A hemicyanine compound, its preparation method and application

Abstract

This invention discloses a hemicyanine compound, its preparation method, and its applications. The compound includes anion X. ‑ The hemicyanine compound IITC, as shown in the structure below, can initiate a self-destruct mechanism under prolonged light exposure, effectively avoiding potential phototoxic reactions. This characteristic makes it a highly promising candidate for a self-destructive organic photosensitizer, opening up new application avenues in the treatment strategy for triple-negative breast cancer (TNBC). This hemicyanine compound exhibits excellent photocatalytic therapeutic effects against highly aggressive triple-negative breast cancer cell lines (such as 4T1 cells). Its mechanism lies in its activation by light of a specific wavelength, which significantly enhances the growth inhibition effect on target cancer cells.

Description

Technical Field

[0001] This invention relates to the field of medicinal chemistry, and in particular to a hemicyanine compound, its preparation method, and its application. Background Technology

[0002] Breast cancer is a malignant tumor, and optimizing its treatment has always been a key focus in the medical community. Triple-negative breast cancer (TNBC) is a highly aggressive subtype of breast cancer with a poor prognosis. It is characterized by the absence of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2). Due to the lack of specific targets, traditional hormone therapy and targeted therapy have limited efficacy for TNBC, and currently, chemotherapy is the primary treatment option. However, chemotherapy is often accompanied by severe toxic side effects and drug resistance problems; therefore, there is an urgent need to develop novel treatment strategies.

[0003] Photodynamic therapy (PDT), as an emerging cancer treatment method, offers new insights for the treatment of tumor-associated neoplasms (TNBC) due to its high selectivity, minimal invasiveness, and fewer side effects. PDT relies on photosensitizers to generate reactive oxygen species (such as singlet oxygen) under specific wavelengths of light, inducing apoptosis or necrosis of tumor cells. However, the clinical application of photosensitizers is currently limited by several factors, such as slow metabolism after treatment, potentially leading to long-term residues in the body and phototoxic reactions that negatively impact patients' quality of life. Therefore, developing self-degrading or inactivated photosensitizers that can automatically degrade or inactivate after treatment has become a research hotspot in the field of PDT.

[0004] Zwitterionic hememicanine dyes, with their excellent water solubility, biocompatibility, and cell penetration, exhibit unique advantages in phototherapy (PDT) applications. Through precise design, incorporating self-destruct mechanisms into the hememicanine dye structure can achieve rapid inactivation or degradation of the photosensitizer upon light exposure, thereby significantly reducing the risk of phototoxicity and improving treatment safety.

[0005] Using the 4T1 cell line as an in vitro research model for TNBC is highly representative and practical. The highly invasive and metastatic characteristics of this cell line allow it to mimic the complex biological behavior of TNBC in vivo. By testing the photodynamic therapy (PDT) effects of self-destructive hemicyanine dyes in this cell line, its antitumor activity, metastasis inhibition capacity, and safety can be systematically evaluated. The photodynamic effects of this dye under specific light conditions are assessed through cytotoxicity assays and apoptosis / necrosis assays. Furthermore, small animal models (such as BALB / c mice) are used to further validate the in vivo therapeutic efficacy and safety of this dye against TNBC. The results will reveal the high efficiency and safety of self-destructive hemicyanine dyes in PDT, providing a new and safer treatment option for TNBC patients. In addition, the success of this study may also promote the application of PDT technology in the treatment of other types of malignant tumors, contributing to the overall advancement of cancer treatment.

[0006] Therefore, it is of great significance to design a hemicyanine compound that has a photodynamic effect on 4T1 cells and can self-destruct. Summary of the Invention

[0007] This invention aims to at least solve one of the technical problems existing in the prior art. To this end, this invention proposes a hemicyanine compound that exhibits strong photocatalytic therapeutic effects on 4T1 cells and is self-destructible.

[0008] The present invention also proposes a method for preparing the above-mentioned compounds.

[0009] The present invention also proposes applications of the above-mentioned compounds.

[0010] According to one aspect of the present invention, a hemicyanine compound (IITC) is provided, said compound comprising anion X - and cations with the following structures:

[0011]

[0012] According to a preferred embodiment of the present invention, at least the following beneficial effects are achieved: the hemicyanine compounds of the present invention can initiate a self-destruct mechanism under prolonged light exposure, thereby effectively avoiding potential phototoxic reactions. This characteristic makes them highly promising candidates for self-destructive organic photosensitizers, opening up new application avenues in the treatment strategy for triple-negative breast cancer (TNBC). These hemicyanine compounds exhibit excellent photocatalytic therapeutic effects against highly aggressive triple-negative breast cancer cell lines (such as 4T1 cells), the mechanism of which lies in activation by light of a specific wavelength, significantly enhancing the growth inhibition effect on target cancer cells.

[0013] Specifically, under suitable light conditions, the compound exhibited extremely high sensitivity to 4T1 cells, with a half-maximal inhibitory concentration (IC50) as low as 0.081 μM, indicating its ability to effectively inhibit cancer cell proliferation at very low concentrations. This characteristic is significant for reducing drug dosage and minimizing treatment side effects. However, under dark conditions, the compound's cytotoxicity was significantly reduced, with the IC50 value increasing to 18.07 μM. This difference not only ensured the safety of the treatment but also highlighted the crucial role of light in activating its therapeutic activity.

[0014] Furthermore, the calculated phototherapy index (PI) reached as high as 223. This value is an important indicator for measuring the balance between the therapeutic efficiency and safety of photosensitizers. A high PI value means that the compound has extremely high therapeutic efficiency under light irradiation while maintaining good safety, providing strong scientific evidence for photodynamic therapy of triple-negative breast cancer.

[0015] Therefore, the hemicyanine compounds of the present invention not only demonstrate unique advantages as self-destructive photosensitizers, but their experimental results in the treatment of triple-negative breast cancer also indicate that they have important research value and broad application prospects in the future development of efficient and low-toxicity photodynamic therapy strategies.

[0016] In some embodiments of the present invention, the X - For Cl - ,Br - I - or PF6 - PF6 is preferred. - .

[0017] According to another aspect of the present invention, a method for preparing the above-mentioned hemicyanine compounds is provided, comprising the following steps:

[0018] S1. Reaction of 5-iodo-2,3,3-trimethyl-3H-indole with 6-bromohexyltrimethylammonium bromide. The product obtained after the reaction is reacted with anion X. - The compounds undergo ion exchange;

[0019] S2. React the compound obtained in step S1 with 3,4-dihydroxy-3-cyclobutene-1,2-dione and quinoline to obtain the hemicyanine compound.

[0020] The preparation method according to a preferred embodiment of the present invention has at least the following beneficial effects: the preparation method of the present invention is simple to operate and has good prospects for industrial application.

[0021] In some embodiments of the present invention, step S1 specifically includes mixing 5-iodo-2,3,3-trimethyl-3H-indole with 6-bromohexyltrimethylammonium bromide and adding solvent I, and reacting under heating.

[0022] In some embodiments of the present invention, the reaction conditions under heating in step S1 include at least one of the following conditions:

[0023] 1) The temperature is 85–100℃;

[0024] 2) The time is 3-7 days;

[0025] 3) The heating method is heating reflux.

[0026] In some embodiments of the present invention, the reaction conditions under heating in step S1 include at least one of the following conditions:

[0027] 1) The temperature is 90℃;

[0028] 2) The time period is 7 days;

[0029] 3) The heating method is heating reflux.

[0030] In some embodiments of the present invention, the molar ratio of 5-iodo-2,3,3-trimethyl-3H-indole to 6-bromohexyltrimethylammonium bromide is 1 to 1.3:1.

[0031] In some embodiments of the present invention, the molar ratio of 5-iodo-2,3,3-trimethyl-3H-indole to 6-bromohexyltrimethylammonium bromide is 1.2:1.

[0032] In some embodiments of the present invention, 5-iodo-2,3,3-trimethyl-3H-indole and 6-bromohexyltrimethylammonium bromide are added to solvent I and PF6-containing solvent. - The NH4PF6 was reacted, then refluxed at 90°C for 7 days. The reactants were then cooled to room temperature, and solvent I was removed. The reaction equation for this step is as follows:

[0033]

[0034] In some embodiments of the present invention, solvent I is acetonitrile.

[0035] In some embodiments of the present invention, in step S2, the compound obtained in step S1 is ion-exchanged and then mixed with 3,4-dihydroxy-3-cyclobutene-1,2-dione and quinoline, and solvent II is added. The mixture is then reacted under heating to obtain the hemicyanine self-destructive organic photosensitizer IITC.

[0036] In some embodiments of the present invention, solvent II is selected from n-butanol / toluene mixture or n-butanol / benzene mixture (preferably, the volume ratio is 1:1).

[0037] In some embodiments of the present invention, in step S2, the reaction conditions under heating include at least one of the following conditions:

[0038] 1) The temperature is 105–115℃;

[0039] 2) The time is 24–48 hours;

[0040] 3) The heating method is heating reflux.

[0041] In some embodiments of the present invention, the conditions for the reaction under heating include at least one of the following conditions:

[0042] 1) The temperature is 110℃;

[0043] 2) The time is 24 hours;

[0044] 3) The heating method is heating reflux.

[0045] In some embodiments of the present invention, in step S2, the molar ratio of the compound obtained in step S1 to 3,4-dihydroxy-3-cyclobutene-1,2-dione and quinoline is 2 to 2.3:1:10.

[0046] In some embodiments of the present invention, in step S2, the molar ratio of the compound obtained in step S1 to 3,4-dihydroxy-3-cyclobutene-1,2-dione and quinoline is 2.2:1:10.

[0047] In some embodiments of the present invention, in step S2, the compound obtained in step S1 is mixed with 3,4-dihydroxy-3-cyclobutene-1,2-dione and a mixture of n-butanol / toluene (1:1) is added, then refluxed at 110°C for 24 h. The reactants are cooled to room temperature, the solvent is evaporated, and a solution of acetone and ethyl acetate is added to precipitate a solid. After dissolution, a solution of methanol and ethyl acetate is added to precipitate another solid. The reaction formula for this step is as follows:

[0048]

[0049] According to another aspect of the present invention, the use of the above-mentioned compound in the preparation of antitumor drugs is proposed.

[0050] According to a preferred embodiment of the present invention, the application has at least the following beneficial effects: The compounds of the present invention have good water solubility and can generate singlet oxygen ions and reactive oxygen species under light irradiation. Excessive accumulation of singlet oxygen ions and reactive oxygen species in organisms is closely related to oxidative stress; therefore, they can kill tumor cells or inhibit their growth by inducing oxidative stress. The compounds of the present invention have a strong ability to generate superoxide anions after light irradiation, which can induce oxidative stress in tumor cells. Therefore, they have good anti-tumor potential, indicating their promising application prospects in the field of anti-tumor drugs.

[0051] In some embodiments of the present invention, the antitumor drug is an anti-triple-negative breast cancer drug. The compounds of the present invention have a strong inhibitory effect on 4T1 cell proliferation and show good application prospects in the treatment of triple-negative breast cancer.

[0052] In some embodiments of the present invention, the triple-negative breast cancer is a triple-negative breast cancer cell line (4T1 cells) induced in BALB / c mice. The compounds of the present invention show promising application prospects in the preparation of anti-triple-negative breast cancer drugs.

[0053] In some embodiments of the present invention, the drug further includes a pharmaceutically acceptable carrier and / or excipient. That is, the drug or photosensitizer, as the main active ingredient, is mixed with a pharmaceutically acceptable carrier and / or excipient to prepare a composition, and then formulated into a clinically acceptable dosage form.

[0054] 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.

[0055] In some embodiments of the invention, the different pharmaceutical excipients used in the drug dosage form may vary depending on the specific medical application. Pharmaceutical excipients can be used to adjust the solubility and bioavailability of a drug, 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, diluents, solubilizers, stabilizers, binders, fillers, coating agents, disintegrants, lubricants, and sweeteners adapted to the use of photosensitizers. Other pharmaceutical matrices may also be included.

[0056] 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.

[0057] In some embodiments of the present invention, there are no particular limitations on the dosage form of the above-mentioned drugs. 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.

[0058] In other embodiments of the invention, the medicament of the invention may 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.

[0059] In some embodiments of the present invention, the prepared drug 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.

[0060] According to another aspect of the present invention, an antitumor organic photosensitizer is provided, wherein the active ingredient of the antitumor metal photosensitizer comprises the above-mentioned hemicyanine compounds.

[0061] The photosensitizer is the main active ingredient, and other active ingredients may also be added.

[0062] According to another aspect of the present invention, a self-destruction method for the above-mentioned hemicyanine compounds is provided, comprising the following steps: irradiating with light of wavelength 630-640 nm (preferably 635 nm) for more than 2 hours.

[0063] Self-destruction can be achieved through exposure to light for a certain period of time, significantly reducing the risk of phototoxicity and improving the safety of treatment.

[0064] 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

[0065] 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:

[0066] Figure 1This is the 1H NMR spectrum of the hemicyanine compounds obtained in the embodiments of the present invention;

[0067] Figure 2 The ultraviolet absorption spectrum of the hemicyanine compounds obtained in the embodiments of the present invention is shown below.

[0068] Figure 3 The graph shows the changes in the ultraviolet absorption spectrum of the hemicyanine compounds prepared in the embodiments of the present invention under light irradiation in water.

[0069] Figure 4 This is a graph showing the changes in the ultraviolet absorption spectra of hemicyanine compounds obtained in an embodiment of the present invention under light irradiation in cell culture medium;

[0070] Figure 5 The image shows the photocatalytic oxidation of NAPH by hemicyanine compounds obtained in the embodiments of the present invention.

[0071] Figure 6 The image shows the test results of the photocatalytic oxidation of NAPDH by hemicyanine compounds prepared in the embodiments of the present invention;

[0072] Figure 7 The graph shows the test results of the ability of the hemicyanine compounds prepared in the embodiments of the present invention to generate superoxide anions.

[0073] Figure 8 The graph shows the test results of the ability of hemicyanine compounds prepared in the embodiments of the present invention to generate singlet oxygen.

[0074] Figure 9 The figure shows the toxicity test results of the hemicyanine compounds prepared in the embodiments of the present invention against triple-negative breast cancer (4T1 cells).

[0075] Figure 10 The image shows the phototoxicity test results of the hemicyanine compounds obtained in the embodiments of the present invention against triple-negative breast cancer (4T1 cells).

[0076] Figure 11 The image shows the phototoxicity test results of the hemicyanine compounds obtained in the embodiments of the present invention against triple-negative breast cancer (4T1 cells). Detailed Implementation

[0077] 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.

[0078] In the description of this invention, the terms "one embodiment," "some embodiments," "illustrative embodiment," "example," "specific example," or "some examples," etc., refer to specific features, structures, materials, or characteristics described in connection with that embodiment or example, which are included in at least one embodiment or example of the invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0079] 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.

[0080] Example

[0081] In this example, a hemicyanine compound was prepared, the structure of which is shown in the formula:

[0082]

[0083] The specific preparation process is as follows:

[0084] S1. 5-Iodo-2,3,3-trimethyl-3H-indole (0.342 g, 1.2 mmol) was mixed with 6-bromohexyltrimethylammonium bromide (0.303 g, 1 mmol), and 10 mL of acetonitrile was added. The mixture was stirred at 90 °C for 7 days. After the reaction was completed, the mixture was cooled to room temperature, and the acetonitrile was evaporated to dryness. Methanol and ethyl acetate were added to precipitate the solid. After filtration, methanol and a saturated aqueous solution of hexafluorophosphate were added to precipitate the solid again. The solid was purified by alumina column chromatography to obtain a purple solid powder (yield 22.18%).

[0085] The chemical reaction equations for the above reactions are shown below:

[0086]

[0087] S2. The compound obtained in step S1 (57.47 mg, 80 μmol) was mixed with 3,4-dihydroxy-3-cyclobutene-1,2-dione (4.34 mg, 38.1 μmol) and quinoline (49.2 mg, 380.95 μmol) and a mixture of n-butanol / toluene (18 ml, v:v = 1:1) was added. The mixture was then refluxed at 110 °C for 24 h. The reactants were cooled to room temperature, the solvent was evaporated, and a solid was precipitated by adding acetone and ethyl acetate solution. After dissolving the solid, a blue-black solid was precipitated by adding methanol and ethyl acetate solution. The yield was 42.3%.

[0088] The chemical reaction equations for the above reactions are shown below:

[0089]

[0090] The solid powder obtained by the above operation was subjected to 1H NMR spectroscopy. The test results are as follows: Figure 1 The high-resolution mass spectrometry information of this compound is shown below.

[0091] 1HNMR (500MHz, DMSO-d6) δ7.90 (s, 2H), 7.68 (d, J=8.3Hz, 2H), 7.19 (d, J=8.3Hz, 2H), 5.78 (s, 2H) , 4.03 (d, J=11.3Hz, 4H), 3.29 (s, 4H), 3.03 (s, 18H), 1.66 (s, 18H), 1.38 (dq, J=39.6, 7.6Hz, 10H).

[0092] To verify the application effect of the above photosensitizer, its performance was tested, as follows:

[0093] 1. Absorption spectroscopy determination

[0094] Using the photosensitizer prepared as a sample, a 10 mM stock solution was prepared. This stock solution was then placed in solvents of different polarities (methanol (EtOH), N,N-dimethylformamide (DMF), water, acetonitrile (MeCN), dichloromethane (DCM), and dimethyl sulfoxide (DMSO)) to achieve a final concentration of 10 μM. The absorbance of the solution was measured using a UV spectrophotometer. The UV-Vis absorption spectra are shown below. Figure 2 As shown in the figure, it exhibits good light absorption in organic solvents. The maximum absorption wavelength in aqueous solution is 640 nm, which may be due to the strong hydrogen bonding and polar interactions of water molecules, resulting in a large energy difference between the ground and excited states of the photosensitizer molecules. In N,N-dimethylformamide and dimethyl sulfoxide solutions, the high polarity and strong polar interactions of both solutions make the excited state energy level of the photosensitizer most stable, leading to a redshift in the absorption wavelength.

[0095] 2. Changes in the UV absorption spectrum of self-destructing photosensitizers in water and culture medium under light irradiation.

[0096] The photosensitizer was dissolved to prepare a 10 mM stock solution, which was then placed in pure water and cell culture medium (DMEM) to achieve a final concentration of 10 μM. The absorbance of the solution was measured using a UV spectrophotometer. A quartz cuvette was placed under 635 nm illumination, and the cuvette was removed at regular intervals to rapidly measure the UV absorption spectrum. The data were recorded, and the results are shown below. Figure 3 (in aqueous solution) and Figure 4 (In DMEM cell culture medium) as shown.

[0097] from Figure 3 and Figure 4 As can be seen, under illumination, the ultraviolet peak absorption of this photosensitizer gradually decreases over time when irradiated by a 635nm light source.

[0098] 3. Determination of the photocatalytic oxidation capacity of NADH

[0099] In aqueous solution, the absorbance of NADH at the 339 nm absorption peak was controlled to be 1, and this concentration was used for subsequent measurements. The above compound was added to the above solution to a concentration of 10 μM as the experimental group, with a solution of the same concentration without NADH as the reference. The light group was irradiated at 635 nm, while the dark group was not irradiated. The UV-Vis spectrum of the solution was measured at regular intervals, and the changes in UV absorption at 339 nm were observed. The results are as follows: Figure 5 As shown.

[0100] from Figure 5 As can be seen, the absorption peak of the photosensitizer at 339 nm decreases under illumination, indicating a strong photocatalytic oxidation effect and good photoselectivity.

[0101] 4. Determination of the photocatalytic oxidation capacity of NADPH

[0102] In aqueous solution, the absorbance of NADH at the 339 nm absorption peak was controlled to be 1, and this concentration was used for subsequent measurements. A compound was added to the above solution to a concentration of 10 μM as the experimental group, with a solution of the same concentration of the compound without added NADPH as the reference. The light group was irradiated at 635 nm, while the dark group was not irradiated. The UV-Vis spectrum of the solution was measured at regular intervals, and the changes in UV absorption at 339 nm were observed. The results are as follows: Figure 6 As shown.

[0103] from Figure 6 As can be seen, the absorption peak of the photosensitizer at 339 nm decreases under illumination, indicating a strong photocatalytic oxidation effect and good photoselectivity.

[0104] 5. Determination of the ability to generate superoxide anions in solution

[0105] The ability of a photosensitizer to generate superoxide anions was determined using the superoxide anion probe dihydrorhodamine 123 (DHR123). The change in fluorescence intensity of the test sample and the DHR123 mixture under different illumination times was monitored using a fluorescence spectrophotometer to reflect the superoxide anion generation capacity. At λex = 465 nm, a mixture of photosensitizer (10 μM) and DHR123 (10 μM) was excited in a 1 cm quartz tube. The blank control was a probe solution of the same concentration without the compound. The entrance and exit slits were set to 2.5 nm. The superoxide anion generation capacity was measured under 635 nm illumination. The test results are shown below. Figure 7 As shown in the figure, the fluorescence of DHR123 is significantly enhanced after illumination, while the fluorescence enhancement of DHR123 in the blank control group and the dark group is not obvious, indicating that this photosensitizer has a strong ability to generate superoxide anions and has a certain degree of photoselectivity.

[0106] 6. Determination of the ability to generate singlet oxygen in solution

[0107] To detect the ability of the photosensitizer synthesized in the examples to generate singlet oxygen under light irradiation, the singlet oxygen probe 9,10-anthrayl-bis(methylene)dimalonic acid (ABDA) was used to determine this ability. When singlet oxygen is generated in the solution, ABDA immediately captures it, reacting to generate an endogenous oxidation product, causing a decrease in the characteristic absorption peak of ABDA. The rate of decrease in the ABDA absorption peak is the singlet oxygen generation rate. Therefore, by monitoring the changes in the UV-Vis absorption spectra of the test sample and the ABDA mixture solution under different irradiation times using a UV-Vis spectrophotometer, the singlet oxygen generation ability can be reflected.

[0108] Two aqueous solutions containing the same photosensitizer (10 μM) and ABDA reagent (100 μM) were placed in cuvettes, and their singlet oxygen generation capacity under 635 nm light irradiation was measured. The test results are as follows: Figure 8 As shown in the figure, after illumination, the characteristic absorption peak of ABDA at 378 nm is significantly reduced, while the characteristic absorption peak in the dark group remains almost unchanged, indicating that this compound has a strong ability to generate singlet oxygen and exhibits photoselectivity.

[0109] 7. Results of dark toxicity and phototoxicity tests

[0110] 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).

[0111] The experimental steps for the azure blade are as follows:

[0112] (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.

[0113] (2) When the cells reach the logarithmic growth phase, seed them into two 96-well plates at a density of 6000 cells / well (each well is used to culture cells with 90 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.

[0114] (3) After the photosensitizer adheres to the wall, add 10 μL of the corresponding concentration of photosensitizer to each well (100, 50, 10, 1, 0.1, 0.01 μM), shake gently, and incubate in a carbon dioxide incubator (37℃, 5% CO2) in the dark.

[0115] (4) After 6 hours of incubation, the cell culture plates of the light-illuminated group were placed under a 635nm light source for 30 minutes (light dose of 44J / cm²). 2 Then, the cells were returned to the incubator and incubated in the dark for another 24 hours (the cells in the dark control group were kept in the incubator in the dark).

[0116] (5) After 24 hours of incubation, 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 2.5–4 hours. The EX540 / EM590 ratio was detected using an ELISA reader, and the cell proliferation inhibition rate was calculated. The IC50 value was then determined. 50 Value (the concentration of the compound when the inhibition rate is equal to 50%).

[0117] Test results are as follows Figure 9 As shown in the figure, the resazurin assay was used to detect the cytotoxic effect of different concentrations of photosensitizers on the triple-negative breast cancer cell line (4T1 cells) under dark and light treatment conditions. Under dark conditions, the IC50 value for the triple-negative breast cancer cell line (4T1 cells) was [data missing]. 50 The IC50 concentration was 18.07 μM, and under light conditions, it had an IC50 value of 18.07 μM for triple-negative breast cancer cell line (4T1 cells). 50The concentration was 0.081 μM, and the phototherapy index (PI) was as high as 223, indicating that the photosensitizer of the present invention has a strong photocatalytic therapeutic effect.

[0118] 8. Results of toxicity tests of photo-exposed products against triple-negative breast cancer (4T1 cells)

[0119] (1) When 4T1 cells are cultured to the logarithmic growth phase, they are seeded into two 96-well plates at a cell density of 6000 cells / well (90 μL of culture medium is used to culture cells in each well, one plate is the light group and the other is the dark control group), and then placed in a 37°C, 5% CO2 incubator until the cells adhere.

[0120] (2) After placing the corresponding concentrations of photosensitizers and commercially available photosensitizers Ce6 and verteporfen (100, 50, 10, 1, 0.1, 0.01 μM) under a 635 nm light source for 2 h, the corresponding photoproducts were generated.

[0121] (3) After the cells adhere to the wall, add 10 μL of the corresponding concentration of light product to each well, shake gently, and incubate in a carbon dioxide incubator (37℃, 5% CO2) in the dark.

[0122] (4) After 6 hours of incubation, the cell culture plates of the light-illuminated group were placed under a 635nm light source for 30 minutes (light dose of 44J / cm²). 2 Then, the cells were returned to the incubator and incubated in the dark for another 24 hours (the cells in the dark control group were kept in the incubator in the dark).

[0123] (5) After 24 hours of incubation, 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 2.5–4 hours. The EX540 / EM590 ratio was detected using an ELISA reader, and the cell proliferation inhibition rate was calculated. The IC50 value was then determined. 50 Value (the concentration of the compound when the inhibition rate is equal to 50%).

[0124] Test results are as follows Figure 10 (lighting) and Figure 11 (Darkness) is shown. As can be seen from the figure, the photodegradation products of the three photosensitizers after being exposed to light for the same period did not produce toxicity when placed in the dark. However, under light, the products of this hemicyanine photosensitizer showed almost no cytotoxicity. This indicates that the compound can automatically degrade or lose its activity after light exposure. Furthermore, compared to Ce6 and verteporfen, this hemicyanine photosensitizer has a faster metabolic rate, which helps reduce the residence time of the photosensitizer in the body, thereby reducing the accumulation of toxic effects and minimizing damage to normal cells or tissues.

[0125] In summary, the compound structure of this invention exhibits good inhibitory effects against triple-negative breast cancer cells under light irradiation. Furthermore, it can undergo self-destruction after prolonged continuous light irradiation, reducing phototoxicity and improving drug safety. It can be prepared into anti-tumor photosensitizers and other drugs, showing promising application prospects in the field of photodynamic therapy for tumors.

[0126] 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. A hemicyanine compound, characterized in that: The compound includes anion X. - and cations with the following structures: 。 2. The hemicyanine compound according to claim 1, characterized in that: The X - For Cl - ,Br - I - or PF6 - .

3. The hemicyanine compound according to claim 1, characterized in that: The X - For PF6 - .

4. The method for preparing hemicyanine compounds according to claim 1, 2, or 3, characterized in that: Includes the following steps: S1. Reaction of 5-iodo-2,3,3-trimethyl-3H-indole with 6-bromohexyltrimethylammonium bromide. The product obtained after the reaction is reacted with anion X. - The compounds undergo ion exchange; S2. React the compound obtained in step S1 with 3,4-dihydroxy-3-cyclobutene-1,2-dione and quinoline to obtain the hemicyanine compound.

5. The preparation method according to claim 4, characterized in that: Step S1 specifically includes mixing 5-iodo-2,3,3-trimethyl-3H-indole with 6-bromohexyltrimethylammonium bromide and adding solvent I. The reaction under heating conditions include at least one of the following: 1) The temperature is 85~100℃; 2) The time is 3-7 days; 3) The heating method is reflux heating; 4) Solvent I is acetonitrile.

6. The preparation method according to claim 4, characterized in that: In step S2, the compound obtained in step S1 is ion-exchanged and then mixed with 3,4-dihydroxy-3-cyclobutene-1,2-dione and quinoline, and solvent II is added. The mixture is then reacted under heating to obtain the hemicyanine self-destructive organic photosensitizer. The reaction conditions under heating include at least one of the following: 1) The temperature is 105~115℃; 2) The time is 24~48 hours; 3) The heating method is reflux heating; 4) Solvent II is selected from n-butanol / toluene mixture or n-butanol / benzene mixture.

7. The use of the hemicyanine compounds according to claim 1, 2, or 3 in the preparation of antitumor drugs; characterized in that: The anti-tumor drug is an anti-triple-negative breast cancer drug.

8. The application according to claim 7, characterized in that: The triple-negative breast cancer is a triple-negative breast cancer cell line induced in BALB / c mice.

9. The application according to claim 7, characterized in that: The drug also includes a pharmaceutically acceptable carrier.

10. An antitumor organic photosensitizer, characterized in that: The active ingredient of the antitumor metal photosensitizer includes the hemicyanine compounds as described in claim 1, 2, or 3.

11. The self-destruction method for hemicyanine compounds as described in claim 1, 2, or 3, characterized in that: The process includes the following steps: irradiating with light of wavelength 630~640nm for more than 2 hours.

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