Embolic microsphere capable of self-fusing in vivo, and preparation and use thereof

Abstract

The present invention discloses an embolic microsphere capable of self-fusing in vivo, and preparation and use thereof. The method comprises: reacting a first prepolymer, a second prepolymer, a coupling agent, and a catalyst at 60-130 °C to obtain polyurethane; mixing the polyurethane obtained in step (1) with a drug and dissolving the mixture in an organic solvent to obtain an oil phase; adding the oil phase to an aqueous polyvinyl alcohol solution and stirring to fully emulsify the oil phase, followed by preparing the embolic microsphere capable of self-fusing in vivo by means of an emulsification-volatilization method under a pressure of -0.9 MPa to -0.01 MPa. The present invention addresses the problem that existing embolic agents are easily fragmented by blood flow impact, failing to achieve complete vascular occlusion and long-term embolization. The embolic microsphere of the present invention has high strength and self-fusing properties, enabling the effects of complete vascular occlusion and long-term embolization, and can also serve as a carrier to load a hydrophobic drug.

Description

Embolic microsphere capable of self-fusion in vivo and its preparation and application Technical Field

[0001] The present invention relates to an embolic microsphere, in particular to an embolic microsphere capable of self-fusion in vivo and the preparation and application thereof. Background Art

[0002] Transcatheter arterial embolization delivers spherical or non-spherical embolic agents to the target vessel, thereby achieving localized blood supply occlusion. Currently, there are two main types of embolic agents used clinically. The first is a mechanical vascular occlusion device, which is deployed within the target vessel in hemorrhagic or aneurysmal situations. The other is a flow-directed embolic agent, which includes particles, polymers, or in situ gels. These are transferred via a catheter to the surrounding vessels of the target tissue, blocking the target tissue's blood supply and causing it to atrophy. Of all commercially available embolic agents, spherical embolic particles are the most advantageous choice.

[0003] Embolic microspheres are often used to embolize various hypervascular tumors, primarily for the treatment of primary liver cancer and uterine fibroids. They can also be used to treat urinary tract obstruction caused by prostatic hyperplasia, bleeding disorders, or hyperthyroidism. To match the size and caliber of the target blood vessels for more effective embolization, embolic microspheres typically come in various sizes. Furthermore, the microspheres must exhibit good elasticity, be easily compressed, and be able to deform through extrusion to achieve a tighter fit between microspheres and between microspheres and blood vessels. However, to achieve good elasticity, the microspheres typically require a low storage modulus. When the storage modulus is below 800 Pa, the microspheres are unable to withstand the pressure of blood flow and are therefore easily broken. These broken microspheres may then migrate with the blood flow to other organs, leading to unintended vascular blockage.

[0004] Compared to spherical embolic agents, liquid embolic agents such as iodized oil, ethanol, and Onyx can be delivered via microcatheters. They can quickly penetrate and completely fill the target vessel, ultimately forming an embolism that completely blocks blood flow. Therefore, they are widely used in clinical practice. However, liquid embolic agents are typically rapidly metabolized by the body, making them ineffective in forming long-lasting embolic blocks. Consequently, the main challenges faced by embolic microspheres currently used in clinical practice include: 1. Low microsphere strength and brittleness, making them easily shattered by blood flow; 2. The presence of gaps between microspheres aggregated within a vessel prevents complete occlusion and blood flow blockage.

[0005] Summary of the Invention

[0006] The purpose of the present invention is to provide embolic microspheres that can self-fuse in vivo, as well as their preparation and application, to solve the problem that existing embolic agents are easily broken by blood flow impact and cannot achieve complete blood vessel occlusion and long-term embolism. Amphiphilic polyurethane material is used to prepare high-strength embolic microspheres loaded with oil-soluble organic matter through vacuum negative pressure combined with emulsification. These microspheres have self-fusing properties in vivo, completely cut off blood flow supply, and achieve long-term embolism.

[0007] In order to achieve the above object, the present invention provides a method for preparing embolic microspheres capable of self-fusion in vivo, the method comprising:

[0008] (1) reacting a first prepolymer, a second prepolymer, a coupling agent, and a catalyst at 60 to 130° C. under inert gas protection to obtain a polyurethane;

[0009] (2) mixing the polyurethane obtained in step (1) with a drug and dissolving them in an organic solvent to obtain an oil phase; adding the oil phase to a polyvinyl alcohol aqueous solution and stirring at 4 to 10° C. to fully emulsify the oil phase; stirring at a pressure of -0.9 MPa to -0.01 MPa and a temperature of 4 to 10° C., then heating to 25° C. to obtain the embolic microspheres capable of self-fusion in vivo by an emulsification and volatilization method;

[0010] The melting point of microspheres is relatively low, at only 33°C. If the reaction temperature is too high, the microspheres may fuse during the preparation process, resulting in the inability to obtain well-dispersed microspheres. If the reaction temperature is too low, the volatilization rate of the organic reagent will decrease, affecting the yield of the microspheres. If the pressure is too high, the microspheres tend to agglomerate. If the pressure is too low, the volatilization rate of the organic solvent is too slow, ultimately resulting in fibrous microspheres and a reduced yield.

[0011] Wherein, the first prepolymer is any one of polyethylene glycol and polypropylene glycol;

[0012] The second prepolymer is any one of polycaprolactone, polylactic acid, polytrimethylene carbonate, poly(caprolactone-trimethylene carbonate), poly(lactic acid-caprolactone) and poly(lactic acid-trimethylene carbonate); if the first prepolymer and the second prepolymer are replaced with other similar or similar substances, polyurethane can be generated, but the melting point and glass transition temperature of the polyurethane will change, and the degradation rate of the target product will be faster if the second prepolymer is replaced with other prepolymers.

[0013] The coupling agent is any one of hexamethylene diisocyanate, diphenylmethane diisocyanate and 4,4-diisocyanate dicyclohexylmethane.

[0014] Preferably, in step (1), the catalyst is an organic tin salt or an organic bismuth salt

[0015] Preferably, in step (1), the total amount of the first prepolymer and the second prepolymer is in a ratio of 1:(0.8-1.2) to the amount of the coupling agent; the mass ratio of the first prepolymer to the second prepolymer is 1:1; the mass of the catalyst is 0.01% to 0.4% of the total mass of the first prepolymer and the second prepolymer. To ensure that the polyurethane has good hydrophilicity, the mass ratio of the first prepolymer to the second prepolymer is 1:1. Too low a content of the first prepolymer will reduce the hydrophilicity of the target product, while too low a content of the second prepolymer will affect the yield of the microspheres and the final molding. Too much coupling agent will lead to a decrease in the molecular weight of the target product, and cross-linking may occur during the synthesis process, making it impossible to obtain a linear polymer, while too low a content will also reduce the molecular weight of the target product. Too low a molecular weight of the target product will affect the formation and yield of microspheres, and microspheres may not be obtained.

[0016] Preferably, in step (1), the inert gas is any one or more of nitrogen, argon and helium.

[0017] Preferably, in step (2), the drug is any one of oleic acid, linoleic acid, iodized oil and poppy seed oil; and the organic solvent is any one or both of dichloromethane and chloroform.

[0018] Preferably, in step (2), the mass ratio of the polyurethane to the drug is (1-5):(1-5); and the volume ratio of the oil phase to the polyvinyl alcohol aqueous solution is 1:(5-20). If the amount of polyurethane used is too high, the melting point of the microspheres will be high, and fusion may not occur under physiological conditions. If the amount of polyurethane used is too low, the strength of the microspheres will be low, and they may not be able to withstand the impact of blood flow and may shift, resulting in ineffective embolization.

[0019] More preferably, the concentration of the polyvinyl alcohol aqueous solution is (5-20) g / L; the concentration of the polyurethane in the oil phase is (10-100) g / L.

[0020] Preferably, in step (2), the stirring rate is 500-1500 rpm.

[0021] The present invention provides an embolic microsphere capable of self-fusion in vivo, which is prepared by the preparation method.

[0022] The present invention provides a use of the embolic microspheres capable of self-fusion in vivo in the preparation of embolic therapeutic drugs.

[0023] The present invention provides an embolic microsphere capable of self-fusion in vivo, and its preparation and application, which solves the problem that existing embolic agents are easily broken by blood flow impact and cannot achieve complete blood vessel occlusion and long-term embolization. It has the following advantages:

[0024] 1. The embolic microspheres of the present invention have high strength and will not break when impacted by blood flow.

[0025] 2. The polyurethane multi-block compound prepared by the present invention has a more uniform distribution of hydrophilic and hydrophobic segments and better hydrophilicity. The prepared embolic microspheres have self-fusion properties in the body. After being fused into one, they can effectively block blood vessels and completely cut off blood supply, achieving complete blockage of blood vessels and long-term embolism.

[0026] 3. The embolic microsphere material prepared from the amphiphilic polyurethane material of the present invention has amphiphilicity, and the microspheres can be used as carriers to load hydrophobic drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 is a diagram of the polyurethane prepared in Example 1 of the present invention. 1 HNMR spectrum.

[0028] FIG2 is a photo of the embolic microspheres prepared in Example 1 of the present invention and Comparative Example 1 under an optical microscope.

[0029] FIG3 is a DSC curve of the embolic microspheres prepared in Example 1 of the present invention and Comparative Example 1.

[0030] FIG4 is a graph showing rheological measurements of the embolic microspheres prepared in Example 1 of the present invention and Comparative Example 1.

[0031] FIG5 is a diagram showing an in vitro fusion experiment of the embolic microspheres prepared in Example 1 of the present invention and Comparative Example 1.

[0032] FIG6 is a graph showing the in vivo biocompatibility evaluation of the embolic microspheres prepared in Example 1 and Comparative Example 1 of the present invention.

[0033] FIG7 is a histological section diagram of the in vivo embolism of the embolic microspheres prepared in Example 1 of the present invention and Comparative Example 1. DETAILED DESCRIPTION

[0034] The following is a clear and complete description of the technical solutions in the embodiments of the present invention. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0035] The manufacturers, models, specifications or preparation methods of the reagents involved in the following examples are as follows:

[0036] Prepolymer PEG (Mn = 2000, polyethylene glycol) was purchased from Shanghai for testing and was chemically pure;

[0037] 1,6-Hexanediisocyanate (HDI) was purchased from Huaxia Reagent with a purity of 99%;

[0038] PCL and PLA were prepared in-house. PCL was prepared by adding ethylene glycol, caprolactone, and stannous octoate to a reaction flask; deaeration was performed using a vacuum oil pump at room temperature (approximately 10 to 30 minutes), followed by nitrogen replacement twice. The vacuum pump was then turned off and nitrogen was applied to maintain a positive pressure in the system. The reaction flask was then placed in an oil bath at 135°C for 48 hours. The product was cooled to room temperature and dissolved in dichloromethane. The product was then purified using a mixed solvent of n-hexane and diethyl ether. The volume ratio of n-hexane to diethyl ether was 4:1. Ethylene glycol was used as an initiator, and the mass ratio of initiator to caprolactone was 1:(31 to 128). Stannous octoate was used as a catalyst, the mass of which was 0.1 wt% of the total mass of ethylene glycol and caprolactone.

[0039] The preparation method of PLA comprises the following steps: adding ethylene glycol, lactide, and stannous octoate to a reaction flask; removing air from the reaction system using a vacuum oil pump at room temperature (approximately 10 to 30 minutes); replacing the reaction system with nitrogen twice; turning off the vacuum pump and applying nitrogen to maintain a positive pressure; and then placing the reaction flask in a 135°C oil bath for 48 hours. The product is cooled to room temperature, dissolved in dichloromethane, and then purified using anhydrous ethanol. Ethylene glycol is used as an initiator in a mass ratio of initiator to lactide of 1:(31 to 128); and stannous octoate is used as a catalyst, the mass of which is 0.1 wt% of the total mass of ethylene glycol and caprolactone.

[0040] Example 1

[0041] A method for preparing embolic microspheres capable of self-fusion in vivo, the method comprising:

[0042] (1) Prepolymer PEG (molecular weight Mn = 2000, polyethylene glycol) and prepolymer PCL (Mn = 2000, polycaprolactone) were dissolved in dichloromethane, stannous octoate was added as a catalyst, and hexamethylene diisocyanate (HDI) was added as a coupling agent. After mixing evenly, the dichloromethane was completely removed by rotary distillation and nitrogen was added for protection. The reaction flask was placed in an oil bath at 90°C for reaction for 24 hours. The obtained polyurethane was recorded as PCEU. Among them, the mass ratio of dichloromethane to the total mass of prepolymer PEG and prepolymer PCL was 3:1, the mass of stannous octoate was 0.4wt% of the total mass of prepolymer PEG and prepolymer PCL, the amount of coupling agent HDI to the total amount of prepolymer PEG and prepolymer PCL was 1:1, and the mass ratio of prepolymer PEG to prepolymer PCL was 1:1.

[0043] (2) Mixing iodized oil and the polyurethane obtained in step (1) in a mass ratio of 1:1 and dissolving them in dichloromethane to obtain an oil phase; slowly adding the obtained oil phase to a 1% PVA (polyvinyl alcohol) aqueous solution at a stirring rate of 200-300 rpm over 3-5 minutes; then stirring at 1000 rpm and 4°C for 20 minutes to fully emulsify the oil phase; maintaining the vacuum degree in the reaction flask at -0.75 MPa by reducing pressure; continuing stirring at 4°C for 20 minutes; then raising the temperature to 25°C and continuing stirring to completely remove the dichloromethane; collecting microspheres by centrifugation; and washing with pure water 5-10 times to remove the residual PVA; and collecting microspheres by centrifugation. The concentrations of polyurethane and iodized oil in the oil phase are both 5% (w / v); and the volume ratio of the oil phase to the PVA aqueous solution is 1:10.

[0044] Comparative Example 1

[0045] A method for preparing embolic microspheres for embolization therapy is basically the same as that in Example 1, except that:

[0046] In step (2), no iodized oil was added;

[0047] The microspheres were collected by centrifugation following the same operation as in Example 1.

[0048] Example 2

[0049] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0050] In step (1), the prepolymer PEG (Mn=2000, polyethylene glycol) and the prepolymer PCL (Mn=2000, polycaprolactone) are adjusted to prepolymer PEG (Mn=2000) and prepolymer PLA (Mn=2000, polylactic acid), and the mass ratio of the prepolymer PEG to the prepolymer PLA is 1:1; the reaction temperature is adjusted from 90°C to 130°C, the mass of stannous octoate is 0.4wt% of the total mass of the prepolymer PEG and the prepolymer PLA, and the molar ratio of the coupling agent HDI to the total molar ratio of the prepolymer PEG and PCL is 1:1.2.

[0051] Example 3

[0052] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0053] In step (1), stannous octoate is adjusted to bismuth octoate, the mass of bismuth octoate is 0.4wt% of the total mass of prepolymer PEG and prepolymer PCL; the amount of coupling agent HDI and the total amount of prepolymer PEG and prepolymer PCL is 1:1.1.

[0054] In step (2), the stirring speed for fully emulsifying the oil phase was adjusted from 1000 rpm to 500 rpm.

[0055] Example 4

[0056] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0057] In step (1), the reaction temperature is adjusted from 90° C. to 130° C., and the reaction time is adjusted from 24 hours to 6 hours; the coupling agent is adjusted from HDI to 4,4′-dicyclohexylmethane diisocyanate (HMDI), and the molar ratio of the coupling agent HMDI to the total molar ratio of the prepolymer PEG and the prepolymer PCL is 1:1.

[0058] Example 5

[0059] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0060] In step (1), the reaction time is adjusted from 24 hours to 6 hours; the molar ratio of the coupling agent HDI to the total molar ratio of the prepolymer PEG and the prepolymer PCL is 1:0.8.

[0061] In step (2), the PVA concentration in the PVA aqueous solution is adjusted from 1% to 0.5%.

[0062] Example 6

[0063] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0064] In step (1), the reaction temperature was adjusted from 90° C. to 130° C.; the mass of stannous octoate was 0.01 wt % of the total mass of the prepolymer PEG and the prepolymer PCL.

[0065] Example 7

[0066] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0067] In step (1), the reaction temperature was adjusted from 90°C to 60°C.

[0068] In step (2), the mass ratio of iodized oil to the polyurethane obtained in step (1) is adjusted from 1:1 to 5:1, and the dichloromethane is adjusted to chloroform.

[0069] Example 8

[0070] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0071] In step (2), the concentrations of polyurethane and iodized oil in the oil phase are adjusted from 5% to 1%; the PVA concentration in the PVA aqueous solution is adjusted from 1% to 0.5%; and the stirring speed for fully emulsifying the oil phase is adjusted from 1000 rpm to 500 rpm.

[0072] Example 9

[0073] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0074] In step (2), the concentrations of polyurethane and iodized oil in the oil phase are adjusted from 5% to 10%; the PVA concentration in the PVA aqueous solution is adjusted from 1% to 2%; and the stirring speed for fully emulsifying the oil phase is adjusted from 1000 rpm to 1500 rpm.

[0075] Example 10

[0076] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0077] In step (2), the volume ratio of the oil phase to the PVA aqueous solution was adjusted from 1:10 to 1:20; and the vacuum degree in the reaction flask was adjusted from -0.75 MPa to -0.02 MPa.

[0078] Example 11

[0079] The method for preparing embolic microspheres capable of self-fusion in vivo is basically the same as that in Example 1, except that:

[0080] In step (2), the mass ratio of iodized oil to the polyurethane obtained in step (1) is adjusted from 1:1 to 5:1; the volume ratio of the oil phase to the PVA aqueous solution is adjusted from 1:10 to 1:5; and the vacuum degree in the reaction flask is adjusted from -0.75 MPa to -0.9 MPa.

[0081] Example 1 Characterization

[0082] 1. Molecular structure characterization

[0083] The polyurethane PCEU prepared in Example 1 was characterized by hydrogen nuclear magnetic resonance spectroscopy. The specific method was as follows: 5 mg of PCEU was dissolved in 600 μL of deuterated chloroform (with 0.5% tetramethylsilane as an internal standard), and tested in a 600 MHz nuclear magnetic resonance spectrometer to obtain a hydrogen nuclear magnetic resonance spectrum.

[0084] As shown in FIG1 , the polyurethane prepared in Example 1 of the present invention 1HNMR spectrum. As shown in Figure 1, the proton peaks of PEG, PCL, and HDI are clearly visible. By calculating the ratio of PEG to PCL, it can be seen that the ratio of PEG to PCL in polyurethane is equivalent to the feed ratio.

[0085] 2. Polyurethane molecular weight detection

[0086] The molecular weight and molecular weight distribution of the polyurethanes prepared in Examples 1 to 7 were characterized by using 2 g / L of the polyurethanes prepared in Examples 1 to 7 for molecular weight determination by gel permeation chromatography to obtain the molecular weight and distribution of the polyurethanes. The results are detailed in Table 1.

[0087] Table 1 Molecular weight and distribution of copolymers prepared in Examples 1 to 7

[0088] As can be seen from Table 1, the polyurethanes prepared in Examples 1 to 7 of the present invention have both relatively high molecular weight (Mn>23000) and uniform molecular weight distribution (PDI<1.73).

[0089] 3. Particle size detection

[0090] The particle size and distribution of the microspheres prepared in Examples 1 to 11 and Comparative Example 1 were characterized. Specifically, the microspheres prepared in Examples 1 to 11 and Comparative Example 1 were fully hydrated and then tested using a particle size analyzer. The results are detailed in Table 2.

[0091] Table 2 Particle size and particle size distribution of microspheres prepared in Examples 1 to 11

[0092] As shown in Table 2, the embolic microspheres prepared in Examples 1 to 11 of the present invention and Comparative Example 1 can all pass through a standard microcatheter (>0.33 mm) and have uniform particle sizes.

[0093] 4. Morphological Characterization of Microspheres

[0094] The morphologies of the microspheres prepared in Example 1 and Comparative Example 1 were characterized as follows: the microspheres prepared in Example 1 and Comparative Example 1 were placed under a microscope for observation and photographing.

[0095] Figure 2 shows optical microscope photographs of the embolic microspheres prepared in Example 1 of the present invention and Comparative Example 1, wherein Example 1 shows the embolic microspheres prepared in Example 1, and Comparative Example 1 shows the embolic microspheres prepared in Comparative Example 1. Figure 2 shows that the embolic microspheres prepared in Example 1 and Comparative Example 1 do not agglomerate and have good dispersion.

[0096] Example 2 Performance Testing

[0097] 1. Thermal analysis

[0098] The microspheres prepared in Example 1 and Comparative Example 1 were characterized using differential scanning calorimetry. Specifically, 6 mg of each of the embolic microspheres prepared in Example 1 and Comparative Example 1 were heated from -70°C to 80°C at a heating rate of 10°C / min in a differential scanning calorimeter in a nitrogen atmosphere to obtain DSC curves.

[0099] Figure 3 shows the DSC curves of the embolic microspheres prepared in Example 1 and Comparative Example 1 of the present invention, where the abscissa represents temperature and the ordinate represents heat flow. Example 1 shows the embolic microspheres prepared in Example 1, and Comparative Example 1 shows the embolic microspheres prepared in Comparative Example 1. As shown in Figure 3, the polyurethane microspheres prepared in Example 1 with iodized oil have a lower melting point, dropping from 44.2°C and 55°C in Comparative Example 1 to 33.2°C and 41.5°C, respectively. This indicates that at the normal physiological temperature of 37°C, some crystals in the polyurethane microspheres prepared in Example 1 can melt.

[0100] 2. Modulus detection

[0101] The storage modulus (G') and loss modulus (G") of the microspheres prepared in Example 1 and Comparative Example 1 were characterized. The specific method is as follows: the microspheres prepared in Example 1 and Comparative Example 1 were immersed in pure water, incubated at 37°C for 48 hours, and then the microsphere monolayers of the Example and Comparative Example were evenly placed on the test bench of the rheometer. The storage modulus (G') and loss modulus (G") with an angular frequency of 0.1 to 100 rad / s were measured at 37°C, and the modulus was tested under a fixed strain (0.1%). The storage modulus represents the stiffness of the material, and it must be greater than 800 Pa to withstand blood pressure and maintain the shape of the microspheres without breaking.

[0102] As shown in Figure 4, the rheological measurement graphs of the embolic microspheres prepared in Example 1 and Comparative Example 1 of the present invention are shown, wherein the horizontal axis represents the angular frequency and the vertical axis represents the coefficient. Example 1 represents the embolic microspheres prepared in Example 1, and Comparative Example 1 represents the embolic microspheres prepared in Comparative Example 1. It can be observed from Figure 4 that the G' of the embolic microspheres prepared in Example 1 and Comparative Example 1 are always greater than G" and greater than 800 Pa, and the two remain consistent at different angular frequencies. This indicates that the microspheres prepared in Example 1 and Comparative Example 1 both have a typical cross-linked hydrogel structure, with high elasticity and relatively high strength. It can also be observed that the G' and G" values ​​of Example 1 are lower than those of Comparative Example 1, which may be due to the addition of iodized oil in Example 1. As a plasticizer, iodized oil can reduce the crystallinity in the microspheres. Therefore, Example 1 is more susceptible to deformation.

[0103] 3. In vitro self-fusion experiment

[0104] The microspheres prepared in Example 1 and Comparative Example 1 were characterized by an in vitro fusion experiment. The specific method was as follows: the microspheres prepared in Example 1 and Comparative Example 1 were placed on a glass slide and placed in an oven, incubated at 37.5, 38.5, 39.5°C, 40.5°C and 41.5°C for 2 h, and then placed under a microscope for observation and photography to obtain microscopic photographs of the microspheres.

[0105] As shown in Figure 5, the in vitro fusion experiment diagram of the embolic microspheres prepared in Example 1 and Comparative Example 1 of the present invention, wherein Example 1 is the embolic microspheres prepared in Example 1, and Comparative Example 1 is the embolic microspheres prepared in Comparative Example 1. After incubation at different temperatures for 2 hours, Example 1 began to fuse with the surrounding microspheres at 37.5°C to form a whole. As the temperature increases, the fusion becomes more complete. However, Comparative Example 1 only shows a mild fusion phenomenon at 41.5°C. This may be because the microspheres are formed by physical cross-linking, in which hydrophobic PCL crystals serve as cross-linking points. When the temperature is higher than T m After that, the chain segments in the PCL crystals are able to move, and the PCL chain segments in Example 1 are able to form new entanglements with adjacent microspheres, enabling them to fuse with each other.

[0106] 4. Biocompatibility testing

[0107] The microspheres prepared in Example 1 and Comparative Example 1 were implanted into SD rats to evaluate the in vivo degradability and biocompatibility of the microspheres. Before the experiment, male SD rats weighing 180-190 g were kept in a research laboratory for at least 1 week. The experimental group (Example 1) was anesthetized by intraperitoneal injection of 0.3% sodium pentobarbital to ensure that the rats were painless. Subsequently, 200 μL (100-300 μm) of the microspheres prepared in Example 1 were implanted into the back of the rats. 1, 2, 3, and 4 weeks after surgery, the implanted microspheres and the skin in contact with the microspheres were stained with hematoxylin-eosin (HE) for histopathological analysis; Control Example 1 (Comparative Example 1) was implanted with the microspheres prepared in Comparative Example 1, and the rest of the operations were the same as those of the experimental group; the rats in the control group were not treated in any way; during the entire experiment, no obvious tissue edema or necrosis was observed on the skin surface.

[0108] Figure 6 shows the in vivo biocompatibility evaluation of the embolic microspheres prepared in Example 1 and Comparative Example 1 of the present invention. The control was untreated, Example 1 was implanted with the embolic microspheres prepared in Example 1, and Comparative Example 1 was implanted with the embolic microspheres prepared in Comparative Example 1. Figure 6 shows that subcutaneous implantation of the embolic microspheres prepared in Example 1 and Comparative Example 1 in rats triggered an immune response, leading to the accumulation of inflammatory cells near the microspheres. As implantation time increased, the number of inflammatory cells surrounding the microspheres gradually decreased, indicating that both the embolic microspheres prepared in Example 1 and Comparative Example 1 exhibited good biocompatibility. It was also observed that the microspheres in Example 1 fused after subcutaneous implantation, no longer maintaining their individual structures but forming a single entity.

[0109] 5. In vivo vascular embolism test

[0110] The microspheres of Example 1 and Comparative Example 1 were characterized for their vascular embolization efficacy. Six healthy rabbits weighing 2.6 to 3.0 kg were dehaired before administration. A 50% (v / v) concentration of the microsphere preparation (microspheres of Example 1 and Comparative Example 1, with a size range of 200 to 300 μm) was slowly injected into the proximal central artery of the right ear, with a volume of 200 μL per ear. A control group received a 200 μL injection of normal saline into the proximal central artery of the right ear. Macroscopic changes in the rabbit ears, including morphology and color, were observed on days 1, 3, 5, 7, and 10 after administration to evaluate the embolization efficacy. The animals were sacrificed 14 days after surgery. The rabbit ears were cleaned and immersed in 4% formaldehyde solution. Tissues from both the unembolized and embolized ears were embedded in paraffin and stained with hematoxylin and eosin. Subsequently, biopsies were observed under an optical microscope.

[0111] The experimental results showed that the central auricular artery and auricular artery branches were visible in the unembolized right ear (control group). On the first day after embolization, the left ears of the experimental group of rats showed varying degrees of blood flow obstruction and different macroscopic phenomena: the central auricular artery swelled significantly after the microspheres of Comparative Example 1 were implanted, and the rabbit ears turned significantly purple after the microspheres of Example 1 were implanted, indicating that we successfully implanted the microspheres into the blood vessels. On the third day after embolization, the microspheres of Example 1 caused thinning of part of the ear cartilage tissue, while the microspheres of Comparative Example 1 showed no significant changes compared with the first day. On the fifth day after embolization, cartilage tissue necrosis was visible in the microsphere group of Example 1, while the microspheres of Comparative Example 1 showed no significant changes. On the seventh day after embolization, obvious cartilage tissue necrosis appeared in the microsphere ear of Example 1, while the microspheres of Comparative Example 1 showed no significant changes compared with the first day. On the tenth day after embolization, large areas of necrotic tissue were visible in the microsphere-embolized ear of Example 1, while the microspheres of Comparative Example 1 showed no significant changes. Therefore, compared with the microspheres of comparative example 1, the microspheres of example 1 of the present invention caused vascular occlusion, resulting in complete ischemic necrosis of tissues around the embolization site, achieving the purpose of complete vascular occlusion and long-term embolism.

[0112] Figure 7 shows histological sections of in vivo embolization of embolic microspheres prepared in Example 1 and Comparative Example 1 of the present invention. The control was implanted with normal saline; Example 1 was implanted with the embolic microspheres prepared in Example 1; and Comparative Example 1 was implanted with the embolic microspheres prepared in Comparative Example 1. As shown in Figure 7 , after implanting normal saline, the microspheres of Example 1, and the microspheres of Comparative Example 1, respectively, HE-stained micrographs revealed that only the fusion of the microspheres of Example 1 produced an embolic agent with a shape that better conformed to the central auricular artery, achieving more effective and long-lasting embolization.

[0113] And the preparation method disclosed in the prior art (patent number CN116102748) is a diblock or triblock copolymer prepared using polyethylene glycol as an initiator. The prepared microspheres should be rigid microspheres, which cannot be deformed and cannot pass through microcatheters and blood vessels into the target blood vessels, and cannot achieve more effective embolization. The polyurethane multi-block compound prepared by the present invention can be deformed and can pass through microcatheters and blood vessels more smoothly into the target blood vessels. At the same time, the microspheres prepared by the present invention can fuse with each other to form a whole under physiological conditions, which can achieve more effective embolization. And compared with the prior art, the polyurethane multi-block compound prepared by the present invention has a more uniform distribution of hydrophilic and hydrophobic segments and has better hydrophilicity.

[0114] Although the present invention has been described in detail through the above preferred embodiments, it should be understood that the above description is not intended to limit the present invention. After reading the above description, various modifications and substitutions of the present invention will become apparent to those skilled in the art. Therefore, the scope of protection of the present invention should be defined by the appended claims.

Claims

1. A method for preparing embolic microspheres capable of self-fusion in vivo, characterized in that: The method includes: (1) reacting a first prepolymer, a second prepolymer, a coupling agent, and a catalyst at 60 to 130° C. under inert gas protection to obtain a polyurethane; (2) mixing the polyurethane obtained in step (1) with a drug and dissolving them in an organic solvent to obtain an oil phase; adding the oil phase to a polyvinyl alcohol aqueous solution and stirring at 4 to 10° C. to fully emulsify the oil phase; stirring at a pressure of -0.9 MPa to -0.01 MPa and a temperature of 4 to 10° C., then heating to 25° C. to obtain the embolic microspheres capable of self-fusion in vivo by an emulsification and volatilization method; Wherein, the first prepolymer is any one of polyethylene glycol and polypropylene glycol; The second prepolymer is any one of polycaprolactone, polylactic acid, polytrimethylene carbonate, poly(caprolactone-trimethylene carbonate), poly(lactic acid-caprolactone) and poly(lactic acid-trimethylene carbonate); The coupling agent is any one of hexamethylene diisocyanate, diphenylmethane diisocyanate and 4,4-diisocyanate dicyclohexylmethane.

2. The preparation method according to claim 1, characterized in that In step (1), the catalyst is an organic tin salt or an organic bismuth salt.

3. The preparation method according to claim 1, characterized in that In step (1), the inert gas is any one or more of nitrogen, argon and helium.

4. The preparation method according to claim 1, characterized in that In step (1), the mass ratio of the first prepolymer to the second prepolymer is 1:1; the mass ratio of the total amount of the first prepolymer and the second prepolymer to the amount of the coupling agent is 1:(0.8-1.2); the mass of the catalyst is 0.01%-0.4% of the total mass of the first prepolymer and the second prepolymer.

5. The preparation method according to claim 1, characterized in that In step (2), the drug is any one of oleic acid, linoleic acid, iodized oil and poppy seed oil; and the organic solvent is any one or both of dichloromethane and chloroform.

6. The preparation method according to claim 1, characterized in that In step (2), the mass ratio of the polyurethane to the drug is (1-5):(1-5); the volume ratio of the oil phase to the polyvinyl alcohol aqueous solution is 1:(5-20).

7. The preparation method according to claim 6, characterized in that The concentration of the polyvinyl alcohol aqueous solution is (5-20) g / L; the concentration of the polyurethane in the oil phase is (10-100) g / L.

8. The preparation method according to claim 1, characterized in that In step (2), the stirring rate is 500-1500 rpm.

9. Embolic microspheres capable of self-fusion in vivo, prepared by the method according to any one of claims 1 to 8.

10. Use of the embolic microspheres capable of self-fusion in vivo as claimed in claim 9 in the preparation of embolic therapy drugs.

Images