Multifunctional anti-stenosis drug coating stent as well as preparation method and application thereof

Abstract

The invention discloses a multifunctional anti-stenosis drug-coated stent which comprises a nickel-titanium stent, the outer surface of the nickel-titanium stent is coated with a PLLA-TPU-based drug-loaded coating, and a drug-loaded film is attached to the inner surface of the nickel-titanium stent. The body of the prepared multifunctional anti-stenosis drug-coated stent is a nickel-titanium stent and has good biocompatibility, the outer surface of the stent is coated with PLLA-TPU-based drug-loaded coating suspension slurry, the stent has good degradation performance and adjustable mechanical performance, inward or outward overgrowth of tumor cells can be effectively inhibited through drug release, and the anti-stenosis drug-coated stent has good anti-stenosis effect on tumor cells. Meanwhile, the medicine carrying membrane on the inner layer of the nickel-titanium stent can effectively inhibit tumor cells and effectively prevent bile mud from being deposited, and the nickel-titanium stent can be used for treating biliary tract stenosis diseases. The invention further provides a preparation method of the multifunctional anti-stenosis drug coating stent, the stent is prepared through a liquid flame spraying technology and a lamination method, the preparation method is simple, and the preparation period is short.

Description

Technical Field

[0001] The present invention relates to the technical field of functional coatings, and in particular to a multifunctional anti-stenosis drug-coated stent and a preparation method and application thereof. Background Art

[0002] Hepatobiliary diseases are among the most common and dangerous threats to human health worldwide, characterized by high mortality and severe morbidity. Biliary stricture, a common complication of hepatobiliary disease, has diverse etiologies, including surgical injury, inflammation, and tumors, significantly impacting patients' quality of life. In recent years, advances in endoscopic and minimally invasive techniques have driven research in early diagnosis and precision treatment, making them key areas of focus in this field.

[0003] Biliary strictures are divided into benign and malignant strictures. Cholangiocarcinoma and pancreatic cancer are one of the main causes of malignant biliary strictures. Strictured bile ducts prevent bile from flowing from the liver and gallbladder to the duodenum, leading to digestive system abnormalities, cirrhosis and jaundice.

[0004] Biliary sludge is a mixture of cholesterol crystals, bilirubin calcium salts, proteins, and other substances precipitated in bile. The accumulation of bile sludge is accompanied by the formation of biofilms, which in turn exacerbate the accumulation of bile sludge. When bile sludge accumulates in the bile duct for a long time, it can not only easily cause blockage within the bile duct but can also accumulate on the surface of bile duct stents, leading to stent obstruction. This obstruction can hinder the normal excretion of bile and can lead to complications such as cholestasis, cholangitis, and even jaundice.

[0005] Biliary stent implantation can reduce pressure and unclog bile duct drainage, making it a palliative treatment option for biliary strictures. Current clinical challenges include restenosis caused by biliary sludge accumulation and inward or excessive tumor growth after bare stent implantation. To address restenosis caused by tumor growth, research on anti-biliary sludge drug-coated stents is urgently needed.

[0006] Plastic and metal stents are typical traditional biliary stents. Plastic stents are susceptible to bacterial biofilm formation due to stent occlusion. Metal stents, whether bare metal or covered, can lead to excessive inward or outward tumor growth, resulting in biofilm formation and biliary sludge accumulation, making the bile duct prone to restenosis. Consequently, stent surface modification is gaining increasing attention, particularly in the research of anti-biofilm coatings and drug-eluting coatings.

[0007] The anti-biliary sludge drug-coated stent consists of four parts: a bare metal stent, a drug carrier, a drug, and a coating material. The bare metal stent is used as an attachment for the coating material, and the drug loaded by the drug carrier is evenly mixed into the coating material to prepare a suspension for spraying and dipping of the coating.

[0008] There are many options for coating polymer materials, such as polyvinyl alcohol (PVA), polycaprolactone (PCL), polylactic acid (PLA), polylactic-co-glycolic acid (PLGA) and polyglycolide (PGA). These materials have good biocompatibility and are basically non-toxic when degraded in the body.

[0009] The coating polymer system selected is poly-L-lactic acid (PLLA) and thermoplastic polyurethane (TPU). PLLA exhibits excellent degradation properties, while TPU exhibits good toughness and ductility. The addition of TPU not only ensures the coating's biodegradability but also provides the coating with elastic mechanical properties compatible with self-expanding metal stents. Both PLLA and TPU exhibit excellent biocompatibility and have widespread applications in drug delivery. The hydrophilic coating and the addition of an antimicrobial agent effectively prevent protein contact with the material, reducing protein adsorption, thereby preventing bacterial biofilm formation and reducing biliary sludge accumulation.

[0010] Currently, the research on biliary stents using traditional non-degradable bare stents is very mature, but the disadvantages and shortcomings of bare stents are also obvious. There is increasing attention on the research on surface-modified biliary stents, such as biodegradable biliary stents, functional coatings, tissue engineering and 3D printed stents, including the prospects for epithelialization-promoting coated stents, multifunctional coated stents, biodegradable shape memory stents and 4D bioprinting.

[0011] Patent document CN109701092A discloses a biodegradable (P3 / 4HBPCL) PU medical biliary stent. The materials include: polytetramethylene glycol (PTMG), hydroxyl-terminated polylactic acid (PLAOH), hexamethylene diisocyanate (HDI), poly-ε-caprolactone (PCL), poly-3-hydroxybutyrate-4-hydroxybutyrate (P3 / 4HB), paclitaxel, lactic acid, non-toxic catalyst, dichloromethane, catalyst and chain extender. The method includes: (1) removing impurities from lactic acid; (2) synthesizing hydroxyl-terminated polylactic acid by direct polycondensation; (3) synthesizing a biodegradable polyurethane medical elastomer; and (4) preparing a biodegradable (P3 / 4HBPCL) PU medical biliary stent material. A degradable TAXOL (P3 / 4HBPCL) PU nanobiliary stent is formed by a coaxial electrospinning process assisted by a special soluble mandrel. The stent has biocompatibility, safety, controlled drug release and the effect of preventing benign biliary stricture. However, this invention mainly forms the stent by modifying and synthesizing the materials and designing the process, and the preparation process is relatively complicated.

[0012] Chinese patent publication CN119524218A discloses a method for preparing a drug-coated stent. The method comprises preparing a NiTi alloy shape memory stent, preparing a PMEA-based drug-coated suspension slurry, and then applying the PMEA-based drug-coated suspension slurry to the outer surface of the NiTi alloy shape memory stent using liquid flame spraying to produce the drug-coated stent. Using liquid flame spraying technology to modify the stent surface, the resulting drug-coated stent exhibits excellent resistance to protein adsorption and can be used clinically to treat hepatobiliary diseases.

[0013] Given the rapid increase in biliary diseases, the research on anti-biliary sludge drug-coated biliary stents is of great significance: it can improve the treatment effect of diseases such as cholangiocarcinoma, especially the control and treatment of tumor recurrence; relieve patients' pain, prolong their survival, and improve their quality of life; promote the application of new drug coating materials, and provide a new direction for the development of stent technology. Summary of the Invention

[0014] In order to solve the above technical problems, the present invention provides a multifunctional anti-stenosis drug-coated stent, which includes a nickel-titanium stent, a PLLA-TPU-based drug-loaded coating on the outer surface of the stent, and a drug-loaded membrane on the inner surface of the stent. It can exert dual effects of anti-tumor and anti-biliary sludge accumulation, and is suitable for the treatment of biliary stenosis diseases.

[0015] A multifunctional anti-stenosis drug-coated stent comprises a nickel-titanium stent, the outer surface of which is coated with a PLLA-TPU-based drug-loaded coating, and the inner surface of which is adhered with a drug-loaded film, wherein the drug-loaded film is sequentially composed of an anti-tumor film, a TPU film and an anti-bile mud film starting from the inner surface of the nickel-titanium stent.

[0016] The multifunctional anti-stenosis drug-coated stent prepared by the present invention has a nickel-titanium stent body with good biocompatibility. The outer surface of the stent is coated with a PLLA-TPU-based drug-loaded coating suspension slurry, which has good degradation performance and adjustable mechanical properties. The release of the drug can effectively inhibit the inward or outward excessive growth of tumor cells. At the same time, the drug-loaded film on the inner layer of the nickel-titanium stent is divided into three layers, which are an anti-tumor membrane, a TPU membrane and an anti-bile mud membrane from the inner surface of the nickel-titanium stent. Tumor cells can be inhibited by the anti-tumor membrane. At the same time, the anti-bile mud membrane can inhibit the production of bacterial biofilms through antibacterial agents, thereby preventing bile duct restenosis caused by bile mud accumulation, and can be used to treat bile duct stenosis diseases.

[0017] Preferably, the PLLA-TPU-based drug-loaded coating is loaded with anti-tumor drug I, and the anti-tumor membrane is loaded with anti-tumor drug II, and the anti-tumor drugs I and II are at least one of cisplatin, carboplatin, oxaliplatin, 5-fluorouracil, methotrexate, doxorubicin, paclitaxel, sorafenib or gemcitabine.

[0018] In the present invention, the PLLA-TPU-based drug-loaded coating on the outer surface of the nickel-titanium stent is loaded with anti-tumor drug I, which can effectively inhibit the inward or outward excessive growth of tumor cells through the release of the drug. The inner surface is attached with an anti-tumor membrane loaded with anti-tumor drug II, which can also synergistically exert an anti-tumor effect.

[0019] Further preferably, the loading amount of the anti-tumor drug I in the PLLA-TPU-based drug-loaded coating is 0.2-5%, and the loading amount of the anti-tumor drug II in the anti-tumor membrane is 0.9-10%.

[0020] In a specific embodiment of the present invention, the anti-tumor drugs I and II are both 5-fluorouracil (5-FU). 5-FU is a pyrimidine antimetabolite drug, and its active metabolite can inhibit thymidylate synthase (TS) and block DNA synthesis; it can also be incorporated into RNA to interfere with the normal function of RNA. The double-strike mechanism enables it to effectively inhibit rapidly proliferating tumor cells, and it has broad-spectrum anti-cancer activity, pharmacokinetic characteristics suitable for local controlled release and mature formulation development, and is low in price.

[0021] Preferably, the anti-bile mud membrane is loaded with an antibacterial agent, and the antibacterial agent is at least one of benzalkonium chloride, chlorhexidine, polymyxin B, vancomycin, gentamicin, erythromycin or levofloxacin.

[0022] Further preferably, the loading amount of the antibacterial agent in the anti-bile mud membrane is 0.9-10%.

[0023] In a specific embodiment of the present invention, the antibacterial agent is chlorhexidine. As a broad-spectrum antibacterial agent, chlorhexidine is widely used in the fields of medical devices, surface modification of biomaterials, etc. due to its high efficiency, safety, and long-lasting effect.

[0024] Preferably, the thickness of the anti-tumor membrane, TPU membrane and anti-bile mud membrane are all 10-30 μm.

[0025] The present invention also provides a method for preparing the multifunctional anti-stenosis drug-coated stent, comprising the following steps: (1) Preparation of drug-loaded film; (2) Preparation of PLLA-TPU based drug-loaded coating suspension slurry: Under the action of a catalyst, amine monomer, aldehyde monomer and anti-tumor drug are placed in acetonitrile to undergo Schiff base reaction to obtain drug-loaded COF suspension, which is then centrifuged, washed and dried to obtain drug-loaded COF powder. The drug-loaded COF powder is dispersed in an oil phase and then added to a mixed solution of PLLA and TPU for ultrasonic treatment to obtain a PLLA-TPU based drug-loaded coating suspension slurry; (3) coating the PLLA-TPU-based drug-loaded coating suspension slurry obtained in step (2) on the outer surface of the nickel-titanium stent by liquid flame spraying to obtain a nickel-titanium stent loaded with the PLLA-TPU-based drug-loaded coating; (4) The drug-loaded film obtained in step (1) is laminated onto the inner surface of the nickel-titanium stent obtained in step (3) to obtain a multifunctional anti-stenosis drug-coated stent.

[0026] This invention utilizes liquid flame spraying technology to modify the stent surface. A PLLA-TPU-based drug-loaded coating suspension is applied to the outer surface of a nickel-titanium stent via liquid flame spraying. The drug-loaded coating is then attached to the inner surface of the nickel-titanium stent via lamination, resulting in a multifunctional, anti-stenosis drug-coated stent. Compared to coatings prepared using traditional coating techniques, thermal spray coatings offer advantages such as improved coating uniformity, superior coating adhesion, and a simpler method. Furthermore, the present invention's simple preparation method and short production cycle offer significant advantages in clinical applications.

[0027] Preferably, the method for preparing the drug-loaded film comprises the following steps: S1, preparing TPU film by TPU granules through a casting machine; S2. The mixed solution of the antitumor drug II and the organic substance I, and the mixed solution of the antibacterial agent and the organic substance II are respectively prepared into an antitumor film and an anti-bile mud film by a casting machine.

[0028] More preferably, in step S1, the operating temperature of the casting machine is 180-200° C., and the pressure is 0.5-2 MPa.

[0029] Further preferably, in step S2, the organic substance I is hydroxypropyl methylcellulose, polyethylene glycol or polyvinyl alcohol, and the organic substance II is thermoplastic polyurethane or polycaprolactone.

[0030] Further preferably, in step S2, the mass ratio of the antitumor drug II to the organic matter I in the antitumor membrane is 1-10:100, and the mass ratio of the antibacterial agent to the organic matter II in the anti-bile mud membrane is 1-10:100.

[0031] More preferably, in step S2, the operating temperature of the casting machine is 40-80° C., the pressure is 0.5-2 MPa, and the time is 5-20 min.

[0032] Preferably, in step (2), the amine monomer and aldehyde monomer are dehydrated and dried before use.

[0033] In the present invention, the amine monomer and the aldehyde monomer must be dehydrated and dried before use, otherwise the Schiff base reaction will be affected, the yield of the drug-loaded COF powder will be reduced, and the structure of the drug-loaded COF powder will be changed.

[0034] Preferably, in step (2), the amine monomer is at least one of 1,3,5-tris(4-aminophenyl)benzene, p-phenylenediamine, 2,5-diamino-p-benzoquinone, 1,3,5-triazine-2,4,6-triamine, 4,4'-diaminobiphenyl, 2,6-diaminopyridine, 2,5-diaminothiophene, 2,4-diaminoimidazole or 5,10,15,20-tetrakis(aminophenyl)porphyrin; and the aldehyde monomer is at least one of terephthalaldehyde, trimesic acid, 2,5-dihydroxy-1,4-benzoquinone, 1,3,5-trialdehyde-1,3,5-triazine, 1,2,4,5-tetraaldehyde benzoquinone, 2,5-diformylthiophene, 2,6-diformylpyridine or 9,9-diformylfluorene.

[0035] In the present invention, an amine monomer is used as a nucleophilic reagent. The nitrogen atom with a lone electron pair in the amine monomer structure attacks the positively charged carbon atom on the carbonyl group in the aldehyde monomer, causing a nucleophilic addition reaction to form an intermediate α-hydroxyamine compound, which is then further dehydrated to form a Schiff base (i.e., COF powder), and at the same time, the anti-tumor drug I is in situ loaded on the COF powder.

[0036] Preferably, in step (2), the molar ratio of the amine monomer, the aldehyde monomer and the anti-tumor drug I is 0.67~1:1:1~4.

[0037] Preferably, in step (2), the catalyst is acetic acid, and the volume of the acetic acid is 1% to 20% of the volume of acetonitrile.

[0038] More preferably, the catalyst is added at a rate of 0.1 to 5 mL / min.

[0039] Preferably, in step (2), the Schiff base reaction time is 1 to 24 h.

[0040] In the present invention, if the reaction time is less than 1 hour, the yield is low and the crystallinity of the product is poor; if the reaction time is greater than 24 hours, the reactants are prone to agglomeration or reversible decomposition with increasing moisture content, thereby reducing the yield.

[0041] Preferably, in step (2), the centrifugation, washing, and drying steps are as follows: centrifuging the drug-loaded COF suspension to obtain a drug-loaded COF colloid, adding a detergent to the drug-loaded COF colloid to remove impurities, and drying the remaining material to obtain a drug-loaded COF powder.

[0042] Further preferably, the detergent is acetonitrile or ethanol, the volume ratio of the drug-loaded COF colloid to the detergent is 1:10-40, and the number of washing times is 5-10 times.

[0043] In the present invention, since the residual amine monomer, aldehyde monomer and catalyst are all in the acetonitrile solution after the Schiff base reaction, detergent needs to be added multiple times for washing and removing impurities.

[0044] More preferably, the centrifugal speed is 8000~12000 r / min.

[0045] More preferably, the drying temperature is 80-120° C., and the drying time is 12-24 h.

[0046] Preferably, in step (2), the oil phase is at least one of petroleum ether, hexane, heptane, silicone oil or mineral oil.

[0047] Preferably, in step (2), the mass ratio of the drug-loaded COF powder to the oil phase is 1:10-100.

[0048] Preferably, in step (2), the mass ratio of PLLA to TPU is 1-10:10.

[0049] Preferably, in step (3), the parameters of the liquid flame spraying process are: O2 as the combustion-supporting gas, the pressure is 0.2~0.8 MPa, the flow rate is 1~4.0 Nm 3 / h; C2H2 is used as the fuel gas, the pressure is 0.05~0.3 MPa, and the flow rate is 0.5~2 Nm 3 / h.

[0050] Preferably, in step (3), during the liquid flame spraying process, the concentric rotation speed of the nickel-titanium stent is 50-200 r / min, the spraying distance of the flame spray gun from the outer surface of the stent is 200-500 mm, the moving speed is 300-1000 mm / s, the spraying angle is 45-135°, and the number of coating spraying times is 5-20 times.

[0051] Preferably, in step (4), the temperature of the lamination method is 80-120°C, the pressure is 0.5-2 MPa, and the time is 5-20 min.

[0052] Preferably, in step (4), the drug-loaded film is subjected to corona treatment at a voltage of 10-15 kV and a treatment time of 1-2 min; the nickel-titanium stent needs to be sandblasted with a sand particle size of 60-120 mesh.

[0053] The present invention also provides the use of the multifunctional anti-stenosis drug-coated stent in the treatment of biliary tract diseases. The drug-coated stent of the present invention has good biocompatibility, good degradation performance, and adjustable mechanical properties. The drug release can effectively inhibit the excessive inward or outward growth of tumor cells. At the same time, the drug-coated membrane on the inner layer of the nickel-titanium stent can effectively inhibit tumor cells and effectively prevent the accumulation of bile sludge. It can be used clinically to treat biliary tract stricture diseases and has good application prospects and economic benefits in the treatment of biliary tract diseases.

[0054] Compared with the prior art, the present invention has the following beneficial effects: (1) The multifunctional anti-stenosis drug-coated stent prepared by the present invention is a nickel-titanium stent with good biocompatibility, good degradation performance and adjustable mechanical properties. The outer surface of the nickel-titanium stent is coated with a PLLA-TPU-based drug-loaded coating loaded with anti-tumor drugs, and the inner surface of the nickel-titanium stent is attached with a three-layer drug-loaded membrane system, which can simultaneously play the dual role of anti-tumor and anti-biliary sludge accumulation. The release of the anti-tumor drug in the outer layer can effectively inhibit the excessive growth of tumor cells inward or outward. At the same time, the drug-loaded membrane in the inner layer of the nickel-titanium stent can effectively inhibit tumor cells and effectively prevent the accumulation of bile sludge. It can be used in the clinical treatment of biliary stenosis and has good application prospects and economic benefits in the treatment of biliary diseases.

[0055] (2) The COF powder prepared by the present invention has a high specific surface area and porosity, adjustable pore size, controllable drug release capability, good biocompatibility and excellent stability, and can release drugs through a pH-responsive mechanism in the weakly acidic tumor microenvironment.

[0056] (3) The present invention adopts liquid flame spraying technology to coat the prepared PLLA-TPU-based drug-loaded coating suspension slurry on the nickel-titanium stent to prepare a drug-loaded coating stent. Compared with the coating prepared by traditional coating technology, the coating can ensure a very high coating deposition efficiency, the coating has better coating uniformity, improves the bonding strength between the coating and the substrate, and can ensure that the drug can still maintain its original efficacy after spraying. BRIEF DESCRIPTION OF THE DRAWINGS

[0057] Figure 1 Schematic diagram of the structure of the multifunctional anti-stenosis drug-coated stent prepared in Examples 1 and 2 of the present invention.

[0058] Figure 2 This is the infrared spectrum of the 5-FU@RT-COF powder prepared in Example 1.

[0059] Figure 3 This is the XRD pattern of 5-FU@RT-COF powder prepared in Example 1.

[0060] Figure 4 This is a microscopic morphology of the 5-FU@RT-COF powder prepared in Example 1.

[0061] Figure 5 The standard curve of 5-FU concentration and absorbance is shown in Figure 2.

[0062] Figure 6 This is a graph showing the toxicity test of 5-FU@RT-COF powder prepared in Example 1 on human hepatobiliary cancer cells.

[0063] Figure 7 These are microscopic images of the outer surfaces of the multifunctional anti-stenosis drug-coated stents prepared in Examples 1 and 2 and the non-drug-loaded nickel-titanium coated stent prepared in Comparative Example 1, where a to c are microscopic images of the outer surfaces of the multifunctional anti-stenosis drug-coated stents prepared in Examples 1 and 2 and the non-drug-loaded nickel-titanium coated stent prepared in Comparative Example 1, respectively.

[0064] Figure 8 These are cross-sectional SEM images of the multifunctional anti-stenosis drug-coated stent prepared in Example 1 and the non-drug-loaded nickel-titanium coated stent prepared in Comparative Example 1, where a and b are cross-sectional SEM images of the multifunctional anti-stenosis drug-coated stent prepared in Example 1 and the non-drug-loaded nickel-titanium coated stent prepared in Comparative Example 1, respectively.

[0065] Figure 9 The 5-FU release curves of the multifunctional anti-stenosis drug-coated stent prepared in Example 2 in PBS with pH=5.5 and PBS with pH=7.4 are shown.

[0066] Figure 10 Graphs showing the inhibitory effects of the multifunctional anti-stenosis drug-coated stents prepared in Examples 1 and 2, the non-drug-loaded nickel-titanium coated stent prepared in Comparative Example 1, and the bare nickel-titanium stent prepared in Comparative Example 2 on human hepatobiliary cancer cells.

[0067] Figure 11 These are scanning electron micrographs of the multifunctional anti-stenosis drug-coated stent prepared in Example 2, the non-drug-loaded nickel-titanium-coated stent prepared in Comparative Example 1, and the bare nickel-titanium stent prepared in Comparative Example 2 after the human cholangiocarcinoma cell attachment experiment. Figures a to c are scanning electron micrographs of the multifunctional anti-stenosis drug-coated stent prepared in Example 2, the non-drug-loaded nickel-titanium-coated stent prepared in Comparative Example 1, and the bare nickel-titanium stent prepared in Comparative Example 2, respectively.

[0068] Figure 12These are the anti-Escherichia coli result graphs of the multifunctional anti-stenosis drug-coated stent prepared in Example 1, the multifunctional anti-stenosis drug-coated stent prepared in Example 2, and the drug-free nickel-titanium coated stent prepared in Comparative Example 3, where a to c are the antibacterial result graphs of the multifunctional anti-stenosis drug-coated stent prepared in Example 1, the multifunctional anti-stenosis drug-coated stent prepared in Example 2, and the drug-free nickel-titanium coated stent prepared in Comparative Example 3, respectively. DETAILED DESCRIPTION

[0069] The present invention will be further described in detail below with reference to the examples, but the embodiments of the present invention are not limited to the following examples.

[0070] The raw materials used in the present invention are all commercially available.

[0071] Example 1 (1) Preparation of TPU film: Weigh 50 g of TPU pellets and use a casting machine to prepare a 20 μm thick TPU film at a temperature of 180 °C and a pressure of 1 MPa.

[0072] (2) Preparation of antitumor membrane and antibile mud membrane: Weigh 5 g of polycaprolactone powder and dissolve it in 50 mL of DMF, then add 0.5 g of 5-FU and disperse it evenly to prepare an antitumor solution; weigh 5 g of polyvinyl alcohol powder and dissolve it in 50 mL of water, then add 0.5 g of chlorhexidine and disperse it evenly to prepare an antibacterial solution; use a casting machine at a temperature of 60 °C and a pressure of 1 MPa to prepare 20 μm thick antitumor membrane and antibile mud membrane, respectively.

[0073] (3) Preparation of PLLA-TPU based drug loading coating suspension slurry: Weigh 1.125 g of 1,3,5-tris(4-aminophenyl)benzene (TAPB) and 0.5189 g of trimesaldehyde (Tf) into a beaker, dry at 60 °C for 12 h, then add 200 mL of acetonitrile to dissolve, mix evenly after dissolution, and add 2 g of 5-FU at the same time. Add 40 mL of acetic acid at a rate of 4 mL / min, react at room temperature and 800 r / min for 12 h to obtain 5-FU@RT-COF suspension; centrifuge at 10000 r / min, wash 5 times with 20 mL of acetonitrile at 10000 r / min, dry in an oven at 80 °C for 24 h to obtain 5-FU@RT-COF powder; weigh 5 g of PLLA and dissolve it in 50 mL of THF, weigh 5 g of TPU and dissolve it in 50 mL of DMF, stir and mix evenly, weigh 0.5 g of 5-FU@RT-COF powder was added thereto, stirred and mixed evenly, 10 g of petroleum ether was added, the ultrasonic power was 60 W, the ultrasonic time was 10 min, and an ice bath was placed to obtain a PLLA-TPU-based drug-loaded coating suspension slurry.

[0074] (4) The PLLA-TPU-based drug-loaded coating suspension obtained in step (3) was evenly sprayed on the outer surface of the stent by liquid flame spraying. The spraying flow rate was 40 mL / min, the compressed air pressure was 0.2 MPa, and O2 was used as the combustion aid at a pressure of 0.4 MPa and a flow rate of 3 Nm 3 / h; C2H2 is used as the fuel gas, the pressure is 0.07 MPa, and the flow rate is 1 Nm 3 / h; the concentric rotation speed of the stent was 100 r / min, the spraying distance between the flame spray gun and the outer surface of the stent was 300 mm, the moving speed was 500 mm / s, the spraying angle was 120°, and the coating spraying times were 20 times, thereby obtaining a nickel-titanium stent loaded with a PLLA-TPU-based drug-loaded coating.

[0075] (5) The TPU film obtained in step (1), the anti-tumor film and the anti-biliary mud film obtained in step (2) were corona treated at a voltage of 10 kV for 2 min; the nickel-titanium stent loaded with the PLLA-TPU-based drug-loaded coating was sandblasted with a sand particle size of 120 mesh; under the conditions of a temperature of 80 °C, a pressure of 1 MPa and a pressing time of 10 min, the three layers of film were pressed onto the inner surface of the nickel-titanium stent loaded with the PLLA-TPU-based drug-loaded coating in the order of the anti-tumor film, the TPU film and the anti-biliary mud film to obtain a multifunctional anti-stenosis drug-coated stent, the structural schematic of which is shown in FIG. Figure 1 shown.

[0076] Example 2 The preparation method is the same as that of Example 1, except that in step (3), the amount of 5-FU@RT-COF powder added to the PLLA-TPU-based drug-loaded coating suspension slurry is 1 g.

[0077] Comparative Example 1 The preparation method is the same as that in Example 1, except that no drug is added to the inner and outer coatings of the nickel-titanium stent. The specific steps are as follows: (1) Weigh 50 g of TPU pellets and use a casting machine to prepare a 20 μm thick TPU film at a temperature of 180 °C and a pressure of 1 MPa.

[0078] (2) Weigh 5 g of polycaprolactone powder and dissolve it in 50 mL of DMF to prepare a polycaprolactone solution; weigh 5 g of polyvinyl alcohol powder and dissolve it in 50 mL of water to prepare a polyvinyl alcohol solution; use a casting machine at a temperature of 60 °C and a pressure of 1 MPa to prepare a 20 μm thick polycaprolactone film and a polyvinyl alcohol film, respectively.

[0079] (3) Weigh 5 g of PLLA and dissolve it in 50 mL of THF, weigh 5 g of TPU and dissolve it in 50 mL of DMF, stir and mix evenly, add 10 g of petroleum ether, ultrasonic power is 60 W, ultrasonic time is 10 min, ice bath, and obtain PLLA-TPU coating suspension slurry.

[0080] (4) Liquid flame spraying was used to evenly spray the PLLA-TPU coating suspension slurry on the outer surface of the bracket. The spraying flow rate was 40 mL / min and the compressed air pressure was 0.2 MPa. O2 was used as the auxiliary gas with a pressure of 0.4 MPa and a flow rate of 3 Nm 3 / h; C2H2 is used as the fuel gas, the pressure is 0.07 MPa, and the flow rate is 1 Nm 3 / h; the concentric rotation speed of the stent was 100 r / min, the spraying distance between the flame spray gun and the outer surface of the stent was 300 mm, the moving speed was 500 mm / s, the spraying angle was 120°, and the coating spraying times was 20 times, thus obtaining a nickel-titanium stent loaded with PLLA-TPU coating.

[0081] (5) The TPU film, polyvinyl alcohol film and polycaprolactone film obtained in steps (1) and (2) were corona treated at a voltage of 10 kV for a treatment time of 2 min; the nickel-titanium stent loaded with PLLA-TPU coating was sandblasted with a sand particle size of 120 mesh; under the conditions of a temperature of 80 °C, a pressure of 1 MPa and a pressing time of 10 min, the three layers of film were pressed onto the inner surface of the sprayed nickel-titanium stent loaded with PLLA-TPU coating in the order of polycaprolactone film, TPU film and polyvinyl alcohol film to obtain a non-drug-loaded nickel-titanium coated stent.

[0082] Comparative Example 2 Bare nickel-titanium stent without any treatment.

[0083] Comparative Example 3 The preparation method is the same as that in Example 1, except that only the drug is added to the outer coating of the nickel-titanium stent. The specific steps are as follows: (1) Weigh 50 g of TPU pellets and use a casting machine to prepare a 20 μm thick TPU film at a temperature of 180 °C and a pressure of 1 MPa.

[0084] (2) Weigh 5 g of polycaprolactone powder and dissolve it in 50 mL of DMF to prepare a polycaprolactone solution; weigh 5 g of polyvinyl alcohol powder and dissolve it in 50 mL of water to prepare a polyvinyl alcohol solution; use a casting machine at a temperature of 60 °C and a pressure of 1 MPa to prepare a 20 μm thick polycaprolactone film and a polyvinyl alcohol film, respectively.

[0085] (3) Preparation of PLLA-TPU based drug loading coating suspension slurry: 1.125 g 1,3,5-tris(4-aminophenyl)benzene (TAPB) and 0.5189 g trimesaldehyde (Tf) were weighed and placed in a beaker, dried at 60 °C for 12 h, and then dissolved in 200 mL acetonitrile. After dissolution, the mixture was mixed evenly. At the same time, 2 g 5-FU was added, and 40 mL acetic acid was added at a rate of 4 mL / min. The mixture was reacted at room temperature and a speed of 800 r / min for 12 h to obtain a 5-FU@RT-COF suspension; centrifuged at 10,000 r / min, washed 5 times with 20 mL acetonitrile at 10,000 r / min, and dried in an oven at 80 °C for 24 h to obtain a 5-FU@RT-COF powder; 5 g PLLA was weighed and dissolved in 50 mL THF, 5 g TPU was weighed and dissolved in 50 mL DMF, and the mixture was stirred evenly. 0.5 g of 5-FU@RT-COF powder was added thereto, stirred and mixed evenly, 10 g of petroleum ether was added, the ultrasonic power was 60 W, the ultrasonic time was 10 min, and an ice bath was placed to obtain a PLLA-TPU-based drug-loaded coating suspension slurry.

[0086] (4) Liquid flame spraying was used to evenly spray the PLLA-TPU coating suspension slurry on the outer surface of the bracket. The spraying flow rate was 40 mL / min and the compressed air pressure was 0.2 MPa. O2 was used as the auxiliary gas with a pressure of 0.4 MPa and a flow rate of 3 Nm 3 / h; C2H2 is used as the fuel gas, the pressure is 0.07 MPa, and the flow rate is 1 Nm 3 / h; the concentric rotation speed of the stent was 100 r / min, the spraying distance between the flame spray gun and the outer surface of the stent was 300 mm, the moving speed was 500 mm / s, the spraying angle was 120°, and the coating spraying times was 20 times, thus obtaining a nickel-titanium stent loaded with PLLA-TPU coating.

[0087] (5) The TPU film, polyvinyl alcohol film and polycaprolactone film obtained in steps (1) and (2) were corona treated at a voltage of 10 kV for a treatment time of 2 min; the nickel-titanium stent loaded with PLLA-TPU coating was sandblasted with a sand particle size of 120 mesh; under the conditions of a temperature of 80 °C, a pressure of 1 MPa and a pressing time of 10 min, the three layers of film were pressed onto the inner surface of the sprayed nickel-titanium stent loaded with PLLA-TPU coating in the order of polycaprolactone film, TPU film and polyvinyl alcohol film to obtain a nickel-titanium coated stent without drug film.

[0088] Sample analysis 1. Detection and Analysis of 5-FU@RT-COF and RT-COF (1) Infrared and XRD detection The 5-FU@RT-COF powder, COF powder, and 5-FU powder synthesized in Example 1 were mixed with potassium bromide at a ratio of 1:100, ground, dried, and pressed into tablets. FT-IR analysis was performed to verify the successful synthesis of 5-FU@RT-COF and COF, and XRD detection was used to verify the successful synthesis and crystallinity of COF.

[0089] Figure 2 and Figure 3 The infrared spectra of 5-FU@RT-COF powder, COF powder and 5-FU powder prepared in Example 1 and the XRD patterns of COF are shown respectively. The FT-IR spectrum confirms that COF is successfully synthesized. The C=N bond is at 1623 cm -1 Stretching vibration at 1505cm -1 The C=C bond at 1223 cm is derived from the vibration of the aromatic ring skeleton. In addition, the XRD spectrum shows a characteristic crystal peak at 2θ=5.7°, which corresponds to the (100) crystal plane, further verifying the effectiveness of the synthesis. -1 The appearance of proves that 5-FU@RT-COF powder was successfully prepared.

[0090] (2) Microscopic morphology observation The COF powder prepared in Example 1 was spread on the conductive adhesive and subjected to gold spraying treatment. The surface micromorphology was observed using SEM.

[0091] Figure 4 This is a microscopic morphology of the COF powder prepared in Example 1, wherein the size of the COF powder is about 1 micron, the size distribution is relatively uniform, and the particles are relatively monodisperse.

[0092] (3) Loading amount of 5-FU in 5-FU@RT-COF powder By drawing a standard curve of 5-FU (such as Figure 5 The 5-FU loading capacity in the 5-FU@RT-COF powder was calculated by measuring the 5-FU concentration in the supernatant using a UV spectrophotometer (as shown). The loading capacity (in g / g) was calculated as follows:

[0093] The test showed that the concentration of 5-FU in the supernatant was 9.8426 mg / mL. The calculation showed that the loading amount of 5-FU in the 5-FU@RT-COF powder prepared in Example 1 was 0.431 g / g.

[0094] (4) Cytotoxicity evaluation of RT-COF powder The toxicity of 5-FU@RT-COF powder on human hepatobiliary carcinoma cells (RBE cells, purchased from Wuhan Pronocell Life Science Co., Ltd., catalog number CL-0191) was evaluated using a CCK8 detection kit.

[0095] Figure 6 The figure shows the toxicity test of the RT-COF powder prepared in Example 1 on human cholangiocarcinoma cells. As shown in the figure, the RT-COF powder prepared in Example 1 has good biocompatibility. When the concentration of the RT-COF powder reaches 1 mg / mL, the inhibition rate of the RT-COF powder on cholangiocarcinoma cells is only 20%.

[0096] 2. Multifunctional Anti-Stenosis Drug-Emulsified Stent Testing (1) Micromorphology test of PLLA-TPU based drug-loaded coating The multifunctional anti-stenosis drug-coated stents prepared in Examples 1 and 2 and the non-drug-loaded nickel-titanium coated stent prepared in Comparative Example 1 were subjected to gold spraying treatment, and their outer surface micromorphologies were observed using SEM.

[0097] Figure 7 Figures a through c show the outer surface micromorphologies of the multifunctional anti-stenotic drug-coated stents prepared in Examples 1 and 2 and the undrug-loaded nickel-titanium-coated stent prepared in Comparative Example 1. As shown in the figures, the outer surfaces of the multifunctional anti-stenotic drug-coated stents prepared in Examples 1 and 2 exhibit a well-melted state, with uniform distribution of 5-FU@RT-COF powder on the coating surface.

[0098] (2) PLLA-TPU based drug-loaded coating thickness test The multifunctional anti-stenosis drug-coated stent prepared in Example 1 and the non-drug-loaded nickel-titanium coated stent prepared in Comparative Example 1 were cut, inlaid, and polished, and then subjected to gold spraying. The coating thickness of the outer surface was measured using SEM.

[0099] Figure 8 These are cross-sectional SEM images of the multifunctional anti-stenosis drug-coated stent prepared in Example 1 and the un-drug-loaded nickel-titanium-coated stent prepared in Comparative Example 1. Figures a and b are cross-sectional SEM images of the multifunctional anti-stenosis drug-coated stent prepared in Example 1 and the un-drug-loaded nickel-titanium-coated stent prepared in Comparative Example 1, respectively. As shown in the figures, Example 1 and Comparative Example 1 produced relatively uniform coatings prepared by flame spraying a suspension material. The coatings were dense, had a small amount of pores, and were approximately 100 μm thick.

[0100] (3) Drug release assay of multifunctional anti-stenosis drug-coated stents Drug release experiments were conducted on the multifunctional anti-stenosis drug-coated stent prepared in Example 2 in PBS with pH=5.5 and PBS with pH=7.4.

[0101] Figure 9 The following are the 5-FU release curves for the multifunctional anti-stenosis drug-coated stent prepared in Example 2 in PBS at pH 5.5 and PBS at pH 7.4. As shown, the multifunctional anti-stenosis drug-coated stent prepared in Example 2 released 5-FU faster in PBS at pH 5.5 than in PBS at pH 7.4, with an overall faster release rate over a 14-day period. This is attributed to the protonated positive charge of COF and 5-FU in an acidic environment, which accelerates 5-FU release, as well as the rapid degradation and molecular conformational expansion of the organic polymer matrix in an acidic environment. This demonstrates the successful implementation of the pH-responsive mechanism of the present invention.

[0102] (4) Evaluation of the tumor cell inhibition effect of multifunctional anti-stenosis drug-eluting stents The multifunctional anti-stenosis drug-coated stents prepared in Examples 1 and 2, the non-drug-loaded nickel-titanium coated stent prepared in Comparative Example 1, the bare nickel-titanium stent prepared in Comparative Example 2, and the drug-free nickel-titanium coated stent prepared in Comparative Example 3 were co-cultured with human hepatobiliary cancer cells for three days and detected using a CCK8 kit.

[0103] Figure 10 Figures show the inhibitory effects of the multifunctional anti-stenosis drug-coated stents prepared in Examples 1 and 2, the non-drug-loaded nickel-titanium coated stent prepared in Comparative Example 1, the bare nickel-titanium stent prepared in Comparative Example 2, and the drug-free nickel-titanium coated stent prepared in Comparative Example 3 on human cholangiocarcinoma cells. As shown in the figure, the inhibition rates of Example 1 on human cholangiocarcinoma cells during 1 day, 2 days, and 3 days of co-culture were 51.5%, 75.5%, and 90.6%, respectively. The inhibition rates of Example 2 on human cholangiocarcinoma cells during 1 day, 2 days, and 3 days of co-culture were 58.4%, 84.1%, and 94.4%, respectively, showing better inhibitory effects on human cholangiocarcinoma cells than Example 1, which is due to the increased drug content. The inhibition rates of comparative example 1 on human cholangiocarcinoma cells during co-culture for 1 day, 2 days and 3 days were 6.6%, 5.2% and 3.3%, respectively. The inhibition rates of comparative example 3 on human cholangiocarcinoma cells during co-culture for 1 day, 2 days and 3 days were 28.3%, 46.5% and 68.4%, respectively. The coating of comparative example 1 showed a slightly better inhibitory effect on human cholangiocarcinoma cells than the bare nickel-titanium stent of comparative example 2. This may be related to the fact that the degradation products of the coating polymer have a certain inhibitory effect on human cholangiocarcinoma cells. Comparative example 3 showed an inhibitory effect on human cholangiocarcinoma cells lower than that of Example 1, which shows that the anti-tumor effect of the inner drug-loaded film is very important.

[0104] (5) Observation of human hepatobiliary cancer cells attached to multifunctional anti-stenosis drug-coated stents The cells attached to and grown on the coating surface were detected using SEM. The specific steps were as follows: Example 2 and Comparative Examples 1 and 2 were sterilized by UV for 2 h and cells were inoculated on the surface of the material at a density of 2000 cells / cm 2 The human cholangiocarcinoma cells were grown in a cell culture incubator environment at 37 °C, 100% humidity, and 5% carbon dioxide content. The human cholangiocarcinoma cells grown on the coating surface for 3 days were washed three times with PBS, dehydrated with 25% ethanol solution for 5 minutes, 50% ethanol solution for 5 minutes, 75% ethanol solution for 5 minutes, 90% ethanol solution for 5 minutes, and 100% ethanol solution for 10 minutes, and then fixed with 4% paraformaldehyde for 4 hours and finally dried at 37 °C for 12 hours.

[0105] Figure 11 The following are scanning electron micrographs of the multifunctional anti-stenosis drug-coated stent prepared in Example 2, the non-drug-loaded nickel-titanium-coated stent prepared in Comparative Example 1, and the bare nickel-titanium stent prepared in Comparative Example 2 after human hepatobiliary cancer cell attachment experiments. Figures a to c are scanning electron micrographs of the multifunctional anti-stenosis drug-coated stent prepared in Example 2, the non-drug-loaded nickel-titanium-coated stent prepared in Comparative Example 1, and the bare nickel-titanium stent prepared in Comparative Example 2, respectively. As shown in the figure, there is essentially no cell attachment on the surface of Example 2, showing good anti-cell attachment properties ( Figure 11 The surfaces of Comparative Examples 1 and 2 also had almost no cell adhesion, showing good anti-cell adhesion properties ( Figure 11 The anti-adhesion property of 2 in Figures b and c) may be due to the fact that cells cannot attach well to this surface.

[0106] (6) Evaluation of the antibacterial effect of multifunctional anti-stenosis drug-eluting stents The antibacterial properties of the stents were determined using the standard plate count method. The specific steps were as follows: the nickel-titanium stents of Examples 1, 2, and Comparative Example 3 were placed in a 6-well plate and sterilized with UV for 30 minutes, and 3 ml of a 1×10 6 The Escherichia coli culture solution with CFU / mL was co-cultured in a 37°C incubator for 24 hours. 100 μL of the culture solution was spread on solid culture medium for 16 hours, and then photos were taken and the data recorded.

[0107] Figure 12Figures 1 and 2 show the antibacterial performance of the multifunctional antistenotic drug-coated stent prepared in Example 1, the multifunctional antistenotic drug-coated stent prepared in Example 2, and the drug-free nickel-titanium-coated stent prepared in Comparative Example 3. Figures a through c show the antibacterial performance of the multifunctional antistenotic drug-coated stent prepared in Example 1, the multifunctional antistenotic drug-coated stent prepared in Example 2, and the drug-free nickel-titanium-coated stent prepared in Comparative Example 3, respectively. As shown in the figures, Examples 1 and 2 showed excellent antibacterial performance, while Comparative Example 3, lacking an inner antibacterial layer, exhibited poor antibacterial performance, demonstrating the necessity of an inner antibacterial layer.

[0108] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art will be able to modify the technical solutions described in the aforementioned embodiments or replace some of the technical features therein with equivalents. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention shall be included within the scope of protection of the present invention.

Claims

1. A multifunctional anti-stenosis drug-coated stent, comprising a nickel-titanium stent, the outer surface of which is coated with a PLLA-TPU-based drug-loaded coating, characterized in that: The inner surface of the nickel-titanium stent is attached with a drug-loaded film, and the drug-loaded film is an anti-tumor film, a TPU film and an anti-bile mud film in order from the inner surface of the nickel-titanium stent.

2. The multifunctional anti-stenosis drug-eluting stent according to claim 1, characterized in that: The PLLA-TPU-based drug-loaded coating is loaded with anti-tumor drug I, and the anti-tumor membrane is loaded with anti-tumor drug II. The anti-tumor drugs I and II are at least one of cisplatin, carboplatin, oxaliplatin, 5-fluorouracil, methotrexate, doxorubicin, paclitaxel, sorafenib or gemcitabine.

3. The multifunctional anti-stenosis drug-eluting stent according to claim 2, characterized in that: The loading amount of the anti-tumor drug I in the PLLA-TPU-based drug-loaded coating is 0.2-5%; the loading amount of the anti-tumor drug II in the anti-tumor membrane is 0.9-10%.

4. The multifunctional anti-stenosis drug-eluting stent according to claim 1, characterized in that: The anti-bile mud membrane is loaded with an antibacterial agent, and the antibacterial agent is at least one of benzalkonium chloride, chlorhexidine, polymyxin B, vancomycin, gentamicin, erythromycin or levofloxacin.

5. The multifunctional anti-stenosis drug-eluting stent according to claim 4, characterized in that: The loading amount of the antibacterial agent in the anti-bile mud membrane is 0.9-10%.

6. The method for preparing a multifunctional anti-stenosis drug-eluting stent according to any one of claims 1 to 5, characterized in that: The following steps are involved: (1) Preparation of drug-loaded film; (2) Preparation of PLLA-TPU based drug-loaded coating suspension slurry: Under the action of a catalyst, amine monomer, aldehyde monomer and anti-tumor drug are placed in acetonitrile to undergo Schiff base reaction to obtain drug-loaded COF suspension, which is then centrifuged, washed and dried to obtain drug-loaded COF powder. The drug-loaded COF powder is dispersed in an oil phase and then added to a mixed solution of PLLA and TPU for ultrasonic treatment to obtain a PLLA-TPU based drug-loaded coating suspension slurry; (3) coating the PLLA-TPU-based drug-loaded coating suspension slurry obtained in step (2) on the outer surface of the nickel-titanium stent by liquid flame spraying to obtain a nickel-titanium stent loaded with the PLLA-TPU-based drug-loaded coating; (4) The drug-loaded film obtained in step (1) is laminated onto the inner surface of the nickel-titanium stent obtained in step (3) to obtain a multifunctional anti-stenosis drug-coated stent.

7. The method for preparing a multifunctional anti-stenosis drug-coated stent according to claim 6, characterized in that: In step (1), the method for preparing the drug-loaded film comprises the following steps: S1, preparing TPU film by TPU granules through a casting machine; S2. The mixed solution of the antitumor drug II and the organic substance I, and the mixed solution of the antibacterial agent and the organic substance II are respectively prepared into an antitumor film and an anti-bile mud film by a casting machine.

8. The multifunctional anti-stenosis drug-eluting stent according to claim 7, characterized in that: In step S2, the mass ratio of the antitumor drug II to the organic matter I in the antitumor membrane is 1-10:100, and the mass ratio of the antibacterial agent to the organic matter II in the anti-bile mud membrane is 1-10:

100.

9. The method for preparing a multifunctional anti-stenosis drug-coated stent according to claim 6, wherein: In step (4), the drug-loaded film is subjected to corona treatment at a voltage of 10-15 kV for 1-2 min; the nickel-titanium stent is subjected to sandblasting treatment with a sand particle size of 60-120 mesh.

10. Use of the multifunctional anti-stenosis drug-eluting stent according to any one of claims 1 to 5 in the treatment of biliary diseases.

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