014] The term "C" as used herein 1-20 The term "alkyl" refers to linear or branched alkyl groups having 1 to 20 carbon atoms, such as methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, etc. 1-4 It contains alkyl, and further includes n-pentyl, 3-methylbutyl, 2-methylbutyl, 1-methylbutyl, 1-ethylpropyl, n-hexyl, 4-methylpentyl, 3-methylpentyl, 2-methylpentyl, 1-methylpentyl, 3-ethylbutyl, 2-ethylbutyl, etc. 1-20Alkyl compounds have 1 to 12 carbon atoms. 1-12 Alkyl, C with 1 to 6 carbon atoms 1-6 Alkyl groups are also included.

[0015] The term "C" as used herein 1-6 The term "alkylcarbonyl" refers to the alkyl part which has already been mentioned as C. 1-6 This refers to alkylcarbonyl groups, such as acetyl, propionyl, n-propylcarbonyl, iso-propylcarbonyl, n-butylcarbonyl, sec-butylcarbonyl, iso-butylcarbonyl, tert-butylcarbonyl, etc. 1-4 It contains "alkylcarbonyl".

[0016] "Amino C" as used herein 2-20 The term "alkyl" refers to a linear or branched alkyl group having 2 to 20 carbon atoms and having an amino group as a substituent. For example, the amino group may be located on the terminal carbon atom of the alkyl group. 2-20 Alkyl compounds include amino acids with 2 to 12 carbon atoms. 2-12 Alkyl compounds are also included.

[0017] The term "hydroxy C" as used herein 2-20 The term "alkyl" refers to a linear or branched alkyl group having 2 to 20 carbon atoms and having a hydroxyl group as a substituent. For example, the hydroxyl group may be located on the terminal carbon atom of the alkyl group. 2-20 Alkyl compounds include hydroxyl atoms with 2 to 12 carbon atoms. 2-12 Alkyl compounds are also included.

[0018] The term "C" as used herein 2-30 The term "alkylene" refers to a linear or branched divalent saturated hydrocarbon group having 2 to 30 carbon atoms, including, for example, ethylene, propylene, etc., and having 2 to 20 carbon atoms. 2-20 Alkylene, C with 2 to 8 carbon atoms 2-8 Alkylene, base "-(CH2)"n - (where n is between 2 and 30, preferably between 2 and 20, and more preferably between 2 and 15).

[0019] The term "C" as used herein 1-5 The term "alkylene" refers to a linear or branched divalent saturated hydrocarbon group having 1 to 5 carbon atoms, including, for example, methylene, ethylene, and propylene.

[0020] The term "C" as used herein 2-8 "Alkenylene" refers to a divalent saturated hydrocarbon group having 2 to 8 carbon atoms, in a linear or branched chain, containing one or more double bonds. Examples include -CH=CH-, -C(CH3)=CH-, 2-butene-1,4-diyl, hepta-2,4-diene-1,6-diyl, octa-2,4,6-triene-1,8-diyl, etc. If geometric isomerism exists, each isomer and mixtures thereof are also included.

[0021] Hyaluronic acid derivative composition The hyaluronic acid derivative composition of this embodiment comprises (A) a hyaluronic acid derivative into which a steryl group has been introduced (hereinafter sometimes referred to as "(A) hyaluronic acid derivative") and (B) an ingredient that promotes hair growth or hair development.

[0022] In the following, (A) hyaluronic acid derivatives into which steryl groups have been introduced and (B) ingredients that promote hair growth or hair development may be referred to as ingredient (A) and ingredient (B), respectively.

[0023] In the hyaluronic acid derivative composition of this embodiment, (B) a component that promotes hair growth or hair nourishment forms a complex encapsulated within (A) the hyaluronic acid derivative. As shown in the examples described later, when the hyaluronic acid derivative composition of this embodiment is administered to the skin of a target animal as a hair growth agent or hair nourishment agent, the complex in which component (B) is encapsulated within component (A) remains in the hair follicles of the target animal for a long period of time, and component (B) is released gradually, thereby effectively delivering component (B) to the hair roots. The hyaluronic acid derivative composition of this embodiment is a topical composition and is useful as a hair growth agent or hair nourishment agent composition.

[0024] The test animals to be administered the hyaluronic acid derivative composition of this embodiment are not particularly limited as long as they are animals with hair, but mammals are preferred. Examples of mammals include humans, monkeys, chimpanzees, marmosets, mice, guinea pigs, rats, rabbits, goats, sheep, dogs, and cats, but humans are preferred among them.

[0025] Next, each component constituting the hyaluronic acid derivative composition of this embodiment will be described in detail below.

[0026] <(A) Hyaluronic acid derivative with introduced steryl group> (A) In hyaluronic acid derivatives, the steryl group may be directly bonded to hyaluronic acid, or it may be bonded after dissociating the linker. The term "linker" as used herein refers to any peptide linker that can be introduced by genetic engineering, or a synthetic compound linker. However, in the case of the (A) hyaluronic acid derivative of this embodiment, a peptide linker is preferred. The length of the peptide linker is not particularly limited and can be appropriately selected by those skilled in the art depending on the purpose. However, a preferred length is 2 amino acids or more (the upper limit is not particularly limited, but is usually 30 amino acids or less, preferably 20 amino acids or less), and particularly preferably 15 amino acids. The peptide linkers contained in the (A) hyaluronic acid derivative may all be peptide linkers of the same length, or peptide linkers of different lengths may be used.

[0027] [Steryl group] The term "steryl group" as used herein is not particularly limited as long as it is a group having a steroid skeleton. Specifically, examples of steroids include cholesterol, cholestanol, campestanol, ergostanol, stigmamanol, coprostanol, stigmasterol, sitosterol, lanosterol, ergosterol, simialenol, bile acids, testosterone, estradiol, progesterone, cortisol, cortisone, aldosterone, corticosterone, deoxycortisterone, etc. Examples of steryl groups include cholesteryl group, stigmasteryl group, lanosteryl group, ergosteryl group, etc., with cholesteryl group (especially cholesta-5-en-3β-yl group) being preferred.

[0028] [Steryl group introduction rate] (A) The rate of steryl group introduction into the hyaluronic acid derivative (hereinafter sometimes simply referred to as "steryl group introduction rate") is preferably 1% or more and less than 60%, more preferably 4% or more and 50%, and even more preferably 8% or more and 45%. When the steryl group introduction rate is within the above range, the hyaluronic acid derivative has the property of forming nanoparticles and dispersing well in water. When the steryl group introduction rate is within the above range, the hyaluronic acid derivative-drug conjugate composition, in which the (A) hyaluronic acid derivative in the hyaluronic acid derivative composition is compounded with a drug, can become a sustained-release formulation that dissolves in water and can be subjected to sterile filtration.

[0029] The rate of steryl group introduction is 1 It can be measured by 1H-NMR measurement. That is, the hyaluronic acid derivative composition 1 The integral value of the peak originating from the steryl group of (A) hyaluronic acid derivative in the 1H-NMR spectrum and the integral value of the peak originating from the acetyl group of N-acetyl-D-glucosamine contained in (A) hyaluronic acid derivative (COCH3, 1.6 ppm to 2.0 ppm, 3H) can be used to calculate the following based on the formula below.H This represents the number of hydrogen atoms corresponding to the peak. Specifically, it can be measured according to the method described in the examples below.

[0030] [Steryl group introduction rate] (%) =[(Peak integral value derived from steryl group × 3 / n H ) / (Peak integral value derived from the acetyl group of N-acetyl-D-glucosamine)] × 100

[0031] [(A) Molecular weight of hyaluronic acid derivative] The molecular weight of the (A) hyaluronic acid derivative is not particularly limited, but from the viewpoint of increasing the number of steryl groups introduced per molecule of the hyaluronic acid derivative and forming a drug conjugate, a (A) hyaluronic acid derivative with a relatively large molecular weight is preferred. From the viewpoint of expecting sterile filtration performance and syringeability when the final dosage form is a solution formulation, a (A) hyaluronic acid derivative with a relatively small molecular weight is preferred. The molecular weight of such a (A) hyaluronic acid derivative is preferably 1,000 (1k) or more and 1,000,000 (1,000k) or less, more preferably 5k or more and 500k or less, and even more preferably 10k or more and 300k or less. When the molecular weight of the (A) hyaluronic acid derivative is above the lower limit, molecular entanglement can be further increased, and the retention of the hyaluronic acid derivative composition in the pores can be further improved. On the other hand, when the molecular weight of the (A) hyaluronic acid derivative is below the upper limit, the increase in viscosity can be suppressed, and the penetration of the hyaluronic acid derivative composition into the pores can be further improved. (A) The molecular weight of hyaluronic acid derivatives can generally be adjusted by using raw materials having the corresponding molecular weight.

[0032] Here, "(A) Molecular weight of the hyaluronic acid derivative" refers to the weight-average molecular weight determined by size exclusion chromatography multi-angle light scattering detector (SEC-MALS). Specifically, it can be measured according to the method described in the examples below.

[0033] Preferred (A) hyaluronic acid derivatives include, for example, hyaluronic acid derivatives having one or more repeating units represented by the following general formula (I) (hereinafter sometimes referred to as "repeating unit (I)").

[0034] [ka]

[0035] (In the formula, R 1 , R 2 , R 3 , and R 4 These are, independently, hydrogen atoms and C 1-6 Alkyl, formyl and C 1-6 Selected from the group consisting of alkylcarbonyls; Z represents a direct linker or a peptide linker consisting of any 2 to 30 amino acid residues; X 1 The formula is as follows: -NR b -R, -NR b -COO-R, -NR b -CO-R, -NR b -CO-NR c -R, -COO-R, -O-COO-R, -SR, -CO-Y a -SR, -O-CO-Y b -SR, -NR b -CO-Y b -SR, and -SSR, A group selected from the group consisting of groups represented by; R a , R b and R c These are, independently, hydrogen atoms and C 1-20 Alkyl, amino C 2-20 Alkyl and hydroxy C 2-20Selected from the group consisting of alkyl, wherein the alkyl portion of the group may have a group selected from the group consisting of -O- and -NR f - inserted therein; R f is a hydrogen atom, C 1-12 alkyl, amino C 2-12 alkyl and hydroxy C 2-12 alkyl, and the alkyl portion of the group may have a group selected from the group consisting of -O- and -NH- inserted therein; R is a sterol group; Y is C 2-30 alkylene, or -(CH2CH2O) m -CH2CH2-, where the alkylene may have a group selected from the group consisting of -O-, -NR g - and -S-S- inserted therein; R g is a hydrogen atom, C 1-20 alkyl, amino C 2-20 alkyl and hydroxy C 2-20 alkyl, and the alkyl portion of the group may have a group selected from the group consisting of -O- and -NH- inserted therein; Y a is C 1-5 alkylene; [[ID=4]]Y b is C 2-8 alkylene or C[[ID=4]] 2-8 alkenylene; m is an integer of 1 or more and 100 or less.)

[0036] (A) The hyaluronic acid derivative preferably includes a hyaluronic acid derivative having one or more repeating units represented by the following general formula (Ia) (hereinafter, may be referred to as "repeating unit (Ia)").

[0037] <ooo0535>

Chemical formula

[0038] (In the formula, R 1 、R2 , R 3 , and R 4 These are, independently, hydrogen atoms and C 1-6 Alkyl, formyl and C 1-6 Selected from the group consisting of alkylcarbonyls; X is -NR a -Y-NR b It is a hydrophobic group represented as -COO-R; R a and R b These are, independently, hydrogen atoms and C 1-6 Selected from the group consisting of alkyl groups; R is a steryl group; Y is C 2-30 Alkylene, or -(CH2CH2O) m It is -CH2CH2-, m is an integer between 1 and 100 (inclusive).

[0039] Here, if (A) the hyaluronic acid derivative contains two or more repeating units (I) or (Ia), these repeating units may be the same or different.

[0040] (A) The hyaluronic acid derivative may be modified at positions other than the repeating unit (I) or the repeating unit (Ia), for example, the hydroxyl group may be -O(C 1-6 Alkyl), -O (formyl), -O (C 1-6 The carboxyl group may be converted to an alkylcarbonyl group, and the carboxyl group may be converted to an amide or ester, and a salt may be formed.

[0041] [Repeating Unit (I)] The base "-Zn(R)" in general formula (I) a )YX 1 The expression is as follows: -NH-(CH2) mz -NH-R; -NH-(CH2) mz -NH-COO-R; -NH-(CH2CH2O) m-CH2CH2-NH-COO-R; -NH-(CH2) mz -COO-R; -NH-(CH2CH2O) m -CH2CH2-COO-R, -NH-(CH2) mz -O-COO-R; -NH-(CH2CH2O) m -CH2CH2-O-COO-R, -NH-(CH2) mz -SR; -NH-(CH2CH2O) m -CH2CH2-SR; -NH-(CH2) mz -O-CO-CH(R 8 )-CH2-SR; -NH-(CH2) mz -NHCO-CH(R 8 )-CH2-SR; -NH-(CH2CH2O) m -CH2CH2-NHCO-CH(R 8 )-CH2-SR; -NH-(CH2CH2O) m -CH2CH2-O-CO-CH(R 8 )-CH2-SR; -NH-(CH2) mz -SSR; and -Z-NR a -Y-NR b -COO-R (Here, mz is an integer between 2 and 30, and R 8 (wherein R is a hydrogen atom or a methyl group, and R and m are as already defined herein.) It includes a group selected from the group consisting of groups represented by . The group in question is, -NH-(CH2) mz -NH-COO-R; -NH-(CH2CH2O) m -CH2CH2-NH-COO-R; and -NH-(CH2) mz -SSR (Here, mz, R, and m are as already defined herein.) A group selected from the group consisting of is preferred.

[0042] (Z) In general formula (I), Z is preferably a direct bond. In another embodiment, when Z is a peptide linker, X 1 -NR b It is preferable that -COO-R. Furthermore, in another embodiment, Z is -NH-[CH(-Z a )-CONH] n-1 -CH(-Z a It may also be a peptide linker represented by )-CO-, where n is an integer between 2 and 30, and Z a Each of these is independently H2N-CH(-Z a This represents a substituent in an α-amino acid, represented as -COOH. The peptide linker is bonded to the carboxyl group of the glucuronic acid moiety at the N-terminus and to the group -N(-R) at the C-terminus. a )-YX 1 It binds to. Examples of amino acids that can be used as amino acid residues of the peptide linker include α-amino acids, such as alanine, arginine, asparagine (Asn), aspartic acid, cysteine, glutamine, glutamic acid, glycine (Gly), histidine, isoleucine, leucine (Leu), lysine, methionine, phenylalanine (Phe), proline, serine, threonine, tryptophan, tyrosine, and valine, as well as their D forms, and all α-amino acids, including synthesized amino acids, can be used. That is, Z a Examples of these include -CH3, H2NC(NH)NH(CH2)3-, H2NCOCH2-, etc. Also, the n Zs may be the same or different. n is an integer between 2 and 30, but is preferably between 2 and 10, and more preferably between 2 and 4. Preferred examples of peptide linkers include, for example, -Gly-Phe-Leu-Gly-, -Asn-Phe-Phe-, -Phe-Phe-, Phe-Gly-, etc.

[0043] (Y) In general formula (I), Y is -(CH2) n1 -and-(CH2CH2O) m1 A base selected from the group consisting of -CH2CH2- (where n1 is an integer between 2 and 20, preferably between 2 and 15, more preferably between 2 and 12, and even more preferably between 2 and 6; m1 is an integer between 1 and 4) is preferred. Specifically, -(CH2)2-, -(CH2)6-, -(CH2)8-, -(CH2) 12 -, or -(CH2CH2O)2-CH2CH2- is preferred. Furthermore, from the viewpoint of achieving high solubility in pure water or low salt concentrations while exhibiting high precipitation ability under physiological salt concentrations, Y is preferably -(CH2)2-, -(CH2)6-, -(CH2)8- and -(CH2) 12 A group selected from the group consisting of - is preferred, and -(CH2)6- is more preferred.

[0044] Y can be, for example, -CH2CH2O-CH2CH2-SS-CH2CH2O-CH2CH2-, -(CH2CH2O)2-CH2CH2-SS-CH2CH2O-CH2CH2-, -CH2CH2O-CH2CH2-SS-(CH2CH2O)2-CH2CH2-, -(CH2CH2O)2-CH2CH2-SS-(CH2CH2O)2-CH2CH2-, etc.

[0045] (Y a ) Y a -CH2- or -CH2-CH2- is preferred.

[0046] (Y b ) Y b The preferred members are -CH2-CH2-, -CH(CH3)CH2-, 2-butene-1,4-diyl, hepta-2,4-diene-1,6-diyl, or octa-2,4,6-triene-1,8-diyl, with -CH2-CH2- or -CH(CH3)CH2- being more preferred.

[0047] Base "-ZN(Ra )YX 1 Specific examples of "" include -NH-(CH2)2-NH-CO-cholesteryl, -NH-(CH2)4-NH-(CH2)3-NH-(CH2)3-NH-COO-cholesteryl, -NH-(CH2)3-NH-(CH2)4-NH-(CH2)3-NH-COO-cholesteryl, -NH-(CH2)4-NH-(CH2)3-NH-COO-cholesteryl, -NH-(CH2)4-N(-(CH2)3-NH2)-COO-cholesteryl, -NH-(CH2)3-NH-(CH2)4-N(-(C Examples include H2)3-NH2)-COO-cholesteryl, -NH-(CH2)3-NH-(CH2)4-N(-(CH2)3-NH-(CH2)3-NH2)-COO-cholesteryl, -NH-(CH2)3-NH-(CH2)4-N(-(CH2)3-NH2)-CO-NH-cholesteryl, -NH-(CH2)3-NH-(CH2)4-N(-(CH2)3-NH2)-CO-cholesteryl, -NH-(CH2)3-NH-(CH2)4-N(-(CH2)3-NH2)-cholesteryl, etc. Preferred group "-Zn(R a )YX 1 " is R a , R b and R c However, it is a hydrogen atom, and Y is a linear C 2-30 Alkylene or -(CH2CH2O) m -CH2CH2- and Y a However, linear C 1-5 It is alkylene, or Y b However, linear C 2-8 Alkylene or linear carbon 2-8 It is alkenylene.

[0048] [Repeating Unit (Ia)] In general formula (Ia), X is -NH-(CH2)2-NH-COO-cholesteryl, -NH-(CH2)6-NH-COO-cholesteryl, -NH-(CH2) 12-NH-COO-cholesteryl or -NH-(CH2CH2O)2-CH2CH2-NH-COO-cholesteryl is preferred, and -NH-(CH2)2-NH-COO-cholesteryl, -NH-(CH2)6-NH-COO-cholesteryl, or -NH-(CH2CH2O)2-CH2CH2-NH-COO-cholesteryl is more preferred.

[0049] (A) In addition to the repeating unit (I), the hyaluronic acid derivative may further contain a repeating unit represented by general formula (II) (hereinafter sometimes referred to as "repeating unit (II)").

[0050] [ka]

[0051] (In the formula, R 1a , R 2a , R 3a , and R 4a These are, independently, hydrogen atoms and C 1-6 Alkyl, formyl and C 1-6 Selected from the group consisting of alkylcarbonyls; X a hydroxy and -OQ + Selected from the group consisting of; where Q + (This is a counter-cation.)

[0052] Here, if (A) the hyaluronic acid derivative contains two or more repeating units (II), these repeating units may be the same or different. In another embodiment, (A) the hyaluronic acid derivative may be a hyaluronic acid derivative substantially comprising repeating unit (I), repeating unit (Ia), and repeating unit (II).

[0053] [Repeating Unit (II)] In general formula (II), Q +The countercation is not particularly limited as long as it forms a salt with a carboxyl group in water, and in the case of a valency of 2 or higher, it forms a salt with multiple carboxyl groups depending on the valency. Examples of countercations include metal ions such as lithium ions, sodium ions, rubidium ions, cesium ions, magnesium ions, and calcium ions; formula: N + R j R k R l R m (In the formula, R j , R k , R l and R m These are, independently, hydrogen atoms and C 1-6 Examples include ammonium ions (selected from the group consisting of alkyl groups). Among them, Q + The preferred ions are sodium ions, potassium ions, or tetraalkylammonium ions (e.g., tetra-n-butylammonium ions). j , R k , R l and R m C 1-6 It is preferable that the group be the same group selected from the group consisting of alkyl groups, and an n-butyl group is preferred.

[0054] R 1 , R 2 , R 3 , and R 4 , and R 1a , R 2a , R 3a , and R 4a It is preferable that all of them are hydrogen atoms. Also, R a and R b Preferably, all of these are hydrogen atoms.

[0055] In particular, (A) the hyaluronic acid derivative is preferably a hyaluronic acid derivative substantially composed of repeating unit (I) and repeating unit (II). In (A) the hyaluronic acid derivative, of the repeating units of the disaccharide composed of D-glucuronic acid and N-acetyl-D-glucosamine contained in the derivative, for example, 80% or more, preferably 90% or more, and more preferably 95% or more are repeating units (I) and repeating unit (II). (A) the hyaluronic acid derivative may consist only of repeating unit (I) and repeating unit (II).

[0056] [(A) Hyaluronic acid derivative content] (A) The content of the hyaluronic acid derivative is preferably 0.01 parts by mass or more and 10 parts by mass or less, and more preferably 1 part by mass or more and 10 parts by mass or less, per 100 parts by mass of the hyaluronic acid derivative composition. (A) When the content of the hyaluronic acid derivative is above the lower limit, molecular entanglement can be further enhanced, and the retention of the hyaluronic acid derivative composition in the pores can be further enhanced. On the other hand, when the content of the hyaluronic acid derivative is below the upper limit, the increase in viscosity can be suppressed, and the penetration of the hyaluronic acid derivative composition into the pores can be further enhanced.

[0057] (B) Ingredients that promote hair growth or hair development (B) The hair growth or hair restoration component is complexed with (A) the hyaluronic acid derivative in water. This allows the hyaluronic acid derivative composition to remain in the hair follicles of the target animal when used as a hair growth agent or hair restoration agent, thereby increasing the hair growth or hair restoration effect derived from component (B).

[0058] In this specification, "promotion of hair growth" means the phenomenon of increasing hair volume. For example, promotion of hair growth includes stimulating hair follicles in the resting phase of the hair cycle to induce cell division of hair matrix cells and regenerate hair cells, or causing hair follicles to appear in areas where there were previously no hair follicles. Furthermore, promotion of hair growth includes, for example, suppressing the transition of hair follicles in the growth phase of the hair cycle to the regression or resting phase.

[0059] (B) The content of the hair growth or hair development promoting ingredient is preferably 0.01 parts by mass or more and 40 parts by mass or less, more preferably 0.01 parts by mass or more and 30 parts by mass or less, even more preferably 0.1 parts by mass or more and 20 parts by mass or less, and particularly preferably 0.1 parts by mass or more and 10 parts by mass or less, per 100 parts by mass of the hyaluronic acid derivative composition. (B) When the content of the hair growth or hair development promoting ingredient is above the lower limit above, the hair growth promoting effect or hair development promoting effect can be more effectively exhibited, while when it is below the upper limit above, skin irritation can be further reduced.

[0060] Furthermore, the ratio of the mass of (B) the hair growth or hair development-promoting component to the mass of (A) the hyaluronic acid derivative is 0.01 % That's all, 10,000 % Preferably, it is 0.01 % The above 500 % It is more preferable that the following conditions be met: 0.1 % The above 400 % More preferably, 1 % The above 300 % The following is particularly preferable: 10 % The above 200 % The following is most preferable: (A) The ratio of the mass of the hair growth or hair development promoting component to the mass of the hyaluronic acid derivative (B) is within the above numerical range, so that component (B) can be retained in the hair follicles of the target animal, and the hair growth or hair development promoting effect derived from component (B) can be increased.

[0061] (B) The content of ingredients that promote hair growth or hair regeneration can be measured using, for example, high-performance liquid chromatography (HPLC), gel permeation chromatography (GPC), or liquid chromatography-mass spectrometry (LC-MS).

[0062] (B) Ingredients that promote hair growth or hair development include (B1) ingredients that promote hair growth and (B2) ingredients that promote hair development.

[0063] (B1) Examples of ingredients that promote hair growth include minoxidil, finasteride, tert-flavanone, quillaja saponin, sarsa saponin, thymosaponin, digitalis saponin, yam saponin, compounds having a sulfonamide group, chitin, chitosan, N-acyl-L-homoserine lactone, etc. Among these, minoxidil or finasteride is preferred.

[0064] (B2) Examples of ingredients that promote hair growth include cephalatin, nicotinamide, benzyl nicotinate, salicylic acid, acetyl pantothenyl ethyl ether, photosensitizer 301, ethinylestradiol, hinokitiol, jojoba oil, glyceryl tri-2-ethylhexanoate, squalane, dl-α-tocopherol, dipotassium glycyrrhizate, pantothenyl ethyl ether, dl-pyrrolidone carboxylate sodium salt, trimethylglycine, sorbitol solution, and glycine.

[0065] The hyaluronic acid derivative composition of this embodiment may contain one of the above-mentioned (B) hair growth or hair development components alone, or it may contain two or more components in combination.

[0066] <Other ingredients> The hyaluronic acid derivative composition of this embodiment may further contain other components, to the extent that they do not affect its effect. Examples of other components include pharmaceutically acceptable excipients, carriers, diluents, surfactants, penetration enhancers, humectants, blood circulation enhancers, anti-inflammatory agents, antioxidants, UV protectants, cell proliferation enhancers, cell differentiation inducers, fragrances, oils and fats, enzymes, cooling agents, vitamins, amino acids, proteins, etc. Examples of excipients include lactose, sucrose, and corn starch. Examples of carriers and diluents may further contain solvents such as sterile water, physiological saline, organic solvents, and corn oil. Examples of organic solvents include ethanol, isopropanol, propylene glycol, butylene glycol, and glycerin.

[0067] <Method for producing hyaluronic acid derivative compositions> The hyaluronic acid derivative composition of this embodiment can be manufactured by (A) producing a hyaluronic acid derivative, and then adding (B) a component that promotes hair growth or hair development to the (A) hyaluronic acid derivative.

[0068] First, as a method for producing (A) hyaluronic acid derivatives, for example, the carboxyl group of glucuronic acid is converted to an amide and a steryl group is introduced to obtain (A) hyaluronic acid derivatives. Furthermore, by adjusting the amount of the compound having a steryl group that is reacted with the raw material hyaluronic acid or its derivative, the steryl group introduction rate can be set to 1% or more and less than 60%.

[0069] Specifically, as a method for converting the carboxyl group of glucuronic acid to an amide and introducing a steryl group, for example, a raw material hyaluronic acid or its derivative, preferably hyaluronic acid or its derivative composed only of repeating units (II), is ion-exchanged with a tetraalkylammonium salt (e.g., tetrabutylammonium (TBA) salt), and in the presence of a suitable condensing agent, the hyaluronic acid salt is mixed with the esteryl group in a solvent, and the formula is "HNR" a -Y-NR b -R, NHR a -Y-NR b -COO-R, HNRa -Y-NR b -COO-R, HNR a -Y-NR b -CO-R, HNR a -Y-NR b -CO-NR c -R, HNR a -Y-COO-R, HNR a -YO-COO-R, HNR a -YSR, HNR a -Y-CO-Y a -SR, HNR a -YO-CO-Y b -SR, HNR a -Y-NR b -CO-Y b -SR, HNR a -YSSR, or -Z-NR a -Y-NR b -COO-R(wherein, R a , R b , R c , Y, Y a , Y b One method involves reacting an amine to which a steryl group (in particular a cholesteryl group) represented by ", where Z and R are as already defined herein" has been introduced with .

[0070] The coupling agent that can be used in the above reaction is not particularly limited and includes, for example, 4-(4,6-dimethoxy-1,3,5-triazine)-4-methylmorphorium (DMT-MM), N,N'-carbonyldiimidazole (CDI), N,N'-dicyclohexylcarbodiimide (DCC), N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ), 2-benzotriazole-1,1,3,3-tetramethyluronium tetrafluoroborate (TBTU), 3,4-dihydro-3-hydroxy-4-oxo-1,2,3-benzotriazine (HODhbt), benzotriazole-1-oxy-tris-pyrrolidino-phosphonium hexafluorophosphate (PyBOP), and benzotriazole-1-yl-oxy-tris(dimethylamino)phosphonium Examples include hexafluorophosphate (BOP), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC), and N-hydroxysuccinimide (NHS).

[0071] In particular, although not limited to, DMT-MM is preferred because the reaction proceeds with high efficiency even in a mixed solvent of water and an organic solvent. Furthermore, by using DMT-MM as a condensing agent, it is possible to selectively form amide bonds between amino groups and carboxyl groups while suppressing ester bond formation in systems where many hydroxyls coexist. By using this condensing agent, it is possible to prevent, for example, the reaction of the solvent alcohol with the carboxyl groups of the hyaluronic acid portion, or the formation of unwanted crosslinks by intramolecular or intermolecular bonding between carboxyl groups and hydroxyls simultaneously present in the hyaluronic acid portion.

[0072] Solvents used in the steryl group introduction reaction include water, DMSO, methanol, ethanol, propanol, butanol, isopropanol, polyhydric alcohols, acetonitrile, DMF, THF, dichloromethane, chloroform, hexane, diethyl ether, ethyl acetate, and mixtures thereof. The polyhydric alcohol may be a dihydric alcohol or a trihydric alcohol. Examples of dihydric alcohols include ethylene glycol, diethylene glycol, propylene glycol, dipropylene glycol, neopentyl glycol, 1,4-butanediol, and 1,6-hexanediol. Examples of trihydric alcohols include glycerin and trimethylolpropane.

[0073] Alternatively, the raw material hyaluronic acid or its derivative may be ion-exchanged with a tetraalkylammonium salt (e.g., tetrabutylammonium (TBA) salt), and the hyaluronic acid salt and the spacer portion may be reacted in a solvent in the presence of a suitable condensing agent (protection and deprotection reactions may be performed as needed) to convert the carboxyl group (-COOH) of the raw material hyaluronic acid or its derivative, and then reacted with a suitable reagent. Examples of combinations of groups derived from the carboxyl group and reaction reagents are shown below. -CONR a -Y-NR b H + Hal-R; -CONR a -Y-NR b H + Hal-COOR; -CONR a -Y-NR b H + HOCO-R; -CONR a -Y-NR b H + Hal-CO-R; -CONR a -Y-NR b -COOH + HNR c -R; -CONR a -Y-NR b -CO-NR c H + Hal-R; -CONR a -Y-NR b H + HOCO-NR c -R; -CONR a -Y-NR b H + Hal-CO-NR c -R; -CONR a -Y-COOH + HO-R; -CONR a -Y-OH + Hal-COO-R; -CONR a -Y-OCOOH + HO-R; -CONR a -Y-OCOOH + Hal-R; -CONR a -Y-OCO-Hal + HO-R; -CONR a -Y-SH + Hal-R; -CONR a -Y-Hal + HS-R; -CONR a -Y-CO-Y a -Hal + HS-R; -CONR a -Y-CO-Y a -SH + Hal-R; -CONR a -Y-O-CO-CH=CH2+ HS-R; -CONR a -Y-NR b -CO-CH(CH3)=CH2+ HS-R; -CONR a -Y-SH + HS-R; -COZ-OH + HNR a -Y-NR b -COO-R; -COZ-NR a -Y-NR b H + Hal-COO-R (wherein, R a 、R b 、R c 、Y、Y a 、Y b(wherein Z is as already defined herein, and Hal represents a halogen atom selected from the group consisting of fluorine, chlorine, bromine, and iodine).

[0074] Reaction modes include dehalogenation reactions, condensation reactions, dehydration reactions, nucleophilic addition reactions such as Michael addition, and oxidative disulfide formation reactions. These are well-known reactions and can be appropriately selected by those skilled in the art, and carried out under preferred reaction conditions. If the converted product or reactant has a carboxyl group, it may be reacted with an N-hydroxysuccinimide (hereinafter also referred to as "NHS") ester.

[0075] Another method involves reacting the carboxyl group of hyaluronic acid or its derivative with 2-aminoethyl 2-pyridyl disulfide to prepare a hyaluronic acid derivative into which a spacer having a mercapto group modified with a leaving group at its terminal end is introduced, and then forming a disulfide bond by nucleophilic substitution of thiocholesterol with this derivative.

[0076] Furthermore, a method can be described in which a portion of the spacer is introduced into the carboxyl group of hyaluronic acid or its derivative, and a portion of the spacer is introduced into the steryl group, and these are reacted. Some specific examples have been described above, but further, if -SS- is inserted into Y, a method can be described in which a hyaluronic acid derivative in which a spacer having a mercapto group at the terminal is introduced into the carboxyl group of hyaluronic acid, and a steryl group in which a spacer having a mercapto group at the terminal is introduced, and these are reacted oxidatively to form a disulfide bond. In this case, one mercapto group can be reacted with 2-mercaptopyridine to form a disulfide, and then substituted with the other mercapto group.

[0077] Furthermore, after preparing the hyaluronic acid derivative of the present invention, other substituents may be introduced. For example, 0.1% to 99.5%, preferably 40% to 65%, of the carboxyl groups in the hyaluronic acid derivative substantially composed of repeating unit (I) and repeating unit (II) may be added as -CO-Xz , where X z is the following group: -NH-(CH2) p1 -O-CO-C(R 17 )=CH2; -NH-(CH2) p1 -O-CO-CH(R 17 )-CH2-S-CH2-CH(OH)-CH(OH)-CH2-SH; -NH-(CH2) p1 -SH; -NH-(CH2) p1 -NH-CO-C(R 17 )=CH2; -NH-(CH2) p1 -NH-C(=NH)-(CH2)3-SH; -NH-(CH2) p1 -NH-CO-(CH2) r -SH; -NH-(CH2) p1 -NH-CO-CH(R 17 )-CH2-S-CH2-CH(OH)-CH(OH)-CH2-SH; -NH-(CH2) p1 -NH-CO-CH(NH2)-CH2-SH; -NH-(CH2) p1 -NH-CO-CH(NH2)-(CH2)2-SH; -NH-NH-CO-(CH2)4-CO-NH-NH-C(=NH)-(CH2)3-SH; -NH-(CH2-CH2-O) q -CH2-CH2-O-CO-C(R 17 )=CH2; -NH-(CH2-CH2-O) q -CH2-CH2-O-CO-CH(R 17 )-CH2-S-CH2-CH(OH)-CH(OH)-CH2-SH; -NH-(CH2-CH2-O) q -CH2-CH2-SH; -NH-(CH2-CH2-O) q -CH2-CH2-NH-CO-C(R 17 )=CH2; -NH-(CH2-CH2-O) q -CH2-CH2-NH-C(=NH)-(CH2)3-SH; -NH-(CH2-CH2-O) q -CH2-CH2-NH-CO-(CH2) r -SH; -NH-(CH2-CH2-O) q -CH2-CH2-NH-CO-CH(R 17 )-CH2-S-CH2-CH(OH)-CH(OH)-CH2-SH; -NH-(CH2-CH2-O) q -CH2-CH2-NH-CO-CH(NH2)-CH2-SH; -NH-(CH2-CH2-O) q -CH2-CH2-NH-CO-CH(NH2)-(CH2)2-SH; -NH-CH(CO2H)-(CH2)-SH; -NH-CH(CO2H)-(CH2)2-SH; and -NH-CH(CO2H)-(CH2)2-CONH-CH(CONH-CH2-CO2H)-CH2-SH (Here, R 17 is a hydrogen atom or C 1-6 [Selected from the group consisting of alkyl groups, where p1 is an integer between 2 and 10, q is an integer between 1 and 200, and r is an integer between 1 and 3] By converting to this, it is also possible to crosslink molecules within the molecule or between molecules including other molecules, thereby causing gelation.

[0078] (A) The step of gelling the hyaluronic acid derivative by chemical crosslinking may be performed under conditions that are appropriate. Conditions for crosslinking include the crosslinking method, polymer concentration, crosslinking agent concentration, solvent, solvent pH, salt concentration, temperature, time, etc.

[0079] (A) In the process of gelling a hyaluronic acid derivative, the crosslink density of the resulting gel can be increased by, for example, increasing the polymer concentration during chemical crosslinking and the introduction rate of groups capable of crosslinking within the reaction conditions for crosslinking.

[0080] (A) When using a crosslinking agent in the process of gelling a hyaluronic acid derivative, it is preferable to add the crosslinking agent at a concentration such that the groups have crosslinking-capable groups at both ends can quickly and adequately participate in the crosslinking reaction. For example, when crosslinking a polymer into which methacryloyl groups (MA groups) have been introduced by a Michael addition reaction using DTT, a MA group:SH group ratio of 3:1 to 1:3 is preferred, and a ratio of 2:1 to 1:2 is particularly preferred.

[0081] (A) The solvent used in the step of gelling the hyaluronic acid derivative is preferably one that can sufficiently dissolve the polymer and the crosslinking agent, and is not particularly limited, but is preferably water, dimethyl sulfoxide (DMSO), dimethylacetamide (DMAc), dimethylformamide (DMF), N-methylpyrrolidone (NMP), or a mixed solvent selected from these. It is also possible to use a mixture of these solvents and an organic solvent that is miscible. Not particularly limited, but examples of miscible organic solvents include methanol, ethanol, propanol, isopropanol, butanol, polyhydric alcohols, acetone, acetonitrile, etc. The polyhydric alcohol may be a dihydric alcohol or a trihydric alcohol. Examples of dihydric alcohols include ethylene glycol, diethylene glycol, propylene glycol, dipropylene glycol, neopentyl glycol, 1,4-butanediol, 1,6-hexanediol, etc. Examples of trihydric alcohols include glycerin, trimethylolpropane, etc.

[0082] (A) Hyaluronic acid derivatives form nanoparticles in aqueous solution, and by crosslinking them in solution, a nano-sized particle gel can be formed. The solution concentration is preferably 0.1 mg / mL or higher, and more preferably 1 mg / mL or higher. On the other hand, the solution concentration is preferably 100 mg / mL or lower, more preferably 50 mg / mL or higher, and even more preferably 20 mg / mL. In other words, the solution concentration is preferably 0.1 mg / mL or more and 100 mg / mL or less, more preferably 1 mg / mL or more and 50 mg / mL or more, and even more preferably 1 mg / mL or more and 20 mg / mL or more. When the solution concentration is above the lower limit, a dilute solution can be created, preventing the microparticle gels from associating with each other, allowing the hyaluronic acid derivative composition to be introduced into the pores as small particles. On the other hand, when the solution concentration is below the upper limit, the aggregation of the microparticle gels improves the retention of the hyaluronic acid derivative composition in the pores.

[0083] (A) The gelation step of the hyaluronic acid derivative may be carried out in bulk, or in a discontinuous phase such as an emulsion or spray droplets. For example, when carried out in a W / O emulsion, the aqueous phase in which the polymer and crosslinking agent are dissolved can be emulsified in a solvent that is immiscible with water, and the gelation reaction can be carried out. The solvent that is immiscible with water is not particularly limited, but examples include hexane, chloroform, dichloromethane, ethyl acetate, medium-chain triglyceride (MCT), liquid paraffin, and soybean oil. A surfactant may be added to stabilize the emulsification. Alternatively, the process may be carried out in a solvent that allows for solvent removal, such as in supercritical carbon dioxide or PEG. In this case, by emulsifying and dispersing the aqueous or organic solvent phase in which the polymer and crosslinking agent are dissolved in the solvent mentioned above, the polymer is concentrated due to solvent removal (solvent diffusion), making it possible to obtain a gel with a higher crosslinking density.

[0084] (A) A step of gelling the hyaluronic acid derivative, and thereafter, an operation to stop the crosslinking reaction and an operation to deactivate or wash away any remaining crosslinkable functional groups may be performed. Crosslinkable functional groups that did not participate in the reaction, groups to which only one end of the crosslinking agent is attached, and remaining crosslinking agents are preferably removed from the viewpoint of safety, stability during storage, and side reactions with the encapsulated drug. Although not particularly limited, for example, if unreacted crosslinking agents remain, they may be removed by washing with excess water. Also, for example, if methacryloyl groups substituted on the polymer remain, they may be removed by adding excess mercaptoethanol to deactivate the methacryloyl groups and then washing away the excess mercaptoethanol with excess water. Furthermore, for example, if mercapto groups remain, they may be removed by adding excess 3-maleimidopropionic acid, iodoacetic acid, etc. to deactivate the mercapto groups and then washing away the excess 3-maleimidopropionic acid, iodoacetic acid with excess water.

[0085] (A) A grinding step may be performed after the step of gelling the hyaluronic acid derivative. Grinding methods include grinding using a mortar and pestle or grinding using a mill, but grinding using a mill is preferred. Examples of mill grinding devices include centrifugal grinders (Nippon Seiki Seisakusho) and rotary disc type grinding devices such as impact mills (Dalton Co., Ltd.), atomizers (Tokyo Atomizer Manufacturing Co., Ltd.), sample mills (Tokyo Atomizer Manufacturing Co., Ltd.), bantam mills (Tokyo Atomizer Manufacturing Co., Ltd.), and screen mills such as SK mills (Tokken), jet grinding devices such as ultra-fine volume lab jet mills (AO jet mills, Seishin Kogyo), and Linrex mills (Liquid Gas Co., Ltd.) which can grind at ultra-low temperatures, but SK mills and Linrex mills are preferred.

[0086] (A) A drying step may be performed after the step of gelling the hyaluronic acid derivative. Examples of drying methods include forced-air drying, drying in a constant-temperature bath, reduced-pressure drying, and hot-air circulation drying. The air velocity, drying time, temperature, pressure, etc., are appropriately selected within a range that does not cause decomposition or deterioration of the gel of the hyaluronic acid derivative (A).

[0087] <Dosage> The dosage of the hyaluronic acid derivative composition in this embodiment is not limited as long as it is an amount that exerts the hair growth or hair restoration effect of the hyaluronic acid derivative composition. For example, it can be an amount that is just enough for the hyaluronic acid derivative composition to come into contact with the application site. The application site here refers to the area where hair growth or hair restoration is desired. The number of doses and frequency of administration are not particularly limited as long as the hair growth promoting effect or hair restoration promoting effect of the hyaluronic acid derivative composition of this embodiment is exerted, but for example, it can be 1, 2, 3, 4, or 5 times a day, or once every 2 days, once every 3 days, once every 4 days, once every 5 days, once every 6 days, once a week, etc.

[0088] <Administration Method> The dosage form of the hyaluronic acid derivative composition of this embodiment is not particularly limited as long as it can be applied to the area where hair growth or hair restoration is targeted, and examples include liquids, powders, gels, creams, etc. Liquids also include suspensions.

[0089] The method of administering the hyaluronic acid derivative composition of this embodiment is not particularly limited, but examples include directly applying the hyaluronic acid derivative composition of this embodiment to the application site of the target animal, or administering the hyaluronic acid derivative composition of this embodiment using a dispensing container without the practitioner directly touching the application site. The dispensing container is not particularly limited as long as it can dispensing the hyaluronic acid derivative composition of this embodiment, but examples include eye dropper containers and spray containers. Alternatively, the hyaluronic acid derivative composition of this embodiment may be patted onto the application site of the target animal. The strength and frequency of patting can be appropriately adjusted depending on the condition of the application site.

[0090] The hyaluronic acid derivative composition of this embodiment can be provided as a pharmaceutical product, a quasi-drug, or a cosmetic product.

[0091] <Treatment method> In one embodiment, the present invention provides a method for treating or preventing alopecia, comprising administering the above-described hyaluronic acid derivative composition to the application site of a target animal. In this case, the above-described hyaluronic acid derivative composition may also be referred to as a pharmaceutical composition for the treatment or prevention of alopecia. The method of administration may be the same as the method exemplified in the "method of administration" above.

[0092] Examples of alopecia include, but are not limited to, wound-induced alopecia, senile alopecia, stress-induced alopecia, male pattern baldness, female pattern baldness, scarring alopecia, drug-induced alopecia, and postpartum alopecia.

[0093] In one embodiment, the present invention provides a method for promoting hair growth or hair development in a target animal, comprising administering the above-described hyaluronic acid derivative composition to the application site of the target animal. The administration method may be the same as the method exemplified in the "Administration Method" above. [Examples]

[0094] The present invention will be described in detail below with reference to examples, but these are not intended to limit the scope of the present invention to these examples.

[0095] [Manufacturing Example 1] (Manufacturing of hyaluronic acid derivative HA-a1) Hyaluronic acid derivatives were prepared according to the following steps 1 to 3.

[0096] 1.Process 1 (Synthesis of cholesteryl 6-aminohexylcarbamate hydrochloride) Cholesteryl 6-aminohexylcarbamate hydrochloride (Chol hydrochloride) was synthesized according to the following steps 1-1, followed by step 1-2.

[0097] (1) Process 1-1 To a solution of cholesteryl chloroformate (3.37 g, 7.5 mmol) in anhydrous dichloromethane (20 mL), triethylamine (TEA, 1.05 mL) was added under an argon atmosphere and stirred. Under ice cooling, 6-(t-butoxycarbonyl)amino-1-aminohexane (1.12 mL, 5 mmol) was added dropwise, and the mixture was stirred under ice cooling for 30 minutes. After raising the temperature to room temperature, the mixture was stirred overnight. The reaction mixture was washed with ultrapure water and saturated brine, dried over anhydrous magnesium sulfate, and the solvent was removed under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: ethyl acetate:n-hexane = 1:4), and the fractions of the target product were combined and the solvent was removed under reduced pressure.

[0098] (2) Process 1-2 The obtained residue was dissolved in ethyl acetate (40 mL), and 40 mL of 4N hydrochloric acid / ethyl acetate solution was added and the mixture was stirred overnight at room temperature. The resulting precipitate was collected by centrifugation. The obtained solid was washed four times with ethyl acetate and dried under reduced pressure to obtain 1.2 g of cholesteryl 6-aminohexylcarbamate hydrochloride (Chol hydrochloride).

[0099] 2.Process 2 (Preparation of tetrabutylammonium (TBA) salt of hyaluronic acid) Hyaluronic acid TBA salt (HA-TBA) was prepared according to the following steps 2-1 and then 2-2.

[0100] (1) Process 2-1 DOWEX® 50WX-8-400 (manufactured by Aldrich) was suspended in ultrapure water, and the resin was washed approximately three times with ultrapure water by decantation. Approximately 1.5 times the molar equivalent of 40 wt% tetrabutylammonium hydroxide aqueous solution (TBA-OH) (manufactured by Aldrich) was added relative to the cation exchange capacity of the resin, and the mixture was stirred for 30 minutes. After removing the excess TBA-OH solution by decantation, the mixture was further washed with excess ultrapure water to obtain a TBA-chlorinated cation exchange resin.

[0101] (2) Process 2-2 Sodium hyaluronate salt (HA-Na) with a molecular weight of 53,000 (53 kDa) was dissolved in ultrapure water at a concentration of 15 mg / mL. The suspension of cation exchange resin, which had been TBA-chlorinated in "(1) Step 2-1", was added in an amount equivalent to 5 times the molar amount of HA units (unit molecular weight 401.3) in terms of the resin's ion exchange capacity. After stirring for 15 minutes, the mixture was filtered using a 0.45 μm filter, and the filtrate was freeze-dried to obtain hyaluronic acid TBA salt (HA-TBA) as a white solid.

[0102] 3.Process 3 An anhydrous DMSO solution (10 mg / mL) of HA-TBA prepared in "2.(2) Step 2-2" was prepared. Then, Chol hydrochloride was added in a molar ratio of 16 / 100 relative to the disaccharide repeating units (HA units) present in the HA-TBA synthesized in "1. Step 1". Next, 4-(4,6-dimethoxy-1,3,5-triazine-2-yl)-4-methylmorpholinium chloride (DMT-MM) was added in a molar ratio of 21 / 100 relative to the HA units, and the mixture was stirred overnight at room temperature. The reaction solution was dialyzed in the following order: 0.3 M ammonium acetate / DMSO solution, 0.15 M NaCl aqueous solution, and ultrapure water (Spectrapore 7, molecular weight cutoff (MWCO): 2,000). The resulting dialysate was freeze-dried to obtain the target product (HA-C6-Chol) as a white solid. 1 In the 1H-NMR spectrum, peaks originating from the acetyl group of N-acetyl-D-glucosamine (COCH3, 1.6 ppm to 2.0 ppm, 3H) and peaks originating from the methyl group in the cholesteryl group (CH3, 0.7 ppm, 3H) were observed, and the cholesterol introduction rate was 15%.

[0103] [Example 1] (Manufacturing of hyaluronic acid derivative compositions) The obtained hyaluronic acid derivative HA-a1 was dissolved in water to a concentration of 15 mg / mL. In a separate container, minoxidil (Tokyo Chemical Industries, Ltd.) was dissolved in water to a concentration of 3 mg / mL. By mixing the aqueous solution of hyaluronic acid derivative HA-a1 and the aqueous solution of minoxidil in a ratio of 2:1, a hyaluronic acid derivative composition containing the hyaluronic acid derivative-minoxidil complex HA-a1-Mino was obtained. The content of the hyaluronic acid derivative was 1 part by mass and the content of minoxidil was 0.1 parts by mass per 100 parts by mass of the hyaluronic acid derivative composition.

[0104] [Comparative Example 1] Minoxidil (Tokyo Chemical Industries, Ltd.) was dissolved in water to a concentration of 1 mg / mL to obtain an aqueous minoxidil solution.

[0105] [Test Example 1] (Test to confirm hair regeneration effect in mouse skin) A test was conducted to confirm the hair regeneration effect on mouse skin using the hyaluronic acid derivative composition obtained in Example 1 and the minoxidil aqueous solution obtained in Comparative Example 1.

[0106] As the animals used, mice that were between 51 and 53 days old at the time of administration were used. Two to three days before the start of administration, the fur on the back of each animal was trimmed using household clippers (DoggyMan Hayashi Co., Ltd.) in a longitudinal direction from the tail ridge to an area of ​​approximately 5 cm and a width of approximately 3 cm, and the hair was removed using a razor (Kai Corporation, Nagae Gold α) to prepare the administration site.

[0107] For all cases in which administration was performed, 0.1 mL of the hyaluronic acid derivative composition obtained in Example 1 or the minoxidil aqueous solution obtained in Comparative Example 1 was dropped onto the back of each animal using a variable continuous dispenser (Nichiryo Co., Ltd. MODEL8100). Then, the solution was applied by moving a finger wearing a finger cot (Sankyo Chemical Industry Co., Ltd., natural rubber powder-free finger cot, Class 1000 compliant) back and forth approximately 5 to 7 times along the body axis. Administration was performed once a day for 18 days, generally in the morning.

[0108] On the administration start date (the first day from the start of administration), the 5th day, the 7th day, the 9th day, the 11th day, the 13th day, the 15th day, the 17th day, and the day after the final administration (the 19th day from the start of administration), the within the evaluation site (about 3 cm × 5 cm) was judged by the hair regeneration score shown in Table 1, and photos showing the hair growth state were taken under certain conditions. The evaluation results are shown in Table 2. In Table 2, the average values for 10 mice each were calculated.

[0109]

Table 1

[0110]

Table 2

[0111] As shown in Table 2, hair growth was promoted in the mouse group to which the hyaluronic acid derivative composition obtained in Example 1 was applied, compared with the mouse group to which the minoxidil aqueous solution obtained in Comparative Example 1 was applied.

[0112] [Example 2] (Production of hyaluronic acid derivative composition) The obtained hyaluronic acid derivative HA-a1 was dissolved in water to a concentration of 12.5 mg / mL. In a separate container, finasteride (Tokyo Chemical Industry Co., Ltd.) was dissolved in ethanol to a concentration of 16.7 mg / mL. By mixing the hyaluronic acid derivative HA-a1 aqueous solution and the finasteride solution at a ratio of 2:3, a hyaluronic acid derivative composition containing the hyaluronic acid derivative-finasteride complex HA-a1-Fina was obtained. The content of the hyaluronic acid derivative was 0.5 parts by mass, the content of finasteride was 1 part by mass, and the content of ethanol was 60 parts by mass with respect to 100 parts by mass of the hyaluronic acid derivative composition.

[0113] [Comparative Example 2] Finasteride (Tokyo Chemical Industry Co., Ltd.) was dissolved in ethanol to a concentration of 16.7 mg / mL. Then, 12 parts by mass of Otsuka Water for Injection were mixed with 18 parts by mass of this ethanol solution of finasteride to obtain a finasteride solution in which the content of finasteride in 100 parts by mass of the test solution was 1 part by mass.

[0114] [Test Example 2] (Test for Confirming Hair Regrowth Effect on Mouse Skin) Using the hyaluronic acid derivative composition obtained in Example 2 and the finasteride solution obtained in Comparative Example 2, a test was conducted to confirm the hair regrowth effect on mouse skin.

[0115] As animals, mice at 8 weeks of age at the time of administration were used. From 2 days before to 3 days before the start of administration, the back hair of each animal was shaved longitudinally from the base of the tail to a width of about 4 cm and a width of about 2.5 cm using a household clipper (dog clipper, Panasonic Corporation), and a depilation cream (Epilat, Kracie Home Products Co., Ltd.) was used to prepare the administration site.

[0116] For all cases of administration, using a variable volume continuous dispenser (Nichijo Co., Ltd. MODEL8100), 0.1 mL of the hyaluronic acid derivative composition obtained in Example 2 or the finasteride solution obtained in Comparative Example 2 was dropped onto the back of each animal. Then, it was applied by reciprocating about 5 to 7 times in the body axis direction with a finger wearing a finger cot (Sankyo Chemical Industry Co., Ltd., natural rubber powder-free finger cot, corresponding to Class 1000). The administration was performed twice a day. For the first administration, a 0.5% testosterone solution was applied. Then, after 15 minutes, the second administration was the hyaluronic acid derivative composition obtained in Example 2 or the finasteride solution obtained in Comparative Example 2. The administration was performed in the morning.

[0117] Observation continued from day 1 to day 26 of administration until hair regrowth was observed in at least one mouse. On day 1, day 26 of administration, and every two days from the time hair regrowth was confirmed, the hair regrowth score within the evaluation area (approximately 2.5 × 4 cm) was assessed using the hair regrowth score shown in Table 3, and photographs showing the hair regrowth status were taken under specific conditions. The evaluation results are shown in Table 4. In Table 4, the average value for 10 mice was calculated for each case.

[0118] [Table 3]

[0119] [Table 4]

[0120] As shown in Table 4, hair growth was promoted earlier in the group of mice treated with the hyaluronic acid derivative composition obtained in Example 2 than in the group of mice treated with the finasteride solution obtained in Comparative Example 2, starting from the beginning of treatment.

[0121] [Manufacturing Example 2] (Manufacturing of hyaluronic acid derivative HA-a1L) Labeled hyaluronic acid derivative HA-a1L was produced using the same method as in Production Example 1, except that in "Step 3" of Production Example 1, 5-aminofluorescein was added in a molar ratio of 4 / 100 before adding Chol hydrochloride, and then DMT-MM was added in a molar ratio of 6 / 100 relative to the HA unit, and the mixture was stirred overnight at room temperature.

[0122] [Test Example 3] (Penetration test into hair follicles in mouse skin) Using the labeled hyaluronic acid derivative HA-a1L obtained in Production Example 2, a penetration test into hair follicles was conducted in mouse skin.

[0123] Specifically, first, labeled hyaluronic acid derivative HA-a1L was dissolved in water to a concentration of 10 mg / mL, and this aqueous solution was applied to the skin of hairless mice and left to stand for 12 hours. After that, the solution was wiped off, and the skin with the solution applied was collected. The collected skin sample was cryopreserved using OCT Compound (Sakura FineTech Japan), and then sections were prepared. Figure 1 shows the results of observing the sections with a fluorescence microscope (magnification: 25x).

[0124] As shown in Figure 1, fluorescence was observed in the pores, confirming the penetration of the labeled hyaluronic acid derivative HA-a1L into the pores. [Industrial applicability]

[0125] The hyaluronic acid derivative composition of this embodiment has excellent hair growth promoting or hair restoration effects and is suitable for use as a hair growth agent or hair restoration agent.

Claims

1. (A) Hyaluronic acid derivatives into which steryl groups have been introduced, (B) Ingredients that promote hair growth or hair development, Includes, The aforementioned (B) ingredient that promotes hair growth or hair development is minoxidil, finasteride, or ethinylestradiol. A hyaluronic acid derivative composition for direct application to the area of ​​a target animal where hair growth or hair restoration is desired, wherein the hyaluronic acid derivative into which the steryl group (A) is introduced has a repeating unit represented by the following general formula (I). 【Chemistry 1】 (In the formula, R1, R2, R3, and R4 are each independently selected from the group consisting of a hydrogen atom, C1-6 alkyl, formyl, and C1-6 alkylcarbonyl; Z represents a direct linker or a peptide linker consisting of any 2 to 30 amino acid residues; X1 is given by the following formula: -NR b -R, -NR b -COO-R, -NR b -CO-R, -NR b -CO-NR c -R, -COO-R, -O-COO-R, -S-R, -CO-Y a -SR, -O-CO-Y b -SR, -NR b -CO-Y b -S-R, and -S-S-R, It is a group selected from the group consisting of groups represented by; R a, R b, and R c are each independently selected from the group consisting of a hydrogen atom, C1-20 alkyl, aminoC2-20 alkyl, and hydroxyC2-20 alkyl, where the alkyl portion of the group may have a group selected from the group consisting of -O- and -NR f- inserted; R f is selected from the group consisting of a hydrogen atom, C1-12 alkyl, aminoC2-12 alkyl, and hydroxyC2-12 alkyl, and the alkyl portion of the group may have a group selected from the group consisting of -O- and -NH- inserted; R is a steryl group; Y is C2-30 alkylene, or -(CH2CH2O)m-CH2CH2-, where the alkylene may have a group selected from the group consisting of -O-, -NRg-, and -S-S- inserted; R g is selected from the group consisting of a hydrogen atom, C1-20 alkyl, aminoC2-20 alkyl, and hydroxyC2-20 alkyl, and the alkyl portion of the group may have a group selected from the group consisting of -O- and -NH- inserted; Y a is a C1-5 alkylene; Yb is a C2-8 alkylene or C2-8 alkenylene; m is an integer between 1 and 100 (inclusive).

2. The hyaluronic acid derivative composition according to claim 1, wherein the content of the hyaluronic acid derivative into which the steryl group is introduced (A) is 0.01 parts by mass or more and 10 parts by mass or less per 100 parts by mass of the hyaluronic acid derivative composition.

3. The hyaluronic acid derivative composition according to claim 1 or 2, wherein the content of the ingredient that promotes hair growth or hair development (B) is 0.01 parts by mass or more and 40 parts by mass or less per 100 parts by mass of the hyaluronic acid derivative composition.

4. The hyaluronic acid derivative composition according to any one of claims 1 to 3, wherein the ratio of the mass of the hair growth or hair development component (B) to the mass of the hyaluronic acid derivative into which the steryl group has been introduced is 0.01% or more and 10,000% or less.

5. The hyaluronic acid derivative composition according to any one of claims 1 to 4, wherein the rate of introduction of the steryl group into the hyaluronic acid derivative into which the steryl group has been introduced is 1% or more and less than 60%.

6. The hyaluronic acid derivative composition according to any one of claims 1 to 5, wherein the molecular weight of the hyaluronic acid derivative into which the steryl group (A) is introduced is 1,000 or more and less than 1,000,000.

7. The hyaluronic acid derivative composition according to any one of claims 1 to 6, wherein R is a cholesteryl group.

8. A hyaluronic acid derivative composition according to any one of claims 1 to 7, which is a hair growth agent composition or a hair tonic composition.

Images