Preparation method and application of PAM-Fe (CO) 5 microspheres for magnetic thermal ablation treatment

Abstract

The invention discloses a preparation method and application of PAM-Fe (CO) 5 microspheres for magnetocaloric ablation treatment, a novel magnetocaloric microsphere is prepared through an SPG membrane emulsification method, and magnetocaloric microsphere mediated noninvasive magnetocaloric ablation treatment of HCC is carried out. A research result shows that the PAM-Fe (CO) 5MSs can effectively heat and kill tumor cells under the action of AMF, and meanwhile, the T cell immune response of an organism is activated, so that the anti-tumor effect is remarkably improved. Especially, when the compound is combined with an immune checkpoint inhibitor PD-1mAb for use, the effector function of immune cells is enhanced and the growth of distant metastatic tumors is inhibited through the displayed synergistic effect.

Description

Technical Field

[0001] The present invention belongs to the field of biology and medicine technology, and specifically relates to a preparation method of PAM@Fe(CO)5 microspheres for magnetothermal ablation treatment and an application method thereof. Background Art

[0002] Primary hepatocellular carcinoma (HCC) is one of the most challenging malignancies worldwide, attracting significant attention due to its high metastatic and recurrence rates. Although image-guided percutaneous ablation plays an important role in HCC treatment, it remains overly reliant on operator experience and can lead to tissue damage. Therefore, this study proposes a novel therapeutic strategy: combining hepatic artery embolization with precise embolization of magnetothermal microspheres into the tumor-feeding artery, and achieving noninvasive magnetothermal ablation of liver tumors by applying an alternating magnetic field in vitro. We successfully prepared polyacrylamide carbonyl iron microspheres (PAM@Fe(CO)5Ms) with excellent magnetothermal effect and biosafety using a SPG membrane emulsification method. PAM@Fe(CO)5Ms effectively ablated primary tumors in mice and activated T cell immune responses. Combined with immune checkpoint blockade (ICB) therapy, they enhanced T cell effector function and exhibited significant inhibitory effects on metastatic tumors. This synergistic mechanism was further validated in Batf3 knockout mice. Experimental results in Batf3 knockout mice suggest that suppression of T cell effector function weakens the anti-tumor efficacy of the combination therapy. Finally, large animal studies verified the safety and efficacy of PAM@Fe(CO)5Ms magnetic thermal ablation in a rabbit VX2 orthotopic liver tumor model, completing preclinical evaluation. Our study provides new insights into the clinical treatment of HCC, promoting the transition from traditional minimally invasive ablation to safer noninvasive ablation methods, and holds significant clinical promise.

[0003] Hepatocellular carcinoma (HCC), the most common malignant liver tumor, faces significant challenges in clinical treatment due to its high metastatic and recurrence rates. In recent years, image-guided percutaneous ablation techniques, particularly microwave ablation (MWA) and radiofrequency ablation, have gradually become one of the mainstays of HCC treatment. These techniques achieve tumor eradication by delivering thermal energy directly to tumor tissue, inducing cell death. Although MWA has demonstrated promising clinical results, it still has several limitations. First, the puncture process is highly dependent on the operator's skill level, potentially causing damage to surrounding organs and blood vessels and posing a risk of metastasis through the tumor puncture tract. Second, for larger or multiple tumor lesions, multiple punctures may be required, further increasing the patient's risk of puncture injury and complications. Therefore, finding safer and more effective ablation treatment strategies is of great significance.

[0004] Magnetic Hyperthermia (MHT) is a treatment method that uses a medium to produce a thermal effect under a high-frequency alternating magnetic field (AMF). As an emerging tumor treatment method, MHT has the advantages of good targeting and no restrictions on tissue depth. Although MHT has shown good therapeutic potential in previous studies, current research mainly focuses on the application of nanoscale magnetic particles or macroscopic metals. The accumulation of these nanoparticles in the tumor site is low, which limits their heating capacity, and their biosafety remains to be verified. Although macroscopic metal materials have better magnetothermal heating effects, they require puncture and implantation, and their practical application is somewhat difficult. Summary of the Invention

[0005] The purpose of this section is to summarize some aspects of the embodiments of the present invention and briefly introduce some preferred embodiments. Some simplifications or omissions may be made in this section and the abstract and title of this application to avoid obscuring the purpose of this section, the abstract and the title of the invention, and such simplifications or omissions should not be used to limit the scope of the present invention.

[0006] In view of the above problems and / or the problems existing in the prior art, the present invention is proposed.

[0007] Therefore, the purpose of the present invention is to overcome the deficiencies in the prior art and provide a method for preparing PAM@Fe(CO)5 microspheres for magnetothermal ablation therapy.

[0008] In order to solve the above technical problems, the present invention provides the following technical solution: a method for preparing PAM@Fe(CO)5 microspheres for magnetothermal ablation treatment, characterized by comprising:

[0009] The nano-scale Fe(CO)5 dispersion, polyethylene glycol PEG, polyacrylamide PAM and SDS aqueous solution are fully stirred and mixed, and introduced into an SPG membrane emulsification device for treatment to obtain the PAM@Fe(CO)5 microspheres for magnetic thermal ablation treatment.

[0010] As a preferred embodiment of the preparation method of the present invention, the nano-scale Fe(CO)5 dispersion, polyethylene glycol PEG, polyacrylamide PAM and SDS aqueous solution are fully stirred and mixed, wherein the mass ratio of the nano-scale Fe(CO)5 dispersion, polyethylene glycol PEG and polyacrylamide PAM is 0.25-1.25:0.5:1; and the concentration of the SDS aqueous solution is 1%.

[0011] As a preferred embodiment of the preparation method of the present invention, the stirring and mixing are performed thoroughly, wherein the stirring rate is 300 to 800 rpm, the stirring temperature is 20 to 25° C., and the stirring time is 30 to 120 min.

[0012] As a preferred embodiment of the preparation method of the present invention, the process is carried out by introducing the raw material into an SPG membrane emulsification device, wherein the emulsification pore size of the SPG membrane emulsification device is 20 microns and the pressure is 0.5 MPa.

[0013] As a preferred embodiment of the preparation method of the present invention, the particle size of Fe(CO)5 in the nano-scale Fe(CO)5 dispersion is 5 to 20 nanometers, and the molecular weight of PEG is 1000 Daltons.

[0014] Another object of the present invention is to overcome the shortcomings of the prior art and provide a PAM@Fe(CO)5 microsphere for magnetothermal ablation treatment, characterized in that the diameter of the PAM@Fe(CO)5 microsphere is 40 to 60 microns, and the content of Fe(CO)5 in the PAM@Fe(CO)5 microsphere accounts for 5% to 25% of the total mass of the microsphere.

[0015] Another object of the present invention is to overcome the deficiencies in the prior art and provide an application of PAM@Fe(CO)5 microspheres.

[0016] As a preferred embodiment of the application of the present invention, the PAM@Fe(CO)5 microspheres are used in magnetic thermal therapy.

[0017] As a preferred embodiment of the application of the present invention, the PAM@Fe(CO)5 microspheres are used in local tumor treatment.

[0018] Beneficial effects of the present invention:

[0019] (1) The present invention prepares a new type of magnetothermal microspheres by the SPG membrane emulsification method, and carries out magnetothermal microsphere-mediated non-invasive magnetothermal ablation treatment for HCC. The results show that PAM@Fe(CO)5MSs, under the action of AMF, can effectively heat up and kill tumor cells, while activating the body's T cell immune response, significantly improving the anti-tumor effect. In particular, when used in combination with the immune checkpoint inhibitor PD-1mAb, the synergistic effect is demonstrated, which enhances the effector function of immune cells and inhibits the growth of distant metastatic tumors. In addition, studies in large animal models have shown that MTA avoids the operational risks and tissue damage brought about by traditional percutaneous puncture ablation, and has good safety and effectiveness.

[0020] (2) The therapeutic strategy of this invention is not only applicable to HCC but may also be extended to the treatment of other types of richly vascular solid tumors. This study provides new ideas for the treatment of HCC and promotes the transition from traditional minimally invasive ablation to safer and more efficient non-invasive ablation methods, with important clinical application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for describing the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. Those skilled in the art can also derive other drawings based on these drawings without inventive effort. Among them:

[0022] Figure 1 The figure shows the synthesis and characterization results of PAM@Fe(CO)5MSs with different Fe(CO)5 contents.

[0023] Figure 2 This is a diagram showing the measurement results of Example 2 of the present invention.

[0024] Figure 3 This is a diagram showing the measurement results of Example 3 of the present invention.

[0025] Figure 4 This is a diagram showing the measurement results of Example 4 of the present invention. DETAILED DESCRIPTION

[0026] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, the specific implementation methods of the present invention are described in detail below in conjunction with the embodiments of the specification.

[0027] In the following description, many specific details are set forth to facilitate a full understanding of the present invention. However, the present invention may also be implemented in other ways different from those described herein. Those skilled in the art may make similar generalizations without violating the connotation of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below.

[0028] Secondly, the term "one embodiment" or "embodiment" herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in various places throughout this specification does not necessarily refer to the same embodiment, nor does it refer to a separate or selective embodiment that is mutually exclusive of other embodiments.

[0029] The raw materials used in the examples of the present invention: nano-Fe(CO)5 dispersion, PEG, and sodium dodecyl sulfate (SDS) were purchased from Sigma-Aldrich. Polyacrylamide (PAM) was purchased from Thermo Fisher Scientific.

[0030] Example 1

[0031] This example provides a method for synthesizing PAM@Fe(CO)5MSs using SPG membrane emulsification technology. During the preparation process, the Fe(CO)5 content is adjusted to optimize its magnetocaloric properties. The method specifically includes the following steps:

[0032] Dispersions containing 10 nanometer Fe(CO)₅ were prepared at different concentrations: 5% Fe(CO)₅ content: 0.25 g Fe(CO)₅ dispersed in 5 ml of deionized water; 15% Fe(CO)₅ content: 0.75 g Fe(CO)₅ dispersed in 5 ml of deionized water; 25% Fe(CO)₅ content: 1.25 g Fe(CO)₅ dispersed in 5 ml of deionized water. PEG (1000 Dalton molecular weight) was used in an amount of 0.5 g. Polyacrylamide (PAM) was used in an amount of 1 g. A 10 ml aqueous solution of sodium dodecyl sulfate (SDS) was used, prepared by dissolving 0.1 g SDS in 10 ml of deionized water. All components (Fe(CO)₅ dispersion, PEG, PAM, and SDS solution) were thoroughly mixed in a container to ensure uniform dispersion. The mixture was then introduced into an SPG membrane emulsifier (MG-20, Jiasheng Technology Co., Ltd.). The emulsification pore size was set to 20 μm and the applied pressure was set to 0.5 MPa.

[0033] After the above treatment, PAM@Fe(CO)5MSs with different Fe(CO)5 contents (5%, 15% and 25%) and a diameter of about 50 microns were finally obtained.

[0034] This embodiment uses nano-sized Fe(CO)5 particles ( Figure 1 A) and micron-sized PAM MSs ( Figure 1 B) PAM@Fe(CO)5MSs with a diameter of about 50 μm were prepared by SPG membrane emulsification method ( Figure 1 C and D). The advantage of the SPG membrane emulsification method is that it can produce uniformly sized, monodispersed microdroplets on a large scale and can flexibly adjust the monomer ratio. By rationally selecting the pore size of the hydrophobic membrane and adjusting the nitrogen pressure in the aqueous phase, PAM@Fe(CO)5MSs with different Fe(CO)5 ratios, including 5%, 15%, and 25%, were successfully prepared. The resulting microspheres had regular morphology and uniform distribution ( Figure 1E). However, when the Fe(CO)5 ratio exceeded 30%, PAM@Fe(CO)5MSs encountered obstacles during catheter arterial embolization, leading to catheter blockage, which may be related to the high density of the microspheres. This finding emphasizes the importance of selecting and optimizing the material composition and ratio during microsphere preparation to ensure their effectiveness and safety in clinical applications.

[0035] Example 2

[0036] This example characterizes the PAM@Fe(CO)5MSs with different Fe(CO)5 contents (5%, 15% and 25%) prepared in Example 1, including the following steps:

[0037] The AMF was provided by a high-frequency induction heating device (SPG-10AB-11, Shenzhen Shuangping Electric Power Technology Co., Ltd.), which was equipped with two different heating coils (diameters: 5.0 and 15.0 cm, f appl =100kHz,H appl =10.0kA m -1 ). Thermal imaging photos were obtained by infrared thermal imager (Fotric 225), and data analysis was performed using Analyz IR software. When analyzing the characteristics of PAM@Fe(CO)5MSs, a transmission electron microscope (TEM, JEOL) was used to take microscopic images, and ImageJ software was used to measure the diameters of microspheres in different groups and draw their size distribution diagrams. At the same time, an X-ray diffractometer (XRD, Bruker D8) was used to analyze the composition of the material. A vibrating sample magnetometer (USM) model PPMS-9T was used to analyze the magnetic properties of the prepared magnetic material. The Diamiand thermal analyzer of PE Company in the United States was used to perform thermal analysis on the prepared material. PAM@Fe(CO)5MSs was stored in PBS (pH 7.4, 10mM) or 10% FBS for 24 hours, stored at 4°C for 30 days, and diluted 100 times in PBS to evaluate its stability.

[0038] The PAM@Fe(CO)5MSs with different Fe(CO)5 contents (5%, 15% and 25%) were placed in AF(f appl =100kHz,H appl =10.0kA m -1 ) were tested in a specific AMF intensity. The temperature change of PAM@Fe(CO)5MSs significantly depends on the Fe(CO)5 content. The results show that MSs containing 25% Fe(CO)5 can quickly heat up to 85°C under specific AMF conditions and maintain stable magnetothermal efficiency, which provides theoretical support for its application in local tumor treatment ( Figure 1 FH).

[0039] In terms of characterization of microspheres, the actual content of Fe(CO)5 in the synthesized microspheres was verified by thermogravimetric analysis (TGA) of MSs with different Fe(CO)5 contents. Figure 1 I). The composition of the material was determined by XRD. From the XRD spectrum, it can be clearly observed that the characteristic peaks of PAM at 2θ value of 21° and Fe(CO)5 at 2θ values of 44.8°, 65.0°, and 82.3° correspond to (100), (220), and (111) of α-Fe, respectively (JCPDS No. 06-0696). The above spectral analysis confirmed the successful preparation of PAM@Fe(CO)5MSs ( Figure 1 J).

[0040] From the hysteresis curve, we can see that the remanence and coercive force of PAM@Fe(CO)5MSs are both 0, which indicates that this magnetic material also exhibits superparamagnetism ( Figure 1 K). Nano-Fe(CO)5 and micro-PAM have the properties of ferromagnetic materials that can produce a large saturation magnetization under a small external magnetic field, and retain the use condition of zero remanence after the external magnetic field disappears. Therefore, this magnetic material has the advantages of superparamagnetism and ferromagnetism. In addition, under simulated long-term physiological conditions, PAM@Fe(CO)5MSs showed good stability. After soaking in PBS (pH 7.4, 10mM) or 10% FBS for 24 hours and storing at 4°C for 30 days, the particle size did not change significantly after 100-fold dilution in PBS ( Figure 1 L).

[0041] Example 3

[0042] This example conducted a cell experiment on the PAM@Fe(CO)5MSs with different Fe(CO)5 contents (5%, 15% and 25%) prepared in Example 1, including the following steps:

[0043] H22 mouse hepatocellular carcinoma cells, 4T1 mouse breast cancer cells, L929 mouse fibroblasts, and rabbit VX2 tumor cells were obtained from the cell bank of the Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, and incubated at 5% CO2 and 37°C. The cells were passaged at a ratio of 1:3 after reaching 90% confluence. In the in vitro magnetic hyperthermia study, these cells were co-exposed to AMF with PAM@Fe(CO)5MSs for 5 minutes. The treatment temperature was set at 42°C (f appl =250kHz,H appl =4.5×10 3 Am -1 )、47℃(f appl =250kHz,H appl =5.5×10 3 A m-1 ) and 52℃(f appl =250kHz,H appl =6.5×10 3 A m -1 To investigate the eddy current heating effects of PAM@Fe(CO)5 MSs containing different Fe(CO)5 contents (5%, 15%, and 25%) on cells, the cells were seeded in culture dishes before testing. After reaching a cell density of approximately 80%, they were directly treated with UV-sterilized PAM@Fe(CO)5 MSs under AMF. The medium temperature was monitored using a thermal imager. The culture dish was gently shaken to ensure uniform heating of the cells.

[0044] To assess apoptosis induced by magnetic hyperthermia, cells were treated with Annexin V-FITC and PI staining for 30 minutes. Collected cells were washed with phosphate-buffered saline and analyzed by flow cytometry. Furthermore, the relative viability of cells in the different treatment groups was determined using a standard methylthiazolyl tetrazolium (MTT) assay.

[0045] For in vitro live / dead double staining assay, cancer cells were stained with Calcein AM (to label live cells) and Propidium Iodide (to label dead cells), and fluorescence images were captured using a confocal laser scanning microscope (CLSM, Zeiss Axio-Imager LSM-800).

[0046] In actual in vivo applications, the biocompatibility of PAM@Fe(CO)5MSs is an important consideration. Therefore, this study selected L929 mouse fibroblasts, H22 mouse hepatoma cells, 4T1 mouse breast cancer cells, and VX2 rabbit hepatoma cells to evaluate the biocompatibility of PAM@Fe(CO)5MSs. The standard MTT cytotoxicity assay ( Figure 2 A) was evaluated, and the results showed that none of the four cell lines showed significant cytotoxicity, indicating that the microspheres have good biocompatibility. This result lays the foundation for the in vivo application of PAM@Fe(CO)5MSs, demonstrating that they can exert therapeutic effects without causing significant cytotoxicity.

[0047] To further explore the killing effect of PAM@Fe(CO)5MSs on tumor cells under the action of AMF, H22 tumor cells were exposed to PAM@Fe(CO)5MSs (diameter 50μm) containing 25% Fe(CO)5 and AMF was applied for 5 minutes. By adjusting the intensity of AMF, the temperature can be controlled at about 42°C, 47°C and 52°C. At a treatment temperature of about 47°C, about 85% of the cells were destroyed, while at 52°C, the killing rate of cancer cells reached about 94% ( Figure 2 B).

[0048] In addition, calcein AM (for labeling live cells) and propidium iodide (for labeling dead cells) were used for staining to further confirm the temperature-dependent toxicity of PAM@Fe(CO)5MSs on H22 cells ( Figure 2 C). No obvious red fluorescence signal was observed in cells treated with AMF or PAM@Fe(CO)5MSs alone. In the group treated with AMF and PAM@Fe(CO)5MSs together, cell death became more obvious with increasing temperature. The results of flow cytometry analysis further confirmed that PAM@Fe(CO)5MSs-mediated magnetic hyperthermia therapy significantly induced apoptosis of H22 cells, with an apoptosis rate of 30.1% at 42°C, 51.3% at 47°C, and 80.4% at 52°C ( Figure 2 D and E).

[0049] Example 4

[0050] In this example, an in vivo MTA experiment was conducted on the PAM@Fe(CO)5MSs with different Fe(CO)5 contents (5%, 15% and 25%) prepared in Example 1, comprising the following steps:

[0051] The BALB / c female mice (4-6 weeks old) used in the experiment were provided by Suzhou Sinosai Biotechnology Co., Ltd. Each mouse was inoculated with 2×10 6 H22 cells were used to establish primary tumors, and 0.5 × 10 6 H22 cells were used to simulate distant metastasis. When the primary tumor volume reached 200 mm 3 The mice were randomly divided into four groups (n=5): control group, PD-1 mAb group (200 μg), PAM@Fe(CO)5MSs+AMF group (f appl =250kHz,H appl =10.0kA m -1 ), PAM@Fe(CO)5MSs+AMF+PD-1mAb (200μg). Groups 3 and 4 received PAM@Fe(CO)5MSs intravenously injected into the left tumor and underwent magnetic hyperthermia for complete ablation. Group 4 received 200μg of PD-1mAb intravenously on days 1, 3, and 5 after MTA. Group 2 mice received only three intravenous injections of PD-1mAb.

[0052] For the left primary tumor, the first and second groups of mice were completely removed by surgery, while the third and fourth groups of mice were completely ablated by MTA. The length and width of the right metastatic tumor were measured every three days, and the tumor volume was calculated (volume = length × width 2 / 2). The temperature changes during AMF exposure were monitored using an infrared thermal imager. The survival time of mice in each group was counted, and survival analysis was performed (n = 7). After euthanasia of the mice, the tumors were collected for hematoxylin and eosin staining for histological analysis.

[0053] Thermal ablation therapy has been shown to be effective in stimulating anti-tumor immune responses. However, increased PD-L1 expression in tumor tissue after thermal ablation induces immunosuppression, which creates conditions for thermal ablation combined with immune checkpoint inhibitors to treat tumors [29,30]. Given the superior performance of PAM@Fe(CO)5MSs in MTA, we constructed a bilateral subcutaneous tumor model in mice based on H22 cells and conducted a series of in vivo animal experiments. The research focuses on evaluating the effect of in situ tumor ablation using magnetic thermal therapy, as well as the inhibitory effect of the combined strategy of MTA and PD-1mAb on distant tumors ( Figure 3 A). It is noteworthy that under the action of AMF, the temperature of the in situ tumor injected with PAM@Fe(CO)5MSs rapidly increased to approximately 80°C within about 5 minutes, while the temperature of the group using AMF alone did not change significantly ( Figure 3 B and C).

[0054] In the control group, distant metastases grew rapidly, while the growth rate of distant metastases in the PD-1mAb and MTA groups was significantly slowed down. In the MTA and PD-1mAb combined treatment group, after 12 days of treatment, the volume of distant metastases was significantly smaller than that in the other groups ( Figure 3 D and E). Mice receiving the combined treatment showed the most significant inhibition of metastatic tumors and a significantly prolonged survival time ( Figure 3 F). The results of hematoxylin-eosin (H&E) staining further confirmed that the combined treatment had a good inhibitory effect on H22 distant metastasis ( Figure 3 G). This finding demonstrates the therapeutic potential of MTA in combination with immune checkpoint inhibitors, and future studies could explore the combination of other immunotherapy strategies with MTA.

[0055] Example 5

[0056] This example is a test for the efficacy and safety of MTA in rabbit VX2 orthotopic liver tumors, comprising the following steps:

[0057] MTA is not limited by the penetration depth. After arterial embolization, the magnetic thermal therapy microspheres can be evenly distributed in the tumor tissue. This feature is expected to overcome the limitations of image-guided percutaneous thermal ablation. To verify the feasibility and safety of PAM@Fe(CO)5MSs-mediated MTA in the treatment of liver malignancies, we established a large animal orthotopic liver tumor model in rabbits and conducted preclinical evaluation studies ( Figure 4 A).

[0058] Under ultrasound guidance (Philips), we implanted VX2 tumor tissue into the left lobe of the liver in New Zealand white rabbits. Approximately 12 days later, CT scans confirmed tumor formation. Subsequently, a microcatheter was inserted into the right femoral artery. Digital subtraction angiography (DSA) revealed the hepatic artery and the tumor-feeding artery. The microcatheter was advanced to the tumor-feeding artery, and PAM@Fe(CO)5MSs were slowly injected until the artery was completely embolized.

[0059] To evaluate the therapeutic effect visually, we used 18F-fluorodeoxyglucose (18F-FDG) for positron emission tomography (PET) imaging ( Figure 4 A and B). 18F-FDG PET imaging results showed strong 18F-FDG signals and high standardized uptake values (SUVs) in HCC tissues of tumor-bearing rabbits before treatment. Seven days after treatment, the 18F-FDG signal in the control group increased due to tumor growth, while the active area in the microsphere embolization group decreased relatively. Notably, the MSs+AMF group showed only a weak 18F-FDG signal in the rabbit liver tumors, indicating that this magnetic hyperthermia treatment has a good inhibitory effect on liver tumors.

[0060] To further verify the therapeutic effect of MTA, we performed H&E staining analysis on tumor tissues. The results showed that the degree of tumor tissue necrosis in the MSs+AMF group was the highest compared with the other groups, proving the effectiveness of MTA treatment ( Figure 4 C). In addition, we also evaluated the safety of MTA treatment, and the results showed that the liver function of the MS embolization group and the MSs+AMF group was affected to a certain extent. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) peaked on the 4th day after surgery and returned to normal levels around the 12th day after surgery ( Figure 4 D and E). During the safety evaluation, no toxicity to important organs such as the heart, liver, spleen, lungs, and kidneys was observed ( Figure 4 F), indicating good feasibility and safety of MTA treatment.

[0061] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the spirit and scope of the technical solutions of the present invention, and all of these should be included in the scope of the present invention.

Claims

1. A method for preparing PAM@Fe(CO)5 microspheres for magnetothermal ablation therapy, characterized by: include, The nano-scale Fe(CO)5 dispersion, polyethylene glycol PEG, polyacrylamide PAM and SDS aqueous solution are fully stirred and mixed, and introduced into an SPG membrane emulsification device for treatment to obtain the PAM@Fe(CO)5 microspheres for magnetic thermal ablation treatment.

2. The preparation method according to claim 1, wherein: The nano-scale Fe(CO)5 dispersion, polyethylene glycol PEG, polyacrylamide PAM and SDS aqueous solution are fully stirred and mixed uniformly, wherein the mass ratio of the nano-scale Fe(CO)5 dispersion, polyethylene glycol PEG and polyacrylamide PAM is 0.25-1.25:0.5:1; and the concentration of the SDS aqueous solution is 1%.

3. The preparation method according to claim 2, wherein: The stirring is performed to mix the mixture thoroughly, wherein the stirring rate is 300-800 rpm, the stirring temperature is 20-25° C., and the stirring time is 30-120 min.

4. The preparation method according to claim 1, wherein: The method is to introduce the emulsification material into an SPG membrane emulsification device for treatment, wherein the emulsification pore size of the SPG membrane emulsification device is 20 microns and the pressure is 0.5 MPa.

5. The preparation method according to claim 2, wherein: The particle size of Fe(CO)5 in the nano-scale Fe(CO)5 dispersion is 5 to 20 nanometers, and the molecular weight of PEG is 1000 Daltons.

6. PAM@Fe(CO)5 microspheres prepared by the preparation method according to any one of claims 1 to 5, characterized in that: The diameter of the PAM@Fe(CO)5 microspheres is 40 to 60 microns.

7. The PAM@Fe(CO)5 microspheres according to claim 6, wherein: The content of Fe(CO)5 in the PAM@Fe(CO)5 microspheres accounts for 5% to 25% of the total mass of the microspheres.

8. Use of the PAM@Fe(CO)5 microspheres prepared by the preparation method according to any one of claims 1 to 5.

9. The use according to claim 8, characterized in that: Application of the PAM@Fe(CO)5 microspheres in magnetic hyperthermia therapy.

10. The use according to claim 9, characterized in that: Application of the PAM@Fe(CO)5 microspheres in local tumor treatment.

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