A multi-component co-loaded nanoparticle for inhibiting paclitaxel crystallization and a preparation method thereof

Abstract

This invention discloses a multi-component co-loaded nanoparticle for inhibiting paclitaxel crystallization and its preparation method, belonging to the field of nanomedicine technology. The nanoparticles comprise a hydrophobic core and a hydrophilic crown surrounding it. The hydrophobic core contains paclitaxel at a mass ratio of 1% to 50%, and at least two other antitumor active components with conjugated structures (selected from anthracyclines, camptothecins, P-glycoprotein inhibitors, and coumarin compounds). Paclitaxel and other components form an amorphous complex through non-covalent interactions, ensuring its concentration in the core is below the critical concentration for crystallization. This invention utilizes the steric hindrance and physical co-intercalation between heterogeneous molecules to disrupt the continuous stacking of paclitaxel molecules from a thermodynamic perspective, achieving efficient steady-state loading and crystallization inhibition of paclitaxel without the need for highly toxic solubilizers. The nanoparticle preparation process is flexible, possesses intelligent stimulus-response release capabilities, and can achieve synergistic targeted therapy with multiple drugs, effectively reversing multidrug resistance in tumors.

Description

Technical Field

[0001] This invention relates to the interdisciplinary fields of biomedical engineering, pharmaceutics and nanomedicine, specifically to a multi-component co-loaded nanoparticle for inhibiting paclitaxel crystallization and its preparation method. Background Technology

[0002] Malignant tumors are one of the major diseases that seriously threaten human life and health. Chemotherapy, as a first-line treatment for solid tumors, still faces many challenges in clinical application. On the one hand, the high heterogeneity of tumor cells makes it difficult for a single drug to completely eliminate tumor lesions; on the other hand, long-term single-drug chemotherapy is prone to inducing multidrug resistance (MDR) in tumor cells, resulting in a significant decrease in the effective concentration of drugs in tumor tissues, which seriously restricts the efficacy of chemotherapy (Nature Reviews Clinical Oncology, 2018, 15(2): 81-94).

[0003] Lipid-based nanoparticles (LNPs) are considered ideal delivery carriers for multidrug co-loading due to their excellent biocompatibility and structural tunability. However, the stable encapsulation of paclitaxel in multidrug co-loading systems has long been a technical bottleneck in the field. Paclitaxel has a large, rigid hydrophobic framework and extremely low water solubility. Its molecules exhibit strong π-π stacking interactions, making it prone to self-aggregation and needle-like crystal formation after entering the hydrophobic core of nanoparticles. Studies have shown that even in the presence of surfactants, PTX molecules are still thermodynamically driven to crystallize (International Journal of Pharmaceutics, 2010, 390(2): 242-249). Once crystals precipitate, they not only destroy the structural integrity of the nanoparticles, leading to premature drug leakage, but also reduce the systemic drug loading and increase the risk of systemic toxicity. Molecular dynamics simulations further revealed that the compatibility between the drug and the carrier material directly affects the encapsulation stability of PTX, and that simple physical encapsulation is insufficient to maintain the long-term homogeneity of the system (Journal of Pharmaceutical Sciences, 2021, 110(6): 2483-2492). Therefore, how to break the intermolecular stacking of PTX from the physicochemical level without using highly toxic solubilizers has become a key problem that urgently needs to be solved in the field of pharmaceutics.

[0004] To address the problem of paclitaxel crystallization inhibition, researchers both domestically and internationally have conducted extensive research. One common strategy is to chemically modify paclitaxel to prepare it as a prodrug molecule, thereby altering its crystallization tendency (ACS Nano, 2024, 18(38):26690-26703). However, the chemical modification route has inherent drawbacks, including cumbersome synthetic steps, high costs, and the potential to alter drug activity and metabolic behavior. Another strategy is to introduce solubilizers or surfactants to maintain the solubility of PTX, but traditional solubilizers such as polyoxyethylene castor oil (Cremophor EL) pose serious risks of allergic reactions and neurotoxicity (Journal of Clinical Oncology, 2001, 19(5): 1382-1392). In recent years, researchers have begun to explore inhibiting PTX crystallization through multi-component co-assembly. For example, cyclodextrin can be used to form inclusion complexes with paclitaxel, or amorphous complexes can be formed by co-assembling amphiphilic polymers with PTX (Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2024, 683: 133026). However, most existing co-assembly strategies are limited to binary systems between a single drug and a carrier. When faced with complex multi-drug co-loaded systems containing up to four or five heterogeneous molecules, the complex intermolecular interactions between the components make it difficult for traditional empirical trial-and-error methods to efficiently screen for formulation ratios that can maintain thermodynamic equilibrium. This leads to multi-drug co-assembly systems being prone to phase separation, crystallization, or structural collapse.

[0005] Furthermore, an ideal multi-drug co-loaded nanosystem not only needs to possess high structural stability but should also be able to achieve precise drug release under specific stimuli in the tumor microenvironment (TME). In recent years, by introducing pH-sensitive or glutathione (GSH)-sensitive functional groups, the targeted release of nanomedicines at tumor sites has been achieved, thereby effectively reducing systemic toxicity (RSC Advances, 2024, 14(13): 7117-7130). However, no nanoplatforms combining paclitaxel crystallization inhibition, multi-drug synergistic co-loading, environmental stimulus-responsive release, and multidrug resistance reversal functions have been reported in existing research.

[0006] In summary, the existing technology has the following technical defects: (1) Paclitaxel is prone to crystallization in multi-drug co-loaded nanosystems, which leads to system instability. Existing chemical modification or solubilization strategies have problems such as high cost, high toxicity, and alteration of drug activity; (2) There is a lack of multifunctional integrated nanoparticles that can simultaneously achieve paclitaxel crystallization inhibition, multi-drug synergy, drug resistance reversal, and release in response to environmental stimuli.

[0007] To address the aforementioned technical deficiencies, this invention provides a multi-component co-loaded nanoparticle for inhibiting paclitaxel crystallization and its preparation method. This invention constructs a formulation library containing multiple candidate ratios by fixing the paclitaxel feeding ratio and exhaustively allocating the remaining components. The optimal formulation is then selected that can effectively disrupt the ordered stacking of PTX molecules and form a stable amorphous complex through supramolecular physical co-intercalation between heterogeneous molecules (such as π-π stacking and hydrophobic interactions). Furthermore, by combining this with a p-gp protein inhibitor, the free drug is modified into a stimulus-responsive prodrug lipid molecule, constructing multifunctional nanoparticles that integrate multi-drug synergy, drug resistance reversal, and intelligent release, enabling personalized precision treatment. Summary of the Invention

[0008] Purpose of the invention To address the shortcomings of existing technologies, such as the tendency of paclitaxel in multi-drug co-loaded nanosystems to readily undergo ordered molecular stacking and precipitate needle-like crystals, poor thermodynamic stability of multi-component systems, and the high toxicity and potential alteration of drug activity associated with existing solubilization or chemical modification strategies, the primary objective of this invention is to provide multi-component co-loaded nanoparticles that inhibit paclitaxel crystallization. These nanoparticles do not require highly toxic solubilizers; instead, they achieve highly uniform and stable multi-drug loading by disrupting the lattice growth of paclitaxel at a physicochemical level through non-covalent interactions between heterogeneous molecules in a specific ratio. Another objective of this invention is to provide a method for preparing these nanoparticles, enabling the acquisition of a multi-drug co-loaded system with predetermined stability by adjusting the ratio of active components and the self-assembly process.

[0009] Technical solution To achieve the above objectives, the present invention adopts the following technical solution: The present invention provides a multi-component co-loaded nanoparticle for inhibiting paclitaxel crystallization. The nanoparticle comprises a hydrophobic core and a hydrophilic canopy surrounding the core. The hydrophobic core of the nanoparticle contains paclitaxel and at least two other antitumor active components with conjugated structures. In the nanoparticle, paclitaxel and the other antitumor active components form an amorphous complex through non-covalent interactions. In the nanoparticle, the mass percentage of paclitaxel is 1%-50%, and the concentration of paclitaxel in the hydrophobic core is lower than its critical crystallization concentration. The other antitumor active components include at least two of anthracycline antitumor antibiotics, camptothecin-type topoisomerase inhibitors, P-glycoprotein inhibitors, and coumarin compounds.

[0010] Furthermore, in the above technical solution, preferably, the other antitumor active components include: (I) At least one anthracycline antitumor antibiotic or a pharmaceutically acceptable salt, prodrug, or derivative thereof; (II) At least one camptothecin-type topoisomerase inhibitor or a pharmaceutically acceptable salt, prodrug, or derivative thereof; (III) At least one P-glycoprotein inhibitor or a pharmaceutically acceptable salt, prodrug, or derivative thereof; (IV) At least one coumarin compound or a pharmaceutically acceptable salt, prodrug, or derivative thereof.

[0011] Furthermore, in the above technical solution, specifically, the anthracycline antitumor antibiotic is selected from at least one of doxorubicin, epirubicin, daunorubicin, idarubicin, arubicin, pirarubicin, and mitoxantrone; the camptothecin-type topoisomerase inhibitor is selected from at least one of camptothecin, 10-hydroxycamptothecin, irinotecan, topotecan, belotetan, eczetine, and SN-38; the P-glycoprotein inhibitor is selected from at least one of α-tocopherol succinate, α-tocopherol polyethylene glycol succinate, verapamil, taribedin, cyclosporine A, and veneclade; and the coumarin compound is selected from at least one of coumarin, warfarin, dicumarin, 4-hydroxycoumarin, aesculin, psoralen, and umbelliferone.

[0012] Furthermore, in the above technical solution, preferably, the mass ratio of paclitaxel and two or more other antitumor active components such as anthracycline antitumor antibiotics, camptothecin topoisomerase inhibitors, P-glycoprotein inhibitors, and coumarin compounds in the nanoparticles is as follows: the mass ratio of paclitaxel is 1% to 50%, and the total mass ratio of the other antitumor active components is 50% to 99%; when there are two or more other antitumor active components, the proportion between each component can be adjusted.

[0013] This invention also provides a method for preparing the above-mentioned multi-component co-supported nanoparticles that inhibit paclitaxel crystallization, specifically including the following steps: (1) Preparation of stock solution: Each component drug is dissolved in its corresponding solvent according to its solubility to prepare a stock solution. Then, the amount of drug taken is calculated according to the mass ratio and the corresponding organic phase and aqueous phase are prepared. Among them, the water-soluble solvent used to prepare the drug stock solution and the hydration process is one of ultrapure water, deionized water, PBS buffer, and Tris buffer; the organic solvent used to prepare the drug stock solution is one or a combination of two or more of methanol, ethanol, dichloromethane, trichloromethane, and dimethyl sulfoxide.

[0014] (2) Self-assembly process: The organic phase and aqueous phase of the drug calculated according to the ratio are self-assembled into multi-component nanoparticles by one or more of the following methods: thin film hydration, nanoprecipitation, microfluidics, emulsification or solvent injection.

[0015] (3) Incubation treatment: Nanoparticles prepared by different self-assembly methods are placed in an environment of 37°C and incubated for 4 hours or more.

[0016] (4) Homogenization treatment: The incubated self-assembled nanoparticles are subjected to particle size homogenization treatment through an extruder to obtain multi-component nanoparticles with uniform particle size distribution.

[0017] Beneficial effects The core of this invention lies in obtaining a highly stable multifunctional composite nanomaterial through precise proportioning and supramolecular assembly of multiple heterogeneous components. This invention has the following significant advantages compared to existing technologies: (1) Breakthrough in the underlying mechanism of crystal inhibition (advantage of material structure): The nanoparticle product protected by this invention breaks away from the traditional approach of relying on external solubilizers. It cleverly utilizes the unique large conjugated structure and spatial configuration of other antitumor components to conduct strong non-covalent interactions and spatial intercalation with paclitaxel in the hydrophobic core. This molecular-level physical co-chimerism completely breaks the π-π continuous stacking network between paclitaxel molecules, allowing it to exist in the carrier in a highly stable "amorphous complex" state, thus eliminating needle-like crystal precipitation and early drug leakage from the thermodynamic level.

[0018] (2) Multidimensional targeting and drug resistance reversal integrated diagnosis and treatment (product functional advantages): This nanomaterial flexibly co-loads chemotherapeutic drugs (such as PTX combined with anthracyclines, camptothecin, etc.) acting on different cell proliferation phases in a single chassis, and can also combine P-glycoprotein inhibitors that block efflux pumps and imaging tracer molecules. When the prodrug macromolecule breaks down under the specific stimulation of the tumor microenvironment, it can achieve the targeted and synchronous release of multiple drugs, which can greatly improve the synergistic tumor suppression rate in vivo and fundamentally reverse the multidrug resistance of solid tumors.

[0019] (3) High adaptability of preparation process and flexibility of formulation (advantage of process expansion): This invention breaks through and defines a broad and effective mass co-assembly range of paclitaxel from 1% to 50%. By flexibly adjusting the types of drugs and the proportion of feed ingredients, the formulation system of this invention can be perfectly adapted to various mainstream nanomaterial self-assembly processes such as thin film hydration, nanoprecipitation, and microfluidics. The prepared final product can maintain colloidal stability for a long time in a complex physiological simulation environment containing serum without agglomeration and precipitation, showing extremely excellent anti-protein adsorption capacity and long-term in vivo circulation potential, which greatly expands the feasibility of industrial production and clinical translation. Attached Figure Description

[0020] Figure 1 The image shows typical needle-like crystals of paclitaxel (PTX) and the effect of introducing different antitumor active components to inhibit crystal precipitation using an optical microscope. Figure 2This is a microscopic image of the highly dense and uniform spherical structure of multi-component co-loaded nanoparticles under a transmission electron microscope (TEM). Figure 3 The bar chart shows the changes in particle size and polydispersity index (PDI) of multi-component co-loaded nanoparticles under different pH values ​​(7.4, 6.5, 5.0) and high concentrations of glutathione (GSH) stimulation conditions. Figure 4 Microscopic images of multi-component co-loaded nanoparticles and their internal multi-drug loading (DOX, CPT, COU, and Merge) fluorescence images obtained by multi-channel laser confocal microscopy (CLSM). Figure 5 A high-precision laser confocal microscope-specific fluorescence colocalization confirmation image for a single multi-component co-loaded nanoparticle; Figure 6 This is a three-channel spatial distribution co-localization analysis diagram of the fluorescence intensity of three different anticancer drug derivatives (COU, CPT, DOX) in multi-component co-assembled nanoparticles. Detailed Implementation

[0021] The present invention will be further described in detail below through embodiments. However, it should be noted that the following embodiments are only used to further illustrate the present invention and should not be construed as limiting the scope of protection of the present invention.

[0022] Unless otherwise specified, the experimental methods described in the following examples are conventional methods; unless otherwise specified, the experimental reagents and materials are commercially available. Example 1

[0023] (1) Preparation of stock solutions: Weigh paclitaxel (PTX), epirubicin (anthracycline), and irinotecan derivative DSPE-PEG-SN38 (camptothecin), setting the mass percentage of paclitaxel at 1%, and epirubicin and DSPE-PEG-SN38 at 49.5% each. Dissolve PTX in dimethyl sulfoxide to prepare an organic phase stock solution, and dissolve epirubicin and DSPE-PEG-SN38 in PBS buffer to prepare an aqueous phase stock solution (the concentration of each stock solution is 5 mg / mL).

[0024] (2) Nanoprecipitation self-assembly: Under magnetic stirring at 800 rpm, the organic phase mother liquor was added dropwise to the aqueous phase mother liquor at 1 mL / min, allowing it to self-assemble into nanoparticles through solvent displacement effect. The organic solvent was then removed by rotary evaporation, and PBS buffer was added for further hydration.

[0025] (3) Incubation and homogenization: The hydrated suspension was placed in a constant temperature shaker at 37°C and incubated in the dark for 4 h. Subsequently, the self-assembled nanoparticles after incubation were repeatedly pushed and pulled 21 times through a liposome extruder (100 nm polycarbonate filter membrane) to perform a membrane homogenization process, and finally, stable and uniform nanoparticles were obtained. Example 2

[0026] (1) Proportioning and Stock Solutions: The mass percentage of paclitaxel was set at 30%, pH-sensitive liposomal epirubicin (an anthracycline) at 35%, and taribida (a P-glycoprotein inhibitor) at 35%. Stock solutions were prepared using methanol and ultrapure water, with a concentration of 5 mg / mL for each stock solution. (2) Solvent injection method self-assembly: A methanol solution containing PTX and taribida was injected into ultrapure water containing arubicin at a rate of 2 mL / min using a micro-injection pump. The organic solvent was then removed by nitrogen purging and ultrapure water was added for further hydration.

[0027] (3) Incubation and homogenization: Incubate at 37°C for 4 h, and then homogenize the particle size using an extruder (the specific operation steps are the same as in Example 1). Example 3

[0028] (1) Preparation of stock solution: Weigh paclitaxel (PTX), mPX (DOX derivative mPEG-Schiff-DOX, an anthracycline), verapamil (P-glycoprotein inhibitor) and warfarin (coumarin). Set the mass percentage of paclitaxel to 5%, and the mass percentages of mitoxantrone, verapamil and warfarin to 31.67% (equal mass fractions). Dissolve them in their respective solvents (anhydrous ethanol as the organic phase and Tris buffer as the aqueous phase) to prepare stock solutions. The concentration of each stock solution is 5 mg / mL.

[0029] (2) Microfluidic self-assembly: The prepared organic phase and aqueous phase are loaded into syringes and pumped into the microchannel of the microfluidic chip. The parameters are set as follows: FRR = 1:3, organic phase: 2 mL / min, aqueous phase: 1.5 mL / min. The two phase solutions undergo intense vortex mixing in the channel to form nanoparticles. The organic solvent is then removed by rotary evaporation and Tris buffer is added for further hydration.

[0030] (3) Incubation and homogenization: The effluent was collected and incubated at 37°C for 4 h, and then the particle size was homogenized by an extruder (the specific operation steps are the same as in Example 1). Example 4

[0031] (1) Proportioning and stock solution: Paclitaxel was set at 40% by mass, and 10-hydroxycamptothecin (camptothecin class), Veneclare (P-glycoprotein inhibitor), and warfarin derivative (coumarin class) each accounted for 20%. Stock solutions were prepared using dichloromethane and Tris buffer, with a concentration of 5 mg / mL for each stock solution.

[0032] (2) Thin film hydration self-assembly: The mixed organic phase was placed in a rotary evaporator to evaporate and remove the organic solvent, forming a uniform multi-drug blended lipid film at the bottom of the bottle. Then, Tris buffer aqueous phase was added, and ultrasonic hydration was performed by an ultrasonic probe (the parameters were strictly set as follows: output power 100 W, working time 2 s, intermittent time 5 s, total processing time 20 min).

[0033] (3) Incubation and homogenization: The effluent was collected and incubated at 37°C for 4 h, and then the particle size was homogenized by an extruder (the specific operation steps are the same as in Example 1). Example 5

[0034] (1) Preparation of stock solution: Weigh paclitaxel (PTX), daunorubicin, PEG-topotecan, cyclosporine A and aescin, set the mass ratio of paclitaxel to 20%, and the mass ratio of the other 4 drugs to 20% each, dissolve them in chloroform and deionized water to prepare stock solution, the concentration of the stock solution is 5 mg / mL.

[0035] (2) Emulsification self-assembly: The organic phase is added to the aqueous phase and subjected to vigorous ultrasonic emulsification by an ultrasonic probe (the parameters are strictly set as follows: output power 100 W, working time 2 s, intermittent time 5 s, total processing time 20 min). Then, chloroform is removed by rotary evaporation to form multi-component nanoparticles.

[0036] (3) Incubation and homogenization: The effluent was collected and incubated at 37°C for 4 h, and then the particle size was homogenized by an extruder (the specific operation steps are the same as in Example 1). Example 6

[0037] (1) Proportioning and stock solutions: Paclitaxel was set to account for 50% of the mass, and idarubicin, SN-38, α-tocopherol polyethylene glycol succinate and umbelliferone each accounted for 12.5%. SN-38, α-tocopherol polyethylene glycol succinate and umbelliferone were dissolved in dichloromethane and ethanol (1:1, V / V) to prepare the corresponding stock solutions, and idarubicin was dissolved in deionized water to prepare the aqueous stock solutions. The concentration of the stock solutions was 5 mg / mL.

[0038] (2) Thin film hydration self-assembly: The mixed organic phase was placed in a rotary evaporator to evaporate and remove the organic solvent, forming a uniform multi-drug blended lipid film at the bottom of the bottle. Then, deionized water containing idarubicin was added, and the film was ultrasonically hydrated by an ultrasonic probe (the parameters were strictly set as follows: output power 100 W, working time 2 s, intermittent time 5 s, total processing time 20 min).

[0039] (3) Incubation and homogenization: The effluent was collected and incubated at 37°C for 4 h, and then the particle size was homogenized by an extruder (the specific operation steps are the same as in Example 1). Example 7

[0040] (1) Preparation of stock solutions: Weigh paclitaxel (PTX), mPX (DOX derivative mPEG-Schiff-DOX, an anthracycline), mPT (CPT derivative mPEG-SS-CPT, a camptothecin), mFT (TOS derivative FA-mPEG-TOS, a P-glycoprotein inhibitor), and coumarin D1421 (COU, a coumarin). Set the mass percentage of paclitaxel to 20%, and the remaining 90% of the mass of the other four antitumor active components to be equally distributed, i.e., the mass percentage of DOX, CPT, TOS, and COU is 20% each. Based on their solubility, dissolve PTX, CPT, TOS, and COU in dichloromethane, and dissolve DOX in ultrapure water to prepare stock solutions with a concentration of 5 mg / mL.

[0041] (2) Thin film hydration self-assembly: The mixed organic phase was placed in a rotary evaporator to evaporate and remove the organic solvent, forming a uniform multi-drug blended lipid film at the bottom of the bottle. Then, an ultrapure water phase containing DOX was added, and the film was ultrasonically hydrated by an ultrasonic probe (the parameters were strictly set as follows: output power 100 W, working time 2 s, intermittent time 5 s, total processing time 20 min).

[0042] (3) Incubation and homogenization: The hydrated suspension was placed in a constant temperature shaker at 37°C and incubated in the dark for 4 h. Subsequently, the self-assembled nanoparticles after incubation were repeatedly pushed and pulled 21 times through a liposome extruder (100 nm polycarbonate filter membrane) to homogenize the particle size and obtain the final nanoparticles. Example 8

[0043] To verify the preparation effect under different component quantities and ratios, the multi-component co-loaded nanoparticles prepared in Examples 1 to 7 were subjected to optical microscopy crystal observation and physicochemical property characterization.

[0044] (1) Crystal precipitation observation: The solutions of each group of nanoparticles were placed at room temperature and observed at a frequency of 3 days / time for 30 days. The needle-like crystals of paclitaxel were continuously observed through an optical microscope.

[0045] (2) Particle size, PDI and TEM morphology determination: The hydrated particle size and polydispersity index (PDI) of each embodiment were determined by dynamic light scattering instrument (DLS) at 25°C, and TEM imaging was performed after negative staining with 2% phosphotungstic acid.

[0046] (3) Verification of the stimulus-response release and other functions of representative nanoparticles: Incubation media with different pH values ​​and reducing gradients were prepared, including PBS (pH=7.4) simulating the normal physiological environment, PBS (pH=6.5 and pH=5.0) simulating the slightly acidic environment of tumors, and a mixed medium with an additional high concentration of glutathione (GSH) to simulate the highly reducing state inside tumor cells. After co-incubating the nanoparticles with the above different media, the particle size changes were monitored using DLS.

[0047] (4) Multi-channel fluorescence co-localization was performed on the nanoparticles using laser confocal microscopy to further demonstrate the uniformity of drug distribution in the nanoparticles; The test results are as follows: Within the paclitaxel content range of 1% to 50%, regardless of whether the sample contained 2, 3, or 4 anticancer active pharmaceutical ingredients (Examples 1-7), no paclitaxel needle-like crystals were observed in the microscopic field of view during the 30-day observation period. (See attached...) Figure 1 As shown, PTX itself readily precipitates needle-like crystals. However, the addition of different anticancer active drug components can inhibit crystal precipitation to varying degrees, as shown in Figures ①②③. This conclusively proves that by introducing two or more conjugated antitumor components within the scope of the claims, strong non-covalent interactions and steric hindrance can be formed in the hydrophobic core, effectively disrupting the continuous stacking of paclitaxel molecules and allowing it to exist stably in an amorphous complex state. Secondly, the multi-component nanoparticles have a stable hydrated particle size of approximately 110–150 nm and a PDI of only 0.12–0.28, exhibiting excellent monodispersity and colloidal stability. Transmission electron microscopy (TEM) reveals a highly dense and uniform spherical structure (as shown in the attached figure). Figure 2 (As shown).

[0048] The particle size distribution of Examples 1-5 ranged from 110 nm to 135 nm, and the PDI ranged from 0.12 to 0.22, all meeting the clinical requirements for nano-drug delivery systems. Although no crystals precipitated in Example 6 (50% limit), the particle size increased to approximately 150 nm due to the excessively high proportion of the paclitaxel rigid framework, and the PDI rose to 0.28. Despite the high internal stress of the vesicle system, it still maintained basic colloidal stability. Example 7 demonstrated comprehensive multifunctional properties.

[0049] Further functional verification experiments on the multi-component nanoparticles in Example 7 showed that, by loading anticancer active components with different functions, the nanoparticles could possess corresponding functions in the tumor microenvironment. This included the excellent pH / GSH dual-stimulation responsiveness exhibited in Example 7 (test results are attached). Figure 3 As shown in the figure, precise controlled release of drugs can be achieved. Simultaneously, multi-channel fluorescence co-localization analysis is performed using laser confocal microscopy (see attached figure). Figure 4 , 5 (6) can also demonstrate the uniformity of the distribution of different drug components in nanoparticles.

[0050] In summary, the preparation process described in this invention is highly feasible, encompasses a variety of methods, and the defined quantities of different components and the 1%-50% paclitaxel protection range can all successfully inhibit crystallization. By replacing anticancer active components with different functions and prodrug molecules or derivatives with corresponding functions, the ultimate nanoparticle product that best meets the needs of complex multi-drug synergistic therapy can be prepared.

Claims

1. A multi-component co-supported nanoparticle for inhibiting paclitaxel crystallization, characterized in that, The nanoparticles comprise a hydrophobic core and a hydrophilic canopy surrounding the core. The hydrophobic core contains paclitaxel and at least two other antitumor active components with conjugated structures. Paclitaxel and the other antitumor active components in the nanoparticles form an amorphous complex through non-covalent interactions. The mass percentage of paclitaxel in the nanoparticles is 1%-50%, and the concentration of paclitaxel in the hydrophobic core is below its critical concentration for crystallization. The other antitumor active components include at least two of anthracycline antitumor antibiotics, camptothecin-based topoisomerase inhibitors, P-glycoprotein inhibitors, and coumarin compounds.

2. The nanoparticles according to claim 1, characterized in that, The other antitumor active components include: (I) At least one anthracycline antitumor antibiotic or a pharmaceutically acceptable salt, prodrug, or derivative thereof; (II) At least one camptothecin-type topoisomerase inhibitor or a pharmaceutically acceptable salt, prodrug, or derivative thereof; (III) At least one P-glycoprotein inhibitor or a pharmaceutically acceptable salt, prodrug, or derivative thereof; (IV) At least one coumarin compound or a pharmaceutically acceptable salt, prodrug, or derivative thereof.

3. The nanoparticles according to claim 1, characterized in that, The anthracycline antitumor antibiotics are selected from at least one of doxorubicin, epirubicin, daunorubicin, idarubicin, arubicin, pirarubicin, and mitoxantrone; the camptothecin-type topoisomerase inhibitors are selected from at least one of camptothecin, 10-hydroxycamptothecin, irinotecan, topotecan, belotetan, eczetine, and SN-38; the P-glycoprotein inhibitors are selected from at least one of α-tocopherol succinate, α-tocopherol polyethylene glycol succinate, verapamil, taribedin, cyclosporine A, and veneclade; and the coumarin compounds are selected from at least one of coumarin, warfarin, dicoumarin, 4-hydroxycoumarin, aesculin, psoralen, and umbelliferone.

4. The nanoparticles according to claim 1, characterized in that, The mass ratio of paclitaxel and two or more other antitumor active components in the nanoparticles, such as anthracycline antitumor antibiotics, camptothecin topoisomerase inhibitors, P-glycoprotein inhibitors, and coumarin compounds, is as follows: paclitaxel accounts for 1% to 50% of the mass, and the total mass ratio of the other antitumor active components is 50% to 99%. When there are two or more other antitumor active components, the proportions between the components can be adjusted.

5. The nanoparticles according to claim 1, characterized in that, The water-soluble solvent used in the preparation of the drug stock solution and the hydration process in the nanoparticle preparation method is one of ultrapure water, deionized water, PBS buffer, and Tris buffer. The organic solvent used in the preparation of the drug stock solution is one or a combination of two or more of methanol, ethanol, dichloromethane, trichloromethane, and dimethyl sulfoxide.

6. The nanoparticles according to claim 1, characterized in that, The self-assembly methods in the preparation process of nanoparticles include one or more combinations of thin film hydration, nanoprecipitation, microfluidics, emulsification and solvent injection.

7. The nanoparticles and their preparation method according to claims 1-6, characterized in that, The preparation method is as follows: each component drug is dissolved in its corresponding solvent according to its solubility to prepare a mother liquor. Then, the amount of drug taken is calculated according to the mass ratio and prepared accordingly. The organic phase and aqueous phase of the drug calculated according to the ratio are self-assembled into multi-component nanoparticles by one of the following methods: thin film hydration method, nanoprecipitation method, microfluidic method, emulsification method, solvent injection method, etc. The nanoparticles prepared by different self-assembly methods are placed in an environment of 37 degrees Celsius and incubated for 4 hours or more. The incubated self-assembled nanoparticles are subjected to particle size homogenization treatment through an extruder to obtain particles with uniform particle size distribution.

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