Low exothermic stimulus-responsive injectable bone cement and its preparation method and use

Abstract

The invention discloses a low exothermic stimulus-responsive injectable bone cement and its preparation method and use, the method comprising the following steps: double-bond-modified hydroxyapatite and camptothecin prodrug are grafted by free radical polymerization to prepare camptothecin prodrug-modified hydroxyapatite; hydroxyapatite and curcumin molecule are connected by oxalate bond to prepare curcumin-modified hydroxyapatite; using polymethyl methacrylate prepolymer solution as liquid phase, a mixture of camptothecin prodrug-modified hydroxyapatite and curcumin-modified hydroxyapatite as solid phase, after adding dibenzoyl peroxide and N, N-dimethylaniline, fully stirred and dispersed, solidified to obtain bone cement. The present invention promotes the treatment of metastatic tumors and the repair of bone by synergistic action of two drug molecules and bone cement.

Description

Technical Field

[0001] The present invention relates to orthopedic medical materials, and in particular to a low-exothermic stimulus-responsive injectable bone cement, a preparation method thereof, and applications thereof. Background Art

[0002] The incidence of metastatic tumors has continued to increase in recent years. The incidence of metastases, formed by primary malignant tumors in various locations spreading to areas with rich blood supply, has been rising year by year. These primary malignant tumors primarily occur in tissues such as the thyroid, breast, kidney, and liver, with metastases most commonly occurring in the thoracic and lumbar vertebrae. The incidence of spinal metastases in patients with malignant tumors is as high as 10% to 15%, with most cases occurring in middle-aged and elderly individuals aged 50 to 60. When tumors metastasize to the vertebral body, bone corrosion and osteolytic destruction can reduce bone structural stability, leading to pain and limited mobility in the corresponding vertebral segments. Some patients with severe disease may also develop pathological fractures, severely impacting their health and quality of life. Traditional conservative treatments are ineffective in relieving pain and are not conducive to restoring spinal stability. Bone cement vertebroplasty is currently the primary treatment option for these patients and has a positive effect on controlling localized tumors.

[0003] Polymethyl methacrylate (PMMA) exhibits excellent plasticity and mechanical properties and is currently the most commonly used bone filling material in clinical practice. It has also been widely used in the treatment of metastatic tumors. Hydroxyapatite (HA), the primary inorganic component of human and animal bone, has osteogenesis-inducing properties, excellent osteoconductivity, and biocompatibility. It can also promote cell-surface interactions through selective protein adsorption, garnering significant attention in bone tissue engineering applications such as prosthetic implants and artificial bone cement. Nanohydroxyapatite (n-HA) exhibits significantly improved solubility and bioactivity compared to standard hydroxyapatite, resulting in superior therapeutic efficacy in practical applications.

[0004] However, single PMMA bone cement has many defects in clinical use in the treatment of metastatic tumors. The curing temperature of bone cement is generally above 70°C, which can cause necrosis and embolism of adjacent soft tissues, especially in cancer patients with poor treatment effects. In addition, polymethyl methacrylate bone cement has poor bioactivity and is difficult to fit closely with human tissue. Although nanohydroxyapatite has good biocompatibility, it is brittle when used alone as the main body, resulting in insufficient mechanical strength and poor processing performance, which greatly limits its application. When the inorganic filler hydroxyapatite is compounded with the organic polymer methyl methacrylate, the poor interfacial bonding between the two causes a decrease in its mechanical properties, resulting in poor load-bearing performance of the material.

[0005] Furthermore, chemotherapy drugs such as camptothecin (CPT) and curcumin (Cur) dispersed in bone cement have been used to treat metastatic tumors. However, the release characteristics of most drugs are characterized by a burst release within hours or days after surgery, resulting in a short-term effect and lack of long-term anti-tumor efficacy. As a foreign implant, bone cement carries the risk of bacterial infection after injection into the human body. Summary of the Invention

[0006] The purpose of the present invention is to provide a low-exothermic stimulus-responsive injectable bone cement, a preparation method thereof, and a use thereof, which promotes the treatment of metastatic tumors and bone repair through the synergistic effect of two drug molecules and bone cement.

[0007] In one aspect of the present invention, the present invention provides a method for preparing a low exothermic stimulus-responsive injectable bone cement. According to an embodiment of the present invention, the method comprises the following steps:

[0008] (1) Double-bond-modified hydroxyapatite was grafted with camptothecin prodrug via free radical polymerization to prepare camptothecin prodrug-modified hydroxyapatite;

[0009] (2) Curcumin-modified hydroxyapatite was prepared by linking hydroxyapatite and curcumin molecules via oxalate bonds;

[0010] (3) Using a polymethyl methacrylate prepolymer solution as the liquid phase and a mixture of camptothecin prodrug-modified hydroxyapatite and curcumin-modified hydroxyapatite as the solid phase, dibenzoyl peroxide and N,N-dimethylaniline are added, and the mixture is fully stirred and dispersed, and solidified to obtain bone cement.

[0011] In addition, the method for preparing a low-exothermic stimulus-responsive injectable bone cement according to the above embodiment of the present invention may also have the following additional technical features:

[0012] In some embodiments of the present invention, in step (1): the preparation method of double-bond modified hydroxyapatite comprises the following steps: dissolving hydroxyapatite, hexamethylene diisocyanate and dibutyltin dilaurate in anhydrous N'N-dimethylformamide, heating for a certain time under condensation reflux and nitrogen protection conditions, dissolving hydroxyethyl methacrylate in N'N-dimethylformamide, injecting into the above system, stirring overnight and then centrifuging, washing with dichloromethane multiple times and collecting powder, fully drying, and grinding with a mortar to obtain double-bond modified hydroxyapatite, whose structural formula is:

[0013]

[0014] The synthesis route of the double bond modified hydroxyapatite HA-CH=CH2 is:

[0015]

[0016] The preparation method of a camptothecin prodrug comprises the following steps: dissolving camptothecin and 4-dimethylaminopyridine in anhydrous dichloromethane, injecting triphosgene under a nitrogen atmosphere, stirring at room temperature, adding 2-((2-hydroxyethyl)disulfonyl)ethyl methacrylate (HEMA) dropwise, stirring the reaction mixture overnight, filtering and evaporating the solvent, diluting the crude product with ethyl acetate, washing with water and brine several times, collecting the organic layer and drying it over anhydrous sodium sulfate, and separating and purifying it by silica gel column chromatography using ethyl acetate as an eluent to obtain the camptothecin prodrug, which has the structural formula:

[0017]

[0018] The synthetic route of camptothecin prodrug is:

[0019]

[0020] In some embodiments of the present invention, during the preparation of the double-bond modified hydroxyapatite, the molar ratio of hydroxyapatite, hexamethylene diisocyanate, and hydroxyethyl methacrylate is 1:1-3:3-10, the reaction temperature is 45-60°C, the heating time is 6-15 hours under condensation reflux and nitrogen protection conditions, the drying temperature is 35-50°C, and the drying time is 12-36 hours.

[0021] During the preparation of the camptothecin prodrug, the molar ratio of camptothecin, 4-dimethylaminopyridine triphosgene and HAMA is 1:2-5:0.25-3:1-3, and the number of washing times with water and brine is 1-3.

[0022] In some embodiments of the present invention, the preparation method of 2-((2-hydroxyethyl)disulfonyl)ethyl methacrylate (HEMA) comprises the following steps: dissolving bis(2-hydroxyethyl)disulfide and triethylamine in anhydrous tetrahydrofuran, adding a certain amount of methacryloyl chloride dropwise in an ice bath under nitrogen protection, transferring to room temperature and stirring overnight after the addition is complete, filtering and evaporating the solvent, diluting the residue with ethyl acetate, washing with water and brine several times in sequence, collecting the organic layer and drying it with anhydrous Na2SO4, and separating and purifying it by silica gel column chromatography using a mixed solvent of ethyl acetate and petroleum ether as an eluent to obtain the 2-((2-hydroxyethyl)disulfonyl)ethyl methacrylate (HEMA), whose structural formula is:

[0023]

[0024] The synthetic route of described HEMA is:

[0025]

[0026] In some embodiments of the present invention, the molar ratio of bis(2-hydroxyethyl) disulfide, methacryloyl chloride and triethylamine is 1:1:1.5, the washing times with water and brine are 2-3 times respectively, the volume ratio of eluent petroleum ether and ethyl acetate is 2:1, and the ice bath temperature is minus 10-minus 20°C.

[0027] In some embodiments of the present invention, the step (1) specifically includes the following steps: dissolving the double-bond modified hydroxyapatite, camptothecin prodrug and azobisisobutyronitrile in anhydrous N'N-dimethylformamide for packaging, degassing under nitrogen three times, flame sealing under vacuum, heating in an oil bath for reaction, and then cooling to terminate the reaction, washing with dichloromethane multiple times and collecting the powder, drying and crushing to obtain the camptothecin prodrug-modified hydroxyapatite, whose structural formula is:

[0028]

[0029] The synthesis route of the camptothecin prodrug modified hydroxyapatite is:

[0030]

[0031] The step (2) specifically comprises the following steps: dissolving hydroxyapatite and curcumin in anhydrous tetrahydrofuran, adding triethylamine dropwise, injecting oxalyl chloride in an ice bath under nitrogen protection, stirring in an ice bath, transferring to room temperature and stirring overnight, filtering the suspension, washing with dichloromethane until the supernatant is nearly colorless, collecting the precipitate, drying and crushing to obtain the curcumin-modified hydroxyapatite, whose structural formula is as follows:

[0032]

[0033] The synthetic route of curcumin-modified nanohydroxyapatite HA-Cur is:

[0034]

[0035] In the step (3), the method for preparing the polymethyl methacrylate prepolymer solution comprises the following steps: heating the methyl methacrylate solution containing the dissolved initiator in a water bath for a certain period of time, and then cooling the solution to terminate the reaction.

[0036] In some embodiments of the present invention, in step (1), the molar ratio of double-bond modified hydroxyapatite, camptothecin prodrug and azobisisobutyronitrile is 1:2-5:0.1-0.5, the oil bath heating reaction temperature is 65-85°C, the reaction time is 24h-48h, the drying temperature is 50-70°C, the drying time is 12-36h, and the freezing temperature is minus 80-minus 50°C;

[0037] In the step (2), the molar ratio of hydroxyapatite, curcumin, triethylamine and oxalyl chloride is 1:1-3:4.5:2-3, the stirring time in an ice bath at -20--10°C is 30 minutes, and the reaction is carried out in the dark. The reaction temperature is 25-45°C, the reaction time is 24-72 hours, the drying temperature is 30-40°C, and the drying time is 12-24 hours.

[0038] In the step (3), during the preparation of the polymethyl methacrylate prepolymer solution, the initiator is azobisisobutyronitrile or dibenzoyl peroxide, the mass ratio of the initiator to the methyl methacrylate monomer is 0.5%-2%, the water bath heating temperature is 70-90°C, the water bath heating time is 15-30 minutes, the cooling temperature is 5-10°C, and the cooling time is 5-15 minutes.

[0039] In some embodiments of the present invention, the mass ratio of the liquid phase to the solid phase is 1:0.05-0.5, the mass ratio of curcumin-modified hydroxyapatite and camptothecin prodrug-modified hydroxyapatite is 1:1, the mass of dibenzoyl peroxide is 0.2%-1.5% of the mass of the polymethyl methacrylate prepolymer solution, the mass of N,N-dimethylaniline is 1%-3% of the mass of the polymethyl methacrylate prepolymer solution, the curing temperature is 36-37°C, and the curing time is 15-60 min.

[0040] In another aspect of the present invention, the present invention provides a low exothermic stimulus-responsive injectable bone cement prepared according to the preparation method of the low exothermic stimulus-responsive injectable bone cement.

[0041] In another aspect of the present invention, the present invention proposes the use of the low-exothermic stimulus-responsive injectable bone cement in the preparation of implant materials for treating metastatic tumors.

[0042] Compared with the prior art, the present invention has the following beneficial effects:

[0043] 1) In this invention, a hydroxyapatite powder grafted with the drug molecules curcumin and camptothecin was prepared as a solid phase, mixed with a liquid polymethyl methacrylate prepolymer solution, and thoroughly dispersed. Simultaneously, a certain amount of dibenzoyl peroxide (BPO) oxidizing agent and N,N-dimethylaniline (DMA) reducing agent were added, resulting in an injectable bone cement with low exothermicity and tumor microenvironment-responsive release of drug molecules. This cement exhibits both anti-tumor and antibacterial and anti-inflammatory effects, and each reaction step is safe, reliable, simple to operate, and exhibits high conversion rates.

[0044] 1) Double-bond-modified hydroxyapatite was grafted with camptothecin prodrug (CPTM) via free radical polymerization to prepare HA-CPT. The drug molecule and hydroxyapatite were covalently bound via disulfide and oxalate bonds. Under the induction of the excessive glutathione microenvironment at the tumor site, the disulfide bond cleaved, releasing the camptothecin drug molecule. HA-Cur was prepared by linking hydroxyapatite and curcumin molecules via oxalate bonds. The excessive hydrogen peroxide microenvironment at the tumor site induces the cleavage of the oxalate bond, releasing the curcumin drug molecule. At the same time, the connection of the two drug molecules enhances the interfacial binding between hydroxyapatite and the polymer, improving its dispersibility in the polymer. Secondly, the two drug molecules (camptothecin prodrug and curcumin) and bone cement work synergistically to promote the treatment of metastatic tumors and bone repair.

[0045] 2) The present invention establishes a free radical polymerization system of BPO and DMA, which lowers the polymerization temperature while reducing heat release, and synergistically acts with two modified hydroxyapatites as heat dissipators to reduce the maximum polymerization temperature to below 44°C (i.e., the heat released during the polymerization process is below 44°C), thereby reducing the risk of tissue damage caused by large amounts of heat release.

[0046] 3) Dispersing drug-modified hydroxyapatite as a filler in polymethyl methacrylate bone cement not only improves the cement's bioactivity but also slowly releases the two drug molecules by inducing chemical bond breakage within the tumor microenvironment, extending the duration of drug action and effectively reducing dosing frequency for long-term anti-tumor efficacy. Furthermore, curcumin also possesses antibacterial and anti-inflammatory properties, effectively reducing the risk of bacterial infection after bone cement implantation.

[0047] 4) The bone cement prepared by the present invention has good biocompatibility, injectability and drug release properties, and has considerable application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 Schematic diagram of the preparation process of a low-exothermic stimulus-responsive injectable bone cement according to an embodiment of the present invention;

[0049] Figure 2 FT-IR spectra of hydroxyapatite (HA), curcumin (Cur), and curcumin-modified hydroxyapatite (HA-Cur) in Example 1 of the present invention;

[0050] Figure 3 is the hydrogen nuclear magnetic resonance spectrum of HEMA in Example 1 of the present invention;

[0051] Figure 4 is the H NMR spectrum of the camptothecin prodrug (CPTM) in Example 1 of the present invention;

[0052] Figure 5FT-IR spectra of hydroxyapatite (HA), isocyanate-modified hydroxyapatite (HA-NCO), and double-bond-modified hydroxyapatite (HA-CH=CH2) in Example 1 of the present invention;

[0053] Figure 6 FT-IR spectra of double bond-modified hydroxyapatite (HA-CH=CH2), camptothecin prodrug (CPTM), and camptothecin prodrug-modified hydroxyapatite (HA-CPT) in Example 1 of the present invention;

[0054] Figure 7 1 is the thermogravimetric diagram of hydroxyapatite (HA) and curcumin-modified hydroxyapatite (HA-Cur) in Example 1 of the present invention;

[0055] Figure 8 The temperature-time diagrams of the four groups of low-exothermic stimulus-responsive injectable bone cements in Examples 1-4 of the present invention are shown;

[0056] Figure 9 Graphs showing the setting time of four groups of low exothermic stimulus-responsive injectable bone cements in Examples 1-4 of the present invention;

[0057] Figure 10 Graphs showing the compressive strength of four groups of low exothermic stimulus-responsive injectable bone cements in Examples 1-4 of the present invention;

[0058] Figure 11 , (a) is a scanning electron micrograph of the low exothermic stimulus-responsive injectable bone cement in Example 2 of the present invention, (b) is a scanning electron micrograph of the low exothermic stimulus-responsive injectable bone cement in Example 4 of the present invention, (c) is a scanning electron micrograph of the low exothermic stimulus-responsive injectable bone cement in Example 2 of the present invention after being immersed in human simulated body fluid SBF for 10 days, and (d) is a scanning electron micrograph of the low exothermic stimulus-responsive injectable bone cement in Example 4 of the present invention after being immersed in human simulated body fluid SBF for 10 days;

[0059] Figure 12 Graph showing the cumulative release rate of curcumin from the four groups of low-exothermic stimulus-responsive injectable bone cements in Examples 1-4 of the present invention in a simulated tumor H2O2 microenvironment;

[0060] Figure 13 This is a graph showing the antioxidant properties of four groups of low exothermic stimulus-responsive injectable bone cements in Examples 1-4 of the present invention;

[0061] Figure 14 Graphs showing the cumulative release rates of camptothecin from the four groups of low-exothermic stimulus-responsive injectable bone cements in Examples 1-4 of the present invention in a microenvironment simulating excess glutathione in a tumor;

[0062] Figure 15Graphs showing the injectability of four groups of low exothermic stimulus-responsive injectable bone cements in Examples 1-4 of the present invention. DETAILED DESCRIPTION

[0063] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. All other embodiments obtained by ordinary technicians in this field based on the embodiments of the present invention without making any creative efforts shall fall within the scope of protection of the present invention.

[0064] Example 1

[0065] A method for preparing a low-exothermic stimulus-responsive injectable bone cement comprises the following steps:

[0066] (1) Synthesis of curcumin-modified hydroxyapatite (HA-Cur): 1 g of nanohydroxyapatite (HA) and 0.73 g of curcumin (Cur) were dissolved in 20 mL of anhydrous tetrahydrofuran (THF). 0.9 g of triethylamine (TEA) was added dropwise. The mixture was stirred in an ice bath at -15°C under nitrogen for approximately 15 min. 0.75 g of oxalyl chloride was then added by injection. The mixture was stirred for 30 min and then transferred to room temperature and stirred overnight. The suspension was filtered and washed with dichloromethane until the supernatant was almost colorless. The precipitate was collected and dried in a vacuum at 40°C for 24 h. The mixture was ground in a mortar to obtain an orange-red solid powder, HA-Cur.

[0067] like Figure 2 As shown, at 1760cm -1 The newly appeared carbonyl absorption peak at 37° proved the successful synthesis of HA-Cur.

[0068] like Figure 7 As shown, the grafting rate of curcumin was calculated to be 46%.

[0069] (2) Synthesis of double-bond modified hydroxyapatite (HA-CH=CH2): 1 g of hydroxyapatite (HA), 1 g of hexamethylene diisocyanate (HDI), and 2 drops of dibutyltin dilaurate (DBTDL) were dissolved in 10 mL of anhydrous N'N-dimethylformamide (DMF) and reacted in an oil bath at 50°C for 8 h under condensation reflux and nitrogen protection to obtain isocyanate-modified hydroxyapatite (HA-NCO). 1.30 g of hydroxyethyl methacrylate (HEMA) was dissolved in 2 mL of DMF and slowly injected into the above reaction system. After stirring overnight, the mixture was centrifuged and the particles were separated several times using a dichloromethane centrifugation-redispersion cycle. The collected powder was placed in a vacuum oven at 35°C and dried for 24 h. The powder was then ground in a mortar to obtain a white powder, HA-CH=CH2.

[0070] like Figure 5 As shown, by 2273cm -1 The absorption peak of -NCO at 1680 cm-1 proved the successful synthesis of HA-NCO. -1 The absorption peak of carbon-carbon double bond at 1729 cm -1 The appearance of the absorption peak of the carbonyl group and the disappearance of the absorption peak of the isocyanate group proved the successful synthesis of HA-CH=CH2.

[0071] (3) Synthesis of HEMA: Dissolve 2.5 g of bis(2-hydroxyethyl) disulfide and 2.4 g of triethylamine in 50 mL of anhydrous THF. In an ice bath at 0°C and under nitrogen protection, dissolve 1.67 g of methacryloyl chloride in 30 mL of anhydrous THF and dropwise add the mixture to the above system. After the addition is complete, transfer the mixture to room temperature and stir overnight. After filtering and evaporating the solvent, dilute the residue with ethyl acetate and wash it twice with water and brine respectively. Collect the organic layer and dry it over anhydrous Na2SO4. Separate and purify the final product, a light yellow liquid HEMA, by silica gel column chromatography using a mixed solvent of ethyl acetate: petroleum ether = 2:1 (v / v) as the eluent.

[0072] like Figure 3 As shown in FIG, the successful synthesis of the product HEMA is demonstrated by the absorption peaks of the newly generated double bonds and methyl groups after the reaction.

[0073] (4) Synthesis of camptothecin prodrug (CPTM): 0.9 g of camptothecin and 0.75 g of 4-dimethylaminopyridine were dissolved in 8 mL of anhydrous DCM, 0.3 g of triphosgene was injected under a nitrogen atmosphere, and the mixture was stirred at room temperature for 2 h for full activation. 0.47 g of HEMA was added dropwise through a constant pressure dropping funnel, and the reaction mixture was stirred overnight. The solvent was filtered and dried on a rotary evaporator, and the crude product was diluted with ethyl acetate, washed once with water and twice with brine, and the organic layer was collected and dried over anhydrous Na2SO4. The final product, CPTM, was separated and purified by silica gel column chromatography using ethyl acetate as the eluent to obtain a light yellow solid powder.

[0074] like Figure 4 As shown in Figure 3, the successful synthesis of the product CPTM was demonstrated by the absorption peaks of CPT and HEMA after the reaction.

[0075] (5) Synthesis of camptothecin prodrug-modified hydroxyapatite (HA-CPT): 1 g of double-bond-modified hydroxyapatite (HA-CH=CH2), 1.2 g of camptothecin prodrug (CPTM), and 15 mg of azobisisobutyronitrile (AIBN) were dissolved in 20 mL of anhydrous DMF and placed in a sealed tube. The tube was degassed three times at -80°C under a nitrogen atmosphere and flame-sealed under vacuum. The reaction was allowed to proceed in a 75°C oil bath for 24 h, followed by cooling to terminate the reaction. The particles were washed with dichloromethane and separated by a centrifugation-redispersion cycle several times. The collected powder was placed in a 45°C vacuum oven and dried for 18 h. The powder was then ground in a mortar to obtain a light yellow powder, HA-CPTM.

[0076] like Figure 6 As shown, by 1760cm -1 The appearance of carbonyl peaks confirmed the successful polymerization of double-bonded hydroxyapatite and camptothecin prodrug.

[0077] (6) Preparation of polymethyl methacrylate prepolymer solution: Dissolve 100 mg of dibenzoyl peroxide (BPO) in 20 g of methyl methacrylate solution, place in a conical flask and heat in a 75°C water bath for 20 min. Then stop heating and place the conical flask in cold water to cool the reaction.

[0078] (7) Preparation of bone cement: 2 g of methyl methacrylate prepolymer was used as the liquid phase, 10 mg of dibenzoyl peroxide (BPO) and two modified hydroxyapatites were used as the solid phase, where the mass of HA-Cur was 50 mg and the mass of HA-CPT was 50 mg. After adding 10 mg of N,N-dimethylbenzene (DMA), the two phases were thoroughly stirred until mixed and injected into the mold. The mixture was placed in a 37°C oven for heating and curing, and the final bone cement sample was obtained after demolding.

[0079] Example 2

[0080] The method for preparing a low-exothermic stimulus-responsive injectable bone cement in this embodiment is different from that in Example 1 in that, in this embodiment, the mass of HA-Cur is 100 mg, and the mass of HA-CPT is 100 mg.

[0081] Example 3

[0082] The method for preparing a low-exothermic stimulus-responsive injectable bone cement in this embodiment is different from that in Example 1 in that, in this embodiment, the mass of HA-Cur is 150 mg, and the mass of HA-CPT is 150 mg.

[0083] Example 4

[0084] The method for preparing a low-exothermic stimulus-responsive injectable bone cement in this embodiment is different from that in Example 1 in that, in this embodiment, the mass of HA-Cur is 200 mg, and the mass of HA-CPT is 200 mg.

[0085] The contents of the solid phase in Examples 1-4 are shown in the following table:

[0086] Table 1 Solid content in Examples 1-4

[0087]

[0088] like Figure 8 As shown, the maximum temperature during the polymerization process is below 45°C, which is much lower than the 90°C requirement of the international standard for surgical implants - acrylic resin cement (ISO-5833). This can avoid tissue damage caused by large amounts of heat released during the curing process.

[0089] like Figure 9 As shown, according to the method in the international standard for surgical implants - acrylic resin cement (ISO-5833), the setting time of the four groups of bone cement was measured to be between 10 and 13 minutes, which all met the requirement of less than 15 minutes in the international standard for surgical implants - acrylic resin cement (ISO-5833).

[0090] like Figure 10 As shown, the bone cement splines are all cylindrical with a diameter of 6 mm and a height of 12 mm. The compressive mechanical properties of the bone cement splines were measured according to the test requirements of the international standard for surgical implants - acrylic resin cement (ISO-5833). Among them, the strength of the bone cements of Examples 1-3 all met the requirement of 70 MPa in the national standard ISO-5833, showing good mechanical properties and being suitable for load-bearing.

[0091] like Figure 11 As shown in the electron micrographs, after SBF immersion, the surface of the bone cement developed a more coarse crystalline structure, indicating greater calcium phosphate deposition and improved bioactivity. Furthermore, the immersion process resulted in a more porous structure, which is more conducive to capillary colonization and drug release.

[0092] like Figure 12 As shown in the figure, equal mass of bone cement particles were immersed in a mixed solution of ethanol and water with a volume ratio of 1:1 containing 300uM H2O2. By detecting the changes in the absorption peak at 425nm of the UV-visible absorption spectrum at different times, it was calculated that as the content of drug-modified hydroxyapatite increased, the release rate of curcumin also increased. The final release rate of the 20% group after 7 days could reach about 74%.

[0093] like Figure 13As shown, a 100 μM DPPH solution was dissolved in a 300 μM H₂O₂ ethanol solution and an equal mass of bone cement particles was added. The quenching efficiency of DPPH was calculated by changes in the UV-visible absorption peak at 517 nm. The DPPH free radical scavenging rate gradually increased with increasing drug-loaded hydroxyapatite content, demonstrating its excellent antioxidant properties and its potential for scavenging excessive reactive oxygen species in tumor sites.

[0094] like Figure 14 As shown, equal mass of bone cement particles were immersed in 10mM dithiothreitol (DTT) ethanol solution. After different time periods, the change in the absorption peak at 368nm of the UV-visible absorption spectrum was used to calculate that as the content of drug-modified hydroxyapatite increased, the release rate of camptothecin also increased. The final release rate of the 20% group after 7 days could reach about 52%.

[0095] Figure 15 The injectability diagram of the four groups of bone cements in the embodiment of the present invention is shown in FIG. The results show that all four groups of bone cements have good injectability.

[0096] The bone cements prepared in Examples 1-4 and the control group were tested for cytotoxicity against tumor cells. Human osteosarcoma cells and the bone cements from the different groups were injected into culture medium and cultured at 37°C for 24 hours. The cells were then rinsed with PBS buffer and DMEM complete culture medium, and MTT was added to detect cytotoxicity.

[0097] Table 2 Cytotoxicity test results of the bone cement prepared in Examples 1-4 and the control group on tumor cells

[0098] experimental group Human osteosarcoma cell (HOS) cell survival rate control group 100％ Example 1 48.7％ Example 2 34.8％ Example 3 20.3％ Example 4 11.5％

[0099] The results showed that as the modified hydroxyapatite increased, it had a good killing effect on tumor cells.

[0100] The above content is merely an example and explanation of the structure of the present invention. Those skilled in the art may make various modifications or additions to the described specific embodiments or replace them in a similar manner. As long as they do not deviate from the structure of the present invention or exceed the scope defined by the claims, they should all fall within the scope of protection of the present invention.

Claims

1. A method for preparing a low exothermic stimulus-responsive injectable bone cement, characterized in that: The following steps are involved: (1) Double-bond-modified hydroxyapatite was grafted with camptothecin prodrug via free radical polymerization to prepare camptothecin prodrug-modified hydroxyapatite; The preparation method of the camptothecin prodrug comprises the following steps: dissolving camptothecin and 4-dimethylaminopyridine in anhydrous dichloromethane, injecting triphosgene under a nitrogen atmosphere, stirring at room temperature, dropwise adding ethyl 2-((2-hydroxyethyl)disulfonyl)methacrylate, stirring the reaction mixture overnight, filtering and evaporating the solvent, diluting the crude product with ethyl acetate, washing with water and brine several times, collecting the organic layer and drying it with anhydrous sodium sulfate, and separating and purifying it by silica gel column chromatography using ethyl acetate as an eluent to obtain the camptothecin prodrug; The structural formula of the 2-((2-hydroxyethyl)disulfonyl)ethyl methacrylate is as follows: (2) Curcumin-modified hydroxyapatite was prepared by linking hydroxyapatite and curcumin molecules via oxalate bonds. (3) Using polymethyl methacrylate prepolymer solution as the liquid phase and a mixture of camptothecin prodrug-modified hydroxyapatite and curcumin-modified hydroxyapatite as the solid phase, dibenzoyl peroxide and N,N-dimethylaniline were added, and the mixture was fully stirred and dispersed, and solidified to obtain bone cement.

2. The method for preparing a low exothermic stimulus-responsive injectable bone cement according to claim 1, wherein: In step (1): The preparation method of double-bond modified hydroxyapatite comprises the following steps: dissolving hydroxyapatite, hexamethylene diisocyanate and dibutyltin dilaurate in anhydrous N,N-dimethylformamide, heating for a certain time under condensation reflux and nitrogen protection conditions, dissolving hydroxyethyl methacrylate in N,N-dimethylformamide, injecting the solution into the above system, stirring overnight and then centrifuging, washing with dichloromethane multiple times and collecting the powder, fully drying, and grinding with a mortar to obtain double-bond modified hydroxyapatite.

3. The method for preparing a low exothermic stimulus-responsive injectable bone cement according to claim 2, wherein: In the preparation process of the double-bond modified hydroxyapatite, the molar ratio of hydroxyapatite, hexamethylene diisocyanate, and hydroxyethyl methacrylate is 1:1-3:3-10, the reaction temperature of the entire reaction process is 45-60°C, the heating time is 6-15 hours under condensation reflux and nitrogen protection conditions, the drying temperature is 35-50°C, and the drying time is 12-36 hours; During the preparation of the camptothecin prodrug, the molar ratio of camptothecin, 4-dimethylaminopyridine, triphosgene and 2-((2-hydroxyethyl)disulfonyl)ethyl methacrylate is 1:2-5:0.25-3:1-3, and the washing times with water and brine are 1-3.

4. The method for preparing a low exothermic stimulus-responsive injectable bone cement according to claim 2, wherein: The preparation method of ethyl 2-((2-hydroxyethyl)disulfonyl)methacrylate comprises the following steps: dissolving bis(2-hydroxyethyl)disulfide and triethylamine in anhydrous tetrahydrofuran, dropwise adding a certain amount of methacryloyl chloride in an ice bath under nitrogen protection, transferring the mixture to room temperature and stirring overnight after the dropwise addition is completed, filtering and evaporating the solvent, diluting the residue with ethyl acetate, washing the residue with water and brine several times in sequence, collecting the organic layer and drying it with anhydrous Na2SO4, and separating and purifying the ethyl 2-((2-hydroxyethyl)disulfonyl)methacrylate by silica gel column chromatography using a mixed solvent of ethyl acetate and petroleum ether as an eluent to obtain the ethyl 2-((2-hydroxyethyl)disulfonyl)methacrylate.

5. The method for preparing a low exothermic stimulus-responsive injectable bone cement according to claim 4, wherein: The molar ratio of bis(2-hydroxyethyl) disulfide, methacryloyl chloride and triethylamine is 1:1:1.5, and the washing times with water and brine are 2-3 times respectively. The volume ratio of petroleum ether and ethyl acetate as eluent is 2:1, and the ice bath temperature is minus 10-minus 20°C.

6. The method for preparing a low exothermic stimulus-responsive injectable bone cement according to claim 1, wherein: Said step (1) specifically comprises the following steps: dissolving double bond modified hydroxyapatite, camptothecin prodrug and azobisisobutyronitrile in anhydrous N,N-dimethylformamide for packaging, degassing for three times under nitrogen, flame sealing under vacuum, heating in an oil bath for reaction and then cooling to terminate the reaction, washing with dichloromethane for multiple times and collecting powder, drying and crushing to obtain said camptothecin prodrug modified hydroxyapatite; The step (2) specifically comprises the following steps: dissolving hydroxyapatite and curcumin in anhydrous tetrahydrofuran, adding triethylamine dropwise, injecting oxalyl chloride in an ice bath under nitrogen protection, stirring in an ice bath and then transferring to room temperature and stirring overnight, filtering the suspension, washing with dichloromethane until the supernatant is nearly colorless, collecting the precipitate, drying and crushing, thereby obtaining the curcumin-modified hydroxyapatite; In the step (3), the method for preparing the polymethyl methacrylate prepolymer solution comprises the following steps: heating the methyl methacrylate solution containing the dissolved initiator in a water bath for a certain period of time, and then cooling the solution to terminate the reaction.

7. The method for preparing a low exothermic stimulus-responsive injectable bone cement according to claim 6, characterized in that: In the step (1), the molar ratio of double-bond modified hydroxyapatite, camptothecin prodrug and azobisisobutyronitrile is 1:2-5:0.1-0.5, the oil bath heating reaction temperature is 65-85°C, the reaction time is 24 h-48 h, the drying temperature is 50-70°C, the drying time is 12-36 h, and the freeze degassing temperature is -80--50°C; In the step (2), the molar ratio of hydroxyapatite, curcumin, triethylamine and oxalyl chloride is 1:1-3:4.5:2-3, the stirring time in an ice bath at -20--10°C is 30 min, and the reaction is carried out in the dark. The reaction temperature is 25-45°C, the reaction time is 24-72 h, the drying temperature is 30-40°C, and the drying time is 12-24 h. In the step (3), during the preparation of the polymethyl methacrylate prepolymer solution, the initiator is azobisisobutyronitrile or dibenzoyl peroxide, the mass ratio of the initiator to the methyl methacrylate monomer is 0.5%-2%, the water bath heating temperature is 70-90°C, the water bath heating time is 15-30 min, the cooling temperature is 5-10°C, and the cooling time is 5-15 min.

8. The method for preparing a low exothermic stimulus-responsive injectable bone cement according to claim 1, wherein: The mass ratio of the liquid phase to the solid phase is 1:0.05-0.5, the mass ratio of the curcumin-modified hydroxyapatite to the camptothecin prodrug-modified hydroxyapatite is 1:1, the mass of dibenzoyl peroxide is 0.2%-1.5% of the mass of the polymethyl methacrylate prepolymer solution, the mass of N,N-dimethylaniline is 1%-3% of the mass of the polymethyl methacrylate prepolymer solution, the curing temperature is 36-37°C, and the curing time is 15-60 minutes.

9. A low exothermic stimulus-responsive injectable bone cement prepared according to the method for preparing the low exothermic stimulus-responsive injectable bone cement according to any one of claims 1 to 8.

10. Use of the low-exothermic stimulus-responsive injectable bone cement according to claim 9 in preparing an implant material for treating metastatic tumors.

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