A tumor phototherapy preparation and a preparation method and application thereof

Abstract

This application provides a tumor phototherapy formulation, its preparation method, and its application, belonging to the field of biomedical technology. The tumor phototherapy formulation includes a carrier, a pH-responsive polymer chain grafted onto the surface of the carrier, and catalase and indocyanine green loaded onto the pH-responsive polymer chain and the surface of the carrier. The carrier comprises polydopamine nanoparticles with a mesoporous cup-like structure. This formulation not only possesses targeted motility, achieving efficient accumulation at the tumor site by utilizing the hydrogen peroxide concentration gradient and pH-responsive motility behavior between the tumor microenvironment and the normal physiological environment, but also, through effective combination and synergistic effects with the suitable photosensitizer indocyanine green, combines and significantly enhances the photothermal and photodynamic therapeutic effects, achieving highly efficient tumor killing.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to a tumor phototherapy preparation, its preparation method, and its application. Background Technology

[0002] Tumors are diseases caused by the abnormal proliferation of cells in the body, and can be broadly classified into benign tumors and malignant tumors (cancer). Malignant tumors are particularly invasive and seriously endanger human health. Currently, there are various conventional treatments for malignant tumors, such as chemotherapy and phototherapy. Chemotherapy uses cytotoxic drugs to kill rapidly proliferating tumor cells; phototherapy (or photodynamic therapy) uses light to irradiate photothermal agents or photosensitive materials concentrated at the tumor tissue, generating localized high temperatures or cytotoxic reactive oxygen species (ROS), thereby killing tumor cells.

[0003] However, existing treatments still have significant limitations: chemotherapy drugs lack specificity, damaging normal tissues while killing cancer cells, leading to severe side effects and easily inducing tumor drug resistance; phototherapy relies on blood circulation to distribute the photothermal or photosensitive reagents injected into the patient's body to various parts of the body, with low tumor targeting efficiency, often requiring high-dose administration or high-power light irradiation to ensure treatment effectiveness, and significant side effects.

[0004] Therefore, there is an urgent need to develop formulations with highly effective therapeutic effects. Summary of the Invention

[0005] In view of the technical problems existing in the background art, this application provides a tumor phototherapy preparation, its preparation method and application, aiming to provide a tumor treatment preparation with highly effective therapeutic or inhibitory effects.

[0006] In a first aspect, embodiments of this application provide a tumor phototherapy formulation, comprising a carrier, a pH-responsive polymer chain grafted onto the surface of the carrier, and catalase and indocyanine green loaded onto the pH-responsive polymer chain and the surface of the carrier, wherein the carrier comprises polydopamine nanoparticles having a mesoporous bowl-shaped structure.

[0007] Secondly, this application also proposes a method for preparing a tumor phototherapy agent, comprising the following steps: The support, triethylamine and 2-dimethylaminopyridine were dispersed in tetrahydrofuran, and 2-bromoisobutyryl bromide was added to obtain the intermediate product; The intermediate product, pH-responsive polymer monomer molecules and a first solvent are mixed, and 2,2-bipyridine and cuprous bromide are added to carry out a polymerization reaction to obtain a first composite, wherein the support comprises polydopamine nanoparticles with a mesoporous cup-shaped structure. The second complex was obtained by loading catalase onto the surface of the first complex using a glutaraldehyde chemical cross-linking method. The second complex, indocyanine green, and the second solvent are mixed and reacted to obtain a tumor phototherapy preparation.

[0008] Thirdly, this application also proposes the application of the aforementioned tumor phototherapy preparation in the preparation of drugs for treating or inhibiting tumors.

[0009] The technical solution proposed in this application has the following beneficial effects: In this application, polydopamine nanoparticles with a mesoporous bowl-shaped structure are used as a carrier, and pH-responsive polymer chains, catalase, and indocyanine green are loaded on their surface. This enables the formulation to not only have targeted motility, but also achieve efficient enrichment of the formulation at the tumor site by utilizing the hydrogen peroxide concentration gradient and pH-responsive motility behavior between the tumor microenvironment and the normal physiological environment. Furthermore, through effective combination and synergistic effect with the suitable photosensitizer indocyanine green (ICG), the combined and significantly enhanced photothermal and photodynamic therapy effects are achieved, resulting in highly efficient tumor killing.

[0010] The above description is only an overview of the technical solution of this application. In order to better understand the technical means of this application and to implement it in accordance with the contents of the specification, and to make the above and other objects, features and advantages of this application more obvious and understandable, specific embodiments of this application are given below. Attached Figure Description

[0011] To more clearly illustrate the technical solutions of this application, the accompanying drawings used in this application will be briefly described below. Obviously, the drawings described below are merely some embodiments of this application. For those skilled in the art, other drawings can be obtained from these drawings without any creative effort.

[0012] Figure 1 TEM image of PDC@ICG obtained in Example 1; Figure 2 A schematic flowchart illustrating a method for preparing a tumor phototherapy agent according to an embodiment of this application; Figure 3 The nitrogen adsorption-desorption isotherm and pore size distribution curve of the PDC prepared in Example 1 are shown. Figure 4 The particle size distribution curves of PDC prepared in Example 1 under different pH environments are shown. Figure 5 The image shows the photothermal performance test results of the PDC prepared in Example 1 of Experimental Example (IV); Figure 6The UV-Vis-NIR absorption spectra of ICG, PDC, and PDC@ICG in Experimental Example (V); Figure 7 The temperature rise curves of ICG, PDC, and PDC@ICG in Experimental Example (VI) are shown below. Figure 8 The UV-Vis absorption spectra of the sample solution in Experiment Example (VII) under different light exposure times; Figure 9 This is a comparison of the motion diffusion coefficients of PDC@ICG and PDA@ICG in an H2O2-rich environment in Experimental Example (8); Figure 10 The above are fluorescence images of PDC@ICG and PDA@ICG in the tumor cell environment in Experiment Example (IX); Figure 11 The figure shows the changes in tumor volume in 4T1 tumor-bearing mice after phototherapy with PDC@ICG and PDA@ICG in Experiment (10). Detailed Implementation

[0013] The embodiments of the technical solution of this application will be described in detail below. The following embodiments are only used to illustrate the technical solution of this application more clearly, and are therefore only examples, and should not be used to limit the scope of protection of this application.

[0014] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains; the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the application; the terms “comprising” and “having”, and any variations thereof, in the specification and claims of this application are intended to cover non-exclusive inclusion.

[0015] Various embodiments of this application may exist in the form of a range; it should be understood that the description in the form of a range is merely for convenience and brevity and should not be construed as a hard limitation on the scope of this application; therefore, it should be considered that the range description has specifically disclosed all possible sub-ranges and single numerical values ​​within that range. For example, it should be considered that the range description from 1 to 6 has specifically disclosed sub-ranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., and single numbers within the range, such as 1, 2, 3, 4, 5, and 6, regardless of the range. Furthermore, whenever a numerical range is referred to herein, it means including any referenced number (fraction or integer) within the referred range.

[0016] In the description of the embodiments of this application, technical terms such as "first" and "second" are used only to distinguish different objects and should not be construed as indicating or implying relative importance or implicitly specifying the number, specific order, or primary and secondary relationship of the indicated technical features. In the description of the embodiments of this application, "multiple" means two or more, unless otherwise explicitly defined.

[0017] In this document, the term "embodiment" means that a particular feature, structure, or characteristic described in connection with an embodiment may be included in at least one embodiment of this application. The appearance of this phrase in various places throughout the specification does not necessarily refer to the same embodiment, nor is it a separate or alternative embodiment mutually exclusive with other embodiments. It will be explicitly and implicitly understood by those skilled in the art that the embodiments described herein can be combined with other embodiments.

[0018] In the description of the embodiments in this application, the term "and / or" is merely a description of the relationship between related objects, indicating that three relationships can exist. For example, A and / or B can represent: A existing alone, A and B existing simultaneously, and B existing alone. Additionally, the character " / " in this document generally indicates that the preceding and following related objects have an "or" relationship.

[0019] In the description of the embodiments of this application, the term "at least one" refers to one or more, "more than one" refers to two or more (including two), similarly, "multiple sets" refers to two or more (including two sets), and "multiple pieces" refers to two or more (including two pieces). "At least one," "at least one of the following," or similar expressions refer to any combination of these items, including any combination of single or multiple items. For example, "at least one of a, b, or c," or "at least one of a, b, and c," can all represent: a, b, c, ab (i.e., a and b), ac, bc, or abc, where a, b, and c can be single or multiple.

[0020] This application provides a tumor phototherapy preparation, such as... Figure 1 and Figure 2 As shown, the tumor phototherapy formulation includes a carrier, a pH-responsive polymer chain grafted onto the surface of the carrier, and catalase and indocyanine green loaded on the pH-responsive polymer chain and the surface of the carrier, wherein the carrier includes polydopamine nanoparticles with a mesoporous bowl-shaped structure.

[0021] In this application, polydopamine nanoparticles with a mesoporous bowl-shaped structure are used as a carrier, and pH-responsive polymer chains, catalase, and indocyanine green are loaded on their surface. This enables the formulation to not only have targeted motility, but also achieve efficient enrichment of the formulation at the tumor site by utilizing the hydrogen peroxide concentration gradient and pH-responsive motility behavior between the tumor microenvironment and the normal physiological environment. Furthermore, through effective combination and synergistic effect with the suitable photosensitizer indocyanine green (ICG), the combined and significantly enhanced photothermal and photodynamic therapy effects are achieved, resulting in highly efficient tumor killing.

[0022] For ease of description, the complex consisting of a carrier, a pH-responsive polymer chain grafted onto the surface of the carrier, and a catalase loaded onto the pH-responsive polymer chain and the surface of the carrier is referred to as PDC. The tumor phototherapy formulation proposed in this application is abbreviated as PDC@ICG.

[0023] The pH-responsive polymer chains not only exhibit pH responsiveness, promoting the movement of tumor phototherapy agents towards low-pH regions, but also provide loading sites, increasing the loading of catalase (CAT) and indocyanine green (ICG), further enhancing the agent's self-driving ability and phototherapy efficacy. Furthermore, the loaded catalase exhibits chemotaxis towards high-concentration hydrogen peroxide regions, further improving the agent's autonomous target-seeking ability and motility. In addition, the intrinsic symmetry of the mesoporous cup-shaped carrier used in this application results in an asymmetrical distribution of catalase on its surface, further strengthening the agent's autonomous motility. Through the combined effects of these factors, the agent in this application possesses excellent motility and targeting, enabling efficient accumulation at tumor sites.

[0024] Meanwhile, under near-infrared light irradiation, ICG can convert light energy into heat energy and generate a large amount of singlet oxygen. 1 O2) is used to achieve photothermal therapy (PTT) and photodynamic therapy (PDT). At the same time, there is a good match and binding between ICG and the loading sites formed on PDC, which can successfully achieve ICG loading on the surface of PDC. In addition, there is a synergistic effect between the two. When the two are combined, the photothermal and photodynamic effects are significantly enhanced. Compared with the combination of conventional chemotherapy drugs and PDC, the treatment effect is better.

[0025] In addition, catalase can reduce reactive oxygen species in tumor sites, improve the hypoxic microenvironment of tumors, and further enhance the tumor-killing effect of the preparation.

[0026] In some embodiments, the loading of indocyanine green is greater than 0 and less than or equal to 15 wt%. Compared to other photosensitizers, indocyanine green not only exhibits good compatibility and binding with the complex composed of mesoporous cup-shaped polydopamine, pH-responsive polymer chains, and catalase, but the overall tumor phototherapy formulation also demonstrates highly effective therapeutic efficacy. It is understood that in this application, the loading of indocyanine green refers to the percentage of the mass of indocyanine green relative to the mass of the tumor phototherapy formulation.

[0027] In some embodiments, the polymer in the pH-responsive polymer chain includes poly(dimethylaminoethyl methacrylate), which can stretch or contract when the environmental pH changes, thereby affecting the size of the formulation and exhibiting pH responsiveness. The pH-responsive polymer chain has a chain length of 10-100 nm, is controllable and has a wide adjustment range, possesses strong loading capacity, can load a large amount of catalase and indocyanine green, and can adjust the loading amount as needed by controlling the chain length, thereby balancing and improving the motility and phototherapy effect of the formulation. In some embodiments, the catalase loading amount in the formulation is 100-1000 mg / g. It is understood that in this application, the catalase loading amount refers to the percentage of catalase mass relative to the mass of the PDC.

[0028] In some embodiments, the average particle size of the carrier is 150-500 nm; for example, it can be 150 nm, 160 nm, 180 nm, 200 nm, 300 nm, 400 nm, 450 nm, 500 nm, or any value between two of the above. Using a nanoscale carrier helps to facilitate better deep penetration of the formulation into tumor tissue and into tumor cells.

[0029] Furthermore, this application also proposes a method for preparing a tumor phototherapy agent, based on which the aforementioned tumor phototherapy agent can be prepared. The method for preparing the tumor phototherapy agent includes the following steps: S10: The support, triethylamine and 2-dimethylaminopyridine are dispersed in tetrahydrofuran, and 2-bromoisobutyryl bromide is added to obtain the intermediate product.

[0030] S20, the intermediate product, pH-responsive polymer monomer molecules and the first solvent are mixed, and 2,2-bipyridine and cuprous bromide are added to carry out a polymerization reaction to obtain the first composite, wherein the support comprises polydopamine nanoparticles with a mesoporous cup-shaped structure.

[0031] S30, using the glutaraldehyde chemical cross-linking method, load catalase onto the surface of the first complex to obtain the second complex.

[0032] S40, the second complex, indocyanine green and the second solvent are mixed and reacted to obtain a tumor phototherapy preparation.

[0033] Please see Figure 2 In this application, a carrier and 2-dimethylaminopyridine are first mixed in an alkaline environment, and an initiator is introduced to promote the reaction, thereby forming initiation sites on the carrier surface and creating a growth environment for subsequent polymer chain grafting. Then, controlled polymerization (e.g., atom transfer radical polymerization) is carried out at the initiation sites to modify the carrier surface with pH-responsive polymer chains. Next, catalase is linked to the surface of the pH-responsive polymer chains and / or the carrier via glutaraldehyde chemical cross-linking to form PDC. Finally, PDC and ICG are mixed, and through π-π stacking and electrostatic interactions, ICG is successfully linked to the pH-responsive polymer chains and / or the carrier surface, thus obtaining a tumor phototherapy formulation. This preparation method is simple and can effectively and stably synthesize PDC@ICG tumor phototherapy formulations.

[0034] In step S10: The amount of each raw material added can meet the following conditions: for every 8 grams of the carrier, add 45~55 mL of 2-bromoisobutyryl bromide, 55~65 mL of triethylamine and 45~55 mL of 2-dimethylaminopyridine.

[0035] Meanwhile, in order to ensure the effective formation of initiation sites, in step S10, after adding 2-bromoisobutyryl bromide, the reaction can be carried out at room temperature (e.g., 20~30℃) for 24 hours.

[0036] In step S20: The pH-responsive polymer monomer molecule includes dimethylaminoethyl methacrylate.

[0037] The amount of each raw material added can meet the following conditions: for every 2 grams of the intermediate product, 2.5 to 6 mL of the pH-responsive polymer monomer molecules are added; the mass ratio of the intermediate product to 2,2-bipyridine is 2:20 to 24; the mass ratio of 2,2-bipyridine to cuprous bromide is 1 to 6:1; for example, it can be 1:1, 2:1, 3:1, 4:1, 5:1, 6:1, or any value between any two of the above.

[0038] The first solvent includes a mixed solution of methanol and water, wherein the volume ratio of methanol to water is 1 to 3:1; for example, it can be 1:1, 1.5:1, 2:1, 2.5:1, 3:1, or any value between any two of the above.

[0039] The polymerization reaction temperature is 10~30℃, for example, it can be 10℃, 15℃, 20℃, 25℃, 30℃, or any value between two of the above. The polymerization reaction time is 1~3h; for example, it can be 1h, 1.5h, 2h, 2.5h, 3h, or any value between two of the above.

[0040] In some embodiments, step S30 may specifically include: The first complex was washed with PBS buffer, and then the washed first complex was mixed with glutaraldehyde and PBS buffer and reacted at 10-30°C for 1.5-2.5 h. Then catalase was added and reacted at 10-30°C for 12-24 h to obtain the second complex.

[0041] The amount of each raw material can meet the following conditions: 0.05~0.15mL of glutaraldehyde and 0.4~4mg of catalase are added for each milligram of the first complex.

[0042] In step S40: The mass ratio of the second complex to the indocyanine green can be 5 to 20:1; for example, it can be 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, or any value between any two of the above.

[0043] In some embodiments, the reaction temperature is 10~30℃, for example, it can be 10℃, 15℃, 20℃, 25℃, 30℃ and any two of the above values; the reaction time is 6~24h; for example, it can be 6h, 10h, 12h, 15h, 18h, 20h, 22h, 24h and any two of the above values.

[0044] In some embodiments, the second solvent includes PBS buffer.

[0045] In some embodiments, before mixing the second complex with the indocyanine green and the second solvent, the process may further include washing the second complex with the second solvent.

[0046] Furthermore, this application also proposes the application of the aforementioned tumor phototherapy preparation in the preparation of drugs for treating or inhibiting tumors.

[0047] The tumor phototherapy formulation proposed in this application not only possesses targeted motility and can efficiently accumulate at the tumor site, but also, through effective binding and synergistic effects with a suitable photosensitizer, indocyanine green (ICG), it combines and significantly enhances the photothermal and photodynamic therapeutic effects, resulting in a highly effective tumor-killing effect. Based on this, this tumor phototherapy formulation can be used directly as a drug for treating or inhibiting tumors, or as a component or precursor in the preparation of drugs for treating or inhibiting tumors.

[0048] In a preferred embodiment, the tumor is a breast tumor. Studies have shown that the phototherapy formulation proposed in this application has a high therapeutic effect on breast tumors and is suitable for use as a breast tumor drug.

[0049] The following are some specific embodiments. It should be noted that the embodiments described below are exemplary and are only used to explain this application, and should not be construed as limiting this application. Where specific techniques or conditions are not specified in the embodiments, they shall be performed in accordance with the techniques or conditions described in the literature in this field or according to the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be obtained commercially.

[0050] Example 1 (1) 8 mg of mesoporous cup-shaped polydopamine carrier (PDA, with an average particle size of about 213 nm; prepared by the dual-template method, see reference Angew. Chemie Int. Ed. 2018, 57, 6176-6180), 0.06 mL of triethylamine and 0.05 mL of 2-dimethylpyridine were dissolved in 5 mL of tetrahydrofuran, and 0.05 mL of 2-bromoisobutyryl bromide was added dropwise. The reaction was carried out at room temperature (about 25 °C) for 24 h to obtain the intermediate product.

[0051] (2) Take 2 mg of intermediate product, 0.004 mL of dimethylaminoethyl methacrylate (DMAEMA) and 5 mL of methanol aqueous solution (methanol to water volume ratio of 2:1), then add 2,2-bipyridine and copper bromide according to the mass ratio of 2:1, and polymerize at 25°C for 1 h to obtain the first complex.

[0052] (3) Take 1 mg of the first complex, wash it twice with PBS buffer, then sonicate it in 0.9 mL of PBS buffer with a concentration of 10 mmol / L, add 0.1 mL of glutaraldehyde, shake at 25℃ for 2 h, then add 3 mg of catalase, shake at 25℃ for 16 h, wash with water three times to obtain the second complex (PDC).

[0053] (4) Wash 1 mg of the second complex twice with PBS buffer at a concentration of 10 mmol / L, then sonicate it in 3.9 mL of PBS buffer at a concentration of 10 mmol / L, sonicate for 10 min, then add 0.1 mL of indocyanine green at a concentration of 1 mg / mL, stir at 25 °C for 12 h, wash three times with water to obtain the tumor phototherapy preparation (PDC@ICG).

[0054] Example 2 The scheme in this embodiment is basically the same as that in embodiment 1, except that in step (4) of this embodiment, the amount of indocyanine green added is changed to 0.05 mL of indocyanine green with a concentration of 1 mg / mL. Other than that, all other parameters and conditions remain unchanged.

[0055] Example 3 The scheme in this embodiment is basically the same as that in embodiment 1, except that in step (4) of this embodiment, the amount of indocyanine green added is changed to 0.2 mL of indocyanine green with a concentration of 1 mg / mL. Other than that, all other parameters and conditions remain unchanged.

[0056] Comparative Example 1 The comparative example scheme is basically the same as that of Example 1, except that the product obtained in this comparative example is PDA@ICG. Accordingly, the second complex in step (4) is replaced by the mesoporous cup-shaped polydopamine carrier in step (1), and steps (1) to (3) are omitted. Apart from this, all other parameters and conditions remain unchanged.

[0057] Comparative Example 2 This comparative example is basically the same as Example 1, except that the loaded drug is replaced with the chemotherapy drug doxorubicin hydrochloride (DOX solution), and urease is added to improve motility. Accordingly, steps (3) and (4) are changed as follows: (3) Take 1 mg of the first complex, wash it twice with PBS buffer, then sonicate it in 0.9 mL of PBS buffer with a concentration of 10 mmol / L, add 0.1 mL of glutaraldehyde, shake at 25℃ for 2 h, then add 3 mg of a mixture of catalase and urease (mass ratio of 1:3), shake at 25℃ for 16 h, wash three times with water to obtain the third complex (PDCU).

[0058] (4) Wash 1 mg of the third complex twice with PBS buffer, then sonicate it in 1 mL of PBS solution of 1 mg / L doxorubicin hydrochloride (DOX), stir at 25 °C for 12 h, wash with water three times to obtain the tumor chemotherapy preparation (PDCU@DOX).

[0059] Apart from this, all other parameters and conditions remain unchanged.

[0060] Experimental Example (1): The morphology of PDC@ICG prepared in Example 1 was observed using a transmission electron microscope (TEM).

[0061] Results analysis: such as Figure 1 As shown, its overall shape is bowl-shaped and its surface is loaded with ICG.

[0062] Experimental Example (II): The PDC prepared in Example 1 was characterized by TEM under acidic conditions (pH 5.6), and the results are as follows. Figure 3 As shown, the polymer chains were successfully grafted and remained in an extended state under acidic conditions.

[0063] Furthermore, using the Bradford Protein Assay Kit manufactured by Beyotime, and following the detection method given in the kit's instructions, the catalase loading was determined to be 489.28 mg / g.

[0064] Experimental Example (3): The PDC prepared in Example 1 was dispersed in PBS buffer solutions of different pH values ​​(pH 5.6, pH 6.2, pH 6.8, pH 7.4), and the particle size distribution was detected using a dynamic light scattering instrument.

[0065] like Figure 4 As shown in the figure, the horizontal axis size represents the particle size and the vertical axis intensity represents the peak intensity. It can be seen from the figure that as the pH decreases, the particle size of PDC gradually increases. Obviously, the PDC prepared in this embodiment has pH responsiveness.

[0066] Experimental Example (IV): The PDC prepared in Example 1 was dispersed in water to prepare a test sample. The sample was exposed to 808nm near-infrared light, using different light intensities (0.5 W / cm²). 2 1.0 W / cm 2 1.5 W / cm 2 2 W / cm 2 The sample was irradiated with light, and the temperature change of the sample was observed using an infrared thermal imager.

[0067] like Figure 5 As shown, under the same light intensity, the sample temperature increases as the irradiation time gradually increases, indicating that the PDC prepared in this embodiment also has photothermal conversion properties and has the potential for photothermal driving and treatment.

[0068] Experimental Example (5): ICG, PDC, and PDC@ICG from Example 1 were dispersed in water as test solutions and detected using a UV-Vis-NIR absorption spectrometer.

[0069] like Figure 6 As shown in the figure, the horizontal axis represents wavelength, and the vertical axis represents absorbance. It can be seen from the figure that PDC has no significant absorption at 780 nm, but after loading ICG, PDC@ICG exhibits an absorption curve similar to that of ICG, indicating that ICG was successfully loaded onto PDC in this embodiment. Furthermore, calculations based on the absorption curves show that the ICG loading in PDC@ICG is 8.15%.

[0070] Furthermore, referring to the above method, the ICG loading in Examples 2 and 3 was tested. In the product obtained in Example 2, the ICG loading was 4.32%, and in the product obtained in Example 3, the ICG loading was 11.54%.

[0071] Experimental Example (VI): ICG, PDC, and PDC@ICG from Example 1 were dispersed in water to prepare ICG, PDC, and PDC@ICG samples with a concentration of 0.3 mg / mL. The samples were exposed to 808 nm near-infrared light at an intensity of 2 W / cm². 2 The temperature changes around the sample were recorded using an infrared thermal imager at different times (0 to 600 s) of irradiation, and temperature rise curves were plotted.

[0072] like Figure 7 As shown in the figure, the horizontal axis "Time" represents time, and the vertical axis "temperature" represents temperature. The figure shows that the temperature around all three samples increased over time, and the PDC@ICG sample showed a more significant temperature increase compared to the other two samples. This indicates that the composite of PDC and ICG in this application achieved a synergistic effect, resulting in superior photothermal conversion performance.

[0073] Experimental Example (VII): The PDC@ICG prepared in Example 1 was mixed with a solution of 1,3-diphenylisobenzofuran (DPBF, singlet oxygen indicator probe) to prepare a sample solution. A wavelength of 808 nm and an illumination intensity of 2 W / cm² were used. 2 The sample solution was irradiated with near-infrared light, and the absorbance curves of the sample solution in the wavelength range of 300 nm to 600 nm were recorded using a UV-Vis absorption spectrometer at different irradiation times (0 min, 1 min, 2 min, 3 min, 4 min, 5 min).

[0074] like Figure 8As shown in the figure, the horizontal axis represents wavelength, and the vertical axis represents absorbance. The figure shows that the absorption intensity of the sample solution gradually decreases with increasing illumination time, indicating that PDC@ICG has the ability to generate singlet oxygen under 808nm illumination, which can be used for photodynamic therapy of tumors.

[0075] Experimental Example (8) The active PDC@ICG (denoted as active PDC@ICG) prepared in Example 1 and the inert PDA@ICG (denoted as Inert PDA@ICG) prepared in Comparative Example 1 were used as samples, and the test solution was prepared according to the following method: The sample was dispersed in PBS solution of 100 μM H2O2 at a concentration of 0.2 mg / mL as the test solution.

[0076] The test solution was spread on an ultrasonically cleaned coverslip. After the system stabilized, the motion behavior of the sample in the test solution was observed using an inverted dark-field microscope under conditions of no near-infrared light irradiation or under conditions of 808 nm near-infrared light irradiation. The motion trajectory and corresponding coordinates of the sample were recorded using commercial software (Video Spot Tracker), and the diffusion coefficient D was calculated. e .

[0077] like Figure 9 As shown in the figure, NIR(-) represents no near-infrared light irradiation, and NIR(+) represents near-infrared light irradiation. Clearly, active PDC@ICG can be effectively driven by physiologically concentrated H2O2 as fuel, and compared to inert PDA@ICG, it exhibits significantly enhanced diffusion behavior. This mobility can be greatly increased under light irradiation conditions, giving it the potential to be used as a highly efficient phototherapy agent.

[0078] Experimental Example (IX): Active PDC@ICG (denoted as active PDC@ICG) prepared in Example 1 and inert PDA@ICG (denoted as Inert PDA@ICG) prepared in Comparative Example 1 were used as samples. The samples to be tested were mixed with excess cyanin fluorescent dye (Cy5) and shaken to obtain fluorescently modified samples. Well-grown 4T1 tumor cells (Pronosei Biotechnology Co., Ltd.) were then subjected to a 4×10⁻⁶ ion exchange rate. 5 Cells were seeded at a density of 1000 cells in DMEM medium and cultured at 37°C for 24 h. Subsequently, the medium was replaced with a new medium containing the fluorescently modified sample and cultured under conditions without near-infrared light irradiation or under conditions with 808 nm near-infrared light irradiation. After 60 min, the enrichment status of the sample in the cells was observed using a fluorescence microscope.

[0079] like Figure 10As shown, active PDC@ICG exhibits a stronger tumor enrichment effect in mouse breast cancer cells than inert PDA@ICG, and this enrichment effect is significantly enhanced under light conditions.

[0080] Experimental Example (10): PDC@ICG prepared in Example 1, PDA@ICG prepared in Comparative Example 1, and PDCU@DOX prepared in Comparative Example 2 were used as test samples, and PBS buffer was used as a blank control sample to conduct in vivo animal experiments to verify the curative effect on tumors.

[0081] 4T1 tumor cells in good growth condition were formulated into 2×10⁻⁶ cells. 7 A cell suspension of 100 cells / mL was prepared and then injected subcutaneously into the tissues of mice weighing 20–22 g. The tumor volume was increased to 100–150 mm. 3 Mice were injected with 100 μL of different samples (test samples and blank control samples) via the tail vein, with the administration occurring every three days at a dose of 5 mg / kg each time. Mice were sacrificed on day 14 after administration, and subcutaneous tumors were removed. Five replicates were performed for each group.

[0082] like Figure 11 As shown, the tumors treated with PDA@ICG, PDCU@DOX, and PDC@ICG all showed a certain degree of growth inhibition compared to the blank control group. However, compared to the control group, the tumor growth of mice treated with PDC@ICG was significantly inhibited, with an inhibition rate of over 98%, indicating that the tumor phototherapy preparation prepared in this application can effectively kill tumors.

[0083] It should be noted that this application is not limited to the above-described embodiments. The above embodiments are merely examples, and any embodiments with the same structure and effect as the technical concept within the scope of this application are included in the technical scope of this application. Furthermore, various modifications that can be conceived by those skilled in the art to the embodiments, and other ways of constructing by combining some of the constituent elements of the embodiments, without departing from the spirit of this application, are also included in the scope of this application.

Claims

1. A tumor phototherapy preparation, characterized in that, The invention includes a carrier, a pH-responsive polymer chain grafted onto the surface of the carrier, and catalase and indocyanine green loaded onto the pH-responsive polymer chain and the surface of the carrier, wherein the carrier comprises polydopamine nanoparticles having a mesoporous bowl-shaped structure.

2. The tumor phototherapy preparation according to claim 1, characterized in that, The indocyanine green loading is greater than 0 and less than or equal to 15%.

3. The tumor phototherapy preparation according to claim 1, characterized in that, The polymer in the pH-responsive polymer chain includes poly(dimethylaminoethyl methacrylate); and / or, The pH-responsive polymer chain has a chain length of 10-100 nm; and / or, The catalase loading is 100~1000 mg / g; and / or, The average particle size of the carrier is 150~500nm.

4. A method for preparing a tumor phototherapy agent according to any one of claims 1 to 3, characterized in that, Includes the following steps: The support, triethylamine and 2-dimethylaminopyridine were dispersed in tetrahydrofuran, and 2-bromoisobutyryl bromide was added to obtain the intermediate product; The intermediate product, pH-responsive polymer monomer molecules and a first solvent are mixed, and 2,2-bipyridine and cuprous bromide are added to carry out a polymerization reaction to obtain a first composite, wherein the support comprises polydopamine nanoparticles with a mesoporous cup-shaped structure. The second complex was obtained by loading catalase onto the surface of the first complex using a glutaraldehyde chemical cross-linking method. The second complex, indocyanine green, and the second solvent are mixed and reacted to obtain a tumor phototherapy preparation.

5. The preparation method according to claim 4, characterized in that, The mass ratio of the second complex to the indocyanine green is 5-20:1; and / or, The reaction temperature is 10~30℃, and the reaction time is 6~24h; and / or, The second solvent includes PBS buffer; And / or, Before mixing the second complex with the indocyanine green and the second solvent, the method further includes washing the second complex with the second solvent.

6. The preparation method according to claim 4, characterized in that, For every 8 grams of the carrier, add 45-55 mL of 2-bromoisobutyryl bromide, 55-65 mL of triethylamine, and 45-55 mL of 2-dimethylaminopyridine. For every 2 grams of the intermediate product, 2.5 to 6 mL of the pH-responsive polymer monomer molecules are added. The mass ratio of the intermediate product to 2,2-bipyridine is 2:20~24; The mass ratio of 2,2-bipyridine to cuprous bromide is 1~6:1; The pH-responsive polymer monomer molecules include dimethylaminoethyl methacrylate; The polymerization reaction is carried out at a temperature of 10~30℃ for 1~3 hours.

7. The preparation method according to claim 4, characterized in that, The steps for loading catalase onto the surface of the first complex using the glutaraldehyde chemical cross-linking method to obtain the second complex include: The first complex was washed with PBS buffer, and then the washed first complex was mixed with glutaraldehyde and PBS buffer and reacted at 10-30°C for 1.5-2.5 h. Then catalase was added and reacted at 10-30°C for 12-24 h to obtain the second complex.

8. The preparation method according to claim 7, characterized in that, For each milligram of the first complex, add 0.05~0.15 mL of glutaraldehyde and 0.4~4 mg of catalase.

9. The use of a tumor phototherapy preparation according to any one of claims 1 to 3 in the preparation of a medicament for treating or inhibiting tumors.

10. The application according to claim 9, characterized in that, The tumor is a breast tumor.

Images