Osmium complex as well as preparation method and application thereof

Abstract

The invention discloses an osmium complex and a preparation method and application thereof, the osmium complex has the following structural formula: # imgabs0 #, and X-represents an anion. The osmium complex disclosed by the invention has a relatively strong photodynamic treatment effect on a human non-small cell lung cancer cell line (A549 cells). Under the condition of illumination, the compound has very strong ability of inhibiting growth and proliferation of human non-small cell lung cancer cells (IC50 is 4.97 mu M), and under the condition of darkness, the cytotoxicity of the compound is greater than 100 mu M, and the phototherapy index PI is as high as 20. The invention has important significance for researching metal drugs for resisting non-small cell lung cancer.

Description

Technical Field

[0001] The present invention relates to the technical field related to medicinal chemistry, and in particular to an osmium complex and a preparation method and application thereof. Background Art

[0002] Lung cancer, a highly malignant neoplastic disease of the respiratory system, is pathologically classified primarily based on the World Health Organization's cytological classification criteria. Non-small cell lung cancer (NSCLC) accounts for a high proportion of clinically confirmed cases. This subtype encompasses adenocarcinoma, squamous cell carcinoma, and other pathological morphologies with significant heterogeneity. Its complex molecular regulatory networks and tumor microenvironmental characteristics pose significant challenges to traditional treatment models. Existing treatments still have significant limitations, particularly in terms of tumor stem cell regulation, epithelial-mesenchymal transition (EMT), and immune escape mechanisms.

[0003] In recent years, the development of photosensitive materials based on metal-organic frameworks has provided innovative directions for precision tumor treatment. Among them, the complex system of the Group VIII transition metal osmium has attracted much attention due to its unique electronic transition properties. The intersystem crossing phenomenon generated by the spin-orbit coupling effect of Os(II) complexes can significantly improve the quantum conversion efficiency of triplet oxygen to singlet oxygen. Its wide spectral absorption characteristics not only enhance the tissue penetration depth, but also its long-lived phosphorescence properties (μs level) are more conducive to the temporal dimension of photodynamic effect regulation, which provides a physical and chemical basis for the construction of intelligent photosensitized therapeutic systems.

[0004] Although the application of transition metal osmium complexes in photodynamic therapy (PDT) has made some remarkable progress, its treatment for non-small cell carcinoma (NSCLC) still faces many challenges. NSCLC is highly heterogeneous, with tumors from different patients showing significant differences in gene expression, metabolic pathways, and immune microenvironment, making it difficult for a single treatment strategy to achieve ideal results. In addition, the metastasis and recurrence of NSCLC are also urgent issues that need to be addressed. Even after comprehensive treatment such as surgical resection and chemoradiotherapy, some patients will still experience recurrence or distant metastasis, leading to treatment failure.

[0005] Therefore, in order to further improve the treatment effect of non-small cell carcinoma and reduce the risk of recurrence and metastasis, it is necessary to continuously explore new treatment strategies and drugs. Summary of the Invention

[0006] The present invention aims to solve at least one of the technical problems existing in the prior art. To this end, the present invention provides an osmium complex with a novel structure and a significant inhibitory effect on the proliferation of non-small cell carcinoma, providing a new direction for the treatment of non-small cell carcinoma.

[0007] According to one aspect of the present invention, an osmium complex is provided, having the following structural formula:

[0008]

[0009] Among them, X - Represents anions.

[0010] According to a preferred embodiment of the present invention, at least the following beneficial effects are achieved: the osmium complex of the present invention has a strong photodynamic therapy effect on human non-small cell lung cancer cell line (A549 cells). Under light conditions, it has a strong ability to inhibit the growth and proliferation of human non-small cell lung cancer cells (IC 50 The photocatalytic activity of the metallodextrin was 4.97 μM, while in the dark, its cytotoxicity was greater than 100 μM, and the phototherapeutic index (PI) was as high as 20. This is of great significance for the study of metallodrugs for the treatment of non-small cell lung cancer.

[0011] In some preferred embodiments of the present invention, X - PF6 - PF6 anion - It can improve the solubility of osmium complexes and enhance their anti-tumor effects.

[0012] According to another aspect of the present invention, a method for preparing the above-mentioned complex is provided, comprising the following steps:

[0013] S1. Make Os 4+ and 4'-bromo-2,2':6',2"-terpyridine in a solvent under heating to prepare an osmium complex intermediate I;

[0014] S2. Add the osmium complex intermediate I prepared in step S1 to the - to obtain an osmium complex intermediate II;

[0015] S3. The osmium complex intermediate II obtained in step S1 is reacted with 2,5-bis(2-ethylhexyl)-3,6-bis(thiophen-2-yl)pyrrolo[3,4-C]pyrrole-1,4(2H,5H)-dione in the presence of a palladium catalyst, a carbonate, and pivalic acid to generate the osmium complex.

[0016] 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 easy to operate and has good industrial application prospects.

[0017] In some embodiments of the present invention, step S1 specifically comprises mixing ammonium chloroosmate or its hydrate with 4'-bromo-2,2':6',2"-terpyridine, adding a solvent, and reacting under heating.

[0018] In some preferred embodiments of the present invention, in step S1, Os 4+The molar ratio of 4'-bromo-2,2':6',2"-terpyridine is 1:2 to 2.2.

[0019] In some preferred embodiments of the present invention, Os 4+ The molar ratio of 4'-bromo-2,2':6',2"-terpyridine is 1:2.1.

[0020] In some embodiments of the present invention, the solvent is selected from ethylene glycol.

[0021] In some embodiments of the present invention, in step S1, the reaction under heating includes at least one of the following conditions:

[0022] 1) The temperature is 180-200°C;

[0023] 2) Duration is 2 to 4 hours;

[0024] 3) The heating method is heating reflux.

[0025] In some preferred embodiments of the present invention, in step S1, the reaction under heating includes at least one of the following conditions:

[0026] 1) Temperature is 200°C;

[0027] 2) The duration is 3 hours;

[0028] 3) The heating method is heating reflux.

[0029] In some embodiments of the present invention, step S3 specifically includes reacting the osmium complex intermediate II with palladium acetate, potassium carbonate, pivalic acid and 2,5-bis(2-ethylhexyl)-3,6-bis(thiophen-2-yl)pyrrolo[3,4-C]pyrrole-1,4(2H,5H)-dione in an aprotic polar solvent to generate the target osmium complex.

[0030] In some embodiments of the present invention, the aprotic polar solvent is dimethylacetamide.

[0031] In some embodiments of the present invention, the heating temperature in step S3 is selected from 90 to 110° C., and the heating time is selected from 4 to 8 hours.

[0032] In some preferred embodiments of the present invention, the heating temperature in step S3 is 110°C.

[0033] In some embodiments of the present invention, the molar ratio of the osmium complex intermediate II to palladium acetate, potassium carbonate, pivalic acid and 2,5-bis(2-ethylhexyl)-3,6-bis(thiophen-2-yl)pyrrolo[3,4-C]pyrrole-1,4(2H,5H)-dione is 1:0.04-0.06:2-2.2:0.08-0.12:2-2.2.

[0034] In some preferred embodiments of the present invention, the molar ratio of the osmium complex intermediate to palladium acetate, potassium carbonate, pivalic acid and 2,5-bis(2-ethylhexyl)-3,6-bis(thiophen-2-yl)pyrrolo[3,4-C]pyrrole-1,4(2H,5H)-dione is 1:0.05:2:0.1:2.1.

[0035] According to another aspect of the present invention, the use of the above-mentioned complex in the preparation of anti-tumor drugs is proposed.

[0036] The application according to a preferred embodiment of the present invention has at least the following beneficial effects: the complex of the present invention can generate superoxide anions upon illumination. Superoxide anions are often associated with oxidative stress in vivo and can act as signaling molecules involved in various physiological and pathological processes. Therefore, they can kill tumor cells or inhibit their growth by inducing oxidative stress. The complex of the present invention has a strong ability to generate superoxide anions upon illumination, which can induce oxidative stress in tumor cells. Therefore, it has excellent anti-tumor potential, indicating that it has promising application prospects in the field of anti-tumor drugs.

[0037] In some embodiments of the present invention, the anti-tumor drug is an anti-non-small cell lung cancer drug. The complex of the present invention has a strong effect of inhibiting the proliferation of non-small cell lung cancer cells and has good application prospects in the treatment of non-small cell lung cancer.

[0038] In some embodiments of the present invention, the non-small cell lung cancer is A549 cell line lung cancer.

[0039] In some embodiments of the present invention, the anti-non-small cell lung cancer drug is a photodynamic therapy drug.

[0040] In some embodiments of the present invention, the wavelength of the excitation light of the photodynamic therapy drug is 200 to 800 nm.

[0041] In some embodiments of the present invention, the excitation light wavelength is 600-700 nm.

[0042] In some embodiments of the present invention, the excitation light has a wavelength of 630-640 nm.

[0043] According to relevant experimental data, the complex of the present invention has high photodynamic activity under red light excitation.

[0044] In some embodiments of the present invention, the pharmaceutical formulation comprises a therapeutically effective amount of a metal osmium complex as an active ingredient, and is compounded with auxiliary ingredients required for pharmaceutical formulations. Specifically, the composition is formed by physically or chemically combining the photosensitizing active ingredient with conventional pharmaceutical excipients to form a pharmaceutical form suitable for clinical administration.

[0045] In some embodiments of the present invention, the pharmaceutical excipients include processing aids required for solid dosage form processing, including but not limited to diluents, adhesion promoters, disintegration promoters, lubricants, dissolution regulators, and formulation stabilizers. In particular, these excipients can synergistically improve the dispersion uniformity of the photosensitive component, control the release kinetics of the active ingredient, and optimize the rheological properties of the formulation.

[0046] In some embodiments of the present invention, the selection of excipient systems follows the principle of dosage form functionality adaptation: for oral solid dosage forms, fillers with specific pore structures, such as microcrystalline cellulose and croscarmellose sodium, are preferred; for injectable dosage forms, phospholipid bilayer constructs or amphiphilic block copolymers are used as stabilizing agents. These differentiated choices are intended to achieve optimal delivery efficacy of the photosensitive component across different routes of administration.

[0047] In some embodiments of the present invention, the delivery system comprises novel functionalized pharmaceutical materials, particularly biodegradable polymers with environmentally responsive properties. Exemplary materials include temperature-sensitive polycaprolactone-polyethylene glycol blocks, pH-responsive chitosan derivatives, and tumor-targeting hyaluronic acid-polylactic acid complexes. These delivery systems can significantly enhance drug accumulation at the lesion site.

[0048] In some embodiments of the present invention, dosage form design transcends the limitations of traditional methods, specifically developing complex dosage forms suitable for photodynamic therapy. These include implantable sustained-release tablets with optical windows, nanogel microspheres for light-controlled release, and interventional catheter systems integrated with optical fibers. These innovative dosage forms are particularly suitable for the precise treatment of deep-seated tumor lesions.

[0049] In other embodiments of the present invention, the photosensitizing agents can be delivered using novel nanostructured delivery systems, including but not limited to gold nanocage drug delivery systems with photothermal conversion properties, mesoporous silica nanoparticles capable of ROS-responsive release, and lipid nanoparticles with dual targeting capabilities. These systems, through surface functionalization, can break through biological barriers and achieve precise delivery at the organelle level.

[0050] In some embodiments of the present invention, the route of administration is designed to fully consider the anatomical characteristics of the lesion: for superficial lesions, a transdermal patch combined with localized illumination is used; for pulmonary metastases, a dry powder aerosol for inhalation is developed; and for central nervous system lesions, a nasal-brain targeted delivery system is designed. Specifically, enteric coating technology is employed for formulations sensitive to gastric acid, with Eudragit polymers enabling pH-dependent release.

[0051] According to another aspect of the present invention, an anti-tumor metal photosensitizer is provided, wherein the active ingredient of the anti-tumor metal photosensitizer includes the above-mentioned osmium complex.

[0052] According to another aspect of the present invention, there is also provided the use of the above-mentioned osmium complex or photosensitizer in the preparation of a drug for photodynamic therapy of non-small cell lung cancer.

[0053] The application of the compound according to the embodiment of the present invention has at least the following beneficial effects: the compound of the present invention has good application prospects in photodynamic therapy for non-small cell lung cancer-related diseases.

[0054] According to some embodiments of the present invention, the photodynamic therapy comprises the steps of contacting the damaged tissue or pre-damaged tissue with the compound or photosensitizer, and exposing the damaged tissue or pre-damaged tissue to light having a wavelength that can be absorbed by the compound or photosensitizer.

[0055] According to some embodiments of the present invention, the damaged tissue or pre-damaged tissue is lung tissue.

[0056] According to some embodiments of the present invention, the wavelength of the light is 200-700 nm.

[0057] According to some embodiments of the present invention, the wavelength of the light is 600-700 nm.

[0058] According to some embodiments of the present invention, the contacting is used to activate a compound or a photosensitizer, wherein the compound or the photosensitizer is activated when the absorbed light dose is 60 to 70 J / cm 2 The light after use is sufficient to activate it.

[0059] According to some embodiments of the present invention, the light dose is 60-65 J / cm 2 .

[0060] Other features and advantages of the present invention will be set forth in the description which follows, and in part will be obvious from the description, or may be learned by practice of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0061] The above and / or additional aspects and advantages of the present invention will become apparent and readily understood from the following description of the embodiments with reference to the accompanying drawings, in which:

[0062] Figure 1 is the ultraviolet absorption spectrum of the metal osmium complex prepared in the embodiment of the present invention;

[0063] Figure 2 1 is a graph showing the test results of the photocatalytic oxidation ability of the metal osmium complex prepared in an embodiment of the present invention;

[0064] Figure 3 Graph showing the results of a test of the ability of the metal osmium complex prepared in an embodiment of the present invention to generate superoxide anions;

[0065] Figure 4 Graph showing the dark toxicity and phototoxicity test results of the metal osmium complex prepared in an example of the present invention against human non-small cell lung cancer cells (A549). DETAILED DESCRIPTION

[0066] The following will clearly and completely describe the concept of the present invention and the technical effects produced in conjunction with the embodiments, so as to fully understand the purpose, characteristics and effects of the present invention. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, other embodiments obtained by those skilled in the art without creative work are within the scope of protection of the present invention. Unless otherwise specified, the test methods used in the embodiments are all conventional methods; the materials, reagents, etc. used, unless otherwise specified, are reagents and materials obtained from commercial channels. Unless otherwise specified, the same parameter in each embodiment has the same value. The embodiments described below are exemplary and are only used to explain the present invention, and are not to be understood as limitations of the present invention.

[0067] In the description of the present invention, it should be understood that descriptions involving orientations, such as up, down, front, back, left, right, etc., indicating orientations or positional relationships, are based on the orientations or positional relationships shown in the accompanying drawings. They are only for the convenience of describing the present invention and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation. Therefore, they cannot be understood as limitations on the present invention.

[0068] The "room temperature" referred to in the present invention refers to 25±5°C, and is specifically 25°C in the embodiments.

[0069] Example

[0070] In this example, an osmium complex was prepared. The complex can be excited by red light. Its structural formula is as follows:

[0071]

[0072] The specific preparation process is as follows:

[0073] S1. Ammonium chloroosmate (0.439 g, 1 mmol) and 4'-bromo-2,2':6',2"-terpyridine (0.624 g, 2 mmol) were added to a reaction tube and ethylene glycol (8 mL) was added. The mixture was then heated under reflux at 200°C for 20 minutes to obtain osmium complex intermediate I.

[0074] The chemical reaction equation above is as follows:

[0075]

[0076] S2. To the cooled reaction solution was added 10 ml of a saturated aqueous solution of ammonium hexafluorophosphate to precipitate the product. After cooling to room temperature, the red precipitate was filtered off and washed with ice water to obtain the osmium complex intermediate II with a yield of 60.4%.

[0077] S3. The osmium complex intermediate II (0.096 g, 0.1 mmol) obtained in step S2, palladium acetate (0.0013 g, 0.005 mmol), potassium carbonate (0.02 mmol), pivalic acid (0.001 g, 0.01 mmol) and 2,5-bis(2-ethylhexyl)-3,6-bis(thiophen-2-yl)pyrrolo[3,4-C]pyrrole-1,4(2H,5H)-dione (0.109 g, 0.21 mmol) were added to a reaction tube for argon protection and N,N-dimethylacetamide (3 mL) was added. The mixture was then heated under reflux at 110°C for 6 hours. After the reactant was cooled to room temperature, water (10 mL) was added and filtered to obtain a blue precipitate. The precipitate was washed with water, dried and purified to obtain the target osmium complex with a yield of 16.4%.

[0078] The chemical reaction equation above is as follows:

[0079]

[0080] The product obtained in the above step S3 was subjected to structural characterization, and the mass spectrum was as follows: ESI-MS [CH3OH, m / z]: 851.35 [M-2PF6 - ] 2+ .

[0081] The H NMR spectrum of the product was: 1H NMR (500 MHz, DMSO-d6) δ9.52 (s, 4H), 9.21 (d, J = 8.1 Hz, 4H), 8.98 (d, J = 3.9 Hz, 4H), 8.68–8.61 (m, 2H), 8.18 (d, J = 4.9 Hz, 2H), 8.13 (d, J = 7.7 Hz, 4H), 7.64 (d, J = 5.7 Hz, 4H), 7.46 (s, 2H), 7.31 (t, J = 6.7 Hz, 4H), 4.18 (d, J = 7.4 Hz, 4H), 4.04–4.02 (m, 5H), 1.84–1.76 (m, 12H), 1.29 (s, 24H), 0.88–0.83 (m, 24H).

[0082] Test Case

[0083] In order to verify the application effect of the above complex, its performance was tested, as follows:

[0084] 1. Absorption spectrum determination of metal osmium complexes

[0085] The metal osmium complex prepared by the above operation was prepared into a 10 μM sample solution using phosphate-balanced saline (PBS) and acetonitrile (MeCN) as solvents, and then a double-beam UV-visible spectrophotometer was used to record the UV absorption spectrum of the new red-light-excited metal osmium complex to characterize its absorbance in phosphate-balanced saline and acetonitrile. The results are as follows: Figure 1 shown.

[0086] from Figure 1 It can be seen from the figure that the complex prepared in the embodiment of the present invention has good light absorption ability in organic solvents.

[0087] 2. Determination of the ability of metal osmium complexes to photocatalytically oxidize NADH

[0088] Under light irradiation, the metal complex can oxidize the reduced coenzyme I (NADH) into its oxidized form NAD + Therefore, the osmium complex (10 μM) and NADH (A 339 nm≈1.0) in a cuvette and its ability to oxidize NADH under light conditions was measured. Figure 2 shown.

[0089] As can be seen from the figure, the osmium complex has obvious photocatalytic oxidation ability for NADH.

[0090] 3. Determination of the ability of metal osmium complexes to generate superoxide anions

[0091] The superoxide anion probe dihydrorhodamine 123 (DHR123) was used to determine the ability of metal osmium complexes to generate superoxide anions. The changes in fluorescence intensity of the mixed solution of the test sample and DHR123 under different illumination times were monitored by a Techcomp FL970 fluorescence spectrophotometer to reflect the ability to generate superoxide anions.

[0092] A mixed solution of metal osmium complex (10 μM) and DHR123 was excited in a 1 cm quartz tube at λex = 465 nm. The entrance and exit slits were set to 2.5 nm. The superoxide anion generation capacity was measured under 635 nm illumination. Figure 3 It is shown that the metal osmium complex has the ability to generate superoxide anions after light irradiation.

[0093] 4. Photodynamic therapy effects of metal osmium complexes on human non-small cell lung cancer cell lines

[0094] Resazurin solution is blue and is commonly used as an acid-base indicator (orange at pH 3.8 to deep purple at pH 6.5) and a redox indicator. During cell viability assays, resazurin penetrates cells and is irreversibly reduced by living cells to a pink color, accompanied by the appearance of red fluorescent resorufin. The absorbance or fluorescence intensity of resorufin is positively correlated with cell number and reducing capacity, allowing analysis of cell proliferation using an enzyme-linked immunofluorescence assay.

[0095] The resazurin experimental steps are as follows:

[0096] (1) First, revive one tube of A549 tumor cells and culture them with fresh complete culture medium (DMEM medium + 10 vol% fetal bovine serum + 1 vol% penicillin-streptomycin mixture). After passage twice, start the experiment.

[0097] (2) When the cells reached the logarithmic growth phase, they were seeded into two 96-well plates at a density of 5000 cells / well (100 μL of culture medium was used to culture cells in each well, one plate was for the light group and the other plate was for the dark control group), and cultured in a 37°C, 5% CO2 incubator.

[0098] (3) After the cells adhere to the wall, the original culture medium was aspirated and 100 μL of metal osmium complexes at six concentrations of 100, 50, 10, 1, 0.1, and 0.01 μM were added to each well. The cells were gently shaken and incubated in a carbon dioxide incubator (37°C, 5% CO2) in the dark.

[0099] (4) After incubation for 6 h, the cell culture plates in the illumination group were placed under a 635 nm light source for 45 min (light dose of 63.7 J / cm 2), and then returned to the incubator for further incubation in the dark for 42 h (cells in the dark control group were kept in the incubator in the dark for incubation).

[0100] (5) After incubation for 42 h, the culture medium was discarded from each well, and 80 μL of resazurin (100 mg / mL) was added to each well. The cells were incubated in a 37°C incubator for another 4 h. EX540 / EM590 was detected using the fluorescence plate of an enzyme-linked immunosorbent assay (ELISA) to calculate the cell proliferation inhibition rate and obtain the IC 50 value (drug concentration when the inhibition rate is equal to 50%).

[0101] The results are as follows Figure 4 As shown in the figure, the resazurin method was used to detect the killing effect of different concentrations of osmium complexes on human non-small cell lung cancer cell line (A549 cells) under dark and light treatment conditions. It can be seen that in the absence of light, the IC 50 Greater than 100 μM, IC against human non-small cell lung cancer cell lines under light conditions 50 The phototherapy index PI is as high as 20, indicating that the metal osmium complex of the present invention has a strong photodynamic therapy effect.

[0102] The embodiments of the present invention are described in detail above, but the present invention is not limited to the above embodiments. Various changes can be made within the knowledge scope of ordinary technicians in the relevant technical field without departing from the scope of the present invention.

Claims

1. An osmium complex, characterized in that: It has the following structural formula: Among them, X - Represents anions.

2. The osmium complex according to claim 1, characterized in that: In the complex, X - PF6 - .

3. The method for preparing the osmium complex according to claim 1 or 2, wherein: The steps include: S1. Make Os 4+ and 4'-bromo-2,2':6',2"-terpyridine in a solvent under heating to prepare an osmium complex intermediate I; S2. Add the osmium complex intermediate I prepared in step S1 to the - to obtain an osmium complex intermediate II; S3. The osmium complex intermediate II obtained in step S1 is reacted with 2,5-bis(2-ethylhexyl)-3,6-bis(thiophen-2-yl)pyrrolo[3,4-C]pyrrole-1,4(2H,5H)-dione in the presence of a palladium catalyst, a carbonate, and pivalic acid to generate the osmium complex.

4. Use of the osmium complex according to claim 1 or 2 in the preparation of antitumor drugs.

5. The use according to claim 4, characterized in that: The anti-tumor drug is an anti-non-small cell lung cancer drug; preferably, the non-small cell lung cancer is A549 cell line lung cancer.

6. An anti-tumor metal photosensitizer, characterized in that: The active ingredient of the anti-tumor metal photosensitizer includes the osmium complex according to claim 1 or 2.

7. Use of the osmium complex according to claim 1 or 2 or the photosensitizer according to claim 6 in the preparation of a drug for photodynamic therapy of non-small cell lung cancer.

8. The use according to claim 7, characterized in that: The photodynamic therapy comprises the following steps: contacting the damaged tissue or pre-damaged tissue with the compound or photosensitizer, and exposing the damaged tissue or pre-damaged tissue to light having a wavelength that can be absorbed by the compound or photosensitizer.

9. The use according to claim 8, characterized in that: The wavelength of the light is 200-700 nm; preferably, the wavelength of the light is 600-700 nm.

10. The use according to claim 9, characterized in that: The contact is used to activate the compound or photosensitizer, and the compound or photosensitizer is activated when the absorbed light dose is 60 to 70 J / cm 2 The light after use is sufficient to activate it.

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