A near-infrared light-excited iridium complex, its preparation method, and its application in anti-breast cancer treatment.

Abstract

This invention discloses a near-infrared light-excited iridium complex, its preparation method, and its application in anti-breast cancer treatment, comprising the structure shown in formula (I): The complex of the present invention exhibits a strong inhibitory effect on the growth and proliferation of mouse breast cancer cells (IC50). 50 The concentration of the phototoxicity was 0.31 μM, while under dark conditions, its cytotoxicity was only 93.9 μM, and the phototherapy index (PI) was as high as 301. This is of great significance for the research of metal drugs for anti-tumor purposes such as breast cancer.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical chemistry, and in particular to a near-infrared light-excited iridium complex, its preparation method, and its application in the treatment of breast cancer. Background Technology

[0002] Cancer poses a significant threat to human health, especially breast cancer, whose incidence has increased in recent years. Developing novel, highly effective, low-toxicity, and highly selective anti-tumor drugs has become a crucial strategic goal in the drug development strategies of governments worldwide. Photodynamic therapy (PDT), a treatment method based on the combination of photosensitizers and light irradiation at specific wavelengths, is widely considered a preferred approach for achieving highly effective, low-toxicity, and highly selective anti-tumor therapy. It utilizes photosensitizers to generate reactive oxygen species under specific wavelengths of light irradiation, thereby disrupting the structure and function of tumor cells.

[0003] Photosensitizers, when exposed to light, can generate highly reactive oxides, such as singlet oxygen and free radicals. These substances can trigger intracellular oxidative stress, leading to damage and death of tumor cells. Compared to traditional chemotherapy drugs, photodynamic therapy can more directly destroy tumor cells, thereby improving treatment efficacy. Furthermore, photosensitizers are inactive in the absence of light exposure and are only activated by light of specific wavelengths. This means that in the absence of light, photosensitizers do not have toxic effects on normal tissues, thus reducing damage to healthy tissues during treatment. In addition, photosensitizers are typically targeted to tumor cells or tumor tissues using specific targeting strategies, such as targeted antibodies and nanoparticles, to deliver the photosensitizers into the tumor cells. Therefore, photodynamic therapy can act more precisely on tumor tissues, minimizing damage to surrounding normal tissues. In conclusion, developing more photosensitizers plays a crucial role in achieving highly efficient, low-toxicity, and highly selective anti-tumor therapy. Compared to small organic molecules, transition metal iridium complexes possess advantages such as larger Stokes shift, stronger spin coupling, higher luminescence efficiency, longer phosphorescence lifetime, high triplet exciton generation efficiency, and simple and tunable color. They have been proven to be effective photosensitizers (PS) for photodynamic therapy (PDT). While some iridium complexes exist as photosensitizers in existing technologies, different types of tumors may respond differently to specific wavelengths and light irradiation conditions. Therefore, developing more photosensitizers is of great significance. Summary of the Invention

[0004] This invention aims to solve at least one of the technical problems existing in the prior art. To this end, this invention proposes a novel near-infrared light-excited iridium complex.

[0005] The present invention also proposes a method for preparing the above-mentioned iridium complex.

[0006] The present invention also proposes applications of the above-mentioned iridium complexes.

[0007] According to one aspect of the present invention, an iridium complex is provided, comprising the structure shown in formula (I):

[0008]

[0009] According to a preferred embodiment of the present invention, at least the following beneficial effects are achieved: the complex of the present invention exhibits a strong inhibitory effect on the growth and proliferation of mouse breast cancer cells (IC50). 50 The concentration of iridium complex in this invention is 0.31 μM, while under dark conditions, its cytotoxicity is only 93.9 μM, and its phototherapy index (PI) is as high as 301. This is of great significance for the research of metal drugs for anti-tumor purposes such as breast cancer. Compared with the traditional anti-tumor metal drug cisplatin, the iridium complex of this invention has a novel anti-tumor mechanism, which can overcome the drug resistance of traditional drugs. Its potential photodynamic therapy effect can effectively reduce the toxic side effects of the drug itself.

[0010] According to another aspect of the present invention, a method for preparing the above-mentioned iridium complex is provided, comprising the following steps:

[0011] S1. Iridium(III) trichloride and 2-phenylpyridine (bpy) are reacted under reflux to give iridium(III) μ-chloro-bridged dimer;

[0012] S2. React the iridium(III)μ-chloro-bridged dimer obtained in step S1 with 5-ethynyl-2,2'-bipyridine to generate an iridium complex intermediate;

[0013] S3. React the ruthenium complex intermediate from step S2 with 3,6-bis(5-bromo-2-thienyl)-2,5-bis(2-ethylhexyl)-2,5-dihydropyrrolo[3,4-C]pyrrolo-1,4-dione to obtain the iridium complex.

[0014] The preparation method according to a preferred embodiment of the present invention has at least the following beneficial effects: the preparation process of the present invention is simple, easy to operate, and has good prospects for industrial application.

[0015] In some embodiments of the present invention, in step S1, iridium(III) chloride hydrate, 2-phenylpyridine (bpy) and 2-ethoxyethanol-water mixture are mixed, refluxed under heating conditions, and the reaction product is cooled to room temperature.

[0016] In some preferred embodiments of the present invention, the volume ratio of 2-ethoxyethanol to water in the 2-ethoxyethanol-water mixture is 2 to 4:1; more preferably 3:1. 2-Ethoxyethanol is also known as ethylene glycol ethyl ether.

[0017] In some preferred embodiments of the present invention, the heating conditions refer to a temperature of 105–115°C, such as 110°C.

[0018] In some preferred embodiments of the present invention, the reflux time is 8 to 14 hours, such as 10 to 12 hours, more preferably 12 hours.

[0019] In some embodiments of the present invention, the molar ratio of 2-phenylpyridine (bpy) to iridium trichloride (III) in step S1 is 1.5 to 2.5:1; more preferably 2:1.

[0020] In some embodiments of the present invention, the reaction formula for step S1 is as follows:

[0021]

[0022] In some embodiments of the present invention, step S2 specifically involves dissolving iridium(III)μ-chloro-bridged dimer and 5-ethynyl-2,2'-bipyridine in a chloroform / methanol mixture, heating at 50-100°C overnight, cooling to room temperature, and then drying and purifying to obtain the iridium complex intermediate.

[0023] In some embodiments of the present invention, the molar ratio of the iridium(III)μ-chloro-bridged dimer to 5-ethynyl-2,2'-bipyridine in step S2 is 1:1.5 to 2.5; more preferably 1:2.

[0024] In some embodiments of the present invention, the volume ratio of chloroform to methanol in the chloroform / methanol mixed solution is 1.5 to 2.5:1; preferably 2:1.

[0025] In some embodiments of the present invention, the heating temperature in step S2 is 60°C.

[0026] In some embodiments of the present invention, the reaction time is 6-12 hours; preferably 8 hours.

[0027] In some embodiments of the present invention, the reaction formula for step S2 is as follows:

[0028]

[0029] In some embodiments of the present invention, step S3 specifically involves: under a protective atmosphere, the iridium complex, 3,6-bis(5-bromo-2-thienyl)-2,5-bis(2-ethylhexyl)-2,5-dihydropyrrolo[3,4-C]pyrrole-1,4-dione, and Pd(PPh3)2Cl2 are dissolved in toluene / triethylamine (V / V = 1.2-1.8:1, preferably 1.5 / 1), heated under reflux at 50-80°C with stirring for 24-36 hours, and after the reaction is completed, cooled to room temperature, and the solvent is evaporated and the product is purified to obtain the final product.

[0030] In some embodiments of the present invention, the protective atmosphere is an inert gas or nitrogen.

[0031] In some preferred embodiments of the present invention, the inert gas is argon.

[0032] In some embodiments of the present invention, the molar ratio of 3,6-bis(5-bromo-2-thienyl)-2,5-bis(2-ethylhexyl)-2,5-dihydropyrrolo[3,4-C]pyrrole-1,4-dione, iridium complex, and Pd(PPh3)2Cl2 in step S3 is 1:1.5 to 2.5:0.08 to 0.12; more preferably, it is 1:2:0.1.

[0033] In some embodiments of the present invention, the temperature of the heating reflux in step S3 is 60°C and the time is 32 hours.

[0034] In some embodiments of the present invention, the reaction formula for step S3 is as follows:

[0035]

[0036] According to another aspect of the present invention, the application of the above-mentioned iridium complex in the preparation of antitumor drugs is proposed.

[0037] According to a preferred embodiment of the present invention, the application has at least the following beneficial effects: the iridium complex of the present invention has good application prospects in the field of near-infrared excited antitumor drug preparation.

[0038] In some embodiments of the present invention, the tumor is breast cancer.

[0039] In some preferred embodiments of the present invention, the breast cancer is 4T1 cell line breast cancer.

[0040] According to another aspect of the present invention, an antitumor drug is provided, wherein the active ingredient of the antitumor drug comprises the above-mentioned iridium complex.

[0041] In some embodiments of the present invention, the tumor is breast cancer.

[0042] In some embodiments of the present invention, the tumor is triple-negative breast cancer.

[0043] In some embodiments of the present invention, the tumor is a mouse triple-negative breast cancer.

[0044] In some embodiments of the present invention, the cancer cells of the mouse triple-negative breast cancer are 4T1 cancer cells.

[0045] In some embodiments of the present invention, the drug (or photosensitizer) further includes a pharmaceutically acceptable carrier and / or excipient. That is, the drug or photosensitizer uses a binuclear ruthenium complex as the main active ingredient, is mixed with a pharmaceutically acceptable carrier and / or excipient to prepare a composition, and is then formulated into a clinically acceptable dosage form.

[0046] In some embodiments of the present invention, the excipient refers to a diluent, binder, lubricant, disintegrant, solubilizer, stabilizer, and other pharmaceutical matrix that can be used in the pharmaceutical field.

[0047] In some embodiments of the invention, the different pharmaceutical excipients used for the drug dosage form (or photosensitizer) may vary depending on the specific medical application. Pharmaceutical excipients can be used to adjust the solubility and bioavailability of the photocatalyst, increase its stability, modulate the host's immune response, and act as emulsifiers, antioxidants, aerosol propellants, tablet binders, and tablet disintegrants. Preferred pharmaceutical excipients include, but are not limited to, binders / fillers, coating agents, disintegrants, lubricants, and sweeteners adapted to the use of photosensitizers.

[0048] In some embodiments of the present invention, the carrier is a functional pharmaceutical excipient acceptable in the pharmaceutical field, including surfactants, suspending agents, emulsifiers, and some novel pharmaceutical polymers, such as cyclodextrin, chitosan, polylactic acid (PLA), polyglycolic acid-polylactic acid copolymer (PLGA), hyaluronic acid, etc.

[0049] In some embodiments of the present invention, there are no particular limitations on the dosage form of the aforementioned drug (or photosensitizer). The active substance can be administered together with an assimilated edible carrier, an inert diluent, or directly combined with food. Drug dosage forms include, but are not limited to, hard-shell or soft-shell gelatin capsules, tablets, pills, powder for injection, solutions, suspensions, elixirs, syrups, wafers, gels, buccal or sublingual tablets, films, suppositories, and enemas.

[0050] In other embodiments of the invention, the photocatalyst of the invention can be formulated without any formulation adjuvants or using other drug delivery systems known in the prior art, such as forming components with liposomes, vectored and non-vectored proteins, organic and inorganic nanoparticles, nanoemulsions and microemulsions, nanocrystals, individual solvents or suitable solvent mixtures, with components such as lactose, polyvinylpyrrolidone (PVP), etc.

[0051] In some embodiments of the present invention, the prepared drug (or photosensitizer) may be administered orally, via any part of the gastrointestinal tract (e.g., mouth, pharynx, esophagus, stomach, small intestine (duodenum, jejunum, ileum), large intestine (cecum, colon, rectum) and anus), or via parenteral route (e.g., intravenous, subcutaneous, intraperitoneal or local). If certain drugs are unstable under gastric conditions, they may be prepared as enteric-coated tablets.

[0052] According to another aspect of the present invention, an antitumor metal photosensitizer is provided, wherein the active ingredient of the metal photosensitizer comprises the above-mentioned iridium complex.

[0053] The application of a preferred embodiment of the present invention has at least the following beneficial effects: the present invention has good application prospects in the field of preparation of anti-tumor drugs such as anti-triple-negative breast cancer.

[0054] Other features and advantages of the invention will be set forth in the description which follows, and will be apparent in part from the description, or may be learned by practicing the invention. Attached Figure Description

[0055] The above and / or additional aspects and advantages of the present invention will become apparent and readily understood from the description of the embodiments taken in conjunction with the following drawings, in which:

[0056] Figure 1 The image shows the ultraviolet absorption spectrum of the iridium complex obtained in an embodiment of the present invention.

[0057] Figure 2 The figure shows the test results of the photocatalytic oxidation of NADH by the iridium complex prepared in the embodiments of the present invention;

[0058] Figure 3 The figure shows the test results of the photocatalytic oxidation of NADPH by the iridium complex prepared in the embodiments of the present invention;

[0059] Figure 4 This is a graph showing the test results of the ability of the iridium complex prepared according to an embodiment of the present invention to generate singlet oxygen.

[0060] Figure 5The figure shows the results of dark toxicity and phototoxicity tests of the iridium complex prepared in the embodiments of the present invention against mouse breast cancer cells (4T1). Detailed Implementation

[0061] The following will clearly and completely describe the concept and technical effects of the present invention in conjunction with embodiments, so as to fully understand the purpose, features and effects of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, not all of them. Other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are all within the scope of protection of the present invention. Unless otherwise specified, the experimental methods used in the embodiments are conventional methods; the materials and reagents used, unless otherwise specified, are commercially available. Unless otherwise specified, the same parameter value is the same in all embodiments. The embodiments described below are exemplary and are only used to explain the present invention, and should not be construed as limiting the present invention.

[0062] The term "room temperature" as used in this invention refers to any temperature between 25 and 5°C, and specifically 25°C in the embodiments.

[0063] Example 1

[0064] In this example, an iridium complex was prepared, the structural formula of which is shown below:

[0065]

[0066] The specific preparation process is as follows: S1. 2-Phenyridine (bpy) (0.310 g, 2 mmol) and iridium(III) trichloride hydrate (0.299 g, 1 mmol) were added to a round-bottom flask and 2-ethoxyethanol / water (12 mL; 3:1 v / v) were added. The mixture was then refluxed at 110 °C for 12 hours. After cooling the reaction mixture to room temperature, a yellow precipitate was obtained by filtration, with a yield of 33.3%.

[0067] The chemical reaction equations for the above reactions are shown below:

[0068]

[0069] S2. Iridium(III)μ-chloro-bridged dimer (170 mg, 0.15 mmol) and 5-ethynyl-2,2'-bipyridine (54 mg, 0.3 mmol) were dissolved in chloroform / methanol (15 mL; 2:1 v / v), stirred overnight at 60 °C, cooled to room temperature, and dried under vacuum to obtain an iridium complex intermediate (red powder, yield 67.8%).

[0070] The chemical reaction equations for the above reactions are shown below:

[0071]

[0072] S3. An iridium complex (144 mg, 0.202 mmol), 3,6-bis(5-bromo-2-thienyl)-2,5-bis(2-ethylhexyl)-2,5-dihydropyrrolo[3,4-C]pyrrole-1,4-dione (68 mg, 0.1 mmol), and Pd(PPh3)2Cl2 (0.7 mg, 0.01 mmol) were dissolved in 6 mL of toluene and 4 mL of triethylamine. The mixture was stirred at 60 °C for 30 h under argon protection. After the reaction was completed, the mixture was cooled to room temperature, dried under vacuum, and purified to obtain a blue solid powder (yield 24%).

[0073] The chemical reaction equations for the above reactions are shown below:

[0074]

[0075] The mass spectrum of the product is: ESI-MS [CH3OH, m / z]: 941.05 [M-2Cl]. - ] 2+ .

[0076] The 1H NMR spectrum of the product is as follows: 1 H NMR (500MHz, DMSO-d6) δ8.98(dd,J=20.1,8.2Hz,2H),8.70(d,J=3.5Hz,1H),8.49(d,J=8.3Hz,1H),8.28(d, J=7.3Hz,3H),7.97–7.91(m,4H),7.88(d,J=5.5Hz,2H),7.82(d,J=5.5Hz,1H),7.75–7.66(m,2H),7.62(d,J =5.5Hz,1H),7.18(dd,J=12.5,6.3Hz,2H),7.02(d,J=7.5Hz,2H),6.92(q,J=7.9Hz,2H),6.23(d,J=7.4Hz,1 H),6.16(d,J=7.5Hz,1H),3.97–3.80(m,2H),1.25(dd,J=23.4,16.4Hz,11H),0.80(dd,J=17.5,6.4Hz,5H).

[0077] Application examples

[0078] The iridium complexes prepared in the examples were subjected to performance tests, as detailed below:

[0079] 1. Absorption spectroscopy determination

[0080] Using ethanol (CH3OH) as the solvent, 10 μM sample solutions of the iridium complexes prepared in the examples were prepared. The UV absorption spectra of the novel near-infrared light-excited binuclear iridium complexes were then recorded using a double-beam UV-Vis spectrophotometer. The results are as follows: Figure 1 As shown. From Figure 1 As can be seen, the complex exhibits high absorbance in the near-infrared region in ethanol, indicating that it has good light absorption capabilities in organic solvents.

[0081] 2. Determination of the photocatalytic oxidation capacity of NADH and NADPH

[0082] Because under light irradiation, metal complexes can oxidize reduced coenzyme I (NADH) and reduced coenzyme II (NADPH) to their oxidized forms, NAD. + and NADP + Therefore, the iridium-containing complex (5 μM) and NADH or NADPH (A 339nm =1.0) was mixed in a cuvette, and its ability to oxidize NADH / NADPH under light conditions was measured. The results are as follows: Figure 2 , 3 As shown in the figure, the iridium complex exhibits significant photocatalytic oxidation capabilities for NADH and NADPH.

[0083] 3. Determination of the ability to generate singlet oxygen

[0084] To detect the photocatalytic ability of the iridium complex synthesized in the examples to generate singlet oxygen, the singlet oxygen probe 9,10-anthrayl-bis(methylene)dimalonic acid (ABDA) was used to determine the ability of the binuclear ruthenium complex to generate singlet oxygen. When singlet oxygen is generated in the solution, ABDA immediately captures the singlet oxygen in the solution and reacts to generate an endogenous oxidation product, causing the characteristic absorption peak of ABDA to decrease. The rate of decrease of the ABDA absorption peak is the singlet oxygen generation rate. Therefore, the singlet oxygen generation ability can be reflected by monitoring the changes in the UV-Vis absorption spectra of the test sample and the ABDA mixture solution under different illumination times using a UV-Vis spectrophotometer.

[0085] Two aqueous solutions containing the same binuclear iridium complex (5 μM) and ABDA reagent (200 μM) were placed in cuvettes, and their singlet oxygen generation capacity under 635 nm illumination was measured. The results are as follows: Figure 4 As shown in the figure, this binuclear metallic iridium complex has the ability to generate singlet oxygen after illumination.

[0086] 5. Photodynamic therapy efficacy test on mouse breast cancer cell lines

[0087] Resazurin solution is blue and is commonly used as an acid-base indicator (orange to deep purple at pH 3.8) and a redox indicator. In cell viability assays, resazurin can penetrate cells and be irreversibly reduced to pink by living cells, simultaneously producing the red fluorescence of resorufin. The absorbance or fluorescence intensity of resorufin is positively correlated with cell number and reducing capacity; therefore, cell proliferation can be analyzed using an enzyme-linked immunofluorescence assay (ELISA).

[0088] The experimental steps for the azure blade are as follows:

[0089] (1) First, revive one tube of 4T1 tumor cells and culture them in fresh complete culture medium (DMEM medium + 10 vol% fetal bovine serum + 1 vol% penicillin-streptomycin mixture). After passage twice, start the experiment.

[0090] (2) When the cells reach the logarithmic growth phase, seed them into two 96-well plates at a density of 5000 cells / well (each well is cultured with 100 μL of culture medium, one plate is the light group and the other is the dark control group), and incubate them in a 37°C, 5% CO2 incubator.

[0091] (3) After the culture medium adheres to the wall, remove the original culture medium and add 100 μL of iridium complex at seven different concentrations (100, 50, 10, 1, 0.1, 0.01, and 0.001 μM) to each well. Shake gently and incubate in a carbon dioxide incubator (37°C, 5% CO2) in the dark.

[0092] (4) After incubation for 6 hours, the cell culture plates of the light-illuminated group were placed under a 635nm light source for 45 minutes (light dose of 63.7 J / cm²). 2 Then, the cells were returned to the incubator and incubated in the dark for another 42 hours (the cells in the dark control group were kept in the incubator in the dark throughout the incubation).

[0093] (5) After incubation for 42 hours, the culture medium was discarded from each well, and 80 μL of resazurin (100 mg / mL) was added to each well. The cells were then incubated at 37°C for another 4 hours. The EX540 / EM590 ratio was detected using the fluorescence plate of an ELISA reader, and the cell proliferation inhibition rate was calculated. The IC50 value was then determined. 50 Value (drug concentration when the inhibition rate is equal to 50%).

[0094] like Figure 5 As shown, the iridium complex at different concentrations was detected by the resazurite assay under dark and light conditions to kill mouse breast cancer cell line (4T1 cells). It can be seen that under dark conditions, the IC50 value for killing mouse breast cancer cell line (4T1 cells) was significantly lower. 50 The IC50 concentration was 93.9 μM, and under light conditions, it inhibited the activity of breast cancer cell lines. 50The concentration was 0.31 μM, and the phototherapy index (PI) was as high as 303, indicating that the iridium complex prepared in the embodiments of the present invention has a strong photodynamic therapy effect.

[0095] The embodiments of the present invention have been described in detail above, but the present invention is not limited to the above embodiments. Within the scope of knowledge possessed by those skilled in the art, various changes can be made without departing from the spirit of the present invention.

Claims

1. The application of an iridium complex in the preparation of antitumor drugs, characterized in that: The iridium complex is a compound with the following structure: ； The tumor is breast cancer.

2. The application according to claim 1, characterized in that: The breast cancer in question is the 4T1 cell line breast cancer.

3. An antitumor drug, characterized in that: The active ingredient of the antitumor drug includes an iridium complex, which is a compound with the following structure: 。 4. The antitumor drug according to claim 3, characterized in that: The antitumor drug has at least one of the following characteristics: (1) The tumor is breast cancer; (2) The drug also includes pharmaceutically acceptable carriers and / or excipients; (3) The dosage form of the drug includes at least one selected from hard-shell or soft-shell gelatin capsules, tablets, pills, powder injections, solutions, suspensions, elixirs, syrups, dry films, gels, films, suppositories and enemas; (4) The raw materials for preparing the drug are also selected from at least one of liposome forming components, carrierd or uncarrierized proteins, organic or inorganic nanoparticles, nanoemulsions, microemulsions, and solvents; (5) The route of administration of the drug is selected from the gastrointestinal tract or parenteral route.

5. The antitumor drug according to claim 3, characterized in that: The antitumor drug has at least one of the following characteristics: (1) The tumor is triple-negative breast cancer; (2) The drug also includes pharmaceutically acceptable excipients; the excipients refer to at least one of diluents, coating agents, binders, lubricants, disintegrants, solubilizers or stabilizers that can be used in the pharmaceutical field.

6. The antitumor drug according to claim 3, characterized in that: The antitumor drug has at least one of the following characteristics: (1) The tumor is a triple-negative breast cancer in mice; (2) The drug also includes a pharmaceutically acceptable carrier; the carrier is a functional pharmaceutical excipient acceptable in the pharmaceutical field.

7. The antitumor drug according to claim 3, characterized in that: The antitumor drug has at least one of the following characteristics: (1) The tumor is a mouse triple-negative breast cancer; the cancer cells of the mouse triple-negative breast cancer are 4T1 cancer cells; (2) The drug further includes a pharmaceutically acceptable carrier; the carrier is selected from at least one of cyclodextrin, chitosan, polylactic acid, polyglycolic acid-polylactic acid copolymer or hyaluronic acid.

8. An antitumor metal photosensitizer, characterized in that: The active ingredient of the metal photosensitizer includes an iridium complex, which is a compound with the following structure: 。

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