JP7773175B2 - Amyloid-β aggregation inhibitor - Google Patents

Description

Patent Act Article 30, Paragraph 2 applies. Publication name: "Journal of the Japanese Society of Neurology, Vol. 60, Special Issue, 2020, 60:S476, Pe-12-3" Publication date: August 7, 2020. Publication name: "Current Alzheimer Research, 2020, Vol. 17, Issue 17, pp. 589-600" Publication date: November 16, 2020.

The present invention relates to an amyloid-β aggregation inhibitor.

Alzheimer's disease is a type of irreversible, progressive dementia that is more likely to occur in the elderly. The number of Alzheimer's disease patients is extremely high, and attention is being focused on the development of therapeutic drugs for the disease.

Cholinesterase inhibitors (such as Aricept (see Patent Document 1) and Reminyl) are examples of drugs used in clinical settings to treat Alzheimer's disease. Cholinesterase inhibitors increase the amount of acetylcholine in the brain by inhibiting the breakdown of the neurotransmitter acetylcholine, thereby slowing the progression of the symptoms of Alzheimer's disease.

However, while cholinesterase inhibitors can slow the progression of Alzheimer's disease symptoms, they do not act on the underlying mechanism of Alzheimer's disease, and therefore cannot serve as a fundamental treatment for the disease. For this reason, efforts are underway to elucidate the mechanism of Alzheimer's disease onset and to develop drugs based on this mechanism.

Clinical research has revealed that while amyloid beta protein is harmless on its own, the amyloid fibrils formed by the aggregation of amyloid beta protein kill nerve cells in the brain, resulting in the onset of Alzheimer's disease. The "amyloid hypothesis," which posits that the aggregation of amyloid beta protein is important for the onset of Alzheimer's disease, is currently an important hypothesis for explaining the mechanism behind the onset of Alzheimer's disease.

Based on the amyloid hypothesis, drugs that prevent the production of amyloid β protein and drugs that remove amyloid β protein have been developed and _clinically tested. Examples of such drugs include R-flurbiprofen, LY2811376, _{H2NCH2CH2CH2SO3H} , tramiprosate, and semagacestat ( _see , for example, Patent Document ₂ ) _.

Japanese Patent Application Publication No. 1-79151 WO02/40451A2 No.

However, epidemiological studies at the clinical trial stage have statistically shown that drugs based on the conventional amyloid hypothesis are only minimally effective in slowing the progression of Alzheimer's disease symptoms and/or increase the risk of other diseases. Furthermore, data has shown that some of these drugs inhibit the function of normal amyloid beta protein. Therefore, there is a need for the development of new drugs based on the amyloid hypothesis.

One aspect of the present invention aims to provide a novel amyloid β aggregation inhibitor that can suppress the aggregation of amyloid β protein.

[1] An amyloid-β aggregation inhibitor according to one embodiment of the present invention contains, as an active ingredient, a phthalocyanine represented by the following formula 1 or 2, or a salt thereof:

(In formula 1,
^M1 is any metal or any metal having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand; or Formula 1 does not include ^M1 ,
each R ¹ is independently a nonionic substituent, a cationic substituent, an anionic substituent, or a zwitterionic substituent, and at least one R ¹ is a cationic substituent, an anionic substituent, or a zwitterionic substituent);

(In Equation 2,
^M2 is any metal or any metal having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand; or Formula 2 does not include ^M2 ,
each ^R2 is independently a nonionic substituent, a cationic substituent, an anionic substituent, or a zwitterionic substituent, and at least one ^R2 is a cationic substituent, an anionic substituent, or a zwitterionic substituent;
However, all of the eight ^R2s are the following structure 1 or structure 2:

Except that they are the same anionic substituents, including

[2] In one embodiment of the amyloid-β aggregation inhibitor of the present invention, the cationic substituent preferably includes the following structure:

[3] In one embodiment of the amyloid-β aggregation inhibitor of the present invention, the anionic substituent preferably includes the following structure:

[4] In one embodiment of the amyloid-β aggregation inhibitor of the present invention, the zwitterionic substituent preferably includes the following structure:

[5] In the amyloid-β aggregation inhibitor according to one embodiment of the present invention, the metal constituting ^M1 or ^M2 is preferably Li, Na, K, Mg, Ti, Zr, V, Nb, Ta, Cr, Mo, W, Mn, Fe, Co, Ni, Ru, Rh, Pd, Os, Ir, Pt, Cu, Ag, Au, Zn, Cd, Hg, Al, Ga, In, Si, Ge, Sn, Pb, Sb, or Bi.

[6] In the amyloid β aggregation inhibitor according to one aspect of the present invention, the salt is preferably a salt of Na ⁺ , H ⁺ , K ⁺ , NH ⁴⁺ , or a halide ion.

One aspect of the present invention makes it possible to realize a novel amyloid β aggregation inhibitor that can suppress the aggregation of amyloid β protein.

1 is a graph showing the amyloid β protein aggregation inhibitory activity of compound (1) according to an example of the present invention. 1 is a graph showing the amyloid β protein aggregation inhibitory activity of compound (2) according to an example of the present invention. 1 is a graph showing the amyloid β protein aggregation inhibitory activity of compound (3) according to an example of the present invention. 1 is a graph showing the amyloid β protein aggregation inhibitory activity of compound (4) according to an example of the present invention. 1 is a graph showing the amyloid β protein aggregation inhibitory activity of compound (5) according to an example of the present invention. 1 is a graph showing the amyloid β protein aggregation inhibitory activity of compound (6) according to an example of the present invention. 1 is a graph showing a comparison of the amyloid β protein aggregation inhibitory ability of compounds according to examples of the present invention.

One embodiment of the present invention is described below, but the present invention is not limited to this. The present invention is not limited to the configurations described below, and various modifications are possible within the scope of the claims. Embodiments and examples obtained by appropriately combining the technical means disclosed in different embodiments and examples are also included in the technical scope of the present invention. Furthermore, all documents described in this specification are incorporated herein by reference. In this specification, when a numerical range is described as "A to B," this means "greater than or equal to A and less than or equal to B."

1. Overview of amyloid β aggregation inhibitors
An amyloid β (amyloid β protein) aggregation inhibitor according to one embodiment of the present invention can be used as a therapeutic agent for Alzheimer's disease or a preventive agent for Alzheimer's disease.

As used herein, the term "therapeutic agent" refers to a drug that provides a therapeutic effect. Examples of such therapeutic effects include, but are not limited to, the effects listed below.

(1) When the therapeutic agent is administered, the severity of one or more symptoms associated with Alzheimer's disease is reduced compared to when the therapeutic agent is not administered.

(2) When the therapeutic agent is administered, the increase in severity or progression of severity of one or more symptoms associated with Alzheimer's disease is prevented compared to when the therapeutic agent is not administered.

(3) When the therapeutic drug is administered, the rate of increase in severity or the rate of progression of severity of one or more symptoms associated with Alzheimer's disease is reduced compared to when the therapeutic drug is not administered.

As used herein, the term "prophylactic agent" refers to a drug that provides a preventive effect. The preventive effect includes, but is not limited to, the effects exemplified below.

(4) When the prophylactic drug is administered, the onset of one or more symptoms associated with Alzheimer's disease is prevented or the risk of onset is reduced compared to when the prophylactic drug is not administered.

(5) When the prophylactic drug is administered, the recurrence of one or more symptoms associated with Alzheimer's disease is prevented or the risk of recurrence is reduced compared to when the prophylactic drug is not administered.

(6) Administration of the prophylactic drug prevents the onset of, or reduces the risk of the onset of, one or more symptoms of Alzheimer's disease compared to when the prophylactic drug is not administered.

2. Amyloid β Aggregation Inhibitors
An amyloid-β aggregation inhibitor according to one embodiment of the present invention contains, as an active ingredient, a phthalocyanine represented by the following formula 1 or 2, or a salt thereof:

(In formula 1,
^M1 is any metal or any metal having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand; or Formula 1 does not include ^M1 ,
each R ¹ is independently a nonionic substituent, a cationic substituent, an anionic substituent, or a zwitterionic substituent, and at least one R ¹ is a cationic substituent, an anionic substituent, or a zwitterionic substituent);

(In Equation 2,
^M2 is any metal or any metal having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand; or Formula 2 does not include ^M2 ,
each ^R2 is independently a nonionic substituent, a cationic substituent, an anionic substituent, or a zwitterionic substituent, and at least one ^R2 is a cationic substituent, an anionic substituent, or a zwitterionic substituent;
However, all of the eight ^R2s are the following structure 1 or structure 2:

Except that they are the same anionic substituents, including

(2-1. Phthalocyanines or Salts Thereof)
Phthalocyanines are cyclic compounds with an 18-pi electron system. Various metals or metal ions can be inserted into the center of the phthalocyanine ring. By inserting a metal or metal ion into the center of the ring, phthalocyanines can become highly stable planar complexes.

In Formula 1 and Formula 2, ^M1 and ^M2 can be any metal or any metal having a halide ion, carboxylate, sulfate ion, nitrate ion, acetylacetonate, or carbonyl compound as an axial ligand. The phthalocyanines and salts thereof represented by Formula 1 and Formula 2 do not necessarily contain ^M1 and ^M2 . When the phthalocyanines and salts thereof represented by Formula 1 and Formula 2 contain ^M1 and ^M2 , the phthalocyanines and salts thereof can become highly stable planar complexes.

The metal constituting ^M1 or ^M2 is preferably Li, Na, K, Mg, Ti, Zr, V, Nb, Ta, Cr, Mo, W, Mn, Fe, Co, Ni, Ru, Rh, Pd, Os, Ir, Pt, Cu, Ag, Au, Zn, Cd, Hg, Al, Ga, In, Si, Ge, Sn, Pb, Sb, or Bi. According to this configuration, an amyloid β aggregation inhibitory effect can be obtained. From the viewpoint of obtaining a higher amyloid β aggregation inhibitory effect, the metal constituting ^M1 and ^M2 is preferably Zn or Pd, and more preferably Fe, among the above-mentioned metals.

The metal constituting ^M1 and ^M2 may be any metal having a halide ion, carboxylate, sulfate ion, nitrate ion, acetylacetonate, or carbonyl compound as an axial ligand. The axial ligand is a ligand located above and/or below the plane connecting the positions of the atoms constituting the phthalocyanines, and is a ligand coordinated to the metal inserted at the center of the phthalocyanine ring. From the viewpoint of obtaining a higher amyloid-β aggregation inhibitory effect, among these six axial ligands, acetylacetonate or carbonyl compound is preferred, and halide ion is more preferred.

The halide ions (F ⁻ , Cl ⁻ , Br ⁻ , I ⁻ , At ⁻ ) are not limited, but from the viewpoint of obtaining a higher inhibitory effect on amyloid β aggregation, Br ⁻ or I ⁻ is preferred, and Cl ⁻ is more preferred.

The carbonyl compound is not limited to, and examples thereof include, but are not limited to, aldehydes, carboxylic acids, esters, and amides. From the viewpoint of achieving a higher inhibitory effect on amyloid-β aggregation, among these carbonyl compounds, amides or aldehydes are preferred, and carboxylic acids are more preferred.

In Formula 1 and Formula 2, ^R1 and ^R2 can each independently be a nonionic substituent, a cationic substituent, an anionic substituent, or a zwitterionic substituent, and at least one (or at least two, at least three, at least four, at least five, at least six, at least seven, or eight) of ^R1 and ^R2 can be a cationic substituent, an anionic substituent, or a zwitterionic substituent.

From the viewpoint of obtaining a high inhibitory effect on amyloid β aggregation and/or from the viewpoint of easily synthesizing or obtaining phthalocyanines and salts thereof, it is preferred that four or eight of R ¹ and R ² in Formula 1 and Formula 2 are cationic substituents, anionic substituents, or zwitterionic substituents.

In Formula 1 and Formula 2, two ^R1 and ^R2 are bonded to each of the four six-membered rings. In Formula 1 and Formula 2, when at least four ^R1 and ^R2 are cationic substituents, anionic substituents, or zwitterionic substituents, it is preferred that at least one cationic substituent, anionic substituent, or zwitterionic substituent is bonded to each of the four six-membered rings. Furthermore, when at least one cationic substituent, anionic substituent, or zwitterionic substituent is bonded to each of the four six-membered rings, it is preferred that a cationic substituent, anionic substituent, or zwitterionic substituent is bonded to a carbon atom located at the same position on the four six-membered rings. More specifically, when at least one cationic substituent, anionic substituent, or zwitterionic substituent is bonded to each of the four six-membered rings, it is preferred that the structures of the four six-membered rings including ^R1 or ^R2 are the same. This configuration is advantageous in that it makes it possible to realize an amyloid β aggregation inhibitor that has a high amyloid β aggregation inhibitory effect, and/or it makes it possible to easily synthesize or obtain phthalocyanines and salts thereof.

The nonionic substituent is intended to be a substituent that does not have an electric charge. The nonionic substituent is not particularly limited, and examples thereof include H, polyethylene glycol, and crown ether. Among these nonionic substituents, polyethylene glycol and crown ether are preferred due to their advantageous affinity for brain cell membranes.

The cationic substituent is intended to be a substituent having a positive charge (e.g., N ⁺ ). The cationic substituent according to one embodiment of the present invention is not particularly limited, but is preferably a substituent having the following structure. With this structure, it is possible to realize an amyloid-β aggregation inhibitor having a high amyloid-β aggregation inhibitory effect.

The cationic substituent according to one embodiment of the present invention may be a substituent containing the above structure, but preferably contains O, S, or Se as another structure (for example, the structure corresponding to the wavy line on the left side of the above structure). This structure allows the cationic substituent to be easily bonded to phthalocyanines.

The anionic substituent is intended to be a substituent having a negative charge (for example, O ⁻ ). The anionic substituent according to one embodiment of the present invention is not particularly limited, but is preferably a substituent having the following structure. With this structure, it is possible to realize an amyloid β aggregation inhibitor having a high amyloid β aggregation inhibitory effect.

The anionic substituent according to one embodiment of the present invention may be a substituent containing the structure shown above, but preferably contains O, S, or Se as another structure (for example, the structure corresponding to the wavy line on the left side of the structure shown above). This structure allows the anionic substituent to be easily bonded to phthalocyanines.

The zwitterionic substituent is intended to be a substituent having both a positive charge (e.g., N ⁺ ) and a negative charge (e.g., O ⁻ ). With this configuration, it is possible to realize an amyloid β aggregation inhibitor that has a high amyloid β aggregation inhibitory effect. The zwitterionic substituent according to one embodiment of the present invention is not particularly limited, but is preferably a substituent having the following structure:

The dipolar substituent according to one embodiment of the present invention may be a substituent containing the structure shown above, but preferably contains O, S, or Se as another structure (for example, the structure corresponding to the wavy line on the left side of the structure shown above). This structure allows the dipolar substituent to be easily bonded to phthalocyanines.

The phthalocyanine according to one embodiment of the present invention may exist as a salt. The salt is not particularly limited, but is preferably a salt of Na ⁺ , H ⁺ , K ⁺ , NH ⁴⁺ , or a halide ion (F ^- , Cl ^- , Br ^- , I ^- , At ^- ). With this configuration, an amyloid β aggregation inhibitor having a high amyloid β aggregation inhibitory effect can be realized.

The amount of active ingredient in an amyloid-β aggregation inhibitor according to one embodiment of the present invention is not particularly limited, and may be, for example, 0.001% to 100% by weight, 0.01% to 100% by weight, 0.1% to 100% by weight, 0.1% to 95% by weight, 0.1% to 90% by weight, 0.1% to 80% by weight, 0.1% to 70% by weight, 0.1% to 60% by weight, 0.1% to 50% by weight, 0.1% to 40% by weight, 0.1% to 30% by weight, 0.1% to 20% by weight, or 0.1% to 10% by weight, relative to the amyloid-β aggregation inhibitor.

The amyloid-β aggregation inhibitor may contain a component other than the above-mentioned phthalocyanines or salts thereof, as long as the component does not inhibit the amyloid-β aggregation inhibitory ability.

These different components may include buffers, pH adjusters, preservatives, antioxidants, isotonicity agents, high molecular weight polymers, excipients, carriers, diluents, solvents, solubilizers, stabilizers, fillers, binders, and surfactants.

Examples of buffering agents include phosphoric acid, phosphate salts, boric acid, borates, citric acid, citrate salts, acetic acid, acetate salts, carbonic acid, carbonate salts, tartaric acid, tartrate salts, ε-aminocaproic acid, and trometamol. Phosphates include sodium phosphate, sodium dihydrogen phosphate, disodium hydrogen phosphate, potassium phosphate, potassium dihydrogen phosphate, and dipotassium hydrogen phosphate. Borates include borax, sodium borate, and potassium borate. Citrates include sodium citrate, disodium citrate, and trisodium citrate. Acetates include sodium acetate and potassium acetate. Carbonates include sodium carbonate and sodium bicarbonate. Tartrates include sodium tartrate and potassium tartrate.

Examples of pH adjusters include hydrochloric acid, phosphoric acid, citric acid, acetic acid, sodium hydroxide, and potassium hydroxide.

Examples of the isotonicity agent include ionic isotonicity agents (such as sodium chloride, potassium chloride, calcium chloride, and magnesium chloride) and non-ionic isotonicity agents (such as glycerin, propylene glycol, sorbitol, and mannitol).

Examples of such preservatives include benzalkonium chloride, benzalkonium bromide, benzethonium chloride, sorbic acid, potassium sorbate, methyl parahydroxybenzoate, propyl parahydroxybenzoate, and chlorobutanol.

Examples of antioxidants include ascorbic acid, tocopherol, dibutylhydroxytoluene, butylhydroxyanisole, sodium erythorbate, propyl gallate, and sodium sulfite.

Examples of such high molecular weight polymers include methyl cellulose, ethyl cellulose, hydroxymethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxyethyl methyl cellulose, hydroxypropyl methyl cellulose, carboxymethyl cellulose, sodium carboxymethyl cellulose, hydroxypropyl methyl cellulose acetate succinate, hydroxypropyl methyl cellulose phthalate, carboxymethyl ethyl cellulose, cellulose acetate phthalate, polyvinylpyrrolidone, polyvinyl alcohol, carboxyvinyl polymer, polyethylene glycol, and atelocollagen.

The amount of ingredients other than the active ingredient in an amyloid-β aggregation inhibitor according to one embodiment of the present invention is not particularly limited, and may be, for example, 0% to 99.999% by weight, 0% to 99.99% by weight, 0% to 99.99% by weight, 5% to 99.9% by weight, 10% to 99.9% by weight, 20% to 99.9% by weight, 30% to 99.9% by weight, 40% to 99.9% by weight, 50% to 99.9% by weight, 60% to 99.9% by weight, 70% to 99.9% by weight, 80% to 99.9% by weight, or 90% to 99.9% by weight, relative to the amyloid-β aggregation inhibitor.

An amyloid-β aggregation inhibitor according to one embodiment of the present invention will help reduce the threat of Alzheimer's disease, thereby contributing to the achievement of the Sustainable Development Goals (SDGs).

The amyloid-β aggregation inhibitor according to one embodiment of the present invention can be produced using a commercially available active ingredient. The amyloid-β aggregation inhibitor according to one embodiment of the present invention can also be produced by synthesizing the active ingredient and then using the synthesized active ingredient. When synthesizing the active ingredient, the active ingredient may be synthesized according to methods described in, for example, "S. Tabassum, A. Nagai, et al. FEBS J, 2015, 282, 463-476." and "R. Fujishiro, T. Ikeue, et al. J. InorgBiochem, 2019, 192, 7-16."

[3. Other]
The present invention can also be configured as follows:
<1> A method for inhibiting amyloid β aggregation, comprising a step of administering to a subject (e.g., a human or a non-human animal) an amyloid β aggregation inhibitor containing, as an active ingredient, a phthalocyanine represented by the following formula 1 or 2, or a salt thereof:

(In formula 1,
^M1 is any metal or any metal having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand; or Formula 1 does not include ^M1 ,
each R ¹ is independently a nonionic substituent, a cationic substituent, an anionic substituent, or a zwitterionic substituent, and at least one R ¹ is a cationic substituent, an anionic substituent, or a zwitterionic substituent);

(In Equation 2,
^M2 is any metal or any metal having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand; or Formula 2 does not include ^M2 ,
each ^R2 is independently a nonionic substituent, a cationic substituent, an anionic substituent, or a zwitterionic substituent, and at least one ^R2 is a cationic substituent, an anionic substituent, or a zwitterionic substituent;
However, all of the eight ^R2s are the following structure 1 or structure 2:

Except that they are the same anionic substituents, including

<2> The method for inhibiting amyloid-β aggregation described in <1>, wherein the cationic substituent has the following structure:

<3> The method for inhibiting amyloid-β aggregation according to claim 1 or 2, wherein the anionic substituent comprises the following structure:

<4> The method for inhibiting amyloid-β aggregation according to any one of <1> to <3>, wherein the zwitterionic substituent includes the following structure:

<5> The method for inhibiting amyloid-β aggregation according to any one of <1> to <4>, wherein the metal constituting ^M1 or ^M2 is Li, Na, K, Mg, Ti, Zr, V, Nb, Ta, Cr, Mo, W, Mn, Fe, Co, Ni, Ru, Rh, Pd, Os, Ir, Pt, Cu, Ag, Au, Zn, Cd, Hg, Al, Ga, In, Si, Ge, Sn, Pb, Sb, or Bi.

<6> The method for inhibiting amyloid β aggregation according to any one of <1> to <5>, wherein the salt is a salt of Na ⁺ , H ⁺ , K ⁺ , NH ⁴⁺ , or a halide ion.

The above-mentioned method for inhibiting amyloid-β aggregation may be a method for treating Alzheimer's disease or a method for preventing Alzheimer's disease.

The present invention can also be configured as follows:
<7> Use of a phthalocyanine represented by the following formula 1 or formula 2, or a salt thereof, for producing an amyloid β aggregation inhibitor:

(In formula 1,
^M1 is any metal or any metal having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand; or Formula 1 does not include ^M1 ,
each R ¹ is independently a nonionic substituent, a cationic substituent, an anionic substituent, or a zwitterionic substituent, and at least one R ¹ is a cationic substituent, an anionic substituent, or a zwitterionic substituent);

(In Equation 2,
^M2 is any metal or any metal having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand; or Formula 2 does not include ^M2 ,
each ^R2 is independently a nonionic substituent, a cationic substituent, an anionic substituent, or a zwitterionic substituent, and at least one ^R2 is a cationic substituent, an anionic substituent, or a zwitterionic substituent;
However, all of the eight ^R2s are the following structure 1 or structure 2:

Except that they are the same anionic substituents, including

<8> The use according to <7>, wherein the cationic substituent has the following structure:

<9> The use according to <7> or <8>, wherein the anionic substituent has the following structure:

<10> The use according to any one of <7> to <9>, wherein the dipolar substituent has the following structure:

<11> The use according to any one of <7> to <10>, wherein the metal constituting ^M1 or ^M2 is Li, Na, K, Mg, Ti, Zr, V, Nb, Ta, Cr, Mo, W, Mn, Fe, Co, Ni, Ru, Rh, Pd, Os, Ir, Pt, Cu, Ag, Au, Zn, Cd, Hg, Al, Ga, In, Si, Ge, Sn, Pb, Sb, or Bi.

<12> The use according to any one of <7> to <11>, wherein the salt is a salt of Na ⁺ , H ⁺ , K ⁺ , NH ⁴⁺ , or a halide ion.

The present invention will be described in more detail below based on examples, but the present invention is not limited to the following examples.

[Synthesis of Compounds]
(i) Synthesis of Compound (1) Compound (1) was synthesized according to the following synthesis scheme.

A DMF solution (10 mL) containing 3-nitrophthalonitrile (0.82 g, 4.7 mmol) and 5-hydroxy-2-methylpyridine (1.0 g, 9.2 mmol) was heated and stirred at 65°C for 48 hours in the presence of _{K2CO3 (} 2.00 g). The reaction mixture was added to ice water to obtain a white precipitate. The organic layer of the precipitate was extracted using _CHCl3 . The resulting solution was concentrated under reduced pressure and then recrystallized using diethyl ether as a poor solvent to obtain a white solid (I-1) (0.47 g, 42%).
The obtained compound (I-1) (0.41 g, 1.74 mmol) was added to an n-pentanol solution (10 mL) and mixed with Zn(OAc) ₂ ·2H ₂ O (0.11 g, 0.50 mmol). Four drops of DBU were then added to the mixture, which was then heated and stirred at 145°C for 24 hours. The reaction mixture was poured into ice water to obtain a green precipitate. The organic layer was extracted from the precipitate using CHCl _3. The resulting solution was concentrated under reduced pressure and recrystallized using diethyl ether to obtain a green solid (II-1) (0.30 g, 90%). A CH ₃ I solution (5.0 mL) containing the obtained compound (II-1) (0.1 g, 0.01 mmol) was heated and stirred at 40°C for 24 hours. The resulting solution was concentrated under reduced pressure and recrystallized using acetone to obtain the target green solid (III-1) (0.13 g, 80%).
Elemental analysis (C ₆₀ H ₄₈ I ₄ N ₁₂ O ₄ Zn・2H ₂ O): Found = C: 44.58%, H: 3.47%, N: 10.25%,
Calcd.=C: 44.81%, H: 3.26%, N: 10.45%.
Absorption spectrum: UV-vis (DMSO): λ _max (ε/mol ⁻¹ dm ³ cm ⁻¹ )=326 (2.3×10 ⁴ ), 382 (1.8×10 ⁴ ), 620 (1.3×10 ⁴ ), 660 (1.3×10 ⁴ , sh), 689 nm (8.4×10 ⁴ )
(ii) Synthesis of Compound (2) Compound (2) was synthesized according to the following synthesis scheme.

A DMF solution (10 mL) containing 3-nitrophthalonitrile (0.82 g, 4.7 mmol) and 5-hydroxy-2-methylpyridine (1.0 g, 9.2 mmol) was heated and stirred at 65°C for 48 hours in the presence of _{K2CO3 (} 2.00 g). The reaction mixture was added to ice water to obtain a white precipitate. The organic layer of the precipitate was extracted using _CHCl3 . The resulting solution was concentrated under reduced pressure and then recrystallized using diethyl ether as a poor solvent to obtain a white solid (I-1) (0.47 g, 42%).
The obtained compound (I-1) (0.41 g, 1.74 mmol) was added to an n-pentanol solution (10 mL) and mixed with anhydrous GaCl (0.11 g, 0.62 mmol). Four drops of DBU were then added to the mixture, which was then heated and stirred at 145°C for 24 hours. The reaction mixture was poured into ice water to obtain a green precipitate. The organic layer of the precipitate was extracted using CHCl _3. The resulting solution was concentrated under reduced pressure and recrystallized using diethyl ether to obtain a green solid (II-2) (0.37 g, 81%).
A _CH3I solution (5.0 mL) containing the obtained compound (II-2) (0.1 g, 0.01 mmol) was heated and stirred for 24 hours at 40° C. The obtained solution was concentrated under reduced pressure and then recrystallized from acetone to obtain the target product as a green solid (III-2) (0.13 g, 84%).
Elemental analysis (C ₆₀ H ₄₈ I ₄ N ₁₂ O ₄ GaCl・H ₂ O): Found = C: 44.38%, H: 3.15%, N: 10.52%,
Calcd.=C: 44.16%, H: 3.09%, N: 10.36%.
Absorption spectrum: UV-vis (DMSO): λ _max (ε/mol ⁻¹ dm ³ cm ⁻¹ )=326 (2.3×10 ⁴ ), 382 (1.8×10 ⁴ ), 620 (1.3×10 ⁴ ), 660 (1.3×10 ⁴ , sh), 689 nm (8.4×10 ⁴ )
(iii) Synthesis of Compound (3) Compound (3) was synthesized according to the following synthesis scheme.

A DMF solution (10 mL) containing 3-nitrophthalonitrile (0.82 g, 4.7 mmol) and 5-hydroxy-2-methylpyridine (1.0 g, 9.2 mmol) was heated and stirred at 65°C for 48 hours in the presence of _{K2CO3 (} 2.00 g). The reaction mixture was added to ice water to obtain a white precipitate. The organic layer of the precipitate was extracted using _CHCl3 . The resulting solution was concentrated under reduced pressure and then recrystallized using diethyl ether as a poor solvent to obtain a white solid (I-1) (0.47 g, 42%).
The obtained compound (I-1) (0.28 g, 1.2 mmol) was added to an n-pentanol solution (10 mL) and mixed with PdCl(II) (0.07 g, 0.40 mmol). Four drops of DBU were then added to the mixture, which was then heated and stirred at 145°C for 24 hours. The reaction mixture was poured into ice water to obtain a green precipitate. The organic layer of the precipitate was extracted using CHCl _3. The resulting solution was concentrated under reduced pressure and recrystallized using diethyl ether to obtain the target product (II-3) as a green solid (0.30 g, 95%).
A _CH3I solution (5.0 mL) containing the obtained compound (II-3) (0.1 g, 0.01 mmol) was heated and stirred for 24 hours at 40° C. The obtained solution was concentrated under reduced pressure and then recrystallized from acetone to obtain the target product as a green solid (III-3) (0.13 g, 80%).
Elemental analysis (C ₆₀ H ₄₈ I ₄ N ₁₂ O ₄ Pd・3H ₂ O): Found = C: 43.48%, H: 3.25%, N: 9.98%,
Calcd.=C: 43.17%, H: 3.26%, N: 10.07%.
Absorption spectrum: λ _max = UV-vis (DMSO): λ _max (ε/mol ⁻¹ dm ³ cm ⁻¹ ) = 337 (3.0×10 ⁴ ), 607 (2.3×10 ⁴ ), 675 nm (1.0×10 ⁵ )
(iv) Synthesis of Compound (4) Compound (4) was synthesized according to the following synthesis scheme.

A DMF solution (10 mL) containing 3-nitrophthalonitrile (0.82 g, 4.7 mmol) and 5-hydroxy-2-methylpyridine (1.0 g, 9.2 mmol) was heated and stirred at 65°C for 48 hours in the presence of _{K2CO3 (} 2.00 g). The reaction mixture was added to ice water to obtain a white precipitate. The organic layer of the precipitate was extracted using _CHCl3 . The resulting solution was concentrated under reduced pressure and then recrystallized using diethyl ether as a poor solvent to obtain a white solid (I-1) (0.47 g, 42%).
The obtained compound (I-1) (0.20 g, 0.85 mmol) was added to an n-pentanol solution (10 mL) and mixed with Cu(OAc)·H ₂ O (0.11 g, 0.60 mmol). Four drops of DBU were then added to the mixture, which was then heated and stirred at 145°C for 24 hours. The reaction mixture was poured into ice water to obtain a green precipitate. The organic layer of the precipitate was extracted using CHCl _3. The resulting solution was concentrated under reduced pressure and recrystallized using diethyl ether to obtain a green solid (II-4) (0.19 g, 86%).
A _CH3I solution (5.0 mL) containing the obtained compound (II-4) (0.1 g, 0.01 mmol) was heated and stirred for 24 hours at 40° C. The obtained solution was concentrated under reduced pressure and then recrystallized from acetone to obtain the target product (III-4) as a green solid (0.13 g, 83%).
Elemental analysis (C ₆₀ H ₄₈ I ₄ N ₁₂ O ₄ Cu・2H ₂ O): Found = C: 44.58%, H: 3.47%, N: 10.25%,
Calcd.=C: 44.81%, H: 3.26%, N: 10.45%.
Absorption spectrum: UV-vis (DMSO): λ _max (ε/mol ⁻¹ dm ³ cm ⁻¹ )=326 (2.3×10 ⁴ ), 338 (4.0×10 ⁴ ), 618 (2.9×10 ⁴ ), 658 (2.8×10 ⁴ , sh), 695 nm (1.3×10 ⁵ )
(v) Synthesis of Compound (5) Compound (5) was synthesized according to the following synthesis scheme.

A DMF solution (10 mL) containing 3-nitrophthalonitrile (0.83 g, 4.7 mmol) and propyl p _- hydroxybenzoate (1.0 g, 5.6 mmol) was heated and stirred at 65°C for 48 hours in the presence of _K2CO3 (2.00 g). The reaction mixture was added to ice water to obtain a white precipitate. The organic layer of the precipitate was extracted using _CHCl3 . The resulting solution was concentrated under reduced pressure and then recrystallized using methanol as a poor solvent to obtain a white solid (I-2) (1.1 g, 78%).
The obtained compound (I-2) (0.31 g, 0.1 mol) was added to an n-pentanol solution (10 mL) and mixed with Zn(OAc) ₂ ·2H ₂ O (0.21 g, 0.1 mmol). Four drops of DBU were then added to the mixture, which was then heated and stirred at 145°C for 24 hours. After concentration under reduced pressure, the mixture was purified using a silica gel column with a developing solvent (CHCl ₃ /MeOH = 95/5). The resulting solution was concentrated under reduced pressure and recrystallized using hexane as a poor solvent to obtain a green solid (II-5) (0.31 g, 97%).
The obtained compound (II-5) (0.31 g, 0.2 mmol) was added to THF (2 mL), mixed with saturated aqueous sodium hydroxide solution (6 mL), and heated with stirring at 40° C. for 4 hours. The obtained solution was concentrated under reduced pressure and then recrystallized using acetone as a poor solvent to obtain the target product as a green solid (III-5) (0.27 g, 98%).
Elemental analysis (C ₆₀ H ₄₈ I ₄ N ₁₂ O ₄ Zn・2H ₂ O): Found = C: 44.58%, H: 3.47%, N: 10.25%,
Calcd.=C: 44.81%, H: 3.26%, N: 10.45%.
Absorption spectrum: UV-vis (DMSO): λ _max (ε/mol ⁻¹ dm ³ cm ⁻¹ )=330 (3.3×10 ³ ), 625 (2.7×10 3 ), 694 nm (1.5×10 ⁴ )
(vi) Synthesis of compound (6)

A DMF solution (10 mL) containing 4,5-dichlorophthalonitrile (0.9 g, 4.6 mmol) and 5-hydroxy-2-methylpyridine (1.0 g, 9.2 mmol) was heated and stirred at 65°C for 48 hours in the presence of _{K2CO3 (} 2.00 g). The reaction mixture was added to ice water to obtain a white precipitate. The organic layer was extracted using _CHCl3 . The resulting solution was concentrated under reduced pressure and recrystallized using diethyl ether as a poor solvent to obtain a white solid (I-3) (0.8 g, 51%).
The obtained compound (I-3) (0.1 g, 0.43 mmol) was added to an n-pentanol solution (10 mL) and mixed with anhydrous GaCl (0.02 g, 0.09 mmol). Four drops of DBU were then added to the mixture, which was then heated and stirred at 145°C for 24 hours. The reaction mixture was poured into ice water to obtain a green precipitate. The organic layer of the precipitate was extracted using CHCl _3. The resulting solution was concentrated under reduced pressure and then recrystallized using diethyl ether. The resulting solution was concentrated under reduced pressure to obtain a green solid (II-6).
A _CH3I solution (5.0 mL) containing the obtained compound (II-6) (0.11 g, 0.07 mmol) was heated and stirred for 24 hours at 40° C. The obtained solution was concentrated under reduced pressure and then recrystallized from acetone to obtain the target product as a green solid (III-6) (0.15 g, 77%).
Absorption spectrum: UV-vis (H ₂ O): λ _max (ε/mol ⁻¹ dm ³ cm ⁻¹ )=359 (8.5×10 ⁴ ), 612 (3.2×10 ⁴ ), 652 (3.2×10 ⁴ , sh), 679 nm (2.1×10 ⁵ )
^1H NMR( _D2O ): δ=9.37, 8.91, 8.40, 7.95, 4.19, 2.76 (ppm)
Elemental analysis (C ₆₀ H ₈₄ I ₄ N ₁₆ O ₈ GaCl・H ₂ O): Found = C: 39.88%, H: 3.25%, N: 8.29%,
Calcd.=C: 39.94%, H: 3.20%, N: 8.47%.
The synthesis method of compound (6) is also described in "R. Fujishiro, T. Ikeue, et al. J. InorgBiochem, 2019, 192, 7-16."

(vii) Synthesis of the compound "ZnPc(COONa) ₈ "

The compound "ZnPc(COONa) ₈ " was synthesized based on the literature "S. Tabassum, A. Nagai, et al. FEBS J, 2015, 282, 463-476."

(viii) Synthesis of the compound "ZnPc(COONa) ₁₆ "

The compound "ZnPc(COONa) ₁₆ " was synthesized based on the literature "S. Tabassum, A. Nagai, et al. FEBS J, 2015, 282, 463-476."

[Evaluation of Compounds]
(Amyloid β aggregation inhibition study)
Amyloid β protein (human, 1-42) was used, manufactured by Peptide Institute, Inc. The amyloid β protein was dissolved in 0.1% _NH3 aqueous solution to a concentration of 250 μM, then rapidly divided into equal parts and stored at −70°C.

Because phthalocyanines possess photocatalytic properties, all of the following experiments were conducted under dark conditions. First, aqueous solutions containing phthalocyanines or their salts (0, 1.25, 5, 10, and 20 μM) were prepared. The amyloid β protein solution synthesized above was added to a fibril synthesis buffer solution (50 mM phosphate buffer, pH 7.5, 100 mM sodium chloride) containing various concentrations of phthalocyanines or their salts. As a control experiment, the amyloid β solution was added to a fibril synthesis buffer solution that did not contain phthalocyanines or their salts. The reaction mixture was incubated at 37°C for 24 hours without stirring, and then the sample was quickly frozen to terminate the reaction.

The amount of amyloid-β fibrils in the sample was quantified using the fluorescence spectrum of the fluorescent agent thioflavin T (ThT, manufactured by Fujifilm Wako Pure Chemical Industries). ThT is a substance that does not fluoresce in monomeric amyloid-β, but emits strong fluorescence in response to amyloid-β aggregates.

The sample was diluted 10-fold with glycine (pH 8.5) to a final glycine concentration of 50 mM. Fluorescence spectra were measured using an F2500 spectrofluorometer (Hitachi) at an excitation wavelength of 446 nm and an emission wavelength of 490 nm. The ThT fluorescence intensity of the amyloid-β-containing sample was normalized by subtracting the fluorescence intensity measured in buffer alone.

Figures 1 to 6 show the results of the amyloid-β aggregation inhibition experiments for compounds (1) to (6), respectively. Figures 1 to 6 demonstrate that all of compounds (1) to (6) have the ability to inhibit amyloid-β aggregation.

Furthermore, it was shown that phthalocyanines with cationic substituents or their salts have greater amyloid-β aggregation inhibitory activity than phthalocyanines with anionic substituents or their salts.

Furthermore, when compound (4), a Cu-containing phthalocyanine salt known not to generate cytotoxic reactive oxygen species upon light irradiation, was used (Figure 4), the ability to inhibit amyloid-β aggregation was confirmed. Therefore, the mechanism of this aggregation inhibition is thought to be due to the substitution of phthalocyanine for the hydrophobic sites of amyloid-β, rather than through the generation of reactive oxygen species upon light irradiation.

FIG. 7 shows the results of a comparison of the inhibitory ability of amyloid β protein aggregation when compounds (1) to (6), ZnPc(COONa) ₈ , and ZnPc(COONa) ₁₆ were used at a concentration of 5 μM.

Figure 7 shows that when comparing compounds belonging to the same group, the compounds in the examples of the present application have greater inhibitory activity against amyloid β protein aggregation.

The present invention can be widely used in the medical, pharmaceutical, and research fields. More specifically, the present invention can be widely used in the treatment and/or prevention of Alzheimer's disease, the production of therapeutic and/or preventive drugs for Alzheimer's disease, and research into Alzheimer's disease and/or amyloid beta protein and amyloid fibrils.

Claims (3)

An amyloid β aggregation inhibitor comprising, as an active ingredient, a phthalocyanine represented by the following formula 1 or 2, or a salt thereof:
(In formula 1,
^M1 is Cu or Cu having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand;
Each R ¹ is independently a non-ionic substituent or a cationic substituent, and at least one R ¹ is a cationic substituent.
(In Equation 2,
^M2 is Cu or Cu having a halide, carboxylate, sulfate, nitrate, acetylacetonate, or carbonyl compound as an axial ligand;
Each ^R2 is independently a nonionic substituent or a cationic substituent, and at least one ^R2 is a cationic substituent.
Here, the cationic substituent includes the following structure :
The amyloid-β aggregation inhibitor according to claim 1 , wherein the cationic substituent comprises the following structure:
The amyloid-β aggregation inhibitor according to claim 1 or 2 , wherein the salt is a salt of Na ⁺ , H ⁺ , K ⁺ , NH ⁴⁺ or a halide ion.