Rotaxane-type photosensitizer

Abstract

To provide a new rotaxane-type photosensitizer exhibiting high light stability and enabling coexistence of suitable water solubility and cell membrane permeability.SOLUTION: A photosensitizer for photodynamic therapy containing a rotaxane-type reactive oxygen species generator, wherein the rotaxane-type reactive oxygen species generator is a [3]-rotaxane formed by being included by two cyclodextrins or derivatives thereof. Also provided is the photosensitizer which is an anticancer agent.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a rotaxane-type photosensitizer. [Background technology]

[0002] Malignant tumors, or cancer, are the leading cause of death among Japanese people, and research into cancer treatments is actively underway. In recent years, photodynamic therapy (PDT), a light-based treatment, has attracted attention as a cancer treatment with minimal side effects. Photodynamic therapy involves intravenously administering a photosensitizing drug to the patient. After the drug has been distributed throughout the body for several tens of hours, the tumor site is then irradiated with light. When irradiated with light, the sensitizer becomes excited, transferring energy to surrounding oxygen molecules and generating reactive oxygen species, such as singlet oxygen. The generated reactive oxygen species attack and kill cancer cells (Figure 1). This treatment, which selectively targets cancer cells with light, has fewer side effects than chemotherapy. Furthermore, as a non-invasive treatment that does not require the traditional surgical procedure, it is expected to be used in clinical settings.

[0003] One PDT treatment currently used in clinical settings is talaporfin sodium (trade name: Laserphyrin). Talaporfin sodium has a porphyrin-like structure and has the advantages of being highly efficient at generating reactive oxygen species (high sensitizing activity) and being excitable with long-wavelength light. However, due to its high hydrophobicity, it is prone to forming aggregates in the body, and these aggregates may remain in the body.

[0004] To overcome the low water solubility of talaporfin sodium, it is conceivable to introduce an anionic substituent into the molecule. Although such a modification improves water solubility, it also creates a new problem of reduced cell membrane permeability. Thus, it is difficult to achieve both adequate water solubility and cell membrane permeability for a porphyrin molecule.

[0005] Furthermore, because porphyrins are organic molecules, the molecules themselves are destroyed by active oxygen when exposed to light for long periods of time, resulting in the problem of photofading (poor photostability).

[0006] Therefore, Ikeda et al. (Non-Patent Document 1) reported a porphyrin derivative as a cyclic molecule, i.e., a molecule covered with two β-cyclodextrin (CD) derivatives. The molecule is water-soluble, and it has been reported that introducing the molecule into HeLa cells and irradiating them with excitation light induced cell death.

[0007] However, the porphyrin dye reported in Non-Patent Document 1 had the problem that the porphyrin and CD were merely reversibly associated by non-covalent bonds, and easily dissociated in vivo. That is, when the molecule was introduced into cells, the porphyrin skeleton was easily exposed, which was thought to result in reduced photostability and water solubility, similar to talaporfin sodium, and was therefore not sufficient for therapeutic use. [Prior art documents] [Non-patent literature]

[0008] [Non-Patent Document 1] A. Ikeda et al., ACS. Med. Chem. Lett. 2017, 8, 555-559. Summary of the Invention [Problem to be solved by the invention]

[0009] The present invention has been made in view of solving the above-mentioned problems, and aims to provide a rotaxane-type photosensitizer in which an active oxygen generator is encapsulated inside a cyclic molecule, thereby improving water solubility and photostability. At the same time, a synthesis method for the rotaxane-type photosensitizer with an increased yield is also provided. [Means for solving the problem]

[0010] As a result of extensive research, the present inventors have discovered a novel rotaxane photosensitizer. They have found that this novel rotaxane photosensitizer is highly photostable and water-soluble, and that its introduction into cells can induce cell death, leading to the completion of the present invention. They have also found that the rotaxane photosensitizer can be synthesized with high yield. That is, the present invention includes the following aspects.

[0011] Item 1. A photosensitizer for photodynamic therapy containing a rotaxane-type reactive oxygen generator. Item 2. The photosensitizer according to Item 1, wherein the rotaxane-type reactive oxygen generator is a [3]-rotaxane formed by inclusion between two cyclodextrins or derivatives thereof. Item 3. The photosensitizer according to Item 1 or 2, wherein the rotaxane-type reactive oxygen generator is a [5]-rotaxane in which the axis between cyclodextrin or a derivative thereof and the stopper is encapsulated with cucurbituril. Item 4. The photosensitizer according to any one of Items 1 to 3, wherein the cucurbituril is cucurbit[6]uril. Item 5. The photosensitizer according to any one of Items 1 to 4, wherein the active oxygen generator is any one of porphyrin, coumarin, chlorin, bacteriochlorin, acridine, methylene blue, phthalocyanine, cyanine, tolan, and derivatives thereof. Item 6. The axis of the rotaxane-type reactive oxygen generator is represented by the following formulas (1) to (6): [ka] (In the formula, R1 may be the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, A represents an organic group capable of generating active oxygen, n represents 0 or 1, and L represents a divalent hydrocarbon group having 1 to 12 carbon atoms and optionally substituted with a hetero atom.) [ka] (wherein R2 may be the same or different and each represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, and A represents an organic group capable of generating active oxygen.) [ka] (wherein R3 is the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group; A represents an organic group capable of generating active oxygen; o represents an integer of 1 to 12; and p represents 0 or 1.) [ka] (wherein R4 is the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group; A represents an organic group capable of generating active oxygen; q represents an integer of 0 to 4; and r represents 0 or 1.) [ka] (wherein R5 are the same or different and each represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group; A represents an organic group capable of generating active oxygen; s represents an integer of 0 to 6; t represents an integer of 0 to 6; and u represents 0 or 1.) [ka] (wherein R6 is the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group; A represents an organic group capable of generating active oxygen; v represents an integer of 0 to 6; w represents an integer of 0 to 6; and x represents 0 or 1). 6. The photosensitizer according to any one of items 1 to 5, wherein the photosensitizer is represented by any one of the following formulas: Item 7. The stopper of the rotaxane-type active oxygen generator is represented by the following formula (7): [ka] (wherein R7 represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group.) Item 7. The photosensitizer according to any one of items 1 to 6, which is represented by: Item 8. The photosensitizer according to any one of Items 1 to 7, which is an anticancer agent. [Effects of the Invention]

[0012] The present invention provides a novel rotaxane-type photosensitizer. This sensitizer has high photostability and exhibits both moderate water solubility and cell membrane permeability, resulting in a higher efficiency of intracellular introduction compared to conventional photosensitizers. It has a high reactive oxygen generation efficiency, and irradiation of the sensitizer introduced into cells with excitation light generates reactive oxygen, inducing cell death. By applying the novel photosensitizer of the present invention to cancer cells, an anticancer drug that avoids the problems of existing drugs can be provided. [Brief explanation of the drawings]

[0013] [Figure 1] FIG. 1 is a diagram showing a schematic diagram of PDT therapy. [Figure 2] FIG. 1 shows the structure of talaporfin sodium, an existing PDT therapeutic drug. [Figure 3] This is a diagram showing the structure of a porphyrin dye covered with β-CD. [Figure 4] FIG. 1 shows the structure of a rotaxane-type alkynylpyrene. [Figure 5] FIG. 1 shows a high-yield synthesis method for rotaxane-type dyes. [Figure 6] FIG. 1 shows a method for synthesizing raw material molecules that serve as the axes for rotaxane formation. [Figure 7] Schematic diagram of the β- and γ-[5]-rotaxane-type molecules prepared. [Figure 8] FIG. 1 shows the synthesis of the axial acridine starting molecule and the acridine control molecule. [Figure 9] FIG. 1 shows a method for synthesizing a rotaxane-type acridine. [Figure 10] FIG. 1 shows the synthesis of the axial dibromotrane starting material molecule and the dibromotrane control molecule. [Figure 11] FIG. 1 shows a method for synthesizing a rotaxane-type dibromotran. [Figure 12] FIG. 1 shows the optical properties of [3]-rotaxane, β- and γ-[5]-rotaxane. [Figure 13] FIG. 1 shows the evaluation of the singlet oxygen generation efficiency of β- and γ-[5]-rotaxanes. [Figure 14] FIG. 1 is a graph showing the evaluation of the hydroxyl radical generation efficiency of rotaxane-type acridines. [Figure 15] FIG. 1 is a graph showing the evaluation of the singlet oxygen generation efficiency of a rotaxane-type dibromotran. [Figure 16] FIG. 1 is a graph showing the evaluation of the singlet oxygen generation efficiency of rotaxane-type pyrenes. [Figure 17] FIG. 1 is a graph showing the evaluation of the singlet oxygen generation efficiency of rotaxane-type anthracenes. [Figure 18] FIG. 1 is a graph showing the evaluation of the singlet oxygen generation efficiency of rotaxane-type perylene. [Figure 19] FIG. 1 shows the evaluation of the photostability of [3]-rotaxane, β- and γ-[5]-rotaxane. [Figure 20] FIG. 1 shows the evaluation of the photostability of a rotaxane-type acridine and a rotaxane-type dibromotran. [Figure 21] FIG. 1 shows the cell transfection efficiency of β- and γ-[5]-rotaxanes. [Figure 22] FIG. 1 shows the evaluation of the photocytic activity of β- and γ-[5]-rotaxanes. DETAILED DESCRIPTION OF THE INVENTION

[0014] The "rotaxane" of the present invention refers to a compound in which cyclic molecules are enclosed around an axis molecule consisting of an organic group, and both ends of the axis molecule are sealed with stoppers. A rotaxane is obtained by irreversibly binding a bulky raw material molecule called a stopper to the end of the raw material molecule that forms the axis after the cyclic molecules are enclosed around the axis to form an association. This prevents the cyclic molecule from detaching from the axis due to steric hindrance, and the axis molecule and cyclic molecule are bound by a non-covalent bond. A molecule with this structure is called a rotaxane-type molecule.

[0015] Examples of cyclic molecules in the present invention include cyclodextrin and its derivatives, and cucurbituril. By incorporating a poorly water-soluble organic molecule into a highly water-soluble cyclic molecule, the water solubility of the organic molecule can be improved. The organic molecule with improved water solubility does not aggregate in water and is stably dispersed in water.

[0016] The stopper has a bulky structure and is located at the end of the axis molecule, thereby physically preventing dissociation of the axis molecule and the cyclic molecule. Examples of bulky molecules include aryl groups and heterocyclic groups. In the rotaxane molecule of the present invention, the raw material molecule that serves as the stopper is covalently bonded to the end of the axis raw material molecule, thereby irreversibly suppressing dissociation of the axis molecule and the cyclic molecule. Therefore, the axis molecule and the cyclic molecule do not dissociate even in water, improving the stability of the rotaxane molecule in water.

[0017] Furthermore, due to the presence of the stopper, the organic groups of the rotaxane molecules of the present invention are always enclosed by cyclic molecules such as CD, which is thought to make them less susceptible to photobleaching when irradiated with excitation light, thereby improving their photostability.

[0018] The raw material molecule serving as the stopper of the present invention preferably has an azide moiety in the bonding region with the axis raw material molecule and an ammonium cation moiety adjacent to it. An example of such a stopper raw material molecule is a structure such as the stopper raw material molecule 6 shown in Figure 7. The azide moiety of the raw material molecule undergoes a Huisgen cyclization reaction when it comes into close proximity with an alkyne. The stopper of the present invention is preferably one having the structure of the following formula (7) that has undergone such a structural change. [ka] (wherein R7 represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group.)

[0019] The axis molecule of the present invention contains an organic group capable of generating active oxygen. The axis molecule of the present invention preferably has an ammonium cation moiety in the linker region located at both ends of the organic group, and an alkyne at the end of the axis raw material molecule. Examples of such axis molecules include those having the structure of the following formula (1): [ka] (In the formula, R1 may be the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, A represents an organic group capable of generating active oxygen, n represents 0 or 1, and L represents a divalent hydrocarbon group having 1 to 12 carbon atoms and optionally substituted with a heteroatom.)

[0020] Here, the heteroatom is selected from O, N, and S, and is preferably O. The number of heteroatoms is, for example, 1 to 3, preferably 2, and more preferably 1. The hydrocarbon group may be a saturated hydrocarbon or an unsaturated hydrocarbon.

[0021] When the terminal alkyne of the base molecule and the azide moiety of the stopper molecule are brought into close proximity, the Huisgen cyclization reaction proceeds rapidly without a transition metal catalyst, resulting in efficient rotaxane formation and an increased synthesis yield of rotaxane molecules.

[0022] The axis molecule in the present invention may have a pyridinium cation moiety in the linker region located at both ends of the organic group capable of generating active oxygen, and an azide moiety at the end of the axis raw material molecule. Examples of such an axis molecule include those having the structure of the following formula (2). [ka] (In the formula, R2 may be the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, and A represents an organic group capable of generating active oxygen.)

[0023] Other axis molecules of the present invention include, for example, those having structures of the following formulae (3) to (6): The R-CO- group located at the end of these may be referred to as a stopper. [ka] (In the formula, R3 is the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, A represents an organic group capable of generating active oxygen, o represents an integer of 1 to 12, and p represents 0 or 1.) [ka] (In the formula, R4 is the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, A represents an organic group capable of generating active oxygen, q is an integer of 0 to 4, and r is 0 or 1.) [ka] (In the formula, R5 is the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group; A represents an organic group capable of generating active oxygen; s represents an integer of 0 to 6; t represents an integer of 0 to 6; and u represents 0 or 1.) [ka] (wherein R6 is the same or different and represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group; A represents an organic group capable of generating active oxygen; v represents an integer of 0 to 6; w represents an integer of 0 to 6; and x represents 0 or 1.)

[0024] Examples of the aryl group of R1 to R7 include a phenyl group, a naphthyl group, and a terphenyl group, with a phenyl group being preferred. Examples of the nitrogen-containing heterocyclic group include a nitrogen-containing 6-membered heterocyclic group such as pyridine, pyrazine, pyrimidine, pyridazine, and triazine, with a 1,3,5-triazine group being preferred.

[0025] Substituents for the aryl and nitrogen-containing heterocyclic groups R1 to R7 include linear or branched lower alkyl groups having 1 to 6 carbon atoms, such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, pentyl, and hexyl; hydroxy groups; alkoxy groups such as methoxy, ethoxy, isopropoxy, and tert-butoxy; linear or branched lower alkylamino groups having 1 to 6 carbon atoms, such as methylamino, ethylamino, isopropylamino, tert-butylamino, dimethylamino, diethylamino, and diisopropylamino; aryl groups such as phenyl; and carbonyl groups such as acetyl, propionyl, methoxycarbonyl, benzoyl, carbamoyl, and carboxy. Of these, methoxy is preferred for its ease of synthesis.

[0026] A rotaxane in which the total number of axle molecules and cyclic molecules is n is called an [n]-rotaxane. Therefore, a rotaxane in which one axle molecule is enclosed by two cyclic molecules is called a [3]-rotaxane, and a rotaxane in which one axle molecule is enclosed by four cyclic molecules is called a [5]-rotaxane.

[0027] The "active oxygen" of the present invention encompasses all active oxygen species, such as singlet oxygen, superoxide anion, hydroxyl radical, etc. The "active oxygen generator" refers to an organic molecule that generates active oxygen, and is not particularly limited as long as it generates active oxygen. Examples include porphyrin, coumarin, chlorin, bacteriochlorin, acridine, methylene blue, phthalocyanine, cyanine, tolane, pyrene, anthracene, perylene, and derivatives thereof, of which porphyrin, coumarin, chlorin, bacteriochlorin, acridine, methylene blue, phthalocyanine, cyanine, and tolane are preferred. Substituents of the derivatives include linear or branched lower alkyl groups having 1 to 3 carbon atoms, such as methyl, ethyl, and propyl; hydroxyl groups; linear or branched lower alkylamino groups having 1 to 3 carbon atoms, such as methylamino, ethylamino, isopropylamino, tert-butylamino, dimethylamino, diethylamino, and diisopropylamino; aryl groups such as phenyl; carbonyl groups such as acetyl, propionyl, methoxycarbonyl, benzoyl, carbamoyl, and carboxy; methoxy; bromo; and iodo groups. Among these, bromo and iodo groups are preferred, as they are expected to promote intersystem crossing from excited singlet to excited triplet (improving photosensitizing activity). Examples of the derivatives include biphenylanthracene in the case of anthracene and dibromo tolane in the case of tolane. Dibromo tolane is particularly preferred as a tolan derivative. The term "active oxygen generating ability" refers to the ability to generate active oxygen. The efficiency of generating the active oxygen is referred to as the active oxygen generating efficiency.

[0028] The photosensitizer in the present invention refers to a compound that generates active oxygen by irradiating an active oxygen generator with excitation light, causing the active oxygen generator to change from a ground state to an excited state and inducing energy transfer or electron transfer to surrounding oxygen molecules. The photosensitizing effect refers to the effect of generating active oxygen by light irradiation as described above. Having the photosensitizing effect is sometimes referred to as having photosensitizing activity. Photosensitizing activity is synonymous with active oxygen generation efficiency.

[0029] "Photodynamic therapy" in this invention refers to a treatment method in which a photosensitizing drug is administered intravenously to a patient, and after several tens of hours have passed and the drug has been distributed throughout the body, the affected area is irradiated with light. The photosensitizer is excited by light, causing energy transfer to surrounding oxygen molecules, generating reactive oxygen species such as singlet oxygen. The generated reactive oxygen species attacks surrounding cells and has the effect of killing them.

[0030] "Cyclodextrin" refers to a compound in which multiple D-glucose molecules form a cyclic structure. The cyclodextrin (CD) of the present invention includes α-CD, in which six glucose molecules are bonded, β-CD, in which seven glucose molecules are bonded, and γ-CD, in which eight glucose molecules are bonded, but is not limited to these as long as the glucose molecules form a cyclic structure. When the organic group capable of generating active oxygen (active oxygen generator) to be included is a porphyrin, β-CD is preferred.

[0031] The CD of the present invention may be a CD derivative. Examples of CD derivatives include those in which some or all of the hydroxy groups of glucose have been modified with alkyl groups, or those in which some or all of the hydroxy groups have been substituted with bromo, iodo, or amino groups. The alkyl group is preferably a single- or branched-chain lower alkyl group having 1 to 6 carbon atoms. A methyl group is more preferred, but is not limited to these.

[0032] In the present invention, the organic group having active oxygen generating ability is enclosed by CD, which improves the water solubility of the molecule containing the organic group, and as a result, the molecule does not aggregate in water, which is advantageous. In addition, the active oxygen generator is always enclosed by CD, which is advantageous because it improves photostability.

[0033] "Cucabitsuuril" is a cyclic molecule in which glycoluril (=C4H2N4O2=) monomers are connected by methylene groups (-CH2-). Cucabituril (CB) is known for its ability to strongly capture cationic substances because both ends are negatively charged due to the polarization of the carbonyl groups. When the raw material molecules that form the axis and the raw material molecules that form the stopper contain ammonium cations or pyridinium cations, they spontaneously assemble with CB through electrostatic interactions. Cucabituril with m glycolurils is referred to as cucabit[m]uril. In the present invention, cucabit[6]uril is preferred.

[0034] When a base molecule (containing a terminal alkyne) with an ammonium cation moiety, a stopper molecule (containing an azide moiety at the terminal), CD and CB are mixed in aqueous solution, all components spontaneously associate non-covalently through hydrophobic interactions between the CD and the base molecule, interactions between the ammonium cations of the base molecule and the stopper molecule and the CB, and hydrogen bonding between the rims of the CD and CB (the hydroxyl group of CD and the carbonyl group of CB). Furthermore, as shown in Figure 5, the terminal alkyne of the base molecule enclosed by CB and the azide moiety of the stopper molecule are in close proximity, allowing for rapid Huisgen cyclization. This reaction proceeds spontaneously without the need for a transition metal catalyst, which is advantageous for efficient rotaxane formation. As a result of this reaction, rotaxane-type photosensitizers can be synthesized with high yields.

[0035] As described above, when the raw material molecules serving as the axis and the stopper, CD and CB, are used in the present invention, they spontaneously associate, and therefore the rotaxane synthesis method of the present invention has the advantage that rotaxane formation is possible even when the affinity between CD and the reactive oxygen generator is low. Therefore, in the present invention, it is preferable to use CB as the cyclic molecule.

[0036] In photodynamic therapy, when the photosensitizer of the present invention is used for the treatment of cancer, for example, the photosensitizer is administered to a patient, and after a certain period of time, the cancer cells are irradiated with excitation light. This allows the cancer cells to be killed by the photosensitizing effect of the photosensitizer accumulated in the cancer cells. In other words, the photosensitizer containing the rotaxane-type reactive oxygen generator of the present invention can be used as an anticancer agent.

[0037] Because the light energy required to excite the photosensitizer of the present invention is low, it can attack only cancer cells while minimizing the impact on normal cells. Therefore, this drug has fewer side effects than conventional anticancer drugs. Furthermore, this treatment does not require conventional surgical procedures and is less invasive to patients. [Example]

[0038] This invention is further illustrated by the following examples, which should not be construed as further limiting.

[0039] 1. Synthesis of β-[5]-rotaxane (Example 1) and γ-[5]-rotaxane (Example 2) As the rotaxane-type active oxygen generator of the present invention, the rotaxane-type porphyrin can be synthesized, for example, as follows. 1-1. Using the route shown in Figure 6, starting material molecule 1, which serves as the axis having a porphyrin skeleton and an ammonium cation at its terminal, was synthesized in two steps from dipyrrolmethane (compound 2) and benzaldehyde derivative 3. Dipyrrolmethane (compound 2) was synthesized according to a non-patent document (M. Calik et al., J. Am. Chem. Soc. 2014, 136, 17802-17807), and benzaldehyde derivative (compound 3) was synthesized according to a non-patent document (Z.-Y. Sun et al., Org. Lett. 2020, 22, 6214-6219).

[0040] 1-2. After bubbling argon gas through dichloromethane (200 mL) for 30 minutes, dipyrrolethane 2 (146 mg, 1.00 mmol) and the benzaldehyde derivative (compound 3) (188 mg, 1.00 mmol) were dissolved in dichloromethane at room temperature. Trifluoroacetic acid (TFA, 57.4 mg, 38.0 μL, 0.503 mmol) was added to the reaction mixture, and the reaction mixture was stirred at room temperature for 3 hours without exposure to light. 2,3-Dichloro-5,6-dicyano-p-benzoquinone (DDQ, 272 mg, 1.20 mmol) was then added, and the mixture was stirred at room temperature for an additional hour. After neutralization with triethylamine (EtN), the mixture was evaporated to dryness on a rotary evaporator. The residue was purified by silica gel column chromatography (developing solvent: ethyl acetate) to obtain compound 4 (142 mg) having a porphyrin skeleton and a terminal hydroxy group as a purple solid in 45% yield.

[0041] 1-3. The physical properties of compound 4 are as follows: Decomposition point: 300°C or higher. 1 H NMR (500 MHz, (CD3)2SO) δ = 10.64 (s, 2H), 9.66 (d, J = 4.5 Hz, 4H), 9.05 (d, J = 4.5 Hz, 4H), 8.24 (d, J = 7.5 Hz, 4H), 7.88 (d, J = 8.0 Hz, 4H), 3.68 (t, J = 6.25 Hz, 4H), 2.66 (t, J = 7.25 Hz, 4H), 1.83-1.89 (m, 4H). 13 C NMR (500 MHz, (CD3)2SO) δ = 146.2, 144.8, 139.9, 134.8, 132.8, 130.7, 130.2, 123.1, 118.0, 105.9, 92.1, 80.4, 59.5, 31.7, 15.6. IR (KBr) λ max = 3388, 2219, 1799, 1706, 1577 cm -1 . ESI-MS (m / z): [M+H]+ calcd for C42 H 35 N4O2, 627.2760; found, 627.2750. 1-4. Compound 4 (63.0 mg, 0.100 mmol), 2-nitro-N-(2-propyn-1-yl)benzenesulfonate 5 (60.0 mg, 0.250 mmol), and triphenylphosphine (PPh3, 68.0 mg, 0.259 mmol) were dissolved in THF (5.00 mL). Diisopropyl azodicarboxylate (DIAD, 40.4 mg, 0.200 mmol) was then slowly added to the mixture at 0 °C. The reaction mixture was stirred overnight at 0 °C and then evaporated to dryness on a rotary evaporator. The residue was purified by silica gel column chromatography (eluent: dichloromethane / ethyl acetate = 1 / 0 to 9 / 1) to give a purple solid (70.0 mg). To a solution of this solid and LiOH·HO (65.0 mg, 1.55 mmol) in N,N-dimethylformamide (DMF, 6.00 mL) was slowly added a solution of p-toluenethiol (64.0 mg, 0.515 mmol) in DMF (2.00 mL). The reaction mixture was stirred overnight at room temperature under argon and then rotary evaporated to dryness. The residue was purified by silica gel column chromatography (ethyl acetate / EtN = 100 / 1) to give a purple solid. This solid was dissolved in a solution of HCl in MeOH and concentrated on a rotary evaporator to give the starting axial molecule 1 (32.0 mg) as a green solid in 38% yield.

[0042] 1-5. The physical properties of the axial raw material molecule 1 are as follows: Decomposition point 120°C. 1H NMR (500 MHz, CD3OD) δ = 11.33 (s, 2H), 9.87 (d, J = 4.5 Hz, 4H), 9.24 (d, J = 4.5 Hz, 4H), 8.54 (d, J = 7.0 Hz, 4H), 8.10 (d, J = 7.0 Hz, 4H), 4.10 (d, J = 2.5 Hz, 4H), 3.46 (t, J = 7.75 Hz, 4H), 3.35 (t, J = 2.75 Hz, 2H), 2.84 (t, J = 6.75 Hz, 4H), 2.15-2.21 (m, 4H)。 13 C NMR (500 MHz, CD3OD) δ = 147.4, 145.1, 139.9, 138.8, 132.7, 132.6, 131.6, 126.9, 122.4, 108.7, 92.0, 82.6, 79.5, 74.7, 47.3, 37.6, 26.3, 17.7。IR (KBr) λ max = 3400, 2947, 2775, 2225, 2126, 1601, 1560, 1502 cm -1 。ESI-MS (m / z): [M-4Cl-3H] + calcd for C 48 H 41 N6, 701.3393; found, 701.3405。 1-6. The raw material molecule 6 that forms the stopper shown in Figure 7 was synthesized according to a non-patent document (Y. Ohishi et al., Adv. Opt. Mater. 2024, 12, 2301457), 2,3-DM-β-CD was synthesized according to a non-patent document (I. Fejos et al., Electrophoresis 2017, 38, 1869-1877), and 2,3-DM-γ-CD was synthesized according to a non-patent document (K. Takeo et al., Carbohydrate Research 1989, 187, 203-221). A solution of starting molecule 1 (0.770 mg, 0.909 μmol), starting molecule 6 (0.570 mg, 2.10 μmol), which serves as the axis and the stopper shown in Figure 7, and either 2,3-DM-β-CD (n = 7) (19.9 mg, 15.0 μmol) or 2,3-DM-γ-CD (n = 8) (22.8 mg, 15.0 μmol) in HO (250 μL) was stirred at 60 °C for 30 min and then cooled to 25 °C. CB6 (2.09 mg, 2.10 μmol) was added to this solution, and the mixture was stirred at 25 °C for 48 h. The resulting solution was filtered through a membrane filter (PTFE, pore size: 0.22 μm) and then purified by reverse-phase HPLC to give β-[5]-rotaxane (3.90 mg) and γ-[5]-rotaxane (3.80 mg) as purple solids in 71% and 65% yields, respectively.

[0043] 1-7. The physical properties of β-[5]-rotaxane are as follows: Decomposition point: 240°C. 1H NMR (500 MHz, D2O) δ = 10.50 (s, 2H), 9.71 (d, J = 4.0 Hz, 4H), 9.09 (d, J = 4.0 Hz, 4H), 8.41 (s, 4H), 8.38 (d, J = 7.5 Hz, 4H), 7.73 (d, J = 8.0 Hz, 4H), 6.99 (d, J = 2.5 Hz, 4H), 6.68 (t, J = 2.25 Hz, 2H), 6.61 (s, 2H), 5.83 (d, J = 15 Hz, 12H), 5.77 (d, J = 15 Hz, 12H), 5.55 (s, 24H), 5.08 (d, J = 3.0 Hz, 14H), 4.55 (s, 4H), 4.37-4.25 (m, 32H), 4.07-3.94 (m, 28H), 3.91-3.82 (m, 16H), 3.77-3.71 (m, 14H), 3.71-3.64 (m, 4H), 3.59-3.52 (m, 14H), 3.30-3.21 (m, 14H), 3.22-3.08 (m, 4H), 3.02-2.95 (m, 14H), 2.91 (s, 42H), 2.65-2.53 (m, 4H), 2.39 (s, 42H)。 13 C NMR (500 MHz, D2O) δ = 170.8, 160.9, 156.9, 156.5, 141.7, 139.3, 134.5, 134.4, 129.8, 124.1, 120.4, 117.5, 108.0, 105.7, 101.5, 99.0, 93.0, 81.4, 80.5, 79.3, 72.2, 70.5, 60.4, 59.5, 57.7, 55.8, 51.7, 51.6, 47.8, 46.2, 45.6, 42.2, 30.4, 25.2, 16.7。IR (KBr) λ max = 3421, 2931, 2835, 1739, 1598 cm -1 。 ESI-MS (m / z): [M-4HCOO] 4+ calcd for C 254 H 344 N 62 O 98, 1458.3482; found, 1458.3464。

[0044] The physical property data of 1-8.γ-[5]-rotaxane are as follows. Decomposition point: 248 °C. 1 H NMR (500 MHz, D2O) δ = 10.44 (s, 2H), 9.67 (d, J = 4.0 Hz, 4H), 9.00 (d, J = 4.0 Hz, 4H), 8.42 (s, 4H), 8.34 (d, J = 7.5 Hz, 4H), 7.89 (d, J = 7.5 Hz, 4H), 7.00 (d, J = 2.0 Hz, 4H), 6.69 (t, J = 2.0 Hz, 2H), 6.61 (s, 2H), 5.83 (d, J = 15.5 Hz, 12H), 5.78 (d, J = 15.5 Hz, 12H), 5.56 (s, 24H), 5.13 (d, J = 3.0 Hz, 16H), 4.56 (s, 4H), 4.40 - 4.24 (m, 32H), 4.00 - 3.89 (m, 32H), 3.87 (s, 12H), 3.86 - 3.82 (m, 4H), 3.77 - 3.70 (m, 16H), 3.64 - 3.60 (m, 4H), 3.60 - 3.55 (m, 16H), 3.55 - 3.51 (m, 4H), 3.28 - 3.20 (m, 16H), 3.01 - 2.94 (m, 16H), 2.85 (s, 48H), 2.66 (s, 48H), 2.61 - 2.52 (m, 4H). 13 C NMR (500 MHz, D2O) δ = 156.1, 152.1, 151.7, 134.1, 129.7, 103.3, 96.7, 93.2, 92.9, 76.7, 76.0, 72.5, 67.2, 65.7, 55.7, 55.0, 54.8, 53.1, 53.0, 51.1, 46.7. IR (KBr) λ max = 3420, 2931, 2836, 1739, 1599 cm -1 . ESI-MS (m / z): [M - 4HCOO] 4+ calcd for C 270 H372 N 62 O 108 , 1553.3902; found, 1553.3887.

[0045] The rotaxane-type porphyrin obtained as described above exhibited high water solubility, and since the CDs were encapsulated by irreversible bonds, it is believed that the porphyrins remain enclosed in the CDs even in vivo.

[0046] This synthetic method has the advantage that it can be used to form rotaxanes even when the affinity between CD and reactive oxygen generators is low. In other words, by using this synthetic method, it is possible to encapsulate a variety of molecules, not just the porphyrins mentioned above, and it is thought that it will be possible to synthesize a variety of rotaxane-type reactive oxygen generators with different properties, such as the type of reactive oxygen generated, the efficiency of reactive oxygen generation, and the light energy (wavelength) required for excitation. In fact, in addition to porphyrins, we have also succeeded in synthesizing rotaxane-type molecules encapsulated with acridine and dibromotrane. The synthetic methods and results for molecules other than porphyrins are shown below.

[0047] 2. Synthesis of [3]-rotaxane (comparative example) As a comparative example of the present invention, we synthesized a [3]-rotaxane in which the porphyrin-containing axis molecule was not enclosed by CD but was sealed with two CB6s and a stopper. A solution of starting molecule 1 (0.770 mg, 0.909 μmol), starting molecule 6 (0.570 mg, 2.10 μmol), and CB6 (2.09 mg, 2.10 μmol) in HO (250 μL) was stirred at 60 °C for 48 hours. The resulting solution was filtered through a membrane filter (PTFE, pore size: 0.22 μm) and purified by reverse-phase HPLC to obtain [3]-rotaxane (1.50 mg) as a purple solid in 45% yield. The physical properties of the [3]-rotaxane are as follows: decomposition point above 300 °C. 1H NMR (500 MHz, CD3OD) δ = 10.50 (s, 2H), 9.55 (d, J = 4.5 Hz, 4H), 9.09 (d, J = 4.5 Hz, 4H), 8.54 (s, 4H), 8.23 ​​(d, J = 8.0 Hz, 4H), 8.00 (d, J = 8.0 Hz, 4H), 6.94 (d, J = 2.5 Hz, 4H ), 6.63 (s, 2H), 6.56 (t, J = 2.0 Hz, 2H), 5.93 (d, J = 15.5 Hz, 12H), 5.87 (d, J = 15.5 Hz, 12H), 5.45 (s, 24H), 4.57 (s, 4H), 4.39 (s, 4H), 4.33 (t, J = 5.75 Hz, 4H), 4.23-4.29 (m, 24H), 3.82-3.88 (m,16H), 3.73 (t, J = 7.5 Hz, 4H), 3.04 (t, J = 7.0 Hz, 4H), 2.58-2.64 (m, 4H). 13 C NMR (500 MHz, CH3OD) δ = 168.3, 162.8, 157.1, 156.8, 140.5, 135.9, 135.5, 131.7, 125.2, 121.7, 108.7, 102.0, 82.8, 71.1, 56.0, 52.6, 52.4, 52.2, 47.4, 46.6, 43.5, 26.4, 18.0. IR (KBr) 3449, 2926, 1631, 1599 cm -1 ESI-MS (m / z): [M-4HCOO] 4+ calcd for C 142 H 148 N 62 O 28 , 792.5522; found, 792.5530. 3. Synthesis of Rotaxane-Type Acridine (Example 3) As the rotaxane-type active oxygen generator of the present invention, a rotaxane-type acridine can be synthesized, for example, as follows. 3-1. The starting material molecule 9, which serves as the axis having an acridine skeleton and an ammonium cation at its terminal, was synthesized in two steps from 2,7-diiodoacridine (compound 7) using the route shown in Figure 8. 2,7-Diiodoacridine (compound 7) was synthesized according to the non-patent document (M. Calik et al., J. Am. Chem. Soc. 2014, 136, 17802-17807).

[0048] 3-2. THF (20 mL) and diisopropylamine (20 mL) were added to a 100 mL round-bottom flask, and argon gas was bubbled through the solution for 30 min. 2,7-Diiodoacridine (432 mg, 0.998 mmol), 8-nonyn-1-ol (421 mg, 3.0 mmol), Pd(PPh3)4 (117 mg, 101 μmol), and CuI (10.0 mg, 52.5 μmol) were added to the solution. The reaction mixture was stirred at 80 °C under argon for 12 h. The resulting precipitate was removed by filtration through Celite. The filtrate was concentrated on a rotary evaporator and purified by silica gel column chromatography (eluent: CHCl2 / ethyl acetate = 9 / 1) to give compound 8 (312 mg) as a pale yellow solid in 69% yield.

[0049] 3-3. The physical properties of compound 8 are as follows: Melting point: 127.9-130.0°C. 1 H NMR (500 MHz, CDCl3) δ = 8.59 (s, 1H), 8.11 (d , J = 9.5 Hz, 2H), 8.03 (s, 2H), 7.71 (dd, J = 1.75 Hz, 8.75 Hz, 2H), 3.67 (t, J = 6.5 Hz, 4H), 2.49 (t, 7 Hz, 4H), 2.18 (s, 2H), 1.65-1.56 (m, 8H), 1.56-1.47 (m, 4H), 1.47-1.37 (m, 8H). 13C NMR (125 MHz, CDCl3) δ = 148.2, 134.9, 133.5, 131.0, 129.4, 126.6, 121.8, 92.8, 80.6, 63.1, 32.8, 29.0, 28.9, 28.7, 25.7, 19.6 ppm. IR (KBr) λmax = 3403, 2933, 1574 cm -1 . ESI-MS (m / z): [M+H]+ calcd for C 31 H 38 NO2, 456.29025; found, 456.28955.

[0050] 3-4. Compound 8 (135 mg, 0.296 mmol), compound 5 (180 mg, 0.749 mmol), and triphenylphosphine (PPh3, 205 mg, 0.782 mmol) were dissolved in THF (10.0 mL), and diisopropyl azodicarboxylate (DIAD, 164 mg, 0.811 mmol) was slowly added to the mixture at 0 °C. The reaction mixture was stirred under argon at 0 °C for 3 h and then evaporated to dryness using a rotary evaporator. The residue was purified by silica gel column chromatography (eluent: dichloromethane / ethyl acetate = 25 / 1) to obtain a yellow solid (224 mg). To a solution of this solid and LiOH·HO (251 mg, 5.98 mmol) in N,N-dimethylformamide (DMF, 6.00 mL) was slowly added a solution of p-toluenethiol (248 mg, 2.00 mmol) in DMF (12.0 mL). The reaction mixture was stirred under argon at room temperature for 1.5 h and then rotary evaporated to dryness. The residue was purified by silica gel column chromatography (eluent: methanol) to give a yellow solid. This solid was dissolved in THF (5 mL) and 35% aqueous HCl (0.200 mL) was added in small portions to precipitate. The resulting precipitate was collected by filtration to give the starting molecule 9 (154 mg) as a yellow solid in 85% yield (three steps).

[0051] 3-5. The physical properties of the raw material molecules of the axis are as follows: 9. Melting point 117.6-120.3 ℃. 1 H NMR (500 MHz, CH3OD) δ = 9.73 (s, 1H), 8.43 (s, 2H), 8.28 (d, 9.5 Hz, 2H), 8.15 (dd, J = 1.5 Hz, 9 Hz, 2H), 3.95 (d, J = 2.5 Hz, 4H), 3.31 (quin, 1.5 Hz, 4H), 3.23 (t, 2.75 Hz, 2H), 3.12 (t, 7.75 Hz, 4H), 2.56 (t, 6.75 Hz, 4H), 1.78-1.67 (m, 8H), 1.61-1.54 (m, 4H), 1.51-1.45 (m, 8H). 13 C NMR (125 MHz, CH3OD) δ = 146.5, 139.6, 131.6, 126.7, 124.3, 120.5, 95.2, 78.8, 77.9, 73.2, 46.7, 36.0, 28.4, 28.1, 26.1, 25.7, 18.8 ppm. IR (KBr) λmax = 3422, 2933, 1462 cm -1 ESI-MS (m / z): [M+2H] 2+ calcd for C 37 H 45 N3, 265.68067; found, 265.68021.

[0052] 3-6. 2,3-DM-α-CD was synthesized according to a non-patent document (O. Jurcek et al. Growth Des. 2020, 20, 4193-4199.). A solution of the starting molecule 9 (2.45 mg, 4.07 μmol), the starting molecule 6 (2.27 mg, 8.34 μmol), which serves as the stopper shown in Figure 9, and 2,3-DM-α-CD (68.1 mg, 59.7 μmol) in HO (1 mL) was stirred at 60 °C for 30 min and then cooled to 25 °C. CB6 (8.42 mg, 8.45 μmol) was added to this solution, and the mixture was stirred at 25 °C for 48 h. The resulting solution was filtered through a membrane filter (PTFE, pore size: 0.22 μm) and then purified by reverse-phase HPLC to give rotaxane-type acridine (10.3 mg) as a white solid in 49% yield.

[0053] 3-7. The physical properties of rotaxane-type acridine are as follows: Decomposition point: 220°C. 1H NMR (500 MHz, D2O) δ = 9.05 (s, 1H), 8.57 (s, 1H), 8.39-8.32 (m, 5H), 8.27 (t, J = 7.0 Hz, 2H), 7.69 (s, 1H), 7.47 (d, J = 10.0 Hz, 1H), 6.98 (t, J = 2.0 Hz, 4H), 6.68 (t, J = 2.25 Hz, 2H), 6.55 (s, 1H), 6.51 (s, 1H), 5.78-5.66 (m, 24H), 5.52 (s, 24H), 5.30-5.23 (m, 6H), 5.23-5.17 (m, 6H), 4.53 (s, 4H), 4.33-4.25 (m, 28H), 4.25-4.20 (m, 2H), 4.20-4.17 (s, 2H), 4.17-4.09 (m, 6H), 4.06-3.94 (m, 12H), 3.94-3.85 (m, 24H), 3.85-3.75 (m, 10H), 3.75-3.63 (m, 24H), 3.63-3.55 (m, 6H), 3.55-3.44 (m, 24H), 3.44-3.34 (m, 20H), 3.34-3.21 (m, 32H), 3.21-3.16 (m, 6H), 2.72-2.64 (m, 2H), 2.53-2.35 (m, 2H), 2.18-1.92 (m, 6H), 1.81-1.54 (m, 12H)。 13 C NMR (500 MHz, D2O) δ =169.4, 169.3 160.7, 156.7, 156.3, 147.5, 139.2, 134.3, 107.9, 101.3, 99.7, 99.6, 99.4, 81.3, 80.8, 80.7, 80.6, 72.6, 70.3, 70.2, 61.6, 60.8, 60.3, 60.1, 57.4, 55.7, 51.6, 51.4, 30.1, 29.9, 29.4, 25.9。IR (KBr) λmax = 3435, 2929, 1739, 1475 cm -1 。ESI-MS (m / z): [M-4HCOO] 4+calcd for 1320.30894; found, 1320.31091. 4. Synthesis of water-soluble acridine (comparative example) As a comparative example of rotaxane-type acridine, acridine derivatives with water-soluble substituents were synthesized. 4-1. A water-soluble acridine derivative 11 was synthesized using the route shown in Figure 8. It was synthesized in one step from 2,7-diiodoacridine 7.

[0054] 4-2. THF (10 mL) and diisopropylamine (10 mL) were added to a 50 mL round-bottom flask, and argon gas was bubbled through the solution for 30 min. 2,7-Diiodoacridine 7 (112 mg, 0.251 mmol), triethylene glycol mono(2-propynyl) ether 10 (108 mg, 0.57 mmol), Pd(PPh3)4 (28.8 mg, 24.9 μmol), and CuI (2.38 mg, 12.5 μmol) were added to the solution, and the reaction mixture was stirred at 80 °C under argon for 12 h. The reaction mixture was evaporated to dryness on a rotary evaporator, dissolved in dichloromethane (50 mL), and washed with water (50 mL). The organic layer was dried over sodium sulfate, concentrated on a rotary evaporator, and purified twice by silica gel column chromatography (eluent: CH2Cl2 / methanol = 10 / 1) to give acridine derivative 11 (32 mg) as a yellow oil in 23% yield.

[0055] 4-3. The physical properties of acridine derivative 11 are as follows: 1 H NMR (500 MHz, CDCl3) δ = 8.63 (s, 1H), 8.12 (t , J = 9.5 Hz, 4H), 7.75 (dd, 1.75Hz, 8.75Hz, 2H), 4.51 (s, 4H), 3.85-3.82 (m, 4H), 3.78-3.74 (m, 8H), 3.71 (s, 8H), 3.65-3.61 (m, 4H). 13C NMR (125 MHz, CDCl3) δ = 148.6, 135.7, 133.1, 132.0, 130.0, 126.4, 120.5, 87.1, 86.1, 72.6, 70.8, 70.5, 70.4, 69.4, 61.8, 59.3 ppm. IR (KBr) λ max =3401, 2869, 1099 cm -1 . ESI-MS (m / z): [M+H]+ calcd for C 31 H 37 NO8, 552.25974; found, 552.25989. 5. Synthesis of rotaxane-type dibromotrans (Example 4) As the rotaxane-type active oxygen generator of the present invention, a rotaxane-type dibromotrane can be synthesized, for example, as follows. 5-1. The starting material molecule 20, which serves as the axis having a dibromotlane skeleton and an ammonium cation at its terminal, was synthesized in 10 steps from 2-bromo-4-iodophenol (compound 12) using the route shown in Figure 10. 2-Bromo-4-iodophenol 12 was synthesized according to a non-patent document (ACJ Heinrich et al. Org. Lett. 2013, 15, 4666-4669.).

[0056] 5-2. Compound 12 (2.7 g, 6.5 mmol), 1,3-dibromopropane (3.6 mL, 35 mmol), K2CO3 (4.8 g, 35 mmol), and acetone (70 mL) were added to a 200 mL round-bottom flask, and the mixture was refluxed for 24 h. The resulting mixture was cooled to room temperature, and the solvent was evaporated. H2O (100 mL) was added to the resulting residue, which was then extracted with diethyl ether (100 mL x 3). The organic layer was dried over sodium sulfate and concentrated on a rotary evaporator. The resulting residue was purified by silica gel column chromatography (eluent: hexane / CHCl2 = 3:1) to give compound 13 (2.72 g) as a colorless oil in 92% yield.

[0057] 5-3. The physical properties of compound 13 are as follows: 1 H NMR (500 MHz, CDCl3) δ = 7.83 (d, J = 2.5 Hz, 1H), 7.54 (dd, J = 1.5, 9.0 Hz, 1H), 6.68 (d, J = 8.5 Hz, 1H), 4.14 (t, J = 5.5 Hz, 2H), 3.66 (t, J = 6.3 Hz, 2H), 2.38-2.33 (m, 2H). 13 C NMR (125 MHz, CDCl3) δ = 155.1, 141.1, 137.3, 115.1, 113.5, 82.8, 66.5, 32.1, 29.8 ppm. IR (NaCl) λmax = 3064, 2936, 2879, 2532 cm -1 . ESI-MS (m / z): [M+Na]+ calcd for C9H9 79 Br 81 BrIONa, 442.79423; found, 442.3377.

[0058] 5-4. Compound 13 (2.1 g, 7.0 mmol), sodium azide (NaN3, 0.78 g, 12 mmol), and acetone (50 mL) were added to a 200 mL round-bottom flask, and the mixture was refluxed for 24 h. The precipitated solid was removed by filtration, and the solvent was evaporated to give crude product 14. The resulting product 14 was used in the next step without further purification. Triphenylphosphine (PPh3, 2.2 g, 8.5 mmol) and water (0.36 mL, 20 mmol) were added to a solution of crude product 14 in THF (15 mL), and the mixture was stirred at room temperature for 24 h. The solvent was evaporated to give crude product 15. The resulting product 15 was used in the next step without further purification. To a solution of crude 15 in CHCl (30 mL), 2-nitrobenzenesulfonyl chloride (1.4 g, 6.5 mmol) and triethylamine (0.91 mL, 6.5 mmol) were added, and the mixture was stirred at room temperature for 24 h. The resulting mixture was concentrated on a rotary evaporator and purified by silica gel column chromatography (eluent: CHCl) to give compound 16 (784 mg) as a white solid in 22% yield (three steps).

[0059] 5-5. The physical properties of compound 16 are as follows: Melting point 108.0-109.7°C. 1 H NMR (500 MHz, CDCl3) δ = 8.14-8.12 (m, 1H), 7.81-7.78 (m, 2H), 7.70-7.67 (m, 2H), 7.5 (dd, J = 2.0, 10.0 Hz, 1H), 6.57 (d, J = 10.0 Hz, 1H), 5.57 (t, J = 5.0 Hz, 1H), 4.04 (t, J = 10.0 Hz, 2H), 3.41-3.37 (m, 2H), 2.12-2.06 (m, 2H); 13C NMR (125 MHz, CDCl3) δ = 154.8, 148.0, 140.9, 137.3, 133.5, 133.4, 132.7, 131.1, 125.2, 114.8, 113.4, 82.9, 66.1, 40.8, 29.2 ppm。IR (KBr) λmax = 3440, 3091, 2935, 1852, 1547 cm -1 。ESI-MS (m / z): [M+Na] + calcd for C 15 H 14 81 BrIN2O5NaS, 564.8730; found, 564.8951。

[0060] 5-6. THF (20 mL) and diisopropylamine (20 mL) were added to a 100 mL round-bottom flask, and argon gas was bubbled through the solution for 30 min. Compound 16 (784 mg, 1.45 mmol), triphenylsilylacetylene (0.28 mg, 1.0 mmol), Pd(PPh3)4 (57.9 mg, 50.0 μmol), and CuI (4.80 mg, 25.0 μmol) were added to the solution, and the reaction mixture was stirred at room temperature for 168 h. The resulting precipitate was removed by filtration through Celite. The filtrate was concentrated on a rotary evaporator and purified by silica gel column chromatography (eluent: CHCl2) to obtain the crude product. This crude product was used in the next step without further purification. A solution of this crude product in THF (10 mL) was added with a THF solution of tetrabutylammonium fluoride (1 M, 2.0 mL, 2.0 mmol) and stirred at room temperature for 24 h. The resulting mixture was concentrated on a rotary evaporator and purified by silica gel column chromatography (eluent: CHCl) to obtain crude product 17, which was used in the next step without further purification. A 50 mL round-bottom flask was charged with THF (10 mL) and diisopropylamine (10 mL), and the solution was bubbled with argon gas for 30 min. The crude product 17, compound 16 (540 mg, 1.0 mmol), Pd(PPh) (57.9 mg, 50.0 μmol), and CuI (4.80 mg, 25.0 μmol) were added to the solution, and the reaction mixture was stirred at room temperature for 168 h. The resulting precipitate was removed by filtration through Celite. The filtrate was concentrated on a rotary evaporator and purified by silica gel column chromatography (eluent: hexane / ethyl acetate = 1:1) to obtain compound 18 (279 mg) as a white solid in 66% yield (three steps).

[0061] 5-7. The physical properties of compound 18 are as follows: Melting point 195.2-196.3°C. 1H NMR (500 MHz, d6-DMSO) δ = 8.15 (br, 2H), 8.00-7.94 (m, 4H), 7.83-7.80 (m, 4H), 7.74 (d, J = 2.0 Hz 2H), 7.49 (dd, J = 2.3, 8.6 Hz, 2H) 7.04 (d, J = 8.6 Hz, 2H) 4.08 (t, J = 5.7 Hz, 4H) 3.14 (t, J = 6.9 Hz, 4H) 1.94-1.89 (m, 4H). 13 C NMR (125 MHz, d6-DMSO) δ = 155.9, 147.7, 135.3, 134.0, 132.6, 132.4, 132.1, 129.5, 115.7, 113.5, 111.0, 87.6, 65.7, 28.8 ppm. IR (KBr) λmax = 3326, 3095, 2936, 2884 cm -1 . ESI-MS (m / z): [M+Na]+ calcd for C 32 H 28 79 Br 81 BrNO 10 NaS 2, 874.9493; found, 874.8555.

[0062] Compound 18 (279 mg, 0.33 mmol), propargyl bromide (90 μL, 1.2 mmol), KCO (0.14 g, 1.0 mmol), and DMF (2 mL) were added to a 5-mL round-bottom flask and stirred at room temperature for 24 h. The resulting mixture was concentrated on a rotary evaporator and purified by silica gel column chromatography (eluent: hexane / ethyl acetate = 1:1) to give compound 19 (235 mg) as a yellow solid in 77% yield.

[0063] 5-9. The physical properties of compound 19 are as follows: Melting point: 124.6-126.6°C. 1H NMR (500 MHz, CDCl3) δ = 8.09 (d, J = 8.0 Hz, 2H), 7.67 (d, J = 2.0 Hz, 2H), 7.61-7.54 (m, 6H), 7.35 (dd, J = 2.5, 8.6 Hz 2H), 6.70 (d, J = 8.6 Hz, 2H), 4.27 (d, J = 2.3 Hz, 4H), 4.01 (t, J = 5.7 Hz, 4H), 3.70 (t, J = 6.9 Hz, 4H), 2.24 (t, J = 2.3 Hz 2H), 2.19-2.14 (m, 4H)。 13 C NMR (125 MHz, CDCl3) δ = 154.9, 147.9, 136.0, 133.5, 132.4, 131.8, 131.2, 124.1, 116.7, 112.3, 111.8, 87.5, 74.0, 65.3, 43.5, 36.7, 27.0 ppm。IR (KBr) λmax = 3430, 3263, 3095, 2934, 2879, 2117 cm -1 。ESI-MS (m / z): [M+Na]+ calcd for C 38 H 32 79 Br 81 BrN4O 10 NaS2, 950.9807; found, 950.9368。

[0064] 5-10. To a solution of compound 19 (0.13 mg, 0.14 mmol) in DMF (10 mL), p-toluenethiol (0.12 g, 1.0 mmol) and LiOH·HO (0.15 g, 3.0 mmol) were added, and the mixture was stirred at room temperature for 24 h. Water (100 mL) was added to the resulting residue, which was then extracted with CHCl (100 mL). The organic layer was dried over sodium sulfate and concentrated on a rotary evaporator. To a solution of the residue in CHCl (10 mL), a solution of HCl in cyclopentyl methyl ether (4 M, 2.0 mL) was slowly added to form a precipitate. The precipitate was collected by filtration and washed with CHCl (3 mL) to give compound 20 (77 mg) as a tan solid in 88% yield (two steps).

[0065] 5-11. The physical properties of compound 20 are as follows: Melting point 209.8-210.6°C. 1 H NMR (500 MHz, CD3OD) δ = 7.71 (d, J = 1.8 Hz, 2H), 7.48 (dd, J = 2.5, 8.6 Hz, 2H), 7.08 (d, J = 8.6 Hz, 6H), 4.24 (t, J = 5.7 Hz 4H), 4.02 (d, J = 2.3 Hz, 4H) 3.40 (t, J = 7.5 Hz, 4H), 3.28 (t, J = 2.9 Hz, 2H), 2.29-2.24 (m, 4H). 13 C NMR (125 MHz, CD3OD) δ = 133.4, 118.5, 114.2, 112.6, 88.3, ​​67.6, 37.7, 26.9 ppm. IR (KBr) λmax = 3244, 2960, 2783, 2426, 2133 cm -1 ESI-MS (m / z): [M] 2+ calcd for C 26 H 28 79 Br 81 BrN2O2, 280.0250; found, 280.3348.

[0066] 5-12. 6-Me-β-CD was synthesized according to a non-patent document (GU-Barretta et al. Carbohydr. Res. 2005, 340, 271-281). A solution (1 mL) of compound 20 (0.628 mg, 1.00 μmol) and 6-Me-β-CD (18 mg, 15 μmol) shown in Figure 11 was stirred at 60 °C for 30 min and then cooled to 25 °C. Cucurbit[6]uril (11.6 mg, 8.40 μmol) and the raw material molecule 6 (0.57 mg, 2.1 μmol), which serves as the stopper shown in Figure 11, were added to the solution, and the mixture was stirred at 25 °C for 48 h. The resulting solution was filtered through a membrane filter (PTFE, pore size: 0.22 mm) and purified by reverse-phase HPLC to give the rotaxane-type dibromotran (3.5 mg) as a white solid in 64% yield.

[0067] 5-13. The physical properties of rotaxane-type dibromotlane are as follows: Decomposition point: 296.3°C. 1 H NMR (500 MHz, D2O) δ = 7.66 (s, 2H), 7.40 (d, J = 8.5 Hz, 2H), 6.92 (d, J = 8.5 Hz, 2H), 6.85 (d, J = 2.3 Hz 4H), 6.55 (t, J = 2.3 Hz, 2H), 6.44 (s, 2H), 5.72 (d, J = 16.0 Hz, 12H), 5.61 (d, J = 15.5 Hz, 12H), 5.38 (s, 24H), 4.90 (s, 14H), 4.46-4.40 (m, 4H), 4.25-4.10 (m, 32H), 3.80-3.71 (m, 48H), 3.60-3.38 (m, 64H), 3.21 (s, 42H). 13 C NMR (125 MHz, D2O) δ = 174.0, 171.1, 72.1, 70.8, 70.2, 58.3, 55.6, 51.5 ppm. IR (KBr) λmax = 3392, 2935, 2838 cm -1. ESI-MS (m / z): [M+4H]4+ calcd for C 218 H 302 79 Br 81 BrN 58 O 100 , 1373.7189; found, 1373.7222. 6. Synthesis of water-soluble dibromotrane (comparative example) As a comparative example of rotaxane-type dibromotlane, dibromotlane derivatives with water-soluble substituents were synthesized. 6-1. Dibromotolane derivative 25, which had a water-soluble substituent introduced therein, was synthesized in four steps from 2-bromo-4-iodophenol 12 and triethylene glycol monotosylate (compound 21) via the route shown in Figure 10. Triethylene glycol monotosylate (compound 21) was synthesized according to a non-patent document (Z. Hou et al. Org. Lett. 2022, 24, 1448-1453).

[0068] 6-2. Compound 12 (3.80 g, 11.4 mmol), triethylene glycol monotosylate (compound 21) (3.80 g, 12.5 mmol), K2CO3 (4.70 g, 34.2 mmol), and acetone (50 mL) were added to a 100 mL round-bottom flask, and the mixture was refluxed for 20 h. The resulting mixture was cooled to room temperature, and the solvent was evaporated. Water (100 mL) was added to the resulting residue, which was then extracted with ethyl acetate (50 mL × 3). The organic layer was dried over sodium sulfate and concentrated on a rotary evaporator. The resulting residue was purified by silica gel column chromatography (eluent: ethyl acetate) to give compound 22 (3.6 g) as a colorless oil in 73% yield.

[0069] 6-3. The physical properties of compound 22 are as follows: 1H NMR (500 MHz, CDCl3) δ = 7.82 (d, J = 1.7 Hz, 1H), 7.53 (dd, J = 2.5, 8.6 Hz, 1H), 6.68 (d, J = 8.6 Hz, 1H), 4.3, J 2.16 (H0), J = = 4.5 Hz, 2H), 3.79 (t, J = 4.5 Hz, 2H), 3.76–3.68 (m, 4H), 3.62 (t, J = 4.5 Hz, 2H), 2.26 (t, J = 6.3 Hz, 1H)。 13 C NMR (125 MHz, D2O) δ = 155.4, 141.1, 137.3, 115.4, 113.6, 83.0, 72.5, 71.3, 70.6, 69.5, 69.3, 61.9 ppm (C max = 3061, 2935, 2879, 2527, 1860, 1735 cm -1 。ESI-MS (m / z): [M+Na]+ calcd for C 12 H 16 79 BrIO4Na, 452.91743; found, 452.9996。

[0070] 6-4. THF (20 mL) and diisopropylamine (20 mL) were added to a 100 mL round-bottom flask, and argon gas was bubbled through the solution for 30 min. Compound 22 (1.29 g, 3.00 mmol), triphenylsilylacetylene (569 mg, 2.00 mmol), Pd(PPh3)4 (116 mg, 100 μmol), and CuI (7.6 mg, 40 μmol) were added to the solution, and the reaction mixture was stirred at room temperature for 48 h. The resulting precipitate was removed by filtration through Celite. The filtrate was concentrated on a rotary evaporator and purified by silica gel column chromatography (eluent: hexane / ethyl acetate = 1:4) to give crude product 23. This crude product was used in the next step without further purification. A solution of crude product 23 in THF (15 mL) was added with a THF solution of tetrabutylammonium fluoride (1 M, 3.4 mL, 3.4 mmol) and stirred at room temperature for 24 h. The resulting mixture was concentrated on a rotary evaporator and purified by silica gel column chromatography (eluent: ethyl acetate) to give crude product 24, which was used in the next step without further purification. A 50 mL round-bottom flask was charged with THF (10 mL) and diisopropylamine (10 mL), and the solution was bubbled with argon gas for 30 min. To the solution were added crude product 24, 22 (503 mg, 1.17 mmol), Pd(PPh3)4 (68 mg, 58 μmol), and CuI (4.5 mg, 23 μmol). The reaction mixture was stirred at room temperature for 168 h. The resulting precipitate was removed by filtration through Celite, and the filtrate was concentrated on a rotary evaporator and purified by silica gel column chromatography (eluent: methanol / ethyl acetate = 0:1 to 1:10) to give compound 25 (450 mg) as a brown oil in 36% yield (3 steps).

[0071] 6-5. The physical properties of compound 25 are as follows: 1H NMR (500 MHz, CDCl3) δ = 7.70 (d, J = 1.5 Hz, 2H), 7.39 (dd, J = 2.0, 8.6 Hz, 2H), 6.87 (d, J = 8.6 Hz, 2H), 4.21 (t, J = 4.8 Hz, 4H), 3.94 (t, J = 5.0 Hz, 4H), 3.82 (t, J = 4.5 Hz, 4H),3.76-3.70 (m, 8H), 3.63 (t, J = 4.5 Hz, 4H), 2.26 (t, J = 6.3 Hz, 2H). 13 C NMR (125 MHz, D2O) δ= 136.2, 131.8, 112.9 72.4, 71.4, 70.5. 69.4, 69.1, 61.8 ppm. IR (NaCl) λ max = 3413, 2924, 2873 cm -1 . ESI-MS (m / z): [M+Na]+ calcd for C 26 H 32 79 Br 81 BrO8Na, 655.0343; found, 655.1520.

[0072] 7. Optical properties of the synthesized rotaxane The absorption and emission of the β-[5]-rotaxane (Example 1), γ-[5]-rotaxane (Example 2), and [3]-rotaxane (Comparative Example) synthesized in 1 and 2 were evaluated. The results are shown in Figure 12. Measurement of the UV-visible absorption spectra of molecules suspended in water at 2.0 μM with a path length of 10 mm at 25°C revealed no significant differences. Among these, γ-[5]-rotaxane exhibited the sharpest waveform. No significant differences were observed in the emission spectra when molecules suspended in water at 15 μM were irradiated with 543 nm excitation light at 25°C. The emission quantum yields were approximately 1-2%. These results demonstrate that the rotaxane-type porphyrins of Examples 1 and 2 maintain the optical properties of porphyrins, regardless of whether they are inclusively bound to CDs.

[0073] 8. Singlet Oxygen Generation Efficiency Evaluation Next, to evaluate the photosensitizing activity of the rotaxane-type porphyrins of Examples 1 and 2, their singlet oxygen generation efficiency was measured. For the measurement, ABDA (9,10-anthracenediyl-bis(methylene)dimalonic acid; a singlet oxygen detection probe), which loses its absorption and fluorescence by capturing singlet oxygen, was used. As a control molecule, TPPS (Tetraphenylporphyrin Tetrasulfonic Acid), which has a porphyrin skeleton but does not aggregate in water, was used. At 6.0 × 10 -5 M ABDA and 1.0 x 10 -5 The rotaxane porphyrin of Example 1 or the rotaxane porphyrin of Example 2 or TPPS was mixed in M, and each mixed solution was irradiated with light from a 631 nm LED lamp. The singlet oxygen generation efficiency was estimated from the change in absorbance of ABDA at 380 nm after the addition of Examples 1 and 2. The results are shown in Figure 13.

[0074] In the mixed solution to which only ABDA was added, there was no change in the absorption spectrum of ABDA. On the other hand, when the rotaxane-type porphyrins of Examples 1 and 2 were added, the absorbance of ABDA decreased. No significant difference was observed in the degree of disappearance between Examples 1 and 2. The degree of disappearance was similar to that of TPPS, a porphyrin dye that does not aggregate in water, indicating that the molecules of Examples 1 and 2 do not aggregate in water even in the rotaxane-formed state, and energy transfer to oxygen occurs. It was also shown that their singlet oxygen generation efficiency is high, comparable to that of TPPS.

[0075] To evaluate the photosensitizing activity of the rotaxane-type acridine of Example 3, the hydroxyl radical generation efficiency was measured. For the measurement, HPF (hydroxyphenylfluorescein; a hydroxyl radical detection probe), which enhances luminescence by capturing hydroxyl radicals, was used. As a control, water-soluble acridine derivative 11 (Figure 8) was used. 1.0 × 10-5 M HPF and 11(1.0×10 -5 M) or the rotaxane-type acridine of Example 3 (1.2 × 10 -5 The water-soluble acridine derivatives 11 and 12 were mixed so that their absorbances at 365 nm were equal, and each mixed solution was irradiated with light at 365 nm using a UV lamp. The hydroxyl radical generation efficiency was estimated from the change in emission intensity at 515 nm after the addition of water-soluble acridine derivative 11 and Example 3. The results are shown in Figure 14.

[0076] In the mixed solution containing only HPF, there was almost no change in the luminescence intensity. On the other hand, when the rotaxane-type acridine of Example 3 was added, the luminescence intensity at 515 nm increased. The degree of this increase was faster than that of the soluble acridine derivative 11. These results demonstrate that the molecules of Example 3 do not aggregate in water due to rotaxane formation and can achieve high hydroxyl radical generation efficiency.

[0077] The photosensitizing activity of the rotaxane-type dibromotrans of Example 4 was evaluated using ABDA in the same manner as in the evaluation of the photosensitizing activity of the rotaxane-type porphyrins of Examples 1 and 2. As a control, water-soluble dibromotrans derivative 25 (Figure 10) was used. 6.0 x 10 -5 M ABDA and 1.0 x 10 -5 The rotaxane-type dibromotrane of Example 4 or the water-soluble dibromotrane derivative 25 was mixed with M, and each mixed solution was irradiated with 313 nm light from a high-pressure mercury lamp. The singlet oxygen generation efficiency was estimated from the change in absorbance of ABDA at 380 nm after the addition of Example 4. The results are shown in Figure 15.

[0078] When the water-soluble dibromotrans derivative 25 was added, the There was no difference in the change in absorption spectrum between the two cases. The reason why the water-soluble dibromotran derivative 25 did not exhibit photosensitizing activity may be that the molecule aggregated in water or that the molecule decomposed due to the singlet oxygen it generated itself or other factors. On the other hand, when the rotaxane-type dibromotran of Example 4 was added, the absorbance of ABDA decreased compared to the mixed solution containing only ABDA. The molecules of Example 4 did not aggregate in water due to rotaxane formation, demonstrating that they could achieve high singlet oxygen generation efficiency.

[0079] According to a non-patent document (Y. Ohishi et al., Adv. Opt. Mater. 2024, 12, 2301457), a rotaxane-type pyrene having a pyrene backbone (Example 5), a rotaxane-type anthracene having an anthracene backbone (Example 6), and a rotaxane-type perylene having a perylene backbone (Example 7) were synthesized, and their singlet oxygen generation efficiencies were evaluated using ABDA in the same manner as in the evaluation of the sensitizing activity of the rotaxane-type porphyrins in Examples 1 and 2. Water-soluble pyrene control molecules, anthracene control molecules, and perylene control molecules were used as comparative controls. The results of Example 5 are shown in Figure 16, the results of Example 6 in Figure 17, and the results of Example 7 in Figure 18.

[0080] The absorbance of ABDA decreased more rapidly when any of the rotaxanes from Examples 5, 6, and 7 was added than when the corresponding control molecule was added. The molecules from Examples 5, 6, and 7 were shown to be less aggregated in water and more stable through rotaxanation, achieving high singlet oxygen generation efficiency. These results demonstrate that rotaxanation can improve the photosensitizer activity of a variety of molecules.

[0081] 9.Photostability evaluation To evaluate the stability of the porphyrin core against light when exposed to excitation light for a long period of time, the molecules of Examples 1 and 2 and the Comparative Example were each diluted to 2.0 × 10 -6An aqueous solution (25°C) containing 1,000 mg of benzophenone-3 ...

[0082] In the comparative example, the absorption spectrum at 407 nm derived from the porphyrin skeleton was attenuated, whereas the rotaxane-type porphyrins of Examples 1 and 2 encapsulated with CDs showed little change in the absorption spectrum at the same wavelength. In other words, in the comparative example without CDs, the porphyrin core was decomposed by excitation light, whereas in the examples in which the porphyrin skeleton was encapsulated with CDs, it was revealed that stability against excitation light irradiation was improved. Furthermore, the γ-[5]-rotaxane of Example 2 showed the highest stability. This suggests that stability varies depending on the size of the CDs.

[0083] The light stability of the rotaxane-type acridine of Example 3 and the rotaxane-type dibromotran of Example 4 was evaluated in the same manner as in Examples 1 and 2. Aqueous solutions (25°C) containing Examples 3 and 4, and control molecule 9 (Figure 9) and control molecule 25 (Figure 10) were irradiated with light from a high-pressure mercury lamp (Example 3) or a handy UV lamp (Example 4), and the changes in the absorption spectra were monitored. The results are shown in Figure 20.

[0084] The decay of the absorption spectrum derived from the sensitizing organic group was slower when the rotaxanes of Examples 3 and 4 were used than when the corresponding control molecules were used. In other words, the sensitizing organic group in the control molecule without CD decomposed due to the excitation light, whereas the examples in which the sensitizing organic group was encapsulated by CD showed improved stability against excitation light irradiation. These results demonstrate that rotaxane formation can improve the photostability of various molecules.

[0085] 10.Excitation Lifetime Evaluation To investigate the energy transfer to oxygen in more detail, the excitation lifetime was measured before and after degassing. Before degassing, the rate of deactivation by oxygen (energy transfer to oxygen molecules) could be investigated, while after degassing, the rate of deactivation independent of oxygen (thermal deactivation) could be investigated. After excitation with a 532 nm laser, the change in absorption in the excited state was tracked. The results are shown in Table 1.

[0086] [Table 1]

[0087] A significant difference in excitation lifetime was observed before degassing (in the presence of oxygen) and after degassing (in the absence of oxygen). This confirms that even in the rotaxane state where collisions with oxygen are prevented by CD, deactivation by oxygen molecules occurs exclusively in the presence of oxygen molecules. This suggests that rotaxane formation does not inhibit energy transfer to oxygen, and that rotaxane formation does not reduce the efficiency of generating reactive oxygen species. Furthermore, under both pre- and post-degassing conditions, Examples 1 and 2 exhibited longer excitation lifetimes than the Comparative Example, suggesting that rotaxane formation slows the rate of energy transfer to oxygen. From this, it was predicted that in the Examples where the porphyrin skeleton is not exposed, as in the Comparative Example, the porphyrin skeleton is protected from various factors that deactivate molecules, resulting in an extended excitation lifetime, and that the presence of CD reduces the probability of collisions with surrounding oxygen.

[0088] It is believed that for energy transfer to occur to oxygen, the porphyrin skeleton and oxygen molecule must approach each other at a distance of 1 nm or less. The presence of CD between the oxygen molecule and the porphyrin skeleton increases the physical distance between them, reducing the probability that they will approach each other at the distance of 1 nm or less, which is the distance at which energy transfer occurs. This decrease in probability is thought to be a disadvantage in terms of inducing energy transfer to oxygen, but this is compensated for by the extended excitation lifetime of the rotaxane porphyrin, and energy is ultimately transferred to oxygen, so the efficiency of singlet oxygen generation is thought not to have decreased. The large physical distance between the oxygen molecule and the porphyrin skeleton is also thought to have contributed to the improved photostability in the examples, as described in Section 5 above.

[0089] 11. Evaluation of cell transfection efficiency (cell membrane permeability) Using the preparations of Examples 1 and 2 and talaporfin sodium as a control, the extent to which they were introduced into cells was evaluated. -6 Each compound in M ​​was added to HeLa cells and cultured for 24 hours at 37°C under 5% CO2 conditions. After culture, the cells were irradiated with light at an excitation wavelength of 560 ± 40 nm, and fluorescence images at a fluorescence wavelength of 630 ± 75 nm were observed under a fluorescence microscope. The phase contrast images (top) and fluorescence images (bottom) are shown in Figure 21 (left). Next, for more quantitative evaluation, flow cytometry was used. The cells were detached by treatment with 0.25% trypsin-EDTA solution and subjected to flow cytometry. The excitation wavelength used for detection was 488 nm, and the fluorescence wavelength was 670-735 nm. The cell counts versus fluorescence intensity under each condition are shown in Figure 21 (right).

[0090] The fluorescence microscopy images and flow cytometry results showed that the fluorescent intensities of all of the present examples were higher than those of the control, talaporfin sodium, indicating that the rotaxane structure improved cell permeability. This indicates that the rotaxane porphyrin of the present invention has achieved appropriate water solubility and cell membrane permeability. Furthermore, the cells luminesced most brightly in the β-[5]-rotaxane of Example 2, indicating that Example 2 had the highest cell membrane permeability. Quantitative evaluation by flow cytometry also showed that the cells to which Example 2 was added had the highest fluorescence intensity, followed by the cells of Example 1. These results demonstrate that differences in CD size also affect cell permeability.

[0091] These results demonstrate that the present example shows that the porphyrin remains stable without decomposition, photobleaching, or aggregation even after being taken up into cells, demonstrating the advantages of the rotaxane-type molecule of the present invention in which the porphyrin is irreversibly encapsulated in CD.

[0092] 12. Cell Death (Photocytotoxicity) Assessment To examine whether the rotaxane-type porphyrins prepared in Examples 1 and 2 can be used in photodynamic therapy, cells transfected with these porphyrins were exposed to excitation light, and the WST-8 assay was used to confirm whether cell death actually occurred. -8 Talaporfin sodium (control), Examples 1 and 2 were added to HeLa cells at a concentration of 100 μM and cultured for 24 hours at 37°C under 5% CO2. A 490 mW LED lamp with a wavelength of 405 nm was used as the light source, and light irradiation was performed at 25°C for 30 minutes. After light irradiation, the cells were cultured for 2 hours at 37°C under 5% CO2, and then a WST-8 assay was performed. The results are shown in Figure 22.

[0093] When cell viability was compared for each compound concentration, a lower cell viability (higher photocytocidal activity) was observed in the present Examples compared to the existing drug, talaporfin sodium. In particular, in Example 1, which had a high intracellular introduction efficiency as described in 11 above, cell death was observed at low concentrations of the compound. These results demonstrate that the rotaxane molecules of Examples 1 and 2 introduced into cells induce the death of surrounding cells upon light irradiation, and because their photocytocidal activity was higher than that of existing drugs, the rotaxane molecules of the present invention are expected to be useful for PDT in clinical settings.

Claims

1. A photosensitizer for photodynamic therapy containing a rotaxane-type reactive oxygen generator.

2. 2. The photosensitizer according to claim 1, wherein the rotaxane-type reactive oxygen generator is a [3]-rotaxane that is encapsulated by two cyclodextrins or derivatives thereof.

3. The photosensitizer according to claim 2, wherein the rotaxane-type reactive oxygen generator is a [5]-rotaxane in which the axis between cyclodextrin or its derivative and the stopper is encapsulated with cucurbituril.

4. The photosensitizer of claim 3, wherein the cucurbituril is cucurbit[6]uril.

5. 5. The photosensitizer according to claim 1, wherein the active oxygen generator is any one of porphyrin, coumarin, chlorin, bacteriochlorin, acridine, methylene blue, phthalocyanine, cyanine, tolan, and derivatives thereof.

6. The axis of the rotaxane-type reactive oxygen generator is represented by the following formulas (1) to (6): 【Chemistry 1】 (In the formula, R 1 are the same or different and represent an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, A represents an organic group capable of generating active oxygen, n represents 0 or 1, and L represents a divalent hydrocarbon group having 1 to 12 carbon atoms which may be substituted with a hetero atom. 【Chemistry 2】 (In the formula, R 2 are the same or different and represent an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, and A represents an organic group capable of generating active oxygen. 【Transformation 3】 (In the formula, R 3 are the same or different and represent an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, A represents an organic group capable of generating active oxygen, o represents an integer of 1 to 12, and p represents 0 or 1. 【Chemistry 4】 (In the formula, R 4 are the same or different and represent an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, A represents an organic group capable of generating active oxygen, q represents an integer of 0 to 4, and r represents 0 or 1. 【Transformation 5】 (In the formula, R 5 are the same or different and represent an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, A represents an organic group capable of generating active oxygen, s represents an integer of 0 to 6, t represents an integer of 0 to 6, and u represents 0 or 1. 【Transformation 6】 (In the formula, R 6 are the same or different and represent an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group, A represents an organic group capable of generating active oxygen, v represents 0 to 6, w represents an integer of 0 to 6, and x represents 0 or 1).

2. The photosensitizer according to claim 1, wherein the photosensitizer is represented by any one of the following formulas:

7. The stopper of the rotaxane type active oxygen generator is represented by the following formula (7): 【Transformation 7】 (In the formula, R 7 represents an optionally substituted monovalent aryl group or nitrogen-containing heterocyclic group.

2. The photosensitizer of claim 1, wherein

8. The photosensitizer according to claim 1, which is an anticancer agent.

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