JP7332148B2 - Dinucleated ligands or binuclear metal complexes - Google Patents

Description

The present invention relates to a dinucleating ligand or a binuclear metal complex having the dinucleating ligand.

Cisplatin is a metal complex that is clinically used as a chemotherapeutic agent for cancer. Cisplatin exhibits anticancer effects by binding to the DNA of cells and distorting the conformation of the DNA. However, cisplatin also acts on the DNA of normal cells and may show side effects such as vomiting and nephrotoxicity.In recent years, platinum-resistant cancers have also been reported. In order to solve these problems, development of an anticancer agent to replace cisplatin is required.

For example, bleomycin binds to iron in cancer cells to activate oxygen, thereby severing DNA strands and suppressing cancer cell proliferation (Non-Patent Document 1). Bleomycin is widely used in clinical medicine as an excellent chemotherapeutic agent for squamous cell carcinomas of human skin, head and neck, uterine cervix, and malignant lymphoma. However, bleomycin is a water-soluble glycopeptide antibiotic obtained from the actinomycete Streptomyces Verticillus, and there is a demand for a chemotherapeutic drug that can be obtained by a simple synthetic method that does not depend on microorganisms.

An iron complex of the N4Py ligand that mimics the active center of bleomycin has been reported to react with hydrogen peroxide to form an active species that oxidatively cleaves DNA and exhibits anticancer activity. (Non-Patent Document 2). However, since this iron complex exhibits high DNA cleavage activity even in the absence of hydrogen peroxide, it does not exhibit selectivity between normal and cancer cells. Therefore, it is necessary to develop a metal complex that has selectivity for normal cells and cancer cells.

Cancer cells have higher concentrations of ROS such as hydrogen peroxide (H ₂ O ₂ ) than normal cells. Therefore, metal complexes that promote DNA oxidative cleavage by H ₂ O ₂ are attracting attention for the development of anticancer drugs that selectively kill cancer cells without affecting normal cells. The inventors have prepared a new dinucleated ligand dinuclear copper complex [Cu ₂ (μ-OH)(bcamide)](ClO ₄ ) ₂ in which cyclen is introduced at the 2,6-positions of p-cresol through an amide bond. We developed (1) and succeeded in oxidative cleavage of DNA by H ₂ O ₂ (Patent Document 1). However, this complex has low cytotoxicity against HeLa cells, which are cancer cells, and needs to be improved.

Japanese Patent Application Laid-Open No. 2018-135304

H. Umezawa, K. Maeda, T. Takeuchi, Y. Okami, J. Antibiot., 19A, 200 (1966). Q. Li, M. G. P. Wijst, H. G. Kazemier, M. G. Rots, G. Roelfes, ACS Chem. Biol., 9, 1044-1051 (2014).

The present invention has been made in view of such problems, and provides a dinucleated ligand or a binuclear metal complex having the dinucleated ligand, which can be easily synthesized and has a precise anticancer effect. intended to provide

The dinucleating ligand according to the present invention is represented by the following chemical formula (I).

Here, R ₁ , R ₂ , R ₃ , R ₄ , R ₅ and R ₆ are each independently a hydrogen atom or a linear or branched alkyl group having 1 to 8 carbon atoms. R ₇ is an R polycyclic aromatic heterocyclic compound or polycyclic aromatic hydrocarbon compound and n is an integer of 1-8.

A binuclear metal complex according to the present invention is represented by the following chemical formula (III).

Here, M is Cu, Fe, Zn, Co, Mn, or Ce. R is a polycyclic aromatic heterocyclic compound or a polycyclic aromatic hydrocarbon compound, and n is an integer of 1-8.

INDUSTRIAL APPLICABILITY According to the present invention, it is possible to easily obtain a dinucleating ligand or a binuclear metal complex which has little effect on normal cells and has an appropriate nucleic acid cleaving action on cancer cells.

1 is a ¹ H NMR spectrum of a dinucleated ligand according to the present invention; FIG. FIG. 2 is a diagram showing ESI-MS of a binuclear metal complex according to the present invention; FIG. 2 shows the DNA oxidative cleavage activity of the binuclear metal complex according to the present invention. FIG. 2 shows the DNA oxidative cleavage activity of the binuclear metal complex according to the present invention. FIG. 2 shows cytotoxicity of binuclear metal complexes according to the present invention.

Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings. The embodiments are intended to facilitate understanding of the principles of the present invention, and the scope of the present invention is as follows. The scope of the present invention is not limited to the embodiments, and other embodiments in which the configurations of the following embodiments are appropriately replaced by those skilled in the art are also included in the scope of the present invention.

As a result of intensive studies, the present inventors have found as a new finding that a dinucleating ligand represented by the following formula has a high nucleic acid cleaving activity, and have completed the present invention based on this fact.

Here, R ₁ , R ₂ , R ₃ , R ₄ , R ₅ and R ₆ are each independently a hydrogen atom or a linear or branched alkyl group having 1 to 8 carbon atoms. _R7 is a polycyclic aromatic heterocyclic compound or a polycyclic aromatic hydrocarbon compound. A polycyclic aromatic heterocyclic compound is a compound that is tricyclic or more and contains one or more heterocycles, and has a π-conjugated system with a highly planar structure. Polycyclic aromatic hydrocarbon compounds are hydrocarbon compounds in which aromatic rings containing no heteroatoms are condensed, and are preferably tricyclic or higher compounds. n is an integer of 1 to 8, preferably 1 to 3, more preferably 2.

_R7 is set to facilitate cellular entry of the complex and facilitate DNA oxidative cleavage of _H2O2 by the complex _. The polycyclic aromatic heterocyclic compound is not particularly limited, but is, for example, acridine, xanthene or carbazole, preferably acridine shown below. The bonding site of the polycyclic aromatic heterocyclic compound is not particularly limited. It can be a binding site, preferably at the 9-, 1- or 8-position carbon atom.

The polycyclic aromatic hydrocarbon compound is not particularly limited, but is, for example, phenanthrene, pyrene, anthracene, tetracene, pentacene, benzopyrene, chrysene, triphenylene, corannulene, coronene or ovalene, preferably shown below. It is a phenanthrene that is The bonding site of the polycyclic aromatic hydrocarbon compound is not particularly limited. It can be the binding site, preferably at the 9- or 10-position carbon atom as shown in the examples below.

Note that _R7 is set to facilitate cellular _introduction of the complex and DNA oxidative cleavage of _H2O2 by the complex, but the polycyclic aromatic heterocyclic compound or polycyclic aromatic hydrocarbon In addition to the compounds, minor groove binders can also be used, and specific examples include distamycin A and netropsin. Molecules that bind in the minor groove of the DNA duplex have sequence selectivity. Therefore, it is possible to selectively act on abnormal genes, leading to reduction of side effects of anticancer drugs.

The dinucleating ligand represented by the above formula is a compound with a novel structure having an amide bond in the compound structure. As will be described later, the dinucleating ligand and binuclear metal complex according to the present invention are capable of cleaving nucleic acids only by reacting the compound with H ₂ O ₂ , but the amide bond portion is H ₂ O ₂ Since the dinucleating ligand and the binuclear metal complex according to the present invention are not oxidatively degraded by , they are capable of stably cleaving nucleic acids.

In the present invention, a dinucleating ligand represented by the following chemical formula (II) is preferred.

In addition, the present inventors have found, as a new finding, that a binuclear metal complex represented by the following formula has a high nucleic acid cleaving activity. Here, M is Cu, Fe, Zn, Co, Mn or Ce, preferably Cu.

In the present invention, a binuclear metal complex represented by the following chemical formula (VI) is preferred.

In the above, the nucleic acid to be cleaved is DNA or RNA.

In addition, the dinucleating ligand and binuclear metal complex according to the present invention can cleave nucleic acids of cancer cells, and thus can be used as anticancer agents. Cancer cells have higher H ₂ O ₂ concentrations than normal cells due to mitochondrial dysfunction and abnormal metabolism. In addition, in normal cells, catalase, an H ₂ O ₂ degrading enzyme, reduces the H ₂ O ₂ concentration. However, cancer cells lack catalase and cannot break down H ₂ O ₂ like normal cells. In addition, superoxide dismutase (SOD), which dismutates superoxide ions to produce H ₂ O ₂ , is highly activated, and cancer cells have a higher H ₂ O ₂ concentration than normal cells. The binucleating ligand and binuclear metal complex according to the present invention are capable of cleaving nucleic acids only by the reaction between the compound and H ₂ O ₂ , and are capable of specifically cleaving nucleic acids of cancer cells. Therefore, according to the present invention, there is little effect on normal cells. In addition, the dinucleated ligand and binuclear metal complex according to the present invention bind H ₂ O ₂ with two metal ions (for example, two copper ions), and thus have high affinity for H ₂ O ₂ . Therefore, even when used in vivo, it reacts with a small amount of H ₂ O ₂ and exhibits high nucleic acid cleaving activity.

In addition, although the oxidative cleavage of DNA by H 2 O 2 was high with only the dinuclear copper complex of the new dinucleated ligand in which cyclen was introduced at the 2 and 6 positions of p-cresol with amide groups, it was found that the oxidative cleavage of DNA by H ₂ O ₂ was high. Although the cytotoxicity to cells was low, in the dinucleating ligand represented by the above chemical formula (I), introduction of an aromatic group such as phenanthrene, pyrene, acridine, etc. that can be used for cell introduction and DNA targeting at R ₇ exhibits higher cytotoxicity and higher DNA oxidative cleavage activity.

The dinucleated ligand and binuclear metal complex according to the present invention can be used for various cancers, and are not particularly limited. Examples include colon cancer, stomach cancer, esophageal cancer, Colon cancer, liver cancer, pancreatic cancer, breast cancer, lung cancer, gallbladder cancer, bile duct cancer, biliary tract cancer, rectal cancer, ovarian cancer, uterine cancer, kidney cancer, bladder cancer, prostate cancer cancer, osteosarcoma, brain tumor, leukemia, sarcoma, skin cancer, malignant melanoma, malignant lymphoma, tongue cancer, myeloma, thyroid cancer, skin metastasis, skin melanoma, etc. .

The dosage form of the anticancer agent having a dinuclear ligand and a binuclear metal complex according to the present invention is not particularly limited, and may be either oral or parenteral dosage form. In addition, suitable dosage forms can be prepared according to the dosage form, for example, oral preparations such as injections, capsules, tablets, granules, powders, pills, fine granules, rectal preparations, and oily suppositories. , can be prepared into various formulations such as aqueous suppositories.

Various formulations can be prepared by appropriately adding pharmacologically acceptable additives such as excipients, binders, lubricants, disintegrants, surfactants, flow enhancers and the like. Excipients such as lactose, fructose, glucose, corn starch, sorbit, etc. Binders such as methyl cellulose, ethyl cellulose, gum arabic, gelatin, hydroxypropyl cellulose, polyvinylpyrrolidone, etc. Lubricants such as talc, magnesium stearate, polyethylene glycol. Disintegrants such as starch, sodium alginate, gelatin, calcium carbonate, calcium citrate, dextrin, magnesium carbonate, synthetic magnesium silicate, etc. Surfactants such as sodium lauryl sulfate, soybean lecithin, sucrose fatty acid ester, polysorbate 80. For example, light anhydrous silicic acid, dried aluminum hydroxide gel, synthetic aluminum silicate, magnesium silicate, etc. can be used as a fluidity enhancer.

The dosage of the anticancer agent having a dinuclear ligand and a binuclear metal complex according to the present invention is appropriately determined in consideration of usage, patient's age, sex, severity of symptoms, and the like. The daily dosage for adults is 10-800 mg, preferably 100-200 mg, which can be administered once or in several divided doses per day.

(1-1) 1,10-ditosyl-1,4,7,10-tetraoxadecane
A rotor, triethylene glycol (31.0 mL, 0.233 mol), p-toluenesulfonyl chloride (87.0 g, 0.456 mol), and CH ₂ Cl ₂ (750 mL) were added to a 2000 mL three-necked reaction vessel. This was immersed in an ice bath and stirred, powdered KOH (110 g, 1.96 mol) was added little by little, a balloon filled with N ₂ was attached, and the mixture was stirred for 3 hours while being kept at 0°C. H ₂ O (450 mL) was added to the reaction vessel, this was separated with CH ₂ Cl ₂ (3 × 225 mL), the organic layer was dehydrated by adding Na ₂ SO ₄ , filtered through Nutsche, and a small amount was collected. of CH ₂ Cl ₂ and the filtrate was collected and concentrated on a rotary evaporator to give a white solid. This was dissolved in hot acetone and recrystallized to give a white solid (92.5 g, Yield 88%).

(1-2) 1,8-diazido-3,6-dioxaoctane
A rotator, 1,10-ditosyl-1,4,7,10-tetraoxadecane (46.7 g, 0.102 mol), tetrabutylammonium iodide (TBAI, 1.94 g, 5.26 mmol), sodium azide (27.1 g, 0.417 mol) and DMF (150 mL) were added, shielded from light, degassed and replaced with nitrogen, and stirred at 80° C. for 24 hours. After returning the reaction vessel to room temperature, the DMF was removed with a rotary evaporator. Et ₂ O (340 mL) was added and insoluble salts were filtered off with a Nutsche, the filtrate was partitioned with H ₂ O (3 × 135 mL), the organic layer was dehydrated with Na ₂ SO ₄ , and filtered with a Nutsche. Filtered, rinsed with a little Et ₂ O, and rotary evaporated the filtrate to give a yellow oil (18.5 g, Yield 91%).

(1-3) 1-amino-8-azido-3,6-dioxaoctane
A rotor, 1,8-diazido-3,6-dioxaoctane (18.5 g, 92.4 mmol), EtOAc (130 mL), 1 M HCl (164 mL) was added to a 1000 mL eggplant flask. A 200 mL pressure equalizing dropping funnel was attached to this, and a solution of triphenylphosphine (PPh ₃ , 23.2 g, 88.5 mmol) in EtOAc (130 mL) was added thereto and slowly added dropwise with vigorous stirring. After 15 hours, the EtOAc layer was removed and the remaining aqueous layer was partitioned with EtOAc (3 x 100 mL). The EtOAc layer was removed, the pH _{of this} aqueous layer was adjusted to 14 with 1 M NaOH aqueous solution, and the mixture was separated with CHCl ₃ (3 × 240 mL). Filter and concentrate the filtrate on a rotary evaporator to give a yellow oil (13.6 g, Yield 84%).

(1-4) 3,5-diformyl-4-hydroxybenzoic acid
A rotator, CF ₃ COOH (TFA, 180 mL), 4-hydroxybezoic acid (10.1 g, 73.1 mmol), and hexamethylenetetramine (HNT, 84.1 g, 0.600 mol) were added to a 1 L three-necked reaction vessel and placed in an oil bath at 110°C. The mixture was shielded from light and refluxed for 2 days. After returning to room temperature, 4 M HCl (450 mL) was added, stirred overnight at 30° C., and filtered through Nutsche while washing with H ₂ O to give an orange filtrate. Vacuum drying gave a yellow solid (9.88 g, Yield 70%).

(1-5) N-(8-azido-3,6-dioxaoctyl)-3,5-diformyl-4-hydroxybenzamide
A 2 L three-necked reaction vessel was equipped with a three-way cock and a balloon, immersed in an ice bath, and dissolved by adding a rotor, 3,5-diformyl-4-hydroxybenzoic acid (4.57 g, 23.5 mmol), and CHCl ₃ (700 mL). 1-amino-8-azido-3,6-dioxaoctane (13.6 g, 78.1 mmol) was dissolved in CHCl ₃ (100 mL) and added, EDC·HCl (15.0 g, 78.2 mmol), Et ₃ N (10.7 mL, 76.8 mmol) was added. After CHCl ₃ (120 mL) was added to degas and replace with nitrogen, the mixture was stirred for 24 hours. After that, 1 M HCl (480 mL) was added and vigorously stirred at room temperature. The reaction was followed by TLC (silica gel, developing solvent: EtOAc/MeOH = 10:1) to confirm that there were almost no by-products, then transferred to a 2 L separatory funnel, the organic layer was taken out, and Na ₂ SO ₄ was added. After dehydration with , it was filtered with Nutsche and washed with a small amount of CHCl ₃ . The filtrate was collected, concentrated with a rotary evaporator, and dried under vacuum. This was dissolved in CHCl ₃ (120 mL) and separated and washed with H ₂ O (3×40 mL). After dehydrating the organic layer with Na ₂ SO ₄ , it was filtered with a Nutsche, and the filtrate was concentrated with a rotary evaporator and dried under vacuum to obtain a brown solid (6.52 g, Yield 79%).

(1-6) 5-(8-azido-3,6-dioxaoctyl)carbamoyl)-2-hydroxyisophthalic acid
A rotor, Ag ₂ O (13.1 g, 56.4 mmol), and N-(8-azido-3,6-dioxaoctyl)-3,5-diformyl-4-hydroxybenzamide (6.52 g, 18.6 mmol) were added to a 300 mL eggplant flask. NaOH (6.34 g, 158 mmol) dissolved in H ₂ O (130 mL) was added thereto and stirred overnight at 60°C. This was filtered through a Kiriyama funnel while washing with a minimal amount of hot H ₂ O (25 mL), and the filtrate was brought to pH 1 using 12 M HCl while immersed in an ice bath to give a white precipitate. This was filtered through a Kiriyama funnel and dried in vacuo to give a white solid (4.72 g, Yield 66%).

(1-7) N,O,O-tritosyl diethanolamine
A rotor was placed in a 300 mL eggplant flask, a three-way cock and a balloon were attached, and vacuum drying was performed. The reaction vessel was immersed in an ice bath, and p-toluenesulfonyl chloride (57.4 g, 0.301 mol) and CH ₂ Cl ₂ (70 mL) were added and dissolved. A 200 mL equal pressure dropping funnel, 3-way stopcock and balloon were attached to this and 2,2'-iminodiethanol (10.7 g, 0.102 mol), benzyltriethylammonium chloride (9.14 g, 40.1 mmol) and NaOH (12.0 g, 0.300 mol) were added to H2O. The solution in ₂ O (75 mL) was placed in a pressure equalizing addition funnel and slowly added dropwise. After completion of dropping, the mixture was stirred at room temperature until heat generation subsided. The reaction solution was transferred to a 300 mL separatory funnel, the organic layer was taken, and it was separated and washed with H ₂ O (3×50 mL). The organic layer was dehydrated by adding Na ₂ SO ₄ , filtered through Nutsche, washed with a small amount of CH ₂ Cl ₂ , the filtrate was collected, concentrated with a rotary evaporator, and dried under vacuum to obtain a clear oily substance. A suitable amount of MeOH was added to this and vigorously stirred to precipitate a white solid, which was filtered through a Nutsche to obtain a white solid (35.1 g, Yield 61%).

(1-8) 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetratosylate
A rotor was placed in a 1 L three-necked reaction vessel, and a 200 mL pressure equalizing dropping funnel, a three-way cock and a balloon were attached, followed by vacuum drying. The reaction vessel was immersed in an ice bath, diethylene-1,4,7-triamine tritosylate (36.1 g, 63.8 mmol), DMF (150 mL), and NaH (approximately 60% in oil, 5.08 g, 0.212 mol) were added for desorption. After purging with gas and nitrogen, the mixture was stirred until generation of H ₂ gas ceased. After that, the reaction vessel was immersed in an oil bath, the temperature was slowly raised to 70°C, the mixture was stirred for about 30 minutes, then the temperature was slowly raised to 90°C, and the mixture was stirred for about 30 minutes. A solution of N,O,O-tritosyl diethanolamine (36.1 g, 63.6 mmol) in DMF (150 mL) was added to a pressure equalizing dropping funnel and slowly added dropwise to the reaction solution. After the dropwise addition was completed, stirring was continued overnight. After removing DMF under reduced pressure, it was filtered through Nutsche while washing with copious amounts of H ₂ O, Et ₂ O, EtOH, and dried in vacuo to give a white solid (48.2 g, Yield 96%).

(1-9) 1,4,7,10-tetraazacyclododecane tetrahydrochloride
A rotor was placed in a 1 L eggplant flask, a reflux tube, a three-way cock, and a balloon were attached, followed by vacuum drying. 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetratosylate (48.1 g, 61.0 mmol) and conc. H ₂ SO ₄ (35 mL) were added to the reactor and stirred at 150°C for 1 hour. did. After returning to room temperature, EtOH (260 mL) and Et ₂ O (420 mL) were slowly added dropwise in this order. After filtering the resulting gray precipitate with a Nutsche, the resulting solid was transferred to a 500 mL Meyer and dissolved in a minimal amount of _H2O . This was filtered through celite, rinsed with a small amount of H ₂ O, and the filtrate was collected and concentrated to give a black oil. To this was added 12 M HCl (50 mL), EtOH (300 mL) and stirred to give a gray solid (9.41 g, Yield 48%).

(1-10) 1,4,7-tetraazacyclododecane
Dowex ^™ 1×8 200-400 mesh strongly basic type I anion exchange resin (Cl type) was placed in a 300 mL beaker and dissolved in 1 M HCl. After filtering this with a Nutsche, the obtained filtrate was dissolved in distilled water and poured into a chromatographic tube packed with cotton and sea sand. 1 M HCl was added until the pH of the drops was 1 and then the pH was brought to 5 using distilled water. Subsequently, 1 M NaOH was added until the pH reached 12, and then the pH was brought to 7 using distilled water. A solution of 1,4,7,10-tetraazacyclododecane tetrahydrochloride (1.50 g, 4.72 mmol) dissolved in a minimum amount of distilled water was added thereto, and pH basic drops were collected. The solution was concentrated on a rotary evaporator and dried under vacuum to give a yellow solid. Dissolve this in _CHCl3 , add _Na2SO4 to dehydrate, filter with Nutsche, rinse with a small amount of _CHCl3 , collect _the filtrate, concentrate with a rotary evaporator, vacuum dry, and leave in the refrigerator. A yellow solid was then obtained (0.772 g, Yield 95%).

(1-11) N,N',N”-4,7,10-tris(tert-butoxycarbonyl)-1,4,7,10-tetraazacyclododecane
A rotor was placed in a 300 mL eggplant flask, a three-way cock and a balloon were attached, and vacuum drying was performed. The reaction vessel was immersed in an ice bath and 1,4,7,10-tetraazacyclododecane (3.22 g, 18.7 mmol), _CHCl3 (90 mL), _Et3N (7 mL, 50.2 mmol) were added. A 100 mL equal pressure dropping funnel was attached to this and a solution of di-tert-butyl dicarbonate (Boc ₂ O, 11.0 g, 50.4 mmol) in CHCl ₃ (70 mL) was added and added dropwise over 6 hours. Separate with H ₂ O (3 × 50 mL), add Na ₂ SO ₄ to the organic layer to dehydrate, filter with Nutsche, rinse with a small amount of CHCl ₃ , collect the filtrate, and concentrate on a rotary evaporator. and vacuum dried to give a white solid. This was purified by silica gel column chromatography (CHCl ₃ /MeOH 100:1), and fractions containing the desired product were collected and concentrated with a rotary evaporator to give a white solid (3.62 g, Yield 41%).

(1-12) 5-(8-azido-3,6-dioxaoctyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris(tert-butoxycarbonyl)-1 ,4,7,10-tetraazacyclododecanyl)-1-carbamoyl)-2-hydroxybenzene
A rotor was placed in a 300 mL eggplant flask, a three-way cock and a balloon were attached, and vacuum drying was performed. 5-(8-azido-3,6-dioxoaoctyl)carbamoyl)-2-hydroxyisophthalic acid (2.36 g, 6.17 mmol) and SOCl ₂ (90 mL) were added to a reaction vessel and reacted at 60°C for 4 hours. After removing SOCl ₂ under reduced pressure, it was dissolved in CH ₂ Cl ₂ (120 mL) and N,N',N”-4,7,10-tris(tert-butoxycarbonyl)-1,4,7,10- Tetraazacyclododecane (6.77 g, 14.3 mmol), _K2CO3 (7.67 g _, 55.5 mmol), and _CH2Cl2 (240 mL) were added to a 500 mL three-neck reaction vessel under _{N2 atmosphere} , followed by degassing and nitrogen replacement. After stirring overnight at room temperature, the reaction mixture was filtered through a Kiriyama funnel and rinsed with a small amount of CH ₂ Cl ₂ , the filtrate was collected, washed with H ₂ O (3 × 200 mL) and washed with an organic solvent. The layer was dehydrated with Na ₂ SO ₄ , filtered through a Nutsche and rinsed with a small amount of CH ₂ Cl ₂ , the filtrate was collected, concentrated and dried in vacuo to give a brown solid which was flashed. Purified by column chromatography (silica gel, developing solvent: gradient CHCl ₃ /MeOH from 150/1 to 100/1) Collecting the eluent containing the desired product and concentrating with a rotary evaporator gave a white solid. (2.59 g, Yield 32%).

(1-13) 5-(8-amino-3,6-dioxaoctyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris(tert-butoxycarbonyl)-1 ,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene
A 200 mL eggplant flask was equipped with a three-way cock with a nitrogen-purged balloon, and the rotor and raw material 5-(8-azido-3,6-dioxaoctyl)carbamoyl)-1,3-di((N,N',N ”-4,7,10-tris(tert-butoxycarbonyl)-1,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene (414.1 mg, 0.32 mmol) and 10% Pd-C (350.9 mg) were added. , dry MeOH (25 mL) was added.The entire reaction vessel was thoroughly degassed and replaced with nitrogen, then degassed and replaced with hydrogen, and stirred overnight under a hydrogen atmosphere.The progress of the reaction was monitored by TLC (silica gel, developing solvent: CHCl ₃ /MeOH = 100 : 1) and ESI-MS spectrum tracking to confirm the absence of raw materials, Celite filtration was performed, the filtrate was concentrated with a rotary evaporator, and dried in vacuo to obtain a brown solid. (341.4 mg, Yield 84%).

(1-14) phenanthrene-9-carboxylic acid
A 500-mL three-necked flask was equipped with a three-way cock equipped with a nitrogen-purged balloon, and a rotor and raw material 9-bromophenanthrene (5.18 g, 20.1 mmol) were placed. A stopper was attached, deaeration was performed, and a nitrogen atmosphere was established. Dry Et ₂ O (200 mL) was added using a syringe under nitrogen and cooled to −30° C. using an EtOH bath. The temperature was kept at -30°C and n-BuLi (15 mL) was added using a syringe. After reacting with stirring for 1 hour, the nitrogen balloon was replaced with a CO ₂ gas balloon, and the mixture was vigorously stirred for about 1 hour and 30 minutes to react. The reaction vessel was then transferred to an ice bath, H ₂ O (90 mL) was added and allowed to stir overnight. Stirring was stopped, 1 M NaOH was added to the reaction mixture to adjust the pH to 11, and the mixture was separated and washed with EtOAc (2 × 200 mL) and hexane (2 × 200 mL). M HCl was added to adjust the pH to 1 and a white solid precipitated out. This was collected by suction filtration and dried in vacuo to give a white solid (1.23 g). Considering that the amount of product was small, the organic layer at the time of liquid separation was concentrated with a rotary evaporator and dried under vacuum to obtain a white solid (2.64 g). When each ¹ HNMR spectrum was measured and confirmed, all of them were found to be the desired product (Fig. 1). These were collected to give a white solid (3.87 g, Yield 86%).

(1-15) phenanthrene-9-carbonyl chloride
Add 2 drops of rotator, phenanthrene-9-carboxylic acid (199.8 mg, 89.9 mmol), SOCl ₂ (3.5 mL), and DMF to a 30 mL eggplant flask using a Pasteur, then attach a Dimroth and calcium chloride tube. , and stirred in an oil bath at 60 °C. After stirring for 3 hours, the mixture was returned to room temperature, and SOCl ₂ was distilled off using an aspirator to produce a yellow solid. When a small amount of benzene is added to the solid, an insoluble salt is generated. Using a Kiriyama funnel and a suction bottle, suction filtration is performed. 95%) was obtained.

(1-16)5-(9-phenanthrene-8-carbamoyl-3,6-dioxaoctyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris(tert- butoxycarbonyl)-1,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene
5-(8-amino-3,6-dioxaoctyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris(tert -butoxycarbonyl)-1,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene (341.4 mg, 0.27 mmol) was added, dissolved in dry THF (25 mL), and then Et ₃ N (0.1 mL) was added. The solution turned orange, and phenanthrene-9-carbonyl chloride (62.6 mg, 0.26 mmol) dissolved in THF (5 mL) was slowly added with Pasteur in an ice bath with vigorous stirring. After the addition, a three-way cock equipped with a nitrogen-purging balloon was attached, and after degassing and purging with nitrogen, the mixture was stirred while immersed in an ice bath, stirred for 1 hour, returned to room temperature, and stirred overnight. Evaporation yielded an orange oil, which was dissolved in CHCl ₃ (30 mL) and H ₂ O (15 mL), partitioned with CHCl ₃ (2×30 mL) and separated into 3 CHCl ₃ layers ( 90 mL) was further washed with H ₂ O (2×30 mL), the organic layer was dried by adding Na ₂ SO ₄ , suction filtered using Nutsche, and dried in vacuo to give a brown solid.

This was purified by silica gel column chromatography (CHCl ₃ :MeOH=100:1), concentrated by a rotary evaporator, and dried under vacuum to give a white solid (257.7 mg, Yield 63%).

(1-17) [3,5-di(1,4,7,10-tetraazacyclododecane-1-carboxyamide)-4-hydroxybenzenecarboxy]-(phenanthrene-9-carboxy)-3,6-dioxaoctane-1,8- N,N'-diamide・6HCl
5-(9-phenanthrene-8-carbamoyl-3,6-dioxaoctyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris (tert-butoxycarbonyl)-1,4,7,10-tetraazacyclo-dodecanyl)carbamoyl)-2-hydroxybenzene (248.6 mg, 0.17 mmol) was added and dissolved in EtOH (10 mL). M HCl (3.5 mL) was slowly added, and the mixture was stirred for 2 days.After confirming that no starting material remained by ESI-MS, it was concentrated by rotary evaporator to obtain a white solid (180.6 mg, Yield 98 %).

(1-18) 1,13-ditosyl-1,4,7,10,13-pentaoxadecane
A rotator was placed in a 500 mL eggplant flask, and tetraethylene glycol (19.6 g, 0.10 mol), p-toluenesulfonyl chloride (40.5 g, 0.21 mol), and CH ₂ Cl ₂ (300 mL) were added to the reaction vessel. This was stirred while immersed in an ice bath, powdered KOH (43.1 g, 0.77 mol) was added little by little, a three-way cock with a nitrogen-purging balloon was attached, and after deaeration and nitrogen substitution, the temperature was lowered to 0°C. Stirring was continued for 3 hours. H ₂ O (160 mL) was added to the reaction vessel, this was separated with CH ₂ Cl ₂ (3 × 100 mL), the organic layer was dehydrated by adding Na ₂ SO ₄ , filtered through Nutsche, and a small amount was collected. of CH ₂ Cl ₂ , the filtrate was collected, and the filtrate was concentrated on a rotary evaporator to give a colorless oil (56.6 g, Yield quant.).

(1-19) 1,11-diazido-3,6,9-trioxaundecane
A rotor, 1,13-ditosyl-1,4,7,10,13-pentaoxaundecane (56.6 g, 113 mmol), tetrabutylammonium iodide (TBAI, 2.04 g, 5.52 mmol), sodium azide (22.8 g , 351 mmol) and DMF (180 mL) were added, and the mixture was shielded from light and replaced with nitrogen. After the reaction vessel was returned to room temperature, DMF was distilled off using a rotary evaporator. Et ₂ O (350 mL) was added and insoluble salts were filtered through Nutsche, the filtrate was separated with H ₂ O (3 × 120 mL), Na ₂ SO ₄ was added to the organic layer to dehydrate, and then Nutsche was added. was suction filtered using a small amount of Et ₂ O, the filtrate was concentrated on a rotary evaporator and dried in vacuo to give a yellow oil (22.4 g, Yield 81%).

(1-20) 1-amino-11-azido-3,6,9-trioxaundecane
A rotor, 1,11-diazido-3,6,9-trioxaundecane (22.4 g, 91.8 mmol), EtOAc (130 mL), 1 M HCl (160 mL) was added to a 500 mL eggplant flask. A 200 mL pressure equalizing dropping funnel was attached to this, and a solution of triphenylphosphine (PPh ₃ , 24.0 g, 91.5 mmol) in EtOAc (130 mL) was added thereto and slowly added dropwise with vigorous stirring. After 15 hours, the EtOAc layer was removed and the remaining aqueous layer was partitioned with EtOAc (2 x 100 mL). The EtOAc layer was removed, the pH of _this aqueous layer _was adjusted to 11 with 1 M NaOH aqueous solution, and the mixture was separated with CHCl ₃ (3 × 120 mL). The filtrate was concentrated on a rotary evaporator and dried in vacuo to give a yellow oil (16.5 g, Yield 82%).

(1-21) N-(11-azido-3,6,9-trioxaundecanyl)-3,5-diformyl-4-hydroxybenzamide
A 1 L eggplant flask was immersed in an ice bath, and a rotor, 3,5-diformyl-4-hydroxybenzoic acid (2.64 g, 13.6 mmol) and CHCl ₃ (370 mL) were added and dissolved. 1-amino-11-azido-3,6,9-trioxaundecane (16.5 g, 75.4 mmol) was dissolved in CHCl ₃ (150 mL) and added, EDC·HCl (8.15 g, 42.5 mmol), Et ₃ N (6.0 mL, 43.1 mmol) was added. A three-way cock with a nitrogen-purged balloon was attached, and after deaeration and nitrogen substitution, the mixture was stirred for 24 hours. After that, 1 M HCl (270 mL) was added and vigorously stirred at room temperature. After tracking the reaction with TLC (silica gel, developing solvent: EtOAc/MeOH = 10:1) and confirming that there was almost no starting material, the organic layer was removed by transferring to a 2 L separatory funnel, and Na ₂ SO ₄ was added. After dehydration, it was filtered through Nutsche, washed with a small amount of CHCl ₃ , and the filtrate was collected, concentrated in a rotary evaporator, and dried in vacuum. This was dissolved in CHCl ₃ (90 mL) and separated and washed with H ₂ O (3×30 mL). After dehydrating the organic layer with Na ₂ SO ₄ , it was filtered with a Nutsche, and the filtrate was concentrated with a rotary evaporator and dried under vacuum to obtain a brown solid (3.57 g, Yield 67%).

(1-22) N-(11-azido-3,6,9-trioxaundecanyl)carbamoyl)-2-hydroxyisophthalic acid
Rotor, N-(11-azido-3,6,9-trioxaundecanyl)-3,5-diformyl-4-hydroxybenzamide (3.57 g, 9.05 mmol), Ag ₂ O (6.51 g, 28.1 mmol) in a 300 mL eggplant flask. ) to which NaOH (3.39 g, 84.8 mmol) dissolved in H ₂ O (65 mL) was added and stirred at 60° C. overnight. This was washed with the minimum amount of hot H ₂ O (13 mL) and suction filtered using a Kiriyama funnel. The filtrate was immersed in an ice bath and adjusted to pH 1 with 12 M HCl to give a white precipitate. was taken. This was suction filtered using a Kiriyama funnel and dried in vacuo to give a white solid (1.94 g, Yield 50%).

(1-23)5-((11-azido-3,6,9-trioxaundecanyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris(tert-butoxycarbonyl )-1,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene
A rotor was placed in a 100 mL two-necked flask, a three-way cock and a balloon were attached, and the mixture was vacuum-dried. N-(11-azido-3,6,9-trioxaundecanyl)carbamoyl)-2-hydroxyisophthalic acid (280.3 mg, 0.66 mmol), 1-[bis(dimethylamino)methylene]-1H-benzotriazolium 3-oxide tetrafluoroborate were added to the reaction vessel. (TBTU, 666.1 mg, 2.07 mmol), dry DMF (20 mL) was added and dissolved. Boc ₃ cyclen (1.04 g, 2.20 mmol) was dissolved in dry DMF (10 mL), added to the reaction vessel, and N,N-diisopropylethylamine (DIPEA, 0.8 mL, 4.46 mmol) was added. After degassing and purging with N ₂ , the mixture was stirred overnight at room temperature while shielded from light. After confirming the absence of starting material by ¹ HNMR, a brown liquid was obtained by concentrating with a rotary evaporator. H ₂ O (60 mL) and CHCl ₃ (60 mL) were added to this, and after separation with CHCl ₃ (3 × 60 mL), separation and washing with H ₂ O (3 × 60 mL) were carried out, and the organic layer was _Na2SO4 was added _for dehydration. Nutsche filtration and vacuum drying gave a brown foamy solid. This was purified by silica gel column chromatography (CHCl ₃ :MeOH=150:1→100:1), concentrated by a rotary evaporator, and dried under vacuum to give a brown solid (383.6 mg, Yield 44%).

(1-24)5-(11-amino-3,6,9-trioxaundecanyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris(tert-butoxycarbonyl) -1,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene
5-(11-azido-3,6,9-trioxaundecanyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris(tert -butoxycarbonyl)-1,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene (363.4 mg, 0.27 mmol) and Pd-C (308.3 mg) were added, and dry MeOH (20 mL) was added. A cock and a balloon were attached, and the entire reaction vessel was thoroughly degassed and replaced with nitrogen, and then degassed and replaced with hydrogen, followed by stirring overnight under a hydrogen atmosphere _. : 1) and after confirming the absence of raw materials by tracing with ESI-MS spectrum, Celite filtration was performed, and the filtrate was concentrated with a rotary evaporator and dried under vacuum to obtain a brown solid (331.8 mg, Yield 93% ).

(1-25) 5-(9-phenanthrene-11-carbamoyl-3,6,9-trioxaundecanyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris( tert-butoxycarbonyl)-1,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene
A rotor, 5-(11-amino-3,6,9-trioxaundecanyl)carbamoyl)-1,3-di((N,N',N”-4,7,10-tris (tert-butoxycarbonyl)-1,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene (331.8 mg, 0.25 mmol) was added and dissolved in dry THF (20 mL), followed by Et ₃ N (0.15 mL). While vigorously stirring in an ice bath, a solution of phenanthrene-9-carbonyl chloride (53.8 mg, 0.22 mmol) in THF (5 mL) was slowly added with a Pasteur. , A three-way cock and a balloon were attached, and after purging with degassed nitrogen, the mixture was stirred while immersed in an ice bath.After stirring for 1 hour, the mixture was returned to room temperature and stirred overnight.When THF was distilled off with a rotary evaporator, an orange oily substance was obtained. The material was dissolved in CHCl ₃ (30 mL) and H ₂ O (15 mL), partitioned with CHCl ₃ (2×30 mL), and the CHCl ₃ layer (90 mL) was further washed with H ₂ O ( 2 × 30 mL), and the organic layer was dehydrated by adding Na ₂ SO ₄ , filtered through Nutsche, and dried under vacuum to obtain a brown solid, which was subjected to silica gel column chromatography (CHCl ₃ :MeOH = 100 :1 → 80:1), rotary evaporated and dried in vacuo to give a white solid (309.2 mg, Yield 81%).

(1-26) [3,5-di(1,4,7,10-tetraazacyclododecane-1-carboxyamide)-4-hydroxybenzenecarboxy]-(phenanthrene-9-carboxy)-3,6,9-trioxaundecane-1, 11-N,N'-diamide・6HCl
A rotor, 5-(9-phenanthrene-11-carbamoyl-3,6,9-trioxaundecanyl)carbamoyl)-1,3-di((N,N',N”-4,7,10 -tris(tert-butoxycarbonyl)-1,4,7,10-tetraazacyclododecanyl)carbamoyl)-2-hydroxybenzene (302.1 mg, 0.20 mmol) was added and dissolved in EtOH (10 mL). M HCl (4.5 mL) was slowly added, and the mixture was stirred for two days.After confirming that no raw material remained by ESI-MS, it was concentrated using a rotary evaporator and dried under vacuum to obtain a white solid (192.5 mg , Yield 85%).

(2-1) Synthesis of binuclear metal complex [Cu ₂ (μ-OH)(bcamide-PEG3-phen)](ClO ₄ ) ₂ (2)
[3,5-di(1,4,7,10-tetraazacyclododecane-1-carboxyamide)-4-hydroxybenzenecarboxy]-(phenanthrene-9-carboxy)-3,6-dioxaoctane-1,8- N,N'-diamide.6HCl (90.4 mg, 0.0831 mmol) was added. Weigh NaOH (193.4 mg), dissolve it in Mill-Q, add 507 μL of the solution adjusted to 1 M NaOHaq to a 100 mL round-bottomed flask, confirm that the pH is neutral, and dry in a vacuum. A white solid formed. This solid was dissolved in CH ₂ Cl ₂ and inorganic salts were removed by celite filtration. The filtrate was concentrated by an evaporator and dried under vacuum to produce a white solid.

Cu(ClO ₄ ) ₂ (72.4 mg, 0.195 mmol) was weighed and dissolved in MeOH (2 mL). A solution was put in a 100 mL eggplant flask, and a rotor was put therein. While stirring this solution, the dried white solid dissolved in MeOH (3 mL) was slowly added with a pasteur, and the solution changed from blue to pale green. A green precipitate formed at the same time. When 1 M NaOHaq was added little by little, the solution changed from light green to dark green. The dark green solution was sucked with Pasteur so as not to suck the precipitate, and transferred to another 100 mL eggplant flask. A rotator was placed in this eggplant flask, and Et ₂ O was added while stirring to produce a bluish-green insoluble matter. After confirming that no starting material remained by ESI-MS (Fig. 2), the insoluble matter was collected by suction filtration and dried in vacuo to give a bluish-green solid (82.1 mg, Yield 82%).

(2-2) Synthesis of binuclear metal complex [Cu ₂ (μ-OH)(bcamide-PEG4-phen)](ClO ₄ ) ₂ (3)
[3,5-di(1,4,7,10-tetraazacyclododecane-1-carboxyamide)-4-hydroxybenzenecarboxy]-(phenanthrene-9-carboxy)-3,6,9-trioxaundecane-1, 11-N,N'-diamide.6HCl (29.0 mg, 0.0256 mmol) was added. Weigh NaOH (380.4 mg), dissolve it in Mill-Q, adjust it to 1 M NaOHaq, add 162 μL of the solution to a 100 mL eggplant flask, confirm that the pH is neutral, and dry it in a vacuum. A white solid formed. This solid was dissolved in CH ₂ Cl ₂ and inorganic salts were removed by celite filtration. The filtrate was concentrated by an evaporator and dried under vacuum to produce a white solid.

Cu(ClO ₄ ) ₂ (21.6 mg, 0.582 mmol) was weighed and dissolved in MeOH (1 mL). A solution was put in a 100 mL eggplant flask, and a rotor was put therein. While stirring the solution, the dried white solid dissolved in MeOH (2 mL) was slowly added with a pasteur, and the solution turned from blue to pale green. A green precipitate formed at the same time. When 1M NaOHaq was added little by little, the solution changed from light green to dark green. The dark green solution was sucked with Pasteur so as not to suck the precipitate, and transferred to another 100 mL eggplant flask. A rotator was placed in this eggplant flask, and Et ₂ O was added while stirring to produce a bluish-green insoluble matter. The insolubles were collected by suction filtration and dried in vacuo to give a blue-green solid (14.2 mg, Yield 44%).

(3) Dinuclear copper complex [ _Cu2 (μ- _OH2 )bcamide]( _ClO4 ) ₂ (1), [ _Cu2 (μ-OH)(bcamide-PEG3-phen)]( _ClO4 ) ₂ (2) ) and [Cu ₂ (μ-OH)(bcamide-PEG4-phen)](ClO ₄ ) ₂ (3) on the H ₂ O ₂ concentration of the oxidative cleavage reaction. Oxidative cleavage of DNA with H ₂ O ₂ was performed as indicated. [NaCl] = 10 mM, [buffer] = 10 mM (pH 6.0 (MES)), [complex] = 10 μM, [pUC19 DNA] = 10 μM bp, [H ₂ O ₂ ] = 0 for this assay. A solution was prepared at 37°C so as to be -500 µM, and the measurement was performed. A portion of the reaction solution was taken every 0, 10, 20, 40, and 60 minutes in the reaction at a hydrogen peroxide concentration of 0-500 μM, and the state of pUC19 DNA cleavage at each time was analyzed by agarose gel electrophoresis. It was measured.

The results of the H ₂ O ₂ concentration dependence of the oxidative cleavage reaction of each complex are shown in FIGS. 3 and 4. FIG. In FIG. 3, (a) is blank, (b) is complex 1, (c) is complex 2, (d) is complex 3, and Form I over time (minutes) at pH 6.0. Decrease. In FIG. 4, (a) is complex 1, (b) is complex 2, and (c) is complex 3, showing the increase in Form II over time (minutes) at pH 6.0. In FIG. 4, (d) is complex 1, (e) is complex 2, and (f) is complex 3, showing the increase in Form III over time (minutes) at pH 6.0. It was found that no DNA was cleaved in the blank experiment with H ₂ O ₂ only. For each complex, it was found that the cleavage activity was greatly improved depending on the concentration of H ₂ O ₂ . Since complex 1 has _a dinuclear _structure , two copper ions bind to two oxygen atoms of _H2O2 , easily forming dinuclear copper-bridged hydroperoxo complexes even at low concentrations of _H2O2 . It has been found in our laboratory that it cuts DNA oxidatively. For the same reason, it is considered that complexes 2 and 3 also readily activate H ₂ O ₂ to oxidatively cleave DNA.

It was also found that the formation ratio of Form III was 14.2% for complex 1, 26.8% for complex 2, and 23.7% for complex 3 at an H ₂ O ₂ concentration of 500 μM. These results suggest that the dinuclear copper complexes 2 and 3, into which the DNA intercalator phenanthrene has been introduced, bind more strongly to DNA than complex 1, and their positions are fixed, and the DAN near the first cleaved position is blocked. It is thought that the high DNA oxidative cleavage activity was exhibited because a structure that could be cleaved again was formed. Furthermore, when comparing the ratios of formation of Form III between complexes 2 and 3, it was found that complex 2 showed a higher ratio. This is thought to be due to the length of the spacer introduced to bind phenanthrene. The spacer of complex 2 was shorter than that of complex 3, which allowed the bridging hydroperoxo complexes involved in DNA cleavage to be more proximal, resulting in improved double-stranded DNA cleavage activity, resulting in higher It is thought that they exhibited DNA oxidative cleavage activity.

Bleomycin catalyzes the oxidative cleavage of DNA. Bleomycin is an antibiotic (anticancer antibiotic) that exhibits high anticancer activity. This is because bleomycin oxidatively cleaves cancer cell DNA, causing cancer cell death. Here, bleomycin is an iron complex, and in the iron (III) state, it reacts with H ₂ O ₂ to rapidly produce active bleomycin called active bleomycin. It is believed to be a hydroperoxo complex of iron(III). Furthermore, it is presumed that cleavage of the OO bond of hydroperoxo generates oxidative species. Bleomycin also has a DNA minor groove binder and a spacer site for binding it. Thus, complexes 2 and 3, which have similar structures, are considered useful as a new anticancer agent to replace bleomycin, which uses H ₂ O ₂ as an oxidizing agent.

(4) Cytotoxicity of binuclear metal complexes 1, 2 and 3 Cytotoxicity of each complex was evaluated by MTT assay. MTT [3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide] taken up into cells is reduced by dehydrogenase in mitochondria to produce formazan dye. . Since the amount of dye correlates with the number of metabolically active cells, the number of viable cells was measured by quantifying this using a colorimetric method (absorbance at 570 nm). HeLa cells treated with MTT/culture solution (0.25 mg/0.5 mL) at 37° C. for 180 minutes were placed in a microtube, and the formalin pigment produced was extracted. After the extraction was completed, data was obtained by measuring absorbance at 570 nm using a spectrophotometer. The results are shown in FIG. Twenty-four hours after exposure of HeLa cells to each complex, the order of cytotoxicity was 2>3>>1 for complexes (Table 1).

As mentioned above, the difference between complexes 1, 2 and 3 is the presence or absence of phenanthrene. Thus, the introduction of phenanthrene significantly improved the cytotoxicity of complexes 2 and 3, probably because the hydrophobicity of the compounds was improved and the cell membrane permeability was increased. Also, 2 showed higher cytotoxicity than 3, probably due to the length of the spacer. This result is consistent with DNA oxidative cleavage experiments. That is, complex 2, which is capable of cleaving double-stranded DNA at a low intracellular concentration of H ₂ O ₂ , is considered to have exhibited the highest cell-specificity. Furthermore, these results suggest that complexes 2 and 3 permeate the HeLa cell membrane, enter the cell, and further enter the nucleus, thereby cleaving DNA and expressing cytotoxicity.

It can be used as an anticancer agent.

Claims (10)

A binucleated ligand represented by the following chemical formula (I) (wherein R ₁ , R ₂ , R ₃ , R ₄ , R ₅ and R ₆ are each independently a hydrogen atom or a linear or branched alkyl group having 1 to 8 carbon atoms, R ₇ is a polycyclic aromatic heterocyclic group or a polycyclic aromatic hydrocarbon group , and n is an integer of 1 to 8; .).

2. The binucleating ligand of claim 1, wherein said polycyclic aromatic heterocyclic group is acridine, xanthene, carbazole or porphyrin and derivatives thereof. 2. The binucleating ligand according to claim 1 , wherein the polycyclic aromatic hydrocarbon group is phenanthrene, pyrene, anthracene, tetracene, pentacene, benzopyrene, chrysene, triphenylene, corannulene, coronene or ovalene. . 2. The dinucleating ligand according to claim 1, characterized by being represented by the following chemical formula (II).

A nucleic acid cleaving agent comprising the dinucleating ligand according to any one of claims 1 to 4. An anticancer agent comprising the dinucleated ligand according to any one of claims 1 to 4. A binuclear metal complex characterized by being represented by the following chemical formula (III) (wherein (i) M is Cu, Fe, Zn, Co, Mn, or Ce, and (ii) R is a multi- a cyclic aromatic heterocyclic group or a polycyclic aromatic hydrocarbon group , and n is an integer of 1 to 8).

8. The binuclear metal complex according to claim 7, characterized by being represented by the following chemical formula (IV).

A nucleic acid cleaving agent comprising the binuclear metal complex according to claim 7 or 8. An anticancer agent comprising the binuclear metal complex according to claim 7 or 8.