125 I. Nanoparticles, their preparation methods, and applications

Abstract

This invention relates to the fields of brachytherapy and radiosensitization, specifically providing a... 125 I-nano implantable particles, their preparation methods, and applications. 125 The 1-nanometer implanted particles have a core-shell structure, consisting of a particle core and particles adsorbed on the outer surface of the particle core. 125 I and the particles encapsulated in the nucleus and 125 The outer shell of the I-type photosensitizer is a covalent organic framework. This invention... 125 The core of the I-nanometer implanted particle is used to enhance the energy deposition of gamma rays and X-rays, generating more hydroxyl radicals; the shell containing a type I photosensitizer is used to absorb the energy of Auger electrons and internal conversion electrons and excite the photosensitizer to generate singlet oxygen, effectively improving the response to... 125 The efficiency of I decay energy utilization. 125 I-nano implanted particles, combined with internal radiation therapy and radiodynamic therapy, destroy tumor cell DNA and cell membranes, enhance tumor killing and induce immunogenic cell death, thereby activating anti-tumor immunity and inducing remote effects.

Description

Technical Field

[0001] This invention relates to the fields of brachytherapy and radiosensitization technology, specifically to a... 125 I-nano implantable particles, their preparation methods, and applications. Background Technology

[0002] Radiation therapy is a first-line treatment for cancer, benefiting over 50% of cancer patients. Depending on the type of radiation source, radiation therapy can be divided into two types: external beam radiation therapy, which uses external ionizing radiation (X-rays, gamma rays, heavy ion rays, etc.) to kill tumors; and internal beam radiation therapy, which uses radioactive nuclides (…). 177 Lu、 225 Ac、 131 I, 125 Iodine-containing radioactive sources (I-type) decay within the body, releasing ionizing radiation to destroy tumors. Although external beam radiotherapy (primarily X-ray external beam radiotherapy) has matured significantly, minimizing damage to normal tissues remains a challenge. Precise and personalized internal beam radiotherapy represents a new trend in future radiotherapy development. Brachytherapy, a type of internal beam radiotherapy, uses imaging techniques such as CT and ultrasound to guide the precise implantation of a radiation source into the tumor area according to the preoperative plan, utilizing the continuous release of ionizing radiation from the decay of radioactive isotopes to kill the tumor. Unlike external beam radiotherapy, internal beam radiotherapy destroys tumors from within; the ionizing radiation does not pass through normal tissues, resulting in fewer side effects on the body. Traditional iodine seed sources generally refer to... 125 I particle, 125 I-particles are a commonly used interstitial brachytherapy technique in my country, consisting of adsorption... 125 The substrate (such as a silver rod) and the titanium shell are sealed by welding. In clinical practice, 125 I-particles can continuously release X-rays and gamma rays of 27.4 keV to 31.4 keV to kill tumors. Their effective radius in tissue is 1.7 cm, thus enabling the delivery of extremely high radiation doses to the tumor site while avoiding damage to normal tissues. Compared to external beam radiotherapy, radioactive particle implantation overcomes the limitation of re-radiotherapy in increasing the tumor dose, offering advantages such as high local dose, minimal damage to normal tissues, minimal invasiveness, and good patient tolerance. Currently, 125 I-particles are suitable for patients with various types of cancer, especially gynecological tumors and prostate cancer.

[0003] However, while traditional brachytherapy techniques are relatively easy to perform on small, single tumors, as the tumor size and number of lesions increase, the number of needles and implanted particles required for the surgery increases significantly, leading to longer operation times and greater difficulty. This results in a substantial increase in treatment costs and limits the mastery of complex surgeries to a select few highly skilled physicians. Furthermore, as the operation time lengthens, patient tolerance decreases significantly, and weak patients may be unable to tolerate the treatment and lose the opportunity for surgery. Therefore, it is necessary to further optimize the therapeutic efficiency of traditional iodine seed sources, and exploring new, more efficient, and easier-to-operate iodine seed sources has become an urgent clinical need.

[0004] Therefore, this invention is proposed. Summary of the Invention

[0005] The main objective of this invention is to propose a... 125 I-nano implantable particles, their preparation methods, and applications, as described in this invention. 125 The 1-nanometer implanted particles are designed with a core-shell structure for more efficient utilization. 125 The radiant energy released by type I decay. A covalent organic framework containing type I photosensitizers serves as... 125 The outer shell of the 1-nanometer implanted particle is used to absorb the energy of Auger electrons and internal conversion electrons and excite photosensitizer molecules to generate singlet oxygen. 1 O2), effectively improving the 125 The efficiency of I decay energy utilization.

[0006] To achieve the above objectives, the present invention provides the following technical solutions.

[0007] The first aspect of the present invention provides 125 I-nanometer implanted particles, the 125 The 1-nanometer implantable particles have a core-shell structure, comprising an implantable particle core and adsorbed particles on the outer surface of the implantable particle core. 125 I, and the core of the implanted particle and the 125 I. The external covalent organic framework shell; the covalent organic framework shell is a covalent organic framework shell containing a type I photosensitizer.

[0008] In some embodiments of the present invention, the covalent organic framework shell containing a type I photosensitizer includes a porphyrin-containing organic framework shell.

[0009] Preferably, the thickness of the covalent organic framework shell is 10 nm to 50 nm.

[0010] In some embodiments of the present invention, the implanted particle core has a core-shell structure, comprising nanocore particles and a silver shell covering the nanocore particles; 125 I is adsorbed on the outer surface of the silver shell.

[0011] Preferably, the radius of the nanocore particles is 25nm to 50nm, and the thickness of the silver shell is 3nm to 10nm.

[0012] In some embodiments of the present invention, the nanocore particles include at least one of gold nanoparticles and heavy metal particles.

[0013] Preferably, the heavy metal particles are selected from at least one of bismuth sulfide, hafnium oxide, and nano-platinum.

[0014] The second aspect of the present invention provides a solution as described in the first aspect. 125 A method for preparing nanoparticles, the method comprising the following steps:

[0015] Preparation of implanted particle cores;

[0016] The implanted particle core modified with the first surface was subjected to a covalent organic framework synthesis reaction to obtain an implanted particle core@covalent organic framework shell;

[0017] After the implanted particle core@covalent organic framework shell is modified with a second surface, it is placed in a container 125 After standing in solution I, and then centrifuged and washed, the product was obtained. 125 I-nano implanted particles.

[0018] In some embodiments of the present invention, the containing 125 In solution of I 125 I activity ranges from 50 μCi to 500 μCi. 100 μg of implanted particle core@covalent organic framework shell contains 50 μCi to 500 μCi of [unclear - possibly a specific ingredient or substance]. 125 I can be, for example, one of 50μCi, 80μCi, 100μCi, 150μCi, 200μCi, 250μCi, 300μCi, 350μCi, 400μCi, 450μCi, 500μCi or any value that satisfies the above range.

[0019] In some embodiments of the present invention, including 125 The solution of I includes sodium iodide solution.

[0020] In some embodiments of the present invention, the implanted particle kernel includes an Au@Ag kernel.

[0021] Under acidic conditions and inert gas protection, the Au@Ag core modified with the first surface is reacted with 5,10,15,20-tetra(4-aminophenyl)-21H,23H-porphyrin and 2,5-dihydroxyterephthalaldehyde at 120°C in the dark for 24-72 hours to obtain Au@Ag@TAPP-COF.

[0022] In some embodiments of the present invention, the second surface modification uses a solution containing polyethylene glycol; the first surface modification uses a solution containing polyvinylpyrrolidone.

[0023] In some embodiments of the present invention, the preparation of the Au@Ag kernel includes:

[0024] The Au@Ag core particles were prepared by mixing the gold nanoparticles with a solution containing polyethyleneimine and a solution containing ascorbic acid, and then adding a solution containing silver ions until the solution turned orange-red.

[0025] The third aspect of the present invention provides a solution as described in the first aspect. 125 I. Nanoparticles or prepared by the method described in the second aspect. 125 Application of I-nano implantable particles in brachytherapy.

[0026] Compared with the prior art, the present invention achieves the following technical effects:

[0027] 1. In this invention 125 The 1-nanometer implanted particles are designed with a core-shell structure for more efficient utilization. 125 The radiant energy released during type I decay. Organic frameworks containing type I photosensitizers serve as... 125 The outer shell of the 1-nanometer implanted particle is used to absorb the energy of Auger electrons and internal conversion electrons and excite photosensitizer molecules to generate singlet oxygen. 1 O2).

[0028] 2. In this invention 125 I-nano implantable particles, as radiosensitizers, enhance the... 125 Based on the radiotherapy effects of I-particles themselves, traditional 125 I particles cannot be utilized 125 The electron energy (AEs and ICEs) produced by I decay is converted into cytotoxicity. 1 O2, effectively improves the 125 The efficiency of I decay energy utilization.

[0029] 3. In this invention, gold nanoparticles are used as... 125 The core of the I-nanometer implanted particles promotes the deposition of X-ray and gamma-ray energy to enhance the radiolysis of water and generate additional hydroxyl radicals (·OH).

[0030] 4. In this invention 125 I-nano implanted particles, combined with internal radiation therapy and radiodynamic therapy, destroy the DNA and cell membrane of tumor cells, achieving the interaction of radiation with reactive oxygen species (·OH) and... 1The dual tumor-killing effect of O2 greatly enhances the killing of tumor cells; at the same time, the disruption of cell membrane integrity will lead to the release of intracellular mediators, thereby activating anti-tumor immunity and showing a certain systemic therapeutic effect in animal models, thus realizing the induction of remote effects by local treatment and expanding the application of brachytherapy in cancer treatment.

[0031] 5. In this invention 125 The 1-nanometer implantable particles have a diameter of approximately 130 nanometers, allowing them to disperse well in saline solution. This enables them to be delivered to the tumor area via a long needle, allowing for less invasive surgery. The good dispersion also facilitates uniform dose deposition within the tumor area, reducing the required radiation dose and the complexity of the procedure.

[0032] The above description is merely an overview of the technical solution of the present invention. In order to better understand the technical means of the present invention and to implement it in accordance with the contents of the specification, and in order to make the above and other objects, features and advantages of the present invention more apparent and understandable, specific embodiments of the present invention are described below. Attached Figure Description

[0033] Various other advantages and benefits will become apparent to those skilled in the art upon reading the following detailed description of preferred embodiments. The accompanying drawings are for illustrative purposes only and are not intended to limit the invention. In the drawings:

[0034] Figure 1 for 125 I-nano implanted particles ( 125 Flowchart of the preparation process of I-Au@Ag@TAPP;

[0035] Figure 2 for 125 I-nano implanted particles ( 125 Transmission electron microscope images of I-Au@Ag@TAPP;

[0036] Figure 3 A graph showing the yield of hydroxyl radicals under different conditions;

[0037] Figure 4 Plots showing singlet oxygen production under different conditions;

[0038] Figure 5 for 125 I-nano implanted particles ( 125 Uptake of 4T1 cells by I-Au@Ag@TAPP;

[0039] Figure 6 for 125 I-nano implanted particles ( 125Cell-killing power map of I-Au@Ag@TAPP against 4T1 and B16F10 cells;

[0040] Figure 7 for 125 I-nano implanted particles ( 125 Diagram of I-Au@Ag@TAPP-induced cell membrane lipid peroxidation.

[0041] Figure 8 for 125 I-nano implanted particles ( 125 I-Au@Ag@TAPP) induced cell CRT exposure diagram;

[0042] Figure 9 for 125 I-nano implanted particles ( 125 Figure 1; I-Au@Ag@TAPP induces HMGB1 release from cells.

[0043] Figure 10 for 125 I-nano implanted particles ( 125 I-Au@Ag@TAPP) treatment of a mouse bilateral tumor model, combined with immunotherapy to induce remote separation effect. Detailed Implementation

[0044] Hereinafter, embodiments of the present invention will be described with reference to the accompanying drawings. However, it should be understood that these descriptions are merely exemplary and not intended to limit the scope of the invention. Furthermore, descriptions of well-known structures and techniques are omitted in the following description to avoid unnecessarily obscuring the concept of the invention.

[0045] 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 invention pertains; the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the invention; the terms “comprising” and “having”, and any variations thereof, in the specification, claims, and foregoing description of the invention, are intended to cover non-exclusive inclusion.

[0046] In the description of the embodiments of this invention, 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 invention, "multiple" means two or more, unless otherwise explicitly defined.

[0047] 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 the invention. 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.

[0048] In the description of the embodiments of this invention, 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.

[0049] In the description of the embodiments of the present invention, the term "multiple" 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).

[0050] Traditional 125 I-particles have shown good therapeutic effects on single small tumors and are suitable for patients with various types of cancer, especially gynecological tumors and prostate cancer. However, traditional... 125 The I particle is facing many challenges:

[0051] (1) 125 I nuclides decay through electron capture, producing gamma rays, X-rays, Auger electrons (AEs, 12.2 keV), and internal conversion electrons (ICEs, 7.2 keV). The electron energies (AEs and ICEs) account for approximately 32% of the total decay energy (61.4 keV), but these electrons have relatively short ranges (AEs range: 1.5 nm–14.0 μm, ICEs range: 0.7–16 μm) and are difficult to penetrate. 125 The titanium shell of the I-particle (50 μm) therefore, traditional 125 I-particles were not fully utilized 125 The radiation energy produced by I decay.

[0052] (2) Traditional 125 I-particle implantation therapy is a localized treatment modality that only has a therapeutic effect on the implanted tumor area. Clinically, no significant regression of tumor lesions in areas without implanted particles has been observed, and over 90% of cancer-related deaths are caused by cancer metastasis. As tumor size and the number of lesions increase, 125The number of puncture needles and implanted particles required for I-particle implantation surgery increases significantly, leading to longer surgery time, increased surgical difficulty, and limited treatment effectiveness. On the one hand, this results in a substantial increase in treatment costs; on the other hand, highly complex surgeries can only be performed by a few highly skilled physicians, and as the surgery time increases, patient tolerance decreases significantly, with weak patients unable to tolerate the treatment and losing the opportunity for surgery.

[0053] (3) Traditional 125 I-particles are implanted into the tumor area via catheter puncture. The procedure is somewhat invasive and is gradually being replaced by other more advanced treatment modalities.

[0054] Nanoparticles containing high atomic number atoms have been used as radiosensitizers. Compared to biological tissues, high atomic number atoms interact more readily with ionizing radiation, thereby enhancing the deposition of radiation energy in the tumor region and improving the tumor-killing effect of radiotherapy. Hafnium dioxide (HfO2) nanoparticles have received clinical approval and have shown good radiotherapy enhancement effects after local injection into the tumor region. In recent years, nanobrachytherapy using nano-radiosensitizers as radionuclide carriers has attracted researchers' attention. Compared with traditional brachytherapy, nanobrachytherapy improves treatment efficacy while facilitating less invasive surgery. Therefore, further exploration is needed. 125 Highly effective radiosensitizers of type I are expected to optimize their clinical application value. 125 I decay produces X-rays, gamma rays, Auger electrons, and internal conversion electrons. Auger electrons and internal conversion electrons account for approximately 32% of the total decay energy, but these electrons, with their high energy transfer density, are... 125 The absorption of I particles by the titanium wall leads to a waste of radiant energy. Especially... 125 Auger electrons produced by I decay have an average energy of approximately 500 eV, enabling them to deposit a large amount of radiation energy within their nanoscale trajectory (<50 nm), which is well-matched to the size of nanomaterials. Therefore, the rational design of radiosensitizers, utilizing traditional... 125 The energy of electrons, which cannot be utilized by I particles, is used to improve... 125 One of the key factors in the effectiveness of I-particle tumor therapy.

[0055] The phenomenon of tumor shrinkage in lesions far from the irradiated site after radiotherapy to a specific tumor lesion is known as the bystander effect or abscopal effect. This phenomenon is generally believed to be related to the release of tumor-associated antigens at the irradiated site after radiotherapy, reactivating the immune system and breaking immune tolerance. Because its incidence is very low and difficult to predict, it was initially only reported in isolated cases and did not attract much attention. However, in the last 20 years, with continuous advancements in radiotherapy technology, especially after the widespread use of stereotactic body radiation therapy (SBRT), reports of abscopal effects have increased, once again drawing attention from the oncology community. The incidence of abscopal effects is higher in tumors with strong immunogenicity, such as melanoma and renal cell carcinoma. The exploration in melanoma has been particularly successful. In a retrospective study, Grimaldi et al. reported that in 21 patients treated with ipilimumab combined with radiotherapy, the incidence of abscopal effects reached 52%. Compared to patients who did not observe abscopal effects, the median survival of patients who experienced abscopal effect increased from 8.3 months to 22.4 months. Currently, the consensus in the oncology community is that combining radiotherapy and immunotherapy can increase the probability of remote effects.

[0056] Photodynamic therapy (PDT) is a clinically approved, virtually non-invasive cancer treatment. Typically, external light is used to activate photosensitizers (such as porphyrins), generating cytotoxic reactive oxygen species (ROS) (most importantly singlet oxygen). 1 O2 kills cancer cells. In addition... 1 O2 is immunogenic and can induce immunogenic cell death in tumor cells, thereby activating anti-tumor immunity. In numerous animal model studies, photodynamic therapy (PDT) has been shown to induce remote effects. However, due to the rapid attenuation of light in biological tissues and limitations in tissue penetration, the clinical application of PDT is usually limited to surface tissues, such as in the treatment of skin cancer. Achieving PDT for deep tumors remains a long-standing challenge in clinical practice.

[0057] In recent years, radiodynamic therapy (RDT) has gained favor among researchers. This treatment modality utilizes nuclear radiation to excite photosensitizers and generate cytotoxic reactive oxygen species (ROS), combining radiotherapy and photodynamic therapy to enhance the anti-tumor effect of radiotherapy. Nuclear radiation has excellent tissue penetration and is currently widely used in the diagnosis and treatment of cancer. Using radiation as the excitation source for photodynamic therapy can fully retain the high clinical relevance of photodynamic therapy while overcoming the depth limitations of traditional photodynamic therapy, enabling photodynamic therapy for deep tumors. In particular, RDT that generates singlet oxygen can induce immunogenic cell death and activate anti-tumor immunity, achieving significant inhibition of tumor growth at the unirradiated end (distance effect). The generation of singlet oxygen may originate from the Auger electron excitation of photosensitizer molecules (such as porphyrins) in the radiosensitizer.

[0058] In summary, this invention develops a method for brachytherapy. 125 I-nano implanted particles. 125 I decay produces X-rays, gamma rays, Auger electrons, and internal conversion electrons. On the one hand, 125 The nanonucleus of the I-nanometer implanted particles can enhance the deposition of long-range photon energy (X-rays, gamma rays) and promote the radiolysis of water to produce more ·OH; on the other hand... 125 The photosensitizer shell of the I-nano implanted particles can utilize traditional... 125 The Auger electron and internal conversion electron energies, which cannot be utilized by I-particles, excite photosensitizers, producing a wider range of action and greater immunogenicity. 1 O2. 125 I-nano implanted particles combined with brachytherapy and radiodynamic therapy enhance... 125 The therapeutic effects of I-particles on tumors and their potential to activate anti-tumor immunity across the province are expected to expand [the scope of research]. 125 Clinical application prospects of I particles.

[0059] One aspect of the present invention provides a method such as Figure 2 shown 125 I-nanometer implanted particles, the 125 The 1-nanometer implantable particles have a core-shell structure, consisting of an implantable particle core and particles adsorbed on the outer surface of the implantable particle core. 125 I, and the encapsulation in the implanted particle core and 125 The outer covalent organic framework shell of type I. The covalent organic framework shell is a covalent organic framework shell containing a type I photosensitizer.

[0060] In some embodiments of the present invention 125 The 1-nanometer implanted particles are designed with a core-shell structure for more efficient utilization. 125 The radiant energy released during the decay of type I photosensitizers. The covalent organic framework shell containing type I photosensitizers serves as... 125The outer shell of the 1-nanometer implanted particle is used to absorb the energy of Auger electrons and internal conversion electrons and excite photosensitizer molecules to generate singlet oxygen. 1 O2).

[0061] 125 I-nano implantable particles, as radiosensitizers, enhance the... 125 Based on the radiotherapy effects of I-particles themselves, traditional 125 I particles cannot be utilized 125 The electron energy (AEs and ICEs) produced by I decay is converted into cytotoxicity. 1 O2, effectively improves the 125 The efficiency of I decay energy utilization.

[0062] In some embodiments of the present invention, the covalent organic framework shell containing a type I photosensitizer may be a porphyrin-containing organic framework shell.

[0063] In some embodiments of the present invention, a covalent organic framework containing porphyrin is used as... 125 The outer shell of the 1-nanometer implanted particle is used to absorb the energy of Auger electrons and internal conversion electrons and excite porphyrin molecules to produce singlet oxygen. 1 O2).

[0064] In some embodiments of the present invention, the photosensitizers in the covalent organic framework shell containing type I photosensitizers include, but are not limited to, 4,4'-(porphyrin-5,15-diyl)diphenylamine, tetraaldehyde phenylporphyrin, 2,6-diiodoporphyry, dihydroporphyrin, hematoporphyrin derivatives, hematoporphyrin monomethyl ether, bamboo red fungus A, hypericin, curcumin, phthalocyanine, and 5-aminolevulinic acid.

[0065] It should be noted that, in the embodiments of the present invention, a covalent organic framework shell containing type I photosensitizers can also be obtained by means of post-modification and adsorption of photosensitizer molecules. For example, photosensitizer molecules can be introduced into the obtained organic framework shell by means of post-modification and adsorption.

[0066] In some embodiments of the present invention, the thickness of the covalent organic framework shell is 10 nm to 50 nm.

[0067] For example, the thickness of the covalent organic framework shell can be one of 10nm, 20nm, 30nm, 40nm, 50nm or any value within the range mentioned above.

[0068] In some embodiments of the present invention, the implanted particle core has a core-shell structure, which includes nanocore particles and a silver shell covering the outside of the nanocore particles. 125 I is adsorbed on the outer surface of the silver shell.

[0069] In some embodiments of the present invention, the radius of the nanocore particles is 25nm to 50nm, and the thickness of the silver shell is 3nm to 10nm.

[0070] For example, the radius of the nanocore particle can be one of 25nm, 30nm, 35nm, 40nm, 45nm, 50nm or any value that meets the above range.

[0071] For example, the thickness of the silver shell can be one of 3nm, 4nm, 5nm, 6nm, 7nm, 8nm, 9nm, 10nm or any value that meets the above range.

[0072] In some embodiments of the present invention 125 In the I-nano implanted particles, the radius of the nanocore particle, the thickness of the silver shell, and the thickness of the covalent organic framework outer shell are all adjustable. The thickness of the covalent organic framework outer shell will significantly affect the yield of singlet oxygen.

[0073] In some embodiments of the present invention, the nanocore particles include, but are not limited to, gold nanoparticles and heavy metal particles.

[0074] In some embodiments of the present invention, nanocore particles are used as 125 The core of the I-nanometer implanted particles promotes the deposition of X-ray and gamma-ray energy to enhance the radiolysis of water and generate additional hydroxyl radicals (·OH).

[0075] For example, gold nanoparticles as 125 The core of the I-nanometer implanted particles promotes the deposition of X-ray and gamma-ray energy to enhance the radiolysis of water and generate additional hydroxyl radicals (·OH).

[0076] In some embodiments of the present invention, the heavy metal particles include, but are not limited to, bismuth sulfide, hafnium oxide, and platinum nanoparticles.

[0077] Another aspect of the present invention provides the aforementioned 125 Preparation method of I nano-implanted particles.

[0078] Reference Figure 1 As shown, with 125 I-nano implanted particles ( 125 The preparation method is illustrated using I-Au@Ag@TAPP as an example.

[0079] Preparation of gold nanoparticles: Mix 5-10 mL of 1 wt% tetrachloroauric acid aqueous solution, 5-15 mL of gold seed sol, and 1-2 mL of 2 wt% polyethyleneimine (MW: 10000) aqueous solution. The solution color changes from light red to orange-yellow. After stirring at room temperature for 3 min, quickly add 5-10 mL of freshly prepared tetrachloroauric acid aqueous solution (equal to the volume of the tetrachloroauric acid aqueous solution) and 0.03 mol / L hydroquinone aqueous solution while stirring rapidly. Continue stirring for 30 min to obtain a deep red gold nanoparticle sol, which is then stored at 4℃ in the dark for later use.

[0080] Preparation of Au@Ag nanoparticles: 500 mL of gold nanoparticle sol, 10 mL–20 mL of 2 wt% polyethyleneimine (MW: 10000) aqueous solution, and 5 mL of 0.1 mol / L ascorbic acid aqueous solution were mixed and stirred at room temperature. 5 mL–10 mL of 0.01 mol / L silver nitrate solution was slowly added dropwise over 10 min, and the solution color gradually changed from deep red to orange-red. The reaction was continued with stirring for 30 min to obtain an orange-red Au@Ag nanoparticle sol, which was then stored at 4°C in the dark for later use.

[0081] Preparation of Au@Ag@TAPP: 500 mL of Au@Ag nanoparticle sol was collected by centrifugation, washed twice with ethanol and n-butanol, and resuspended in 5 mL–10 mL of n-butanol. 5 mL–10 mL of o-dichlorobenzene (n-butanol: o-dichlorobenzene volume ratio = 1:1) and 100 mg–400 mg of polyvinylpyrrolidone (concentration: 10 mg / mL–20 mg / mL) (MW: 58000) were added, and the mixture was sonicated for 1 h. 5,10,15,20-tetratetra(4-aminophenyl)-21H,23H-porphyrin (TAPP, 3.7 mg–22.5 mg) and 2,5-dihydroxyterephthalaldehyde (DHTA, 2.0 mg–12.3 mg) (TAPP:DHTA mass ratio = 75 / 41) were added to the reaction solution, and the mixture was sonicated for 10 min. Then, 100 μL of acetic acid was added, and the mixture was allowed to stand in the dark for 4 h. Subsequently, 300 μL of acetic acid was added, and the mixture was degassed three times using a freeze-pump-thaw cycle. The tube was then sealed with argon gas, and the reaction was carried out at 120°C in the dark for 24–72 hours. After cooling to room temperature, the precipitate was separated by centrifugation. The precipitate was washed successively with tetrahydrofuran and acetone until the supernatant was colorless. The product was resuspended in ethanol and stored at 4°C in the dark for later use.

[0082] Polyethylene glycol modification of Au@Ag@TAPP: 5 mg Au@Ag@TAPP and 25 mg DSPE-PEG2000 were dispersed in 1 mL of dimethyl sulfoxide (Au@Ag@TAPP:DSPE-PEG2000 mass ratio = 1:5, Au@Ag@TAPP concentration = 5 mg / mL), and sonicated for at least 1 h. The mixture was then slowly added dropwise to 10 mL of vigorously stirred water (water volume 10 times that of dimethyl sulfoxide) over 10 min. After stirring for approximately 30 min, the mixture was dialyzed with deionized water (molecular weight cutoff 10 kDa) to remove residual DSPE-PEG2000 and dimethyl sulfoxide. The product was centrifuged and resuspended in deionized water to obtain polyethylene glycol-modified Au@Ag@TAPP, which was stored at 4°C protected from light for later use.

[0083] It is worth mentioning that DSPE-PEG2000 can be replaced with FITC-DSPE-PEG2000 (partial replacement, such as 20%), and fluorescein-PEG modified Au@Ag@TAPP can be prepared according to a similar procedure.

[0084] preparation 125 I-Au@Ag@TAPP: Take Au@Ag@TAPP (100μL, 1mg / mL) into a 1.5mL centrifuge tube, add Na... 125 I aqueous solution (50 μCi ~ 500 μCi), 6 parallel groups, after standing at room temperature for 5 min, 10 min, 20 min, 30 min, 60 min and 90 min respectively, the precipitate was separated by centrifugation and washed 3 times with deionized water. The labeled product was resuspended in 100 μL of deionized water.

[0085] Labeling rate = Product activity / 100 μCi × 100%.

[0086] Another aspect of the present invention provides the aforementioned 125 I. Nanoparticles or prepared by the aforementioned method 125 Application of I-nano implantable particles in brachytherapy.

[0087] In an embodiment of the present invention, 125 I-nanometer implantable particles are used in brachytherapy to improve the effectiveness of tumor treatment.

[0088] The present invention will be described below with reference to specific embodiments. 125 This paper provides a detailed description of the nanoparticles and their preparation methods.

[0089] Unless otherwise specified, the techniques or conditions described in the examples shall be performed in accordance with the techniques or conditions described in the literature in this field, or in accordance with the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased through legitimate channels. Unless otherwise specified, the experimental methods in the following examples are conventional methods. Unless otherwise specified, the experimental materials used in the following examples are all commercially available products.

[0090] Example 1

[0091] Example 1 presents a method for brachytherapy. 125 I-nano implanted particles ( 125 I-Au@Ag@TAPP can improve the effectiveness of tumor treatment.

[0092] Preparation of gold seeds

[0093] Add 100 mL of deionized water and 1 mL of 1 wt% tetrachloroauric acid aqueous solution to a 250 mL two-necked flask, mix, stir at 1000 rpm, heat to 130 °C, and reflux until vigorous boiling. Add 3 mL of freshly prepared 1% (w / v) sodium citrate aqueous solution (reducing agent). The solution color changes from pale yellow to gray, purple, and dark red in sequence. After the color stabilizes, continue stirring for 30 min to obtain the gold seed sol, which is then stored at 4 °C in the dark for later use.

[0094] Preparation of gold nanoparticles

[0095] In a 1L round-bottom flask, 475mL of deionized water, 10mL of 1wt% tetrachloroauric acid aqueous solution, 15mL of gold seed sol, and 1mL of 2wt% polyethyleneimine (PEI) aqueous solution (weight average molecular weight MW: 10000) were added sequentially. The solution color changed from pale red to orange-yellow. After stirring at room temperature for 3 minutes, 10mL of freshly prepared 0.03mol / L hydroquinone (HQ) aqueous solution was quickly added with rapid stirring. The reaction was continued for 30 minutes to obtain a deep red Au nanoparticle sol, which was then stored at 4℃ in the dark for later use.

[0096] Preparation of Au@Ag nanoparticles

[0097] In a 1L round-bottom flask, add 500mL of Au nanoparticle sol, 20mL of 2wt% polyethyleneimine (MW: 10000) aqueous solution, and 5mL of 0.1mol / L ascorbic acid aqueous solution sequentially. Stir and mix at room temperature. Slowly add 10mL of 0.01mol / L silver nitrate solution dropwise over 10min, causing the solution color to gradually change from deep red to orange-red. Continue stirring for 30min to obtain an orange-red Au@Ag nanoparticle sol, which is then stored at 4℃ in the dark for later use.

[0098] Preparation of Au@Ag@TAPP

[0099] 500 mL of Au@Ag nanoparticle sol was collected by centrifugation, washed twice with ethanol and n-butanol, and resuspended in 5 mL of n-butanol. 5 mL of o-dichlorobenzene and 200 mg of polyvinylpyrrolidone (PVP, MW: 58000) were added, and the mixture was sonicated for 1 h. The reaction solution was transferred to a 25 mL Schlenk flask, and 5,10,15,20-tetratetra(4-aminophenyl)-21H,23H-porphyrin (TAPP, 7.5 mg, 0.011 mmol) and 2,5-dihydroxyterephthalaldehyde (DHTA, 4.1 mg, 0.025 mmol) were added. After sonication for 10 min, 100 μL of acetic acid (AcOH) was added, and the mixture was allowed to stand in the dark for 4 h. Subsequently, 300 μL of acetic acid was added, and the mixture was degassed three times using a freeze-pump-thaw cycle. The flask was then sealed with argon gas and heated at 120 °C in the dark for 72 h. After cooling to room temperature, the precipitate was separated by centrifugation. The precipitate was washed successively with tetrahydrofuran and acetone until the supernatant was colorless. The product was resuspended in 10 mL of ethanol and stored at 4°C in the dark for later use.

[0100] Polyethylene glycol modification of Au@Ag@TAPP

[0101] 5 mg Au@Ag@TAPP and 25 mg distearate phosphatidylethanolamine-polyethylene glycol 2000 (DSPE-PEG2000) were dispersed in 1 mL of dimethyl sulfoxide and sonicated for 1 h. The mixture was then slowly added dropwise to 10 mL of vigorously stirred water over 10 min. After stirring for another 30 min, the mixture was dialyzed with deionized water (molecular weight cutoff of 10 kDa) to remove residual DSPE-PEG2000 and dimethyl sulfoxide. The product was centrifuged and resuspended in 5 mL of deionized water to obtain polyethylene glycol-modified Au@Ag@TAPP, which was stored at 4°C protected from light for later use.

[0102] preparation 125 I-Au@Ag@TAPP

[0103] Take Au@Ag@TAPP (100 μL, 1 mg / mL) into a 1.5 mL centrifuge tube, add Na 125 I aqueous solution (10 μL, 100 μCi) was allowed to stand at room temperature for 60 min, then the precipitate was separated by centrifugation and washed three times with deionized water. The labeled product was then resuspended in 100 μL of deionized water.

[0104] Labeling rate = Product activity / 100 μCi × 100%.

[0105] Example 2

[0106] Example 2 uses the same preparation process as Example 1, except that in the polyethylene glycol modification step, 20wt% DSPE-PEG2000 is replaced with fluorescein polyethylene glycol phospholipid (FITC-DSPE-PEG2000) to obtain fluorescein-polyethylene glycol modified Au@Ag@TAPP.

[0107] The following will demonstrate the experimental results obtained in the embodiments of the present invention. 125 The performance of I-Au@Ag@TAPP was verified.

[0108] one, 125 I-nano implanted particles ( 125 I-Au@Ag@TAPP) produces ·OH and in aqueous solution 1 O2

[0109] 1. Detection of OH: Prepare solutions containing 20 μmol / L APF (hydroxyl radical fluorescent probe, Thermo Fisher) 125 I-Au@Ag@TAPP, TAPP+Na 125 I, Na 125 Solution I (concentration 50 μg / mL) 125 The I activity concentration was 200 μCi / mL. After standing in the dark for 12 h, 24 h, 36 h and 48 h, the sample solution was centrifuged (15000×g, 3 min), the supernatant was collected, and the fluorescence signal intensity (488 nm excitation, 515 nm emission) was measured.

[0110] The results are as follows Figure 3 As shown, 125 I-Au@Ag@TAPP can produce a higher concentration than TAPP+Na. 125 I, Na 125 The solution contains more ·OH.

[0111] 2. 1 O2 detection: Preparations were made containing 20 μmol / L SOSG (singlet oxygen fluorescent probe, Thermo Fisher). 125 I-Au@Ag@TAPP, TAPP+Na 125 I, Na 125 Solution I (concentration 50 μg / mL) 125 The I activity concentration was 200 μCi / mL. After standing in the dark for 1 h, 2 h, 3 h, 4 h and 5 h, the sample solution was centrifuged (15000×g, 3 min), the supernatant was collected, and the fluorescence signal intensity (504 nm excitation, 524 nm emission) was measured respectively.

[0112] The results are as follows Figure 4 As shown,125 I-Au@Ag@TAPP can significantly produce 1 O2, and TAPP+Na 125 I, Na 125 Solution I is almost undetectable. 1 O2.

[0113] two, 125 I-nano implanted particles ( 125 I-Au@Ag@TAPP) generates 1 O2's main energy comes from Auger electrons.

[0114] 125 The density of I-Au@Ag@TAPP is set to 1.0 g / cm³. 3 It is configured with a three-layer structure, consisting of a spherical gold core with a radius of 40nm from the inside out, a silver shell with a thickness of 3nm on the surface, and a TAPP covalent organic framework (COF) with a radius of 25nm on the gold and silver core.

[0115] Using Geant 4.10.7 software, Monte Carlo simulation was performed. 125 Electrons or photons produced by I decay and 125 Interactions between I-Au@Ag@TAPP were investigated. An isotropic particle source generating electrons or photons was uniformly placed 0.1 nm from the surface of a silver shell, with the external space filled with water. Energy deposition at different locations was calculated. The results are shown in Table 1.

[0116] Table 1

[0117]

[0118] The results show that the ionizing radiation energy deposited in the TAPP-COF shell is mainly contributed by Auger electrons, proving that the generation of TAPP is excited. 1 The main energy source of O2 comes from 125 Auger electrons released during I decay.

[0119] three, 125 I-nano implanted particles ( 125 Cellular uptake and cytotoxicity of I-Au@Ag@TAPP

[0120] 1. Cell uptake: 4T1 cells were seeded in 24-well plates (5 × 10⁴ cells per well) and cultured for 12 h. The culture medium was then discarded, and medium containing 100 μg / mL Au@Ag@TAPP was added. After incubation for 1 h, 2 h, 4 h, and 8 h, the cells were washed three times with phosphate-buffered saline (PBS). Trypsin solution was added to separate the cells from the culture dish, and the cell suspension was collected. The supernatant was discarded by centrifugation. The cells were then digested with nitric acid (60 °C, 24 h), diluted with deionized water to 5 mL, and the Au content was determined by inductively coupled plasma mass spectrometry.

[0121] The results are as follows Figure 5 As shown, the uptake of Au@Ag@TAPP by 4T1 cells increased over time.

[0122] 2. Cytotoxicity: 4T1 and B16F10 cells were seeded in 96-well plates (1 × 10⁶ cells per well). 4 After culturing for 12 hours, the culture medium was discarded, and 50 μg / mL of different specific activities were added. 125 Cell viability was assessed using I-Au@Ag@TAPP (0 μCi / mL, 50 μCi / mL, 100 μCi / mL, 150 μCi / mL, 200 μCi / mL) culture medium after 48 h of incubation. The absorbance at 450 nm was measured using the standard CCK8 assay. Untreated cell populations were used as a reference for establishing 100% cell viability.

[0123] The results are as follows Figure 6 As shown, with 125 An increase in I activity concentration, 125 The killing effect of I-Au@Ag@TAPP on 4T1 cells gradually increased.

[0124] Four, 125 I-nano implanted particles ( 125 I-Au@Ag@TAPP induces immunogenic cell death

[0125] 1. Cell membrane lipid peroxidation: 4T1 cells were seeded in 35 mm confocal dishes (2 × 10⁵ / mL), cultured for 12 h, and then the culture medium was discarded. A solution containing Au@Ag@TAPP and Na₂O was added. 125 I, 125 I-Au@Ag@TAPP (material concentration is 50ug / mL) 125Cells were cultured in a medium with an activity concentration of 100 μCi / mL for 24 h, then the medium was discarded, and the cells were washed three times with PBS. Subsequently, medium containing a 5 μmol / L lipid peroxidation sensor (BODIPY 581 / 591C11) was added, and after 1 h of culture, the cells were washed three times with PBS and photographed using a confocal fluorescence microscope. The fluorescence intensity at 591 nm and 510 nm was quantitatively analyzed using ImageJ software.

[0126] The results are as follows Figure 7 As shown, 125 Cells treated with I-Au@Ag@TAPP showed significant cell membrane lipid peroxidation compared to other control groups.

[0127] 2. Calreticulin (CRT) exposure: 4T1 cells were seeded in 35 mm confocal dishes (2 × 10⁵ / mL), cultured for 12 h, and the culture medium was discarded. Then, a solution containing Au@Ag@TAPP and Na₂SO₄ was added. 125 I, 125 Cells were cultured in I-Au@Ag@TAPP medium (material concentration 50ug / mL, 125I activity concentration 100μCi / mL) for 24 h, then the medium was discarded, and the cells were washed three times with PBS. Cells were then blocked in PBS containing 1% BSA at 4°C for 1 h, and the supernatant was discarded. Alexa was added. 488-CRT antibody (diluted 500-fold in PBS containing 1% BSA) was incubated at 4°C for 12 h. After washing three times with PBS, cell nuclei were stained with DAPI, mounted with anti-fluorescence quencher, and photographed using a confocal fluorescence microscope. Fluorescence intensity was quantitatively analyzed using ImageJ software.

[0128] The results are as follows Figure 8 As shown, 125 Cells treated with I-Au@Ag@TAPP showed significant CRT exposure compared to other control groups.

[0129] 3. Release of high-mobility group box 1 (HMGB1): 4T1 cells were seeded in 96-well plates (1×10⁴ cells per well), cultured for 12 h, and then the culture medium was discarded. A solution containing Au@Ag@TAPP and Na₂SO₄ was added. 125 I, 125 I-Au@Ag@TAPP (material concentration is 50ug / mL) 125 After culturing in a culture medium with an activity concentration of 100 μCi / mL for 24 h, the concentration of HMGB1 in the cell supernatant was measured according to the ELISA kit instructions.

[0130] The results are as follows Figure 9 As shown, 125Cells treated with I-Au@Ag@TAPP showed significant release compared to other control groups.

[0131] Auger electrons can be used to excite photosensitizers to produce singlet oxygen. 125 I-nano implanted particles will traditional 125 The electron energy (mainly Auger electrons) that the implanted particles cannot utilize is converted into cytotoxic and immunogenic singlet oxygen, which enhances cell killing ability and induces immunogenic cell death.

[0132] five, 125 I-nano implanted particles ( 125 I-Au@Ag@TAPP induces a distant effect in a mouse bilateral tumor model.

[0133] 1. Establishment of a mouse tumor model: Female BALB / c mice (6-8 weeks old) were subcutaneously injected with a PBS suspension containing 1 × 10⁶ 4T1 cells (primary tumor) in the right shoulder, and 5 days later, a PBS suspension containing 5 × 10⁵ 4T1 cells (distal tumor) was subcutaneously injected into the left shoulder. The length (L) and width (W) of the tumor were measured using digital calipers. The tumor volume (V) was calculated using the formula V = 1 / 2 × L × W².

[0134] 2. Treatment trial: When the volume of the primary tumor reaches ~50mm 3 Mice were randomly divided into 3 groups (n=10), and injected in situ with PBS (40μL), respectively. 125 I-Au@Ag@TAPP-PEG, Au@Ag@TAPP (50mg / kg, 1mCi), and in situ implantation 125 I-particles (0.5mCi×2) were administered to 5 mice in each group after treatment. Anti-PD-L1 antibody (75μg / mouse) was injected intraperitoneally every 3 days. Tumor volume and mouse weight were recorded every two days.

[0135] The results are as follows Figure 10 As shown, 125 I-nanoplastin implants are far more effective in treating tumors than traditional methods. 125 I-implanted particles, combined with immunotherapy, activate systemic anti-tumor immunity, exhibiting significant distal effects, while traditional... 125 I-implanted particles have almost no far-end effects.

[0136] Combined immunotherapy can improve 125 The anti-tumor immune effect of I-nano implanted particles, combined with PD-L1 immune checkpoint blockade in this invention, may have similar effects when using other immunotherapies.

[0137] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features therein. These modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention, and they should all be covered within the scope of the claims and specification of the present invention. In particular, as long as there is no structural conflict, the various technical features mentioned in the embodiments can be combined in any way. The present invention is not limited to the specific embodiments disclosed herein, but includes all technical solutions falling within the scope of the claims.

Claims

1. A kind 125 I-nanometer implantable particles, characterized in that... The 125 The 1-nanometer implantable particles have a core-shell structure, comprising an implantable particle core and adsorbed particles on the outer surface of the implantable particle core. 125 I, and the core of the implanted particle and the 125 I. An external covalent organic framework shell; the covalent organic framework shell is a covalent organic framework shell containing a type I photosensitizer; The implanted particle core has a core-shell structure, comprising nano-core particles and a silver shell covering the nano-core particles; 125 I is adsorbed on the outer surface of the silver shell; the nano-core particles are gold nanoparticles. The type I photosensitizer is 5,10,15,20-tetra(4-aminophenyl)-21H,23H-porphyrin.

2. As described in claim 1 125 I-nanometer implantable particles, characterized in that... The thickness of the covalent organic framework shell is 10 nm to 50 nm.

3. As described in claim 1 125 I-nanometer implantable particles, characterized in that... The radius of the nanocore particles is 25 nm to 50 nm, and the thickness of the silver shell is 3 nm to 10 nm.

4. The method described in any one of claims 1-3 125 The method for preparing 1-nanometer implantable particles is characterized by, Includes the following steps: Preparation of implanted particle cores; The implanted particle core, after undergoing the first surface modification, was subjected to a covalent organic framework synthesis reaction to obtain an implanted particle core@covalent organic framework shell; the first surface modification used a solution containing polyvinylpyrrolidone; After the implanted particle core@covalent organic framework shell is modified with a second surface, it is placed in a container 125 After standing in solution I, and then centrifuged and washed, the product was obtained. 125 I-nanometer implanted particles; The second surface modification uses a solution containing polyethylene glycol.

5. The preparation method according to claim 4, characterized in that, The containing 125 In solution of I 125 I activity ranges from 50 μCi to 500 μCi.

6. The preparation method according to claim 4, characterized in that, The implanted particle kernel includes an Au@Ag kernel. Under acidic conditions and inert gas protection, the Au@Ag core modified with the first surface was reacted with 5,10,15,20-tetra(4-aminophenyl)-21H,23H-porphyrin and 2,5-dihydroxyterephthalaldehyde at 120°C in the dark for 24 h to 72 h to obtain Au@Ag@TAPP-COF.

7. The preparation method according to claim 6, characterized in that, The preparation of the Au@Ag kernel includes: The Au@Ag core particles were prepared by mixing the gold nanoparticles with a solution containing polyethyleneimine and a solution containing ascorbic acid, and then adding a solution containing silver ions until the solution turned orange-red.

Images