PEG-coated europium-based MOFs nano composite material as well as preparation method and application thereof

Abstract

The invention discloses a PEG-coated europium-based MOFs nano composite material as well as a preparation method and application thereof. The nano composite material comprises a core structure and a shell structure, the core structure comprises a europium-based organic framework (Eu-MOF), and silver nanoparticles (AgNPs) and paclitaxel (PTX) which are loaded on the europium-based organic framework; the shell structure is polyethylene glycol (PEG) and wraps the core structure. According to the invention, Eu-MOF, photo-thermal agent silver nanoparticles (AgNPs) and PTX are co-assembled by using metal coordination, physical adsorption and pi-pi action, and the dispersibility of the assembled Eu-MOF, the photo-thermal agent silver nanoparticles (AgNPs) and PTX is enhanced by using PEG (Polyethylene Glycol) for wrapping, so that a multifunctional nano-composite for pH response of a tumor microenvironment is constructed. The nanocomposite shows excellent fluorescence imaging and photo-thermal imaging capabilities, can provide clear tumor localization and monitoring, and significantly improves the treatment effect of breast cancer through dual-mode combined treatment of photo-thermal treatment and chemotherapy.

Description

Technical Field

[0001] The invention relates to the technical field of nanocomposites, and in particular to a PEG-coated europium-based MOFs nanocomposite material and a preparation method and application thereof. Background Art

[0002] As one of the most common malignant tumors among women worldwide, the incidence and mortality of breast cancer have continued to rise in recent years, posing a serious threat to human health. Currently, traditional breast cancer treatments include surgery, chemotherapy, radiotherapy, etc. Although these methods have achieved certain results in treatment, their significant side effects, greater trauma, and easy development of drug resistance are still the main obstacles, affecting the treatment effect of breast cancer. In order to reduce side effects and improve treatment effects, researchers have gradually begun to pay attention to the application of combined treatment strategies in the treatment of breast cancer.

[0003] Combination therapy can complement or enhance the efficacy of different treatment methods by integrating them, while reducing the side effects that may be caused by single therapy. Photothermal therapy (PTT) has shown great application potential in the treatment of breast cancer due to its rapidity, simplicity, and good therapeutic effect. The combination of PTT and chemotherapy can significantly improve the therapeutic effect and inhibit tumor recurrence and metastasis to a certain extent. When these two treatments are used in combination, the local high temperature generated by PTT can destroy the structure of cancer cells, activate the immune response, indirectly enhance the effect of chemotherapy drugs on cancer cells, and reduce the damage of chemotherapy drugs to healthy cells. In addition, the thermal effect of PTT destroys the surrounding microvessels, further limiting the nutrient supply of the tumor and reducing the possibility of recurrence and metastasis.

[0004] Therefore, constructing a nanocomplex that can achieve combined photothermal / chemotherapy treatment of breast cancer can provide new thinking for combined treatment mode, which is of great significance for improving the effect of tumor treatment. Summary of the invention

[0005] The technical problem to be solved by the present invention is to provide a PEG-coated europium-based MOFs nanocomposite material and a preparation method and application thereof. The nanocomposite material can be used for the combined photothermal / chemotherapy treatment of breast cancer to improve the treatment effect.

[0006] To solve the above technical problems, according to one aspect of the present invention, a PEG-coated europium-based MOFs nanocomposite material is provided, comprising a core structure and a shell structure; the core structure comprises a europium-based organic framework (Eu-MOF), and silver nanoparticles (AgNPs) and paclitaxel (PTX) loaded on the europium-based organic framework; the shell structure is polyethylene glycol (PEG), which is wrapped around the outside of the core structure.

[0007] Furthermore, the element ratio of the PEG-coated europium-based MOFs nanocomposite material is C:N:O:Ag:Eu=70.84%:1.25%:23.94%:1.97%:3.25%.

[0008] According to another aspect of the present invention, provided is a method for preparing the above-mentioned PEG-coated europium-based MOFs nanocomposite material, comprising: Step 1, preparation of Eu-MOF; 1,3,5-Benzenetricarboxylic acid (BTC) ethanol-water solution and Eu(NO 3 ) 3 6H 2 O aqueous solution was mixed and reacted at room temperature, and Eu-MOF was obtained after centrifugation and washing; Step 2, preparation of AgNPs / Eu-MOF; Eu-MOF and AgNO 3 Dissolve in deionized water and stir to mix; add NaBH 4 The reaction was carried out under stirring conditions, and AgNPs / Eu-MOF was obtained after washing and centrifugation; Step 3, preparation of AgNPs / PTX / Eu-MOF@PEG; AgNPs / Eu-MOF was dispersed in ethanol, PTX solution was added, and then PEG was added under stirring conditions. The reaction was stirred at room temperature, and AgNPs / PTX / Eu-MOF@PEG was obtained after washing and drying.

[0009] Further, in step 1, 1,3,5-benzenetricarboxylic acid (BTC) ethanol-water solution and Eu(NO 3 ) 3 6H 2 The concentration of O aqueous solution is 0.5 M; the volume ratio of the two is 40:1.

[0010] Furthermore, in step 1, during washing, ethanol and water are used for washing several times each.

[0011] Furthermore, in step 2, Eu-MOF and AgNO 3 The concentrations dissolved in deionized water were 1 mg / ml and 1.5 mg / ml respectively.

[0012] Further, in step 2, NaBH 4 The concentration is 1.4 mmol / L.

[0013] Furthermore, in step three, the concentration of the PTX solution is 10 mg / mL.

[0014] Furthermore, in step three, the concentration of PEG is 10 mg / mL.

[0015] According to another aspect of the present invention, provided is the use of the above-mentioned PEG-coated europium-based MOFs nanocomposite material in the preparation of a drug for treating breast cancer.

[0016] The present invention utilizes metal coordination, physical adsorption and π-π interaction to co-assemble Eu-MOF, photothermal agent silver nanoparticles (AgNPs) and PTX, and uses PEG encapsulation to enhance their dispersibility, thereby constructing a multifunctional nanocomposite (AgNPs / PTX / Eu-MOF@PEG, EAPP) for pH response in the tumor microenvironment, which can be applied to the combined photothermal / chemotherapy treatment of breast cancer.

[0017] The nanocomposite (AgNPs / PTX / Eu-MOF@PEG, EAPP) designed by the present invention exhibits excellent fluorescence imaging and photothermal imaging capabilities, and can provide clear tumor localization and monitoring; EAPP significantly improves the therapeutic effect of breast cancer through the dual-mode combined treatment of photothermal therapy and chemotherapy. The constructed EAPP uses MOFs to load PTX, which solves the defects of poor water solubility and low bioavailability of traditional chemotherapy drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 Schematic diagram of the synthesis of EAPP and chemotherapy / photothermal combined therapy; Figure 2 In the figure, (a) and (b) are TEM images of Eu-MOF and EAPP, the inset in (b) is a schematic diagram of the lattice of AgNPs, and (c) and (d) are SEM images of Eu-MOF and EAPP; Figure 3 is the XRD pattern of Eu-MOF and EAPP; Figure 4 In the figure, (a) is the XPS general spectra of Eu-MOF, EA and EAPP, (b), (c), (d) and (e) are the detailed spectra of C1s, O1s, Ag3d and Eu3d respectively; Figure 5 In the figure, (a), (b), and (c) represent the ultraviolet (UV-Vis) spectrum, fluorescence spectrum, and infrared (FT-IR) spectrum of Eu-MOF and EAPP, respectively, and (d) is the potential characterization result of Eu-MOF, EA, EAPP, and PTX; In Figure 6, (a) is the absorbance standard curve of PTX, and (b) is the PTX drug loading corresponding to different mass ratios; Figure 7 shows the drug release of EAPP in pH 7.4 and pH 5.5 environments; Figure 8 The temperature change curves after irradiation of 2 mL PBS and 2 mL (200 μg / mL) Eu-MOF, EA, and EAPP; Fig. 9 The photothermal images after irradiation of 2 mL PBS and 2 mL (200 μg / mL) Eu-MOF, EA, and EAPP; Fig.10 In the figure, (a) is the temperature change curve after irradiation with different concentrations of EAPP, (b) is the temperature change curve of 200 μg / mL EAPP after irradiation with different powers of laser; Fig.11 The photothermal stability results of EAPP suspension after 5 consecutive irradiation cycles; Fig.12 In the figure, (a) is the temperature rise and fall curve of one irradiation cycle of the EAPP suspension, (b) is the linear relationship diagram between time and -ln(θ) of the EAPP suspension after laser irradiation; Fig.13 (a) shows the survival rates of HUVEC and MCF-7 cells after incubation with different concentrations of EAPP suspension for 4 h (n=4 / group, ns, *, **, *** represent p>0.05, p≤0.05, p≤0.01 and p≤0.001, respectively); (b) shows the survival rates of MCF-7 cells after incubation with Eu-MOF, EA, EA+NIR, PTX, EAPP, and EAPP+NIR for 4 h (n=4 / group, ns, *, **, *** represent p>0.05, p≤0.05, p≤0.01 and p≤0.001, respectively); Fig.14 CLSM images of PBS and EAPP in MCF-7 cells; Fig.15 Cell apoptosis images of ( a ) PBS, ( b ) Laser, ( c ) Eu-MOF, ( d ) EAPP, ( e ) EA +Laser, and ( f ) EAPP + Laser groups; Fig.16 are the cell apoptosis rates of PBS, Laser, Eu-MOF, PTX, EA + Laser, and EAPP + Laser groups (ns, *, **, and *** represent p>0.05, p≤0.05, p≤0.01, and p≤0.001, respectively); Fig.17Live and dead cell staining images of (a) PBS, (b) Laser, (c) Eu-MOF, (d) PTX group, (e) EAPP, (f) EA +Laser, (g) EAPP + Laser group; Fig.18 In the figure, (a) shows the distribution of EAPP in mice at different times after EAPP injection, and (b) shows the fluorescence images of the main organs and tumors of mice 9 hours after EAPP injection; Fig.19 Photothermal images of tumor-bearing mice after injection of different nanocomplexes; Fig. 20 (a) Changes in tumor volume and (b) changes in body weight of mice in different treatment groups within 14 days; Fig.21 (a) Tumor-bearing photos and (b) tumor photos of mice in different treatment groups; Fig. 22 HE staining images of tumor sections of mice in different treatment groups; Fig.23 HE staining images of main organ tissue sections of mice in different treatment groups; Fig.24 Blood indicators of mice in different treatment groups after 14 days of treatment; Fig.25 Liver and kidney indicators of mice in different treatment groups after 14 days of treatment. DETAILED DESCRIPTION

[0019] In this embodiment, Eu 3+ is the metal center, H 3 Lanthanide europium-based MOF was synthesized using BTC as raw material, and AgNPs and PTX were loaded by in-situ reduction and physical adsorption to successfully prepare AgNPs / PTX / Eu-MOF@PEG nanocomposite (EAPP). The microstructure, elemental composition and optical properties of EAPP were analyzed in detail using TEM, SEM, XPS, XRD, FT-IR, fluorescence spectroscopy and other techniques. The photothermal effect of EAPP was tested using a thermocouple probe; the drug loading rate of EAPP and the PTX release rate under different pH conditions were determined using a spectrophotometer; the biocompatibility and tumor site aggregation ability of EAPP were verified by MTT, cell uptake and other experiments; the photothermal and chemotherapy combined therapeutic effects of EAPP on breast cancer cells at the cellular level were confirmed by flow cytometry combined with live / dead cell staining technology; at the animal level, the photothermal / chemotherapy combined therapeutic effect of EAPP in vivo was verified by a tumor-bearing mouse model. 1. Preparation of AgNPs / PTX / Eu-MOF@PEG (EAPP) nanocomposites Preparation of Eu-MOF: Eu(NO 3 ) 3 6H 2 O aqueous solution (0.5 M, 1 mL). After reacting at room temperature for 60 minutes, a milky white suspension was obtained. The milky white suspension was centrifuged at 12000 rpm for 10 min to remove the unreacted precursor and collect the white precipitate, which was washed several times with ethanol and water respectively, and dried in an oven at 70°C overnight. The obtained white powder was stored at 4°C for later use.

[0020] Preparation of AgNPs / Eu-MOF (EA): 10 mg of Eu-MOF and 15 mg of AgNO were added to 10 mL of deionized water. 3 After 1.5 hours of stirring, the mixture was placed in ice water and 1 mL of NaBH was added dropwise. 4 (concentration 1.4 mmol / L) and stirred vigorously for 2 hours to finally obtain a black solution. Then, the black suspension was washed alternately with anhydrous ethanol and deionized water several times to remove unreacted precursors and impurities, and the white precipitate was collected by centrifugation, freeze-dried to obtain EA, and stored at 4 °C for future use.

[0021] Preparation of EAPP: 10 mg of EA nanocomposite was dispersed in 3 mL of ethanol, 1 mL of PTX solution (10 mg / mL) was added, and after stirring evenly under ultrasound, 1 mL of PEG (10 mg / mL) was added. The mixture was stirred at room temperature for 4 h, centrifuged and washed three times with ethanol, and dried in an oven at 70 °C overnight. The obtained black EAPP was refrigerated at 4 °C for later use.

[0022] The microstructures of Eu-MOF and EAPP were characterized by TEM and SEM. Figure 2 As shown in (a), the synthesized Eu-MOF exhibits a typical stick-like structure with an average length of about 350 nm and an average diameter of about 45 nm. Figure 2 (b) The morphology of Eu-MOF shown in Figure 2 does not change significantly after being loaded with AgNPs and PTX, and the bright spherical AgNPs loaded on the surface of Eu-MOF can be clearly observed. The distribution is relatively uniform and no flocculation is formed, indicating that the synthesis and loading methods used can ensure the integrity of the Eu-MOF structure. Figure 2 The inset of (b) shows the lattice fringes of in-situ loaded AgNPs, with a diameter of 0.18 nm. Figure 2 (c) and Figure 2 In (d), the stick-like structure of Eu-MOF and the successful loading of AgNPs can be clearly observed in the SEM image.

[0023] The crystal structures of Eu-MOF and EAPP were analyzed using XRD. 3+ Ion and rigid organic linker H 3 BTC is constructed by coordination bonding, such as Figure 3 As shown, the Eu-MOF sample has a high degree of crystallinity. After loading silver nanoparticles and PTX, the peak intensity of Eu-MOF is slightly weakened, some diffraction peaks between 28°-50° disappear, and diffraction peaks corresponding to the (111), (200), and (220) crystal planes of AgNPs appear at 38.1°, 444.4°, and 64.6°. XRD results prove that the addition of the loading component does not destroy the integrity of the crystal structure of Eu-MOF. The elemental composition of Eu-MOF and EAPP was characterized by XPS. Figure 4 In the XPS spectrum shown in (a), the peaks at 1163 eV and 1133 eV, 530 eV and 283 eV are attributed to the binding energies of Eu 3d, O 1s and C 1s orbitals, respectively. In addition, the nanocomposite loaded with AgNPs showed binding energies at 571 eV and 601 eV, 374 eV and 368 eV attributed to the Ag 3d orbital, proving the successful loading of AgNPs and having no effect on the integrity of the crystal structure of Eu-MOF. After EA was loaded with PTX, the C / O element ratio shown in XPS changed significantly, with a significant increase in the C element, which was attributed to PTX (C 47 H 51 NO 14 ) contains more C elements than O elements, which indirectly proves the successful loading of PTX. In the C element fine map ( Figure 4 (b) The peaks of C at 288.8 eV, 286.5 eV, 284.84 eV and 285.2 eV are respectively attributed to the binding energies of C=O, CO, C=C and CC groups. In the fine spectrum of O element ( Figure 4 (c)), the peaks at 533.2 eV and 531.7 eV are attributed to the C=O and CO bond binding energies. Figure 4 In the Ag element fine spectrum shown in (d), the peaks at 374.4 eV and 368.4 eV are respectively attributed to Ag 3d 3 / 2 and Ag 3d 5 / 2 The binding energy of Figure 4 In (e), multiple characteristic peaks can be observed in the fine spectrum of Eu element, among which 1164.5 eV and 1156.6 eV are attributed to Eu 3d 3 / 2 The binding energy of 1135.3 eV is attributed to Eu 3d 5 / 2The binding energy of the three main peaks is assigned to Eu 3+ Indicates Eu 3+ The peak at 1143.1 eV is attributed to the presence of Eu metal, indicating that there is no oxidized Eu in EAPP. 2 O 3 No redox reaction occurred during the synthesis of EAPP. The XPS total spectrum and fine spectrum results confirmed that EAPP is mainly composed of C, O, Ag and Eu elements, and through calculation, it can be known that the element content ratio of EAPP is approximately C:N:O:Ag:Eu≈70.84%:1.25%:23.94%:1.97%:3.25%.

[0024] UV-Vis spectra of nanocomposites such as EAPP, EA, and Eu-MOF Figure 5 (a) As shown. The maximum absorption peak of pure PTX is at 240 nm, and Eu-MOF exhibits spectral absorption in the visible light region, and the absorption intensity gradually decreases with increasing wavelength. The dip in the absorption curve around 235 nm is attributed to the fact that AgNPs are affected by the localized surface plasmon resonance (LSPR) effect. Their limited volume causes the excited electrons to condense and oscillate under the magnetic field of the incident light point, resulting in strong absorption and scattering effects. The ultraviolet absorption peak of EAPP shows the characteristic peaks of PTX and AgNPs. The fluorescence properties of Eu-MOF, PTX, EA, and EAPP were studied using fluorescence spectroscopy. Figure 5 As shown in (b), PTX has no obvious fluorescence, while Eu-MOF has Eu in the range of 600-720 nm. 3+ After loading with AgNPs and PTX, the fluorescence intensity of EAPP decreased significantly, suggesting that its electron-hole recombination rate is low; the resulting significant reduction in the recombination velocity of photogenerated carriers makes it more suitable for photothermal therapy. Figure 5 (c) shows that Eu-MOF has a peak at 534 cm -1 and 456cm -1 The peak at corresponds to H 3 BTC connects the Eu-O bond vibration of Eu ions. In addition, the -1 , 1708 cm -1 , 2346cm -1 and 706 cm -1The peaks are attributed to C=C, C=O, O=C=O and CH bond stretching vibration. Compared with the FT-IR image of Eu-MOF, the functional groups of EA are basically consistent with those of Eu-MOF, and the loading of AgNPs has no effect on the properties of the functional groups on the surface of Eu-MOF. -1 The newly appeared narrow peak at is consistent with the characteristic peak of PTX. Finally, the zeta potential results of Eu-MOF, PTX, EA and EAPP measured by Zeta potential instrument are shown as follows: Figure 5 (d). As shown in the figure, the potentials of Eu-MOF, EA, PTX and EAPP are 3.37, 11.2, -5.33 and 7.39, respectively. Due to the loading of AgNPs, the H + The potential increases; after the introduction of PTX and PEG with negative surface charge, the EAPP potential decreases further.

[0025] The above characterization results verified the successful synthesis of EAPP.

[0026] 2. EA drug loading determination The standard curve of PTX was drawn by spectrophotometry. Figure 6 (a) is shown. The drug loading results of different mass ratios are shown in Figure 6 (b) shows that when the mass ratio of PTX to EA is 1:1, the drug loading reaches a maximum of 26.84%. As the relative mass of the drug increases, the drug loading will gradually decrease. Therefore, the mass ratio of PTX to EA is 1:1.

[0027] 3. Study on the release of drug PTX The pH-responsive drug delivery mechanism is the most common method to improve the shortcomings of traditional chemotherapeutic drugs, such as poor water solubility and poor bioavailability. Among various stimuli, pH-responsive drug release is one of the most commonly used strategies. Figure 7 As shown in the figure, there are large differences in the release of PTX under different pH conditions. In an environment of pH = 7.4, PTX can only release 19.2%, while at pH = 5.5 (simulating the tumor microenvironment), the PTX release rate reaches 52.4%. The high release rate in the environment of pH 5.5 may be due to the low pH environment Eu 3+ With H 3 The coordination bonds between BTC molecules are broken and disintegrated. In addition, the drug release behavior of PTX is time-dependent and can be gradually released over a period of up to 36 hours.

[0028] 4. EAPP in vitro photothermal effect

[0029] To verify the photothermal effect of EAPP, the temperature changes were recorded after laser irradiation. Figure 8As shown, after 15 minutes of laser irradiation, the temperatures of EA and EAPP rose to 52.3 ℃ and 52.6 ℃, respectively, which were about 28 ℃ higher than before irradiation, and there was no significant difference in the temperature between the two groups. The difference is that the temperature of PBS and Eu-MOF suspension as the control group only rose to 27.6 ℃ and 27.9 ℃ after laser irradiation of the same time and intensity. Eu-MOF and PBS do not have photothermal effect; after loading with AgNPs, EA and EAPP showed good photothermal ability, indicating that their photothermal effect mainly comes from AgNPs. In order to verify the imaging effect of photothermal capacity, an infrared thermal imager was used for shooting. As shown Fig. 9 As shown, EAPP has good photothermal imaging effect and can be applied to photothermal imaging.

[0030] 4.1 EAPP photothermal effect evaluation The photothermal effect of EAPP is affected by different laser irradiation powers and different EAPP concentrations. In order to achieve the hyperthermia effect without affecting the surrounding normal cells, the temperature needs to be controlled at about 50 to 55 °C. Fig.10 As shown, under the same laser irradiation power, the highest temperature of EAPP with different concentrations and the same concentration of EAPP with different irradiation powers can reach 50-50.6 ℃, which proves that EAPP has a good photothermal effect.

[0031] 4.2 Calculation of thermal cycling stability and photothermal conversion efficiency (η) of EAPP After 5 consecutive cycles of irradiation-cooling, the temperature change of the EAPP solution is recorded as follows: Fig.11 After 5 cycles of heating and cooling, the highest temperature of EAPP was basically maintained at 53.3 ℃, showing good photothermal stability.

[0032] Through the heating-cooling curve, a single heating-cooling curve can be obtained ( Fig.12 (a) ) and the linear relationship between time and -ln(θ) ( Fig.12 (b) ), find τ s =324. In addition, the absorbance of EAPP at 660 nm is A=2.276, the laser power is I=1.8 W, ΔT=31.8 ℃, and the specific heat capacity of water is C water =4.2J / (kg*℃), the mass of the solution is m=2.357 g, and the photothermal conversion efficiency of EAPP is calculated to be η=45.70%, which proves its good photothermal effect and can be used as a photothermal material.

[0033] 5. EAPP Cell Experiment

[0034] 5.1 Cytotoxicity assay Good biocompatibility of nanocomposites is an extremely important prerequisite for their potential anti-tumor applications. Since EAPP can release PTX slowly under low pH conditions, experiments were conducted using EAPP and EAP (without PTX) nanocomposites. Fig.13 As shown in (a), the biocompatibility of EAPP was detected by MTT method in cancer cells (MCF-7 cells) and normal cells (HUVEC cells). The EAP nanocomplex did not cause obvious damage to the cells at a high concentration of 200 μg / mL, and the cell viability was always maintained above 90%, proving that the EAP nanocomplex has good biocompatibility. However, due to the presence of the chemotherapeutic drug PTX, EAPP caused varying degrees of cell viability decline in HUVEC cells and MCF-7 cells, but thanks to the pH-responsive release ability of Eu-MOF and the encapsulation of PEG, the cell viability of normal cells decreased less and the impact was lower.

[0035] The phototoxicity of EAPP was evaluated by MTT assay. Fig.13 (b) As shown. The viability of MCF-7 cells after incubation with Eu-MOF was not significantly affected, indicating that Eu-MOF has good biocompatibility; after photothermal therapy (EA+NIR) and chemotherapy (EAPP), cell viability decreased significantly, indicating that photothermal therapy and chemotherapy have therapeutic effects; at the same time, there was no significant difference between the PTX group and the EAPP group (n>0.05), proving that there was no significant difference in the cell killing ability of EAPP and PTX. The combined treatment (EAPP+NIR) showed significant differences compared with the single photothermal therapy (EA+NIR) and chemotherapy (EAPP, PTX) (n≤0.01), indicating that the cell killing ability of the combined treatment was significantly better than the single application of chemotherapy or photothermal therapy, which fully demonstrated the therapeutic effect of the combined treatment.

[0036] 5.2 Cell imaging experiments like Fig.14 As shown, bright red fluorescence can be observed after incubation with MCF-7 cells for 4 hours, proving that EAPP has good cell uptake ability and potential for cell imaging applications. The experiment proved that EAPP can be effectively taken up by MCF-7 cells and can be used in cell imaging.

[0037] 5.3 Cell apoptosis assay Fig.15 and Fig.16The results and apoptosis rates of each treatment group. The apoptosis rate of MCF-7 cells treated with Laser and Eu-MOF was similar to that of the blank control group (PBS group), indicating that laser irradiation and co-incubation with Eu-MOF hardly induced apoptosis. The PTX and EA+Laser groups were chemotherapy or photothermal therapy treatment groups, and their apoptosis rates reached 30.38% and 46.08%, respectively, proving that chemotherapy and photothermal therapy have the ability to induce apoptosis. The apoptosis rate of the EAPP+Laser combined treatment group reached 86.9%, indicating that the combined chemotherapy / photothermal therapy of EAPP can effectively induce apoptosis of cancer cells and has a good killing effect on breast cancer cells.

[0038] 5.4 Cell combination therapy experiments Live and dead cell staining experiments were used to evaluate the therapeutic effects of (a) PBS, (b) Laser, (c) Eu-MOF, (d) PTX, (e) EAPP, (f) EA + Laser, and (g) EAPP + Laser groups. The results are shown in Fig.17 As shown. There was almost no cell death in the PBS group and the Laser group, while only a very small amount of cell death occurred in the Eu-MOF group. In the experiment, some cells in the experimental group that received chemotherapy or photothermal therapy alone died, but in the chemotherapy / photothermal combined therapy group, almost all cells died. These results show that the combined treatment has a significant advantage over single treatment.

[0039] 6. Antitumor effect of EAPP in vivo

[0040] 6.1 In vivo distribution behavior and fluorescence / photothermal imaging like Fig.18 As shown in the figure, in order to analyze the distribution behavior and fluorescence imaging ability of EAPP in mice, a small animal fluorescence imager was used to collect images of mice and organs at different time points. After EAPP was injected into the mouse, it gradually accumulated in the tumor site over time, causing the fluorescence intensity in the tumor area to gradually increase, reaching a peak at 9 hours, and the fluorescence signal almost disappeared at 24 hours, proving that EAPP can effectively accumulate and metabolize in the tumor site. Starting from 3 hours after injection, fluorescent signals gradually appeared in the liver of the mouse and increased with time, which was consistent with the results of the experiment. Fig.18 (b) is consistent with the fluorescence imaging of the liver, a major organ of mice, proving that EAPP is metabolized from the mouse liver.

[0041] Photothermal imaging of mice Fig.19As shown in the figure, under laser irradiation, the photothermal signal of the tumor site of mice increased to varying degrees according to different injections. In the PBS group and Eu-MOF group, the intensity of the photothermal signal of mice did not change significantly, and the final photothermal signal intensity of the two groups was close, indicating that Eu-MOF did not have photothermal capacity. After 15 minutes of 660 nm laser irradiation of EA nanocomposites and EAPP, the photothermal signal of the tumor site was significantly enhanced, and the temperature of mice treated with EAPP rose more evenly, with the highest temperature reaching 51.6 ℃, proving that EAPP has a good photothermal effect.

[0042] 6.2 Antitumor Effect of EAPP The average tumor volume of tumor-bearing mice reached 100 mm 3 Afterwards, the mice were randomly divided into 6 groups, each containing 5 mice. Each mouse was marked separately, and the weight, tumor size and tumor image of each mouse were recorded every other day during the subsequent 14-day intervention period. Fig. 20 The weight changes and relative changes of tumor volume of mice. Compared with the blank control group (PBS group), there was no significant difference in the weight of mice in each experimental group, indicating that 660 nm laser irradiation and EAPP had no obvious toxic side effects on mice. Fig. 20 As can be seen in (b), the relative tumor volumes of mice in each treatment group can be divided into three general trends: the relative tumor volumes of the PBS group, Laser group, and Eu-MOF group increased rapidly within 14 days, and the increase was relatively fast; the tumor volumes of the EAPP and EA+Laser groups, which were chemotherapy and photothermal therapy groups, still increased to varying degrees, but the increase and final tumor volumes were significantly lower than those of the first three groups; finally, the relative tumor volume of the EAPP+Laser combined treatment group showed a downward trend, indicating that the combined application of chemotherapy and photothermal therapy effectively curbed tumor growth and development. Each group of mice underwent different treatment interventions, Fig.21 The following are photos of mice and tumors during the treatment process. During the 14-day treatment period, the tumors of the treated mice were significantly reduced. Fig.21 (b) The tumor image shows that the sizes of tumors treated with PBS, Laser, and Eu-MOF are almost the same. The tumors in the chemotherapy (EAPP) group and the photothermal therapy (EA+Laser) group are relatively smaller, while the tumor in the chemotherapy + photothermal therapy (EAPP+Laser) group is the smallest. The treatment effect is significant, indicating that the combined treatment has the effect of inhibiting tumor growth. After the intervention period, the mice were killed and the tumor sites were taken for HE staining and sectioning. Fig. 22As shown, in the combined treatment group, tumor cells were observed to shrink and change in morphology, and a large number of cells underwent apoptosis. Compared with the experimental group that received only a single treatment, only some cells underwent apoptosis, and no signs of apoptosis or lysis were observed in tumor cells in other groups. This indicates that EAPP combined with photothermal / chemotherapy can induce apoptosis and necrosis of tumor cells, effectively ablate tumor cells, and has the potential to treat breast cancer.

[0043] 6.3 Biosafety Assessment The biosafety of EAPP was verified using the eye blood and major organs of tumor-bearing mice, and blood routine analysis, biochemical index analysis, and HE staining experiments were performed. Fig.23 ), there was no obvious abnormality in the main organs of mice after different intervention treatments compared with the blank control group, and no inflammatory response or allergic symptoms were observed, further proving that EAPP has low toxic side effects in vivo and good biosafety.

[0044] The results of routine blood analysis and blood biochemical index analysis of mice are as follows Fig.24 , 25 There were no significant differences in the blood routine and blood biochemical indicators of mice in each group, which proved that EAPP has good biosafety.

[0045] In summary, the AgNPs / PTX / Eu-MOF@PEG nanocomposite (EAPP) obtained by the present invention has a drug release rate of 52.4% under a pH of 5.5. The loading of AgNPs enables it to exhibit a good photothermal effect under 660 nm laser irradiation, and can be heated to 50-55 °C, with photothermal treatment effect and photothermal imaging capability. 3+ Good fluorescence properties, EAPP has bright red fluorescence and can be used for tumor cell and mouse tumor fluorescence imaging. The MTT method proved that EAPP has good biocompatibility; cell imaging experiments and live-dead cell staining experiments proved that the nanocomplex can effectively enter cancer cells and kill breast cancer cells through chemotherapy / photothermal combined therapy. Animal experiments have proved that EAPP can fully exert its imaging and tumor killing abilities in vivo and has high biosafety.

Claims

1. A PEG-coated europium-based MOFs nanocomposite material, characterized in that: It comprises a core structure and a shell structure; the core structure comprises a europium-based organic framework (Eu-MOF), and silver nanoparticles (AgNPs) and paclitaxel (PTX) loaded on the europium-based organic framework; the shell structure is polyethylene glycol (PEG) wrapped outside the core structure.

2. The PEG-coated europium-based MOFs nanocomposite material according to claim 1, characterized in that: The ratio of elements it contains is C:N:O:Ag:Eu=70.84%:1.25%:23.94%:1.97%:3.25%.

3. The method for preparing the PEG-coated europium-based MOFs nanocomposite material according to claim 1, characterized in that: include: Step 1, preparation of Eu-MOF; 1,3,5-benzenetricarboxylic acid (BTC) ethanol-water solution and Eu(NO3)3·6H2O aqueous solution were mixed and reacted at room temperature, and Eu-MOF was obtained after centrifugation and washing; Step 2, preparation of AgNPs / Eu-MOF; Eu-MOF and AgNO3 were dissolved in deionized water and stirred to mix; NaBH4 was added to react under stirring conditions, and AgNPs / Eu-MOF was obtained after washing and centrifugation; Step 3, preparation of AgNPs / PTX / Eu-MOF@PEG; AgNPs / Eu-MOF was dispersed in ethanol, PTX solution was added, and then PEG was added under stirring conditions. The reaction was stirred at room temperature, and AgNPs / PTX / Eu-MOF@PEG was obtained after washing and drying.

4. The preparation method according to claim 3, characterized in that: In step 1, the concentrations of the 1,3,5-benzenetricarboxylic acid (BTC) ethanol-water solution and the Eu(NO3)3·6H2O aqueous solution are both 0.5 M; the volume ratio of the two is 40:

1.

5. The preparation method according to claim 4, characterized in that: In step 1, during washing, wash with ethanol and water several times each.

6. The preparation method according to claim 3 or 5, characterized in that: In step 2, the concentrations of Eu-MOF and AgNO3 dissolved in deionized water were 1 mg / ml and 1.5 mg / ml, respectively.

7. The preparation method according to claim 6, characterized in that: In step 2, the concentration of NaBH4 is 1.4 mmol / L.

8. The preparation method according to claim 3 or 7, characterized in that: In step 3, the concentration of the PTX solution is 10 mg / mL.

9. The preparation method according to claim 8, characterized in that: In step 3, the concentration of PEG was 10 mg / mL.

10. Use of the PEG-coated europium-based MOFs nanocomposite material according to claim 1 or 2 in the preparation of drugs for treating breast cancer.

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