CN111529510A

CN111529510A - Application of nanoparticles as tumor microenvironment responsive drug or imaging agent

Info

Publication number: CN111529510A
Application number: CN202010387805.1A
Authority: CN
Inventors: 任建丽; 周志益; 罗远利; 乔斌; ***; 冉海涛; 李攀; 郝兰; 曹阳
Original assignee: Chongqing Medical University
Current assignee: Chongqing Medical University
Priority date: 2020-05-09
Filing date: 2020-05-09
Publication date: 2020-08-14

Abstract

The invention belongs to the field of biomedicine, relates to application of nanoparticles, and particularly relates to application of nanoparticles as a tumor microenvironment responsive drug or an imaging agent. The nanoparticles comprise a chelate shell formed by tannic acid and ferric ions, and chemotherapeutic drugs are loaded in the chelate shell. The nano particles can slowly release chemotherapeutic drugs carried in the nano particles in a tumor microenvironment, and iron ions in the nano particles are slowly released to enhance the nuclear magnetic resonance imaging effect. The technical scheme can be applied to the medical practice of tumor imaging and tumor treatment.

Description

Application of nanoparticles as tumor microenvironment responsive drug or imaging agent

Technical Field

The invention belongs to the field of biomedicine, relates to application of nanoparticles, and particularly relates to application of nanoparticles as a tumor microenvironment responsive drug or an imaging agent.

Background

Chemotherapy is a classic and effective tumor treatment technique and has been widely used in clinical practice. However, the chemotherapy medicament lacks targeting, and is easy to be phagocytized and absorbed at non-pathological parts; moreover, some chemotherapeutic drugs have short half-life in vivo and are easy to inactivate in the in vivo transportation process; in addition, the burst release of chemotherapeutic agents also increases their systemic toxicity. The above disadvantages prevent the clinical application of chemotherapy. Therefore, more and more researchers and clinicians are dedicated to the study of loading chemotherapeutic drugs into nanoparticle drug delivery systems, and the rationally designed nanoparticle drug delivery systems are of great significance in improving the efficacy and biological safety of chemotherapy.

In the prior art, researchers encapsulated chemotherapeutic drugs in liposomes or polylactic acid-glycolic acid copolymer shells to form nanoparticles encapsulating chemotherapeutic drugs, thereby reducing the toxicity of chemotherapeutic drugs to normal tissues and preventing the chemotherapeutic drugs from being prematurely inactivated during in vivo operation. However, the prior art solutions have the following drawbacks: after the nanoparticles enter biological tissues, the tissue distribution and aggregation conditions of the chemotherapeutic drugs cannot be monitored in real time, the fixed-point release of the chemotherapeutic drugs at tumor tissues cannot be realized, and the control on the release speed of the chemotherapeutic drugs cannot be mentioned.

Disclosure of Invention

The invention aims to provide application of a nanoparticle as a tumor microenvironment responsive drug or an imaging agent, wherein the nanoparticle can slowly release chemotherapeutic drugs carried in the nanoparticle and iron ions in the nanoparticle so as to enhance nuclear magnetic resonance imaging effect in a tumor microenvironment.

In order to solve the technical problems, the technical scheme of the invention provides application of nanoparticles as a tumor microenvironment responsive drug or an imaging agent, wherein the nanoparticles comprise a chelate shell formed by tannic acid and ferric ions, and chemotherapy drugs are loaded in the chelate shell.

By adopting the technical scheme, the technical principle is as follows: the chelate shell formed by two substances of tannic acid and ferric ion is unstable in an acidic or glutathione environment and gradually dissociates, so that the chemotherapeutic drug wrapped in the chelate shell is gradually released, and ferric ion is released. The tumor microenvironment is an acidic and glutathione-rich environment, while the normal tissue microenvironment is neutral or weakly alkaline and does not contain glutathione. After the nano particles are injected into organisms, the nano particles are stable in nature and are not dissociated in normal tissues, but after the nano particles enter tumor tissues, the stability of the nano particles is reduced under the influence of pH value and glutathione (the nano particles have the characteristics of responding to the pH value and the glutathione), chemotherapeutic drugs entrapped in the nano particles are gradually released, the proliferation of tumor cells is inhibited, ferric ions in chelate shells are also gradually released, and the T1 weighted magnetic resonance imaging effect is enhanced.

Has the advantages that:

in the stimulus-responsive nanoparticle drug delivery system in the prior art, metal nanoparticles or liquid fluorocarbon microbubble substances are generally added to the nanoparticle drug delivery system, and the release of chemotherapeutic drugs in the nanoparticle drug delivery system is usually realized through external light, heat, ultrasonic or other stimuli. However, the additional external stimulation condition may cause some side effects to the living body, such as skin burn, increased radiation exposure, etc., and the process of promoting the release of the chemotherapy drug by using the external stimulation requires additional medical equipment, which increases the complexity of the operation and is not favorable for the clinical popularization of the technology. The inventor researches the response phenomenon of external light and thermal stimulation of the chelate nanoparticle formed by tannic acid and ferric ions, and unexpectedly finds that the chelate nanoparticle can be broken under the external light and thermal stimulation, the chelate nanoparticle can be dissociated in a tumor microenvironment, the dissociation process is carried out step by step, and the chemotherapeutic drug carried in the chelate nanoparticle can be released step by step, so that the slow release of the drug is realized, and the system toxicity of the drug is reduced. The inventor further researches the principle that the chelate nanoparticle formed by tannic acid and ferric ion can be dissociated in the tumor microenvironment. The oxygen content in the tumor cells is low, and the excessive secretion of glutathione into intercellular substance of the tumor cells is stimulated; in addition, in tumor cells, the excessive secretion of lactate or other tumor cell metabolites leads to an acidic microenvironment outside the tumor cell. The low pH value and the high glutathione environment stimulate the dissociation of the chelate nanoparticles, thereby promoting the release of the chemotherapeutic drugs in the chelate nanoparticles. In this scheme, the tumor microenvironment mainly refers to an acidic environment and an environment containing glutathione.

In conclusion, the nanoparticles in the technical scheme are applied to tumor microenvironment responsive drugs or imaging agents, and have the following beneficial effects:

(1) the nano particles in the technical scheme are stable in normal tissues, and release chemotherapeutic drugs in tumor tissues, so that the fixed-point release of the chemotherapeutic drugs at tumor parts is realized, and the fixed-point release does not need external stimulation such as light, heat or ultrasound and the like, is convenient to operate, and is suitable for clinical popularization.

(2) The shell of the nano particles in the technical scheme is gradually dissociated at the tumor tissue, so that the control of the drug release rate is realized, and the system toxicity caused by the burst release of the chemotherapeutic drug is avoided.

(3) The shell of the nano-particle in the technical scheme is gradually dissociated at the tumor tissue to gradually release ferric ions, so that T1 weighted magnetic resonance imaging can be enhanced, and the nano-particle can be used as an imaging agent to be applied to diagnosis and detection of tumors.

(4) The nano-particles in the technical scheme also have the properties of photothermal imaging (PTI) and photoacoustic imaging (PAI). After being chelated, the ferric ions and the tannic acid have the photoacoustic imaging property and can be monitored by a photoacoustic imager; the photo-thermal imaging material has photo-thermal imaging property under illumination and can be monitored by a thermal infrared imager.

(5) The nanoparticles in the technical scheme also have the effect of photothermal therapy (PTT). After chelation of ferric ions and tannic acid, the compound has high photo-thermal conversion efficiency, and can generate thermal effect under the irradiation of 600-900nm laser, thereby realizing the photo-thermal treatment of tumors.

(6) The chelate formed by the tannic acid and the ferric ions coats and modifies the chemotherapeutic drug, reduces the toxicity of the chemotherapeutic drug and increases the circulation time of the chemotherapeutic drug in blood, thereby preventing the chemotherapeutic drug from being prematurely inactivated during in vivo operation.

Further, the ferric ion is provided by ferric chloride.

By adopting the technical scheme, the ferric trichloride is a common chemical reagent, is easy to obtain and has low cost.

Further, the chemotherapeutic agent is doxorubriosin.

By adopting the technical scheme, the Tourethricin is a medicament widely applied to chemotherapy, and has clear pharmacological action and good treatment effect.

Further, the nanoparticles are prepared by the following method:

step (1) preparation of hydrophobic topiroxomicin: dissolving hydrochloric acid carbendazim in dimethyl sulfoxide solution, adding trimethylamine, and uniformly mixing to obtain a mixture;

step (2) preparation of the shell: dripping high-purity water into the mixture obtained in the step (1) and stirring the mixture simultaneously to obtain dispersion liquid; sequentially adding a tannic acid solution and a ferric trichloride hexahydrate solution into the dispersion liquid to obtain a mixed dispersion liquid, and carrying out ultrasonic treatment on the mixed dispersion liquid to obtain a crude product solution, wherein the crude product solution contains nano particles;

and (3) post-treatment: adjusting the pH value of the crude product solution to be neutral, and then washing and collecting the nano particles in the crude product solution.

By adopting the technical scheme, the nanoparticles with uniform particle size, stable property, good dispersion degree and high chemotherapeutic drug encapsulation rate can be prepared. In the step (1), hydrophobic polyoxins nanoparticles (mixture B is a hydrophobic polyoxins nanoparticle solution) are prepared, namely hydrophobic polyoxins self-assemble into a nano inner core template, and then on the basis of the inner core template, a chelate film formed by tannic acid and ferric ions is covered on the inner core template. Tannic acid is a naturally occurring polyphenol that can act as a chelating site for ferric ions at room temperature. The nano particles (DOX-Fe-TA-nanoparticles, DFTNPs) of the technical scheme are formed by the synergistic effects of coordination, hydrophobicity, P-P accumulation and the like on the surface of a nano core template formed by self-assembly of hydrophobic polyoxins. The hydrophobic polyoxins nanoparticles are used as the core, and a chelate film is formed on the core, so that compared with the traditional scheme of preparing an outer film and then wrapping a medicament, the preparation scheme is simpler, and the medicament encapsulation rate is higher.

High purity water (High purity water) is referred to in this case as: water having a conductivity of less than 0.1. mu.s/cm at 25 ℃ and a residual salt content of less than 0.3 mg/L.

Furthermore, in the step (1), the dosage ratio of the hydrochloric acid carbendazim to the dimethyl sulfoxide solution to the trimethylamine is (3-50) mg, (3-50) ml and (0.3-5) ml.

By adopting the technical scheme, the hydrophobic Toxoplasma variegated nanoparticles with stable properties and proper particle size can be obtained (namely, the hydrophobic Toxoplasma variegated nanoparticles are self-assembled into a nano inner core template).

Further, in the step (2), the volume ratio of the mixture, the high-purity water, the tannic acid solution and the ferric chloride hexahydrate solution is 100:980 (10-100) to (10-100); the concentration of tannic acid in the tannic acid solution is 40mg/ml, and the concentration of ferric chloride hexahydrate in the ferric chloride hexahydrate solution is 2-20 mg/ml.

By adopting the technical scheme, the tannic acid and ferric ions can quickly form cross-linking, and a chelate film is formed outside the hydrophobic Toxoplasma nanoparticles.

Further, in the step (2), high pure water is dropped to the mixture at a dropping speed of 100 to 1000. mu.l/min while stirring the mixture at a rotation speed of 300 to 1000 rpm.

By adopting the technical scheme, the mixture is stirred at a high speed, and high pure water is simultaneously dripped, so that hydrophobic multi-ratio soft star can be fully dispersed, nano-scale multi-ratio soft star particles are obtained, and subsequent chelation and attachment of tannic acid and ferric ions are facilitated.

Further, in step (3), the pH of the crude solution was adjusted to neutral using 1mM NaOH.

By adopting the technical scheme, 1mM NaOH is a common pH value adjusting reagent, has stable property and is easy to obtain.

Furthermore, the particle diameter of the nano-particles is 84.0-209.8 nm, and the surface potential is-26.7-16.1 mV.

By adopting the technical scheme, the nanoparticles with the particle sizes have better bioavailability and can pass through the vascular wall to reach the target tissue with higher efficiency.

Further, the polymer dispersion coefficient of the nano particles is 0.100-0.200.

By adopting the technical scheme, the dispersion coefficient of the polymer is not more than 0.2, the nano particles are uniform and dispersed, and the agglomeration tendency is small. The lower the polymer dispersion coefficient pdi (polymer dispersion index), the better the nanoparticle dispersion and the smaller the tendency to agglomerate.

Drawings

FIG. 1 is a transmission electron micrograph of DFTNPs of Experimental example 1;

FIG. 2 is a graph showing particle sizes of DFTNPs in Experimental example 1;

FIG. 3 is a potential diagram of DFTNPs of Experimental example 1;

FIG. 4 is a graph showing UV absorption of DFTNPs of Experimental example 1;

FIG. 5 is a graph of the in vitro drug release profiles (different GSH content) of DFTNPs from Experimental example 2;

FIG. 6 is a graph showing in vitro drug release profiles (different pH values) of DFTNPs of Experimental example 2;

FIG. 7 is a graph showing the results of photothermal effect experiments (different concentrations of DFTNPs) of DFTNPs of Experimental example 3;

FIG. 8 is a graph showing the results of photothermal effect experiments (different irradiation intensities) of DFTNPs of Experimental example 3;

FIG. 9 is a thermal imaging graph of different concentrations of DFTNPs in Experimental example 3 under 808nm laser irradiation;

FIG. 10 is a graph showing temperature change curves of different components in DFTNPs of Experimental example 3 under 808nm laser irradiation;

FIG. 11 shows the results of the CCK-8 method for evaluating cytotoxicity of DFTNPs in Experimental example 4;

FIG. 12 is a result of the CCK-8 method for evaluating phototoxicity of DFTNPs in Experimental example 4;

FIG. 13 is a graph showing the evaluation of apoptosis of DFTNPs by flow cytometry under different treatment conditions in Experimental example 4;

FIG. 14 shows the phagocytosis of DFTNPs by MDA-MB-231 cells of Experimental example 5 at different observation time points;

FIG. 15 shows the results of photoacoustic imaging in vitro using DFTNPs from Experimental example 6;

FIG. 16 shows the results of in vivo photoacoustic imaging experiments using DFTNPs from Experimental example 6;

FIG. 17 is an MRI image of DFTNPs from Experimental example 7 at different GSHs;

FIG. 18 is an MRI image of DFTNPs of Experimental example 7 at different pH;

FIG. 19 is an in vivo MRI image of DFTNPs of Experimental example 7;

FIG. 20 is the photothermographic signals of the tumor sites of tumor-bearing nude mice with DFTNPs of Experimental example 8 at different time points;

FIG. 21 is the temperature changes of the tumor sites of nude mice bearing tumor at different time points for DFTNPs of Experimental example 8;

FIG. 22 shows the tumor volume changes of the DFTNPs of Experimental example 8 in tumor-bearing nude mice at different observation time points;

FIG. 23 shows the body weight changes of tumor-bearing nude mice with DFTNPs of Experimental example 8 at different observation time points.

Detailed Description

The following is further detailed by way of specific embodiments:

example 1: preparation of nanoparticles

10mg DOX & HCl (doxorubicin hydrochloride) was dissolved in 10ml DMSO (dimethyl sulfoxide) solution. Then, the mixture is stirred moderately at room temperature, 1ml of trimethylamine is added, hydrochloric acid is removed to convert hydrophilic DOX & HCl into hydrophobic DOX nano particles, and the mixture is obtained after the mixture is stirred uniformly. Then, 100. mu.l of the prepared mixture (DOX/DMSO solution neutralized with trimethylamine) was dropped into 980. mu.l of high purity water with vigorous stirring (stirring at 700 rpm), the dropping speed of the high purity water being 800. mu.l/min, to obtain a dispersion. Then 10. mu.l of Ta (tannic acid) solution (40mg/ml) and 10. mu.l of FeCl₃·6H₂O solution (20mg/ml) was added to the above dispersion in order, followed by sonication for 50s and neutralization with 1. mu.M NaOH solution. Centrifugally washing the obtained product with deionized water to remove excessive TA and FeCl₃. And re-dispersing the nano particles in high-purity water to obtain the DFTNPs.

In the preparation process, the dosage ratio of DOX & HCl, DMSO solution and trimethylamine can be (3-50) mg, (3-50) ml and (0.3-5) ml. Mixture, high purity water, Ta solution and FeCl₃·6H₂The volume ratio of the O solution can be 100:980, (10-100) to (10-100). FeCl₃·6H₂FeCl in O solution₃·6H₂The concentration of O can be 2-20 mg/ml. Dropping highly pure water into the mixture at a dropping speed of 100 to 1000. mu.l/min while stirring the mixture at a rotation speed of 300 to 1000 rpm.

Example 2

10mg DOX & HCl (doxorubicin hydrochloride) was dissolved in 10ml DMSO (dimethyl sulfoxide) solution. Then, with moderate stirring at room temperature, 1ml of trimethylamine was added and the hydrochloric acid was removed to convert the hydrophilic DOX · HCl into hydrophobic DOX nanoparticles. Then, 100. mu.l of prepared DOX/DMSO was dissolvedThe liquid was dropped into 980. mu.l of high purity water with vigorous stirring (stirring at 1000 rpm) at a rate of 1000. mu.l/min to obtain a dispersion. Then 100. mu.l of Ta (tannic acid) solution (40mg/ml) and 100. mu.l of FeCl₃·6H₂O solution (20mg/ml) was added to the above dispersion in order, followed by sonication for 50s and neutralization with 1. mu.M NaOH solution. Centrifugally washing the obtained product with deionized water to remove excessive TA and FeCl₃. And re-dispersing the nano particles in high-purity water to obtain the DFTNPs.

Example 3

10mg DOX & HCl (doxorubicin hydrochloride) was dissolved in 10ml DMSO (dimethyl sulfoxide) solution. Then, with moderate stirring at room temperature, 1ml of trimethylamine was added and the hydrochloric acid was removed to convert the hydrophilic DOX · HCl into hydrophobic DOX nanoparticles. Then, 980. mu.l of high purity water was dropped into 100. mu.l of the prepared DOX/DMSO solution under vigorous stirring (stirring at 300 rpm), and the dropping rate of the high purity water was 100. mu.l/min, to obtain a dispersion. Then 50. mu.l of Ta (tannic acid) solution (40mg/ml) and 50. mu.l of FeCl₃·6H₂O solution (20mg/ml) was added to the above dispersion in order, followed by sonication for 50s and neutralization with 1. mu.M NaOH solution. Centrifugally washing the obtained product with deionized water to remove excessive TA and FeCl₃. And re-dispersing the nano particles in high-purity water to obtain the DFTNPs.

Experimental example 1: characteristics and physical Properties of the nanoparticles

For the DFTNPs prepared in example 1, the morphology of the nanoparticles was observed using a transmission electron microscope, and the particle size and potential of DOXNPs and DFTNPs were measured using a potentiostat (see fig. 1, 2 and 3). The particle size of DFTNPs is: 146.9 +/-62.9 nm, potential: -21.4 ± 5.3 mV. The preparation method of the DOXNPs comprises the following steps: 10mg DOX & HCl (doxorubicin hydrochloride) was dissolved in 10ml DMSO (dimethyl sulfoxide) solution. Then, with moderate stirring at room temperature, 1ml of trimethylamine was added to the mixture and the hydrochloric acid was removed to convert hydrophilic DOX · HCl into hydrophobic DOX nanoparticles. Detection of DOXNPs, DFTNPs and Fe using UV absorption spectroscopy³⁺UV absorption spectrum of/TA (see FIG. 4). Wherein, Fe³⁺The term/TA means direct conversion of Fe³⁺Chelating with tannic acid and then sonicatingDispersed nanoparticles formed.

Experimental example 2: encapsulation efficiency and in vitro drug release

And (3) determining the encapsulation efficiency: UV-Vis absorption peaks of DFTNPs were measured and standard curves for DOX were plotted by measuring the UV absorption spectra at 480nm of solutions of various concentrations of DOX (300,150,75,37.5,0.3125 μ g/mL). By comparing the ultraviolet absorption of DOX in the supernatant, the encapsulation efficiency of DFTNPs to DOX is calculated to be 76.3% by combining a standard curve.

In vitro drug release of DOX: drug release experiments were performed at different pH and GSH concentrations (glutathione). 5mL of DFTNPs solution was placed in a dialysis bag (MWCO: 3500Da) and immersed in 15mL of buffer and the buffer was stirred at 160rpm for 120 h. The release of DOX from DFTNPs was measured at pH 5.5,6.5,7.4 and GSH concentrations 0mM,2mM,10mM (calculated according to the standard curve). The results are shown in fig. 5 and 6, which demonstrate that the release rate of DOX varies with the pH and GSH concentration. When the nano-particle moves in vivo, the pH value of a cancer part is low, the GSH content is high, and the release of DOX can be promoted, so that the therapeutic effect of the nano-particle is realized. However, in normal tissues, if the pH is higher and the GSH content is lower, the release amount of DOX is reduced, and the side effect of the DOX on the normal tissues is reduced. The nanoparticle DFTNPs have a selective release function, so that side effects are reduced, and the treatment effect is improved.

Experimental example 3: in vitro photothermal effect study

DFTNPs were added to a 96-well plate, and the relationship between the concentration of DFTNPs and the photothermal effect was investigated by irradiating DFTNPs (50,100,200,300,400. mu.g/mL) at different concentrations for 10min with 808nm laser. Irradiation with 808nm laser light of different intensities (0.5,1.0,1.5,2.0W/cm²) And (5) irradiating the DFTNPs for 10min, and researching the relation between the irradiation intensity and the photothermal effect. As shown in fig. 7 and 8, the higher the concentration of DFTNPs and the stronger the irradiation intensity, the stronger the photothermal effect of the nanoparticle. Meanwhile, the photothermal effects of DFTNPs with different irradiation times are also researched, and the experimental result is shown in FIG. 9, wherein the longer the irradiation time is, the higher the concentration of DFTNPs is, and the stronger the photothermal effect of the nanoparticle is.

To investigate the photothermal sources of DFTNPs, DFTNPs (400. mu.g/mL), DOXNPs (400. mu.g/mL), tannic acid (400. mu.g/mL), FeCl₃(400. mu.g/mL) and Fe ³⁺200. mu.L of each of the solutions/TA was added to a 96-well plate as a control group, and a 808nm laser (2.0W/cm)²) The irradiation was carried out for 10min and the temperature change was recorded using a near infrared imager (Fotric 226). The results of the experiment are shown in FIG. 10, FeCl₃Tannic acid and DOXNPs all have no photothermal effect, Fe³⁺the/TA has a certain photothermal effect, and the DFTNPs formed by adding DOX into the nanoparticles have a stronger heat supply effect.

Experimental example 4: cytotoxicity and proliferation assay

The cytotoxicity and photothermal therapeutic effects of DFTNPs were evaluated using the CCK-8 assay. MDA-MB-231 cells were plated on a 96-well plate overnight, 100. mu.L of DFTNPs (8.13-65. mu.g/ml) at various concentrations were added to the plate, and incubated for 36h with the cells, 10. mu.L of CCK-8 solution was added to the plate, and the cell activity was measured using a plate reader. The experimental results are shown in FIG. 11, and the DFTNPs have lower cytotoxicity and are safer than DOXNPs.

To further evaluate the photothermal chemotherapeutic synergistic therapeutic effect of DFTNPs, different concentrations of DFTNPs (25-200. mu.g/ml) were incubated with cells for 36 h. Then the cells were irradiated with laser at 808nm (2.0W/cm)²) Irradiating for 10min, incubating for 12h, and detecting cell activity by CCK-8 method. As shown in fig. 12, after laser irradiation, DFTNPs showed cytotoxicity and were applicable to photothermal chemotherapy of cancer cells.

To further evaluate the apoptosis of MDA-MB-231 cells after DFTNPs treatment, MDA-MB-231 cells were placed in a 6-well plate for 12h to allow complete cell adhesion, and then DOXNPs (50. mu.g/ml) and DFTNPs (50. mu.g/ml) were added to incubate for 24h, followed by laser irradiation of the cells at 808nm (2.0W/cm)²) Irradiating for 10min, and detecting cell activity by flow cytometry. The experimental result is shown in fig. 13, the DFTNPs can promote more apoptosis after laser irradiation, and have the effect of promoting cancer cell apoptosis.

Example 5: endocytosis assay

To investigate that DFTNPs could be efficiently phagocytosed by MDA-MB-231 cells, MDA-MB-231 cells (5 × 10)⁴One) was planted on a glass petri dish and incubated for 12h, after which the existing culture was discardedAnd (3) adding DFTNPs into nutrient, incubating for different times (1h, 2h, 4h and 8h), fully washing cells, and staining cell nuclei for 15min by using DAPI. Cell phagocytosis was observed using confocal laser microscopy. The experimental results are shown in fig. 14, and the experimental results prove that the DFTNPs can be effectively phagocytized by cells.

Example 6: in vivo and in vitro photoacoustic imaging

To investigate the potential of DFTNPs as contrast agents for photoacoustic imaging, photoacoustic imaging evaluations were performed using a photoacoustic imager (Visual sonic). Different concentrations of DFTNPs (40-200 mug/ml) are placed in a gel model, and photoacoustic imaging signals of the DFTNPs are detected by a photoacoustic imager. As shown in fig. 15, the photoacoustic imaging signal becomes stronger as the ion concentration increases.

Tumor-bearing mice were used to evaluate the photoacoustic imaging effect of DFTNPs in vivo. Injecting DFTNPs (5mg/mL) into tail vein, and observing the photoacoustic imaging signal intensity of the DFTNPs at the tumor site at different time points (0h, 2h, 4h and 24 h). As shown in fig. 16, DFTNPs have photoacoustic imaging signals at tumor sites in tumor-bearing nude mice.

Example 7: in vivo and in vitro MRI imaging signals

To investigate the potential of DFTNPs as contrast agents for magnetic resonance imaging, magnetic resonance imaging evaluations were performed using a magnetic resonance imager (simensmrc 406903.0 t MRI imager). Different concentrations of DFTNPs (40-200. mu.g/ml) were placed in a test tube model. The DFTNPs were adjusted to different pH values (4.5,7.4) and different GSH contents (2mM, 10mM), and the magnetic resonance imaging signals of the DFTNPs were detected by a magnetic resonance imager. The experimental results are shown in fig. 17 and 18, and the GSH concentration and the pH value have a certain influence on the nuclear magnetic imaging intensity of the DFTNPs.

Tumor-bearing mice were used to evaluate the MRI imaging effect of DFTNPs in vivo. DFTNPs (5mg/mL) were injected into the tail vein, and the MRI imaging signal intensity of the DFTNPs at the tumor site was observed at different time points (0h, 2h, 4h, and 24 h). The experimental result is shown in fig. 19, and the DFTNPs are proved to have the function of tumor MRI imaging.

Example 8: in vivo antitumor therapeutic study

By subcutaneous injection of MDA-MB-231 cells (100. mu.L, 10) into the right hip of nude mice⁷one/mL) to establish a tumor-bearing mouse model.When the tumor of the mouse grows to 100mm³Tumor-bearing nude mice were randomly divided into 6 groups (n ═ 5): the control group, the simple laser group, the DOXNPs group, the DFTNPs group, the DOXNPs plus laser group and the DFTNPs plus laser group are respectively. Tumor-bearing mice were injected with 5% glucose solution, DOXNPs solution and DFTNPs solution via tail vein, respectively. After 24h, the tumors were irradiated with 808nm laser (2W/cm)²10 min). The temperature change at the tumor site was recorded using a thermal infrared imager. The body weight of the nude mice and the major axis (L) and minor axis (W) of the tumor were recorded every two days. Tumor volume (V) according to V ═ tumor major diameter (L)²× minor diameter (W)/2, relative tumor volume was calculated using V/V0, where V represents tumor volume and V0 represents initial tumor volume the results of the experiments are shown in FIGS. 20-23, and the in vivo experiments show that DFTNPs have better tumor therapeutic effect than DOXNPs.

The foregoing is merely an example of the present invention and common general knowledge in the art of designing and/or characterizing particular aspects and/or features is not described in any greater detail herein. It should be noted that, for those skilled in the art, without departing from the technical solution of the present invention, several variations and modifications can be made, which should also be regarded as the protection scope of the present invention, and these will not affect the effect of the implementation of the present invention and the practicability of the patent. The scope of the claims of the present application shall be determined by the contents of the claims, and the description of the embodiments and the like in the specification shall be used to explain the contents of the claims.

Claims

1. The application of the nanoparticle as a tumor microenvironment responsive drug or an imaging agent is characterized in that the nanoparticle comprises a chelate shell formed by tannic acid and ferric ion, and chemotherapeutic drugs are loaded in the chelate shell.

2. Use according to claim 1, wherein the ferric ions are provided by ferric chloride.

3. The use of claim 2, wherein the chemotherapeutic agent is doxorubriosin.

4. Use according to claims 1-3, wherein the nanoparticles are prepared by a process comprising:

5. The use of claim 4, wherein in the step (1), the dosage ratio of the hydrochloric acid carbendazim to the dimethyl sulfoxide solution to the trimethylamine is (3-50) mg, (3-50) ml, (0.3-5) ml.

6. The use as claimed in claim 5, wherein in the step (2), the volume ratio of the mixture, the high-purity water, the tannic acid solution and the ferric chloride hexahydrate solution is 100:980 (10-100): 10-100); the concentration of tannic acid in the tannic acid solution is 40mg/ml, and the concentration of ferric chloride hexahydrate in the ferric chloride hexahydrate solution is 2-20 mg/ml.

7. The use according to claim 6, wherein in the step (2), the dropping speed of high pure water into the mixture is 100 to 1000. mu.l/min, and the mixture is stirred at a rotation speed of 300 to 1000rpm at the same time.

8. The use as claimed in claim 7, wherein in step (3), 1mM NaOH is used to adjust the pH of the crude solution to neutral.

9. The use according to claim 8, wherein the nanoparticles have a particle size of 84.0 to 209.8nm and a surface potential of-26.7 to-16.1 mV.

10. The use according to claim 9, wherein the nanoparticles have a polymer dispersion factor of 0.100 to 0.200.