CN115317608A - Nanocomposite with X-ray mediated photodynamic therapy effect and preparation method and application thereof - Google Patents

Nanocomposite with X-ray mediated photodynamic therapy effect and preparation method and application thereof Download PDF

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CN115317608A
CN115317608A CN202211263799.4A CN202211263799A CN115317608A CN 115317608 A CN115317608 A CN 115317608A CN 202211263799 A CN202211263799 A CN 202211263799A CN 115317608 A CN115317608 A CN 115317608A
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宋坤
刘宗明
李金凯
孔北华
刘斌
姚舒
刘昶
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Qilu Hospital of Shandong University
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Abstract

The invention discloses a nano composite material with an X-ray mediated photodynamic therapy effect and a preparation method and application thereof, belonging to the technical field of scintillation crystal preparation. The invention is realized by adding the active ingredient into Lu 3 Al 5 O 12 The surface of Mn/Ca nano scintillator is loaded with photosensitizer Ce6, modified with PEG and coated with SiO 2 The water solubility and the biocompatibility of the nano scintillator composite material are increased so as to realize the radiotherapy sensitization and XEffects of radiation-mediated photodynamic therapy. Experiments prove that the prepared nano composite material with the X-ray mediated photodynamic therapy effect can induce tumor cells to generate more active oxygen under the excitation of X-rays, so that more DNA (deoxyribonucleic acid) damage and cell apoptosis are caused, and the tumor inhibition rate of the nano composite material in a mouse model reaches 87 percent (the tumor inhibition rate of a pure X-ray group is 35 percent), so that the nano composite material has a good therapy effect.

Description

Nanocomposite with X-ray mediated photodynamic therapy effect and preparation method and application thereof
Technical Field
The invention relates to the technical field of scintillation crystal preparation, in particular to a nano composite material with an X-ray mediated photodynamic treatment effect, and a preparation method and application thereof.
Background
Tumor radiotherapy is a method of treating tumors with radiation. Radiotherapy is a local treatment method, and the radiation used includes alpha, beta and gamma rays generated by radioactive isotopes, and x-rays, electron beams, proton beams and other particle beams generated by various x-ray treatment machines or accelerators. About 70% of cancer patients need radiotherapy in the process of treating cancer, about 40% of cancers can be cured by radiotherapy, and the radiotherapy is one of the main means for treating malignant tumors.
Radiotherapy is an important approach to cancer treatment. However, the ideal therapeutic effect cannot be achieved by radiotherapy alone, so that a new scheme needs to be designed to enhance the therapeutic effect of radiotherapy.
Photodynamic therapy is a novel cancer treatment means. However, light has poor penetration in tissues, limiting its use in deep tissues. Therefore, devising a new approach to circumvent the drawback of insufficient tissue penetration of light is an urgent problem to be solved to achieve the application of photodynamic therapy in deep tumor tissue.
Disclosure of Invention
The invention aims to provide a nano composite material with an X-ray mediated photodynamic treatment effect, and a preparation method and application thereof. By reaction on Lu 3 Al 5 O 12 The surface of Mn/Ca nano scintillator is loaded with photosensitizer Ce6, modified with PEG and coated with SiO 2 The water solubility and the biocompatibility of the nano scintillator composite material are increased, so that the radiotherapy sensitization and X-ray mediated photodynamic therapy effect are realized.
In order to achieve the purpose, the invention provides the following technical scheme:
the invention adopts one of the technical schemes: provided is a method for preparing a nanocomposite material having an X-ray mediated photodynamic therapy effect, comprising the steps of:
（1）Lu 3 Al 5 O 12 the synthesis of Mn/Ca nano scintillators:
will contain Lu 3+ 、Al 3+ 、Mn 2+ And Ca 2+ Is added dropwise to NH 4 HCO 3 Reacting in the solution, drying and calcining the precipitate obtained after the reaction is finished, and adding the calcined product and polyethylene glycol into water for reaction to obtain Lu 3 Al 5 O 12 Mn/Ca nano-scintillators;
（2）Lu 3 Al 5 O 12 synthesizing a Mn/Ca-Ce6 nano composite material:
the Lu is mixed 3 Al 5 O 12 Mixing Mn/Ca nano scintillator with chlorin e6, N-hydroxysuccinimide (NHS) and N- (3-dimethylaminopropyl) -N' -ethylcarbodiimide hydrochloride (EDC) in a solvent, reacting in the dark, centrifuging, precipitating and drying to obtain Lu 3 Al 5 O 12 Mn/Ca-Ce6 nanocomposites;
（3）Lu 3 Al 5 O 12 :Mn/Ca-Ce6@SiO 2 synthesis of the nano composite material:
mixing the Lu 3 Al 5 O 12 Dispersing a Mn/Ca-Ce6 nano composite material in a solvent, dropwise adding a hexadecyl trimethyl ammonium aqueous solution for reaction, and removing the solvent after the reaction is finished; adding the precipitate and tetraethyl silicate into an ethanol water solution of sodium hydroxide, heating for reaction, centrifuging and washing the precipitate to obtain Lu 3 Al 5 O 12 :Mn/Ca-Ce6@SiO 2 A nanocomposite material.
Mn added according to the invention 2+ Can be used as activator for generating red light emission, ca 2+ Then enhance Mn as a charge compensator 2+ Red light emission of (a). Mn 4+ The ion having 3d 3 By substituting Ge with an octahedral coordination environment in a suitable matrix 4+ ， Al 3+ ， Si 4+ Plasma exists stably, lu in the experiment 3 Al 5 O 12 Al in (1) 3+ The octahedral coordination environment is very suitable for Mn 4+ And Mn 4+ And Al 3+ Having similar ionic radii can be substituted by Mn 4+ Instead, exhibiting red emission and absorption of visible and near ultraviolet light. Mn (Mn) 4+ The superior spectral characteristics provide broadband excitation and narrow-band emission for its unique electronic structure. Due to Mn 4+ The high valence state of the ion gives it a strong crystal field environment, resulting in its confinement with spin and parity 2 E g → 4 A 2 The transition process corresponds to a red emission peak with an emission spectrum around 670 nm. Due to Mn 4+ Non-equivalent substitution of Al 3+ So that the fluorescent particles influence Mn because the fluorescent particles do not satisfy the charge neutrality principle 4+ The fluorescence property of the fluorescent material is greatly reduced due to the transition luminescence of (2). Ca 2+ By introducing the above system in place of Al 3+ Can effectively neutralize Mn 4+ The resulting charges and defects are not equivalently replaced, thereby significantly improving their fluorescence emission.
The addition of NHS and EDC in the invention can adjust the charge property and material surface groups in the solution, so that the ce6 is easily fixed on the material surface. carboxyl-COOH in Ce6 and-OH in polyethylene glycol on the surface of fluorescent particles can be combined with guanidino (NH) on the surface of NHS and EDC 2 ) 2 C = NH (one carbon with 2 amino groups, a double bond with one imine group, with basicity comparable to KOH) amino group reaction, by acid-base neutralization, i.e. hydroxyl dehydridation + Amino radical to H + . The mechanism is that the oxygen induction of carbonyl in carboxyl attracts electrons, so that the electronegativity of-OH oxygen in the carboxyl is weakened to H + Thereby making Ce6 more firmly fixed on the surface of the fluorescent nanoparticle.
Preferably, the Lu in step (1) 3+ 、Al 3+ 、Mn 2+ And Ca 2+ The molar ratio of (a) to (b) is 3; the NH 4 HCO 3 The concentration of the solution was 2M; the drying temperature is 80 ℃; the calcining temperature is 1000 ℃, and the time is 4h; the polyethylene glycol is PEG600, and the mass-to-volume ratio of the calcined product to the PEG600 is 10mg.
Preferably, the solution in step (1) contains Lu 3+ 、Al 3+ 、Mn 2+ And Ca 2+ With said NH 4 HCO 3 The reaction time of the solution is 30min; the reaction time of the calcined product and the polyethylene glycol is 12h.
Preferably, the Lu in step (2) 3 Al 5 O 12 The mass ratio of Mn/Ca nano scintillator, chlorin e6, N-hydroxysuccinimide and N- (3-dimethylaminopropyl) -N' -ethylcarbodiimide hydrochloride is (100); the reaction time is 24h.
Preferably, the Lu in step (3) 3 Al 5 O 12 The mass to volume ratio of the Mn/Ca-Ce6 nano composite material to the tetraethyl silicate is 100mg.
Preferably, the aqueous ethanol solution of sodium hydroxide in step (3) is prepared by mixing water, ethanol, and 2M sodium hydroxide solution in a volume ratio of 40.
Preferably, the concentration of the aqueous solution of cetyltrimethylammonium in step (3) is 10mg/mL, and the dropping speed is 5mL/min.
Preferably, step (3) further comprises subjecting the Lu to 3 Al 5 O 12 :Mn/Ca-Ce6@SiO 2 Removing impurities from the nano composite material, specifically, removing impurities from the Lu 3 Al 5 O 12 :Mn/Ca-Ce6@SiO 2 Nanocomposite with NH 4 NO 3 Dispersing in ethanol, reacting at 50 deg.C for 2 hr, washing with water, and drying to remove impurities.
The second technical scheme of the invention is as follows: there is provided a nanocomposite having an X-ray mediated photodynamic therapy effect prepared according to the above method for preparing a nanocomposite having an X-ray mediated photodynamic therapy effect.
The second technical scheme of the invention is as follows: provides the application of the nano composite material with the X-ray mediated photodynamic treatment effect in the preparation of radiotherapy medicaments.
Preferably, the radiotherapeutic agent is an X-ray mediated adjuvant therapy agent.
The invention has the following beneficial technical effects:
the invention is realized by adding the active ingredient into Lu 3 Al 5 O 12 The surface of Mn/Ca nano scintillator is loaded with photosensitizer Ce6, modified with PEG and coated with SiO 2 The water solubility and the biocompatibility of the nano scintillator composite material are increased, so that the radiotherapy sensitization and X-ray mediated photodynamic therapy effect are realized. Experiments prove that the prepared nano composite material with the X-ray mediated photodynamic therapy effect can induce tumor cells to generate more active oxygen under the excitation of X-rays, so that more DNA (deoxyribonucleic acid) damage and cell apoptosis are caused, and the tumor inhibition rate of the nano composite material in a mouse model reaches 87 percent (the tumor inhibition rate of a pure X-ray group is 35 percent), so that the nano composite material has a good therapy effect.
Drawings
FIG. 1 is a representation of the Ce6 starting material and the LAMC intermediate and the LAMCCS final product of example 1; wherein a is an ultraviolet-visible light absorption spectrum of the monomers LAMCC and Ce6, b is a Fourier transform infrared spectrum of the monomers LAMC and LAMCC, c is an X-ray diffraction spectrum of the monomers LAMC and LAMCC, d is a particle size and Zeta potential detection diagram of the monomers LAMC, LAMCC and LAMCCS, and e is an X-photoluminescence spectrum of the monomers LAMC and LAMCCS and an ultraviolet-visible light absorption spectrum of Ce 6.
FIG. 2 is a representation of the end product LAMCCS of example 1; wherein a is a transmission electron microscope and high-resolution transmission electron microscope microscopic image of the LAMCCS, b is an energy dispersion spectrum of the LAMCCS, and c is an element positioning image of the LAMCCS.
FIG. 3 is a graph of the X-ray mediated photodynamic effects of LAMC, LAMCCS in example 1; wherein a is the singlet oxygen produced by LAMC and LAMCCS under different conditions: ( 1 O 2 ) The intensity of the signal of the excited detection index, b is HeLa cellReactive oxygen species are produced under different stimulation conditions.
FIG. 4 shows the in vitro tumor killing effect of LAMC, LAMCCS in example 1; wherein a is a cell plate cloning experiment of the HeLa cell under different condition stimulations, b is a DNA damage level (blue is cell nucleus and green is DNA double-chain damage) of the HeLa cell under different condition stimulations, and c is an apoptosis level of the HeLa cell under different condition stimulations.
FIG. 5 shows the in vivo tumor killing effect of LAMC, LAMCCS in example 1; wherein a is the tumor image of the mice of different treatment groups, b is the tumor volume growth curve of the mice of different treatment groups, and c is the weight change curve of the mice of different treatment groups.
Detailed Description
Reference will now be made in detail to various exemplary embodiments of the invention, the detailed description should not be construed as limiting the invention but rather as a more detailed description of certain aspects, features and embodiments of the invention. It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.
Further, for numerical ranges in this disclosure, it is understood that each intervening value, between the upper and lower limit of that range, is also specifically disclosed. Every intervening value, to the extent any stated or intervening value in a stated range, and every other stated or intervening value in that stated range, is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included or excluded in the range.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention.
As used herein, the terms "comprising," "including," "having," "containing," and the like are open-ended terms that mean including but not limited to.
Example 1
Preparation of nanocomposite materials with X-ray mediated photodynamic therapeutic effect:
（1）Lu 3 Al 5 O 12 synthesis of Mn/Ca Nanoborant (LAMC):
adding 0.1M Lu 2 O 3 Dispersing into 200mL of deionized water, adding 10mL of concentrated nitric acid and heating to 90% o C was dissolved and then diluted to a final concentration of 0.5M. Lu (NO) 3 ) 3 、Al(NO 3 ) 3 、MnCl 2 And Ca (NO) 3 ) 2 According to the chemical formula: lu (Lu) 3 (Al 0.9996 Mn 0.0002 Ca 0.0002 ) 5 O 12 Proportionally mixing the components in deionized water, and magnetically stirring for 30 minutes to obtain a precursor salt solution. Then, the precursor salt solution was slowly dropped into 150mL of 2M NH at room temperature 4 HCO 3 In solution and the reaction was allowed to continue for 30 minutes. Impurities were removed by washing centrifugation with deionized water and ethanol, and the resulting precipitate was dried at 80 ℃ and calcined at 1000 ℃ for 4 hours, and the calcined product (10 mg) was mixed with PEG600 (4 mL) in 10mL of deionized water for 12 hours in order to increase the water solubility of the calcined product. The mixed solution was centrifuged at 10000 rpm for 5 minutes, and washed with water and centrifuged 3 times to remove excess PEG600, to obtain LAMC, which was stored at 4 ℃ for further use.
（2）Lu 3 Al 5 O 12 Synthesis of Mn/Ca-Ce6 nanocomposite (LAMCC):
LAMC (100 mg), chlorin e6 (Ce 6,1 mg), N-hydroxysuccinimide (NHS, 5 mg) and N- (3-dimethylaminopropyl) -N' -ethylcarbodiimide hydrochloride (EDC, 3 mg) obtained in step (1) were mixed in 100ml ethanol and reacted for 24 hours in the absence of light. The dispersion solution was centrifuged at 8000 for 5 minutes and washed three times with ethanol and water. The reaction was then lyophilized for 24h to obtain LAMCC.
（3）Lu 3 Al 5 O 12 :Mn/Ca-Ce6@SiO 2 Synthesis of nanocomposites (LAMCCS), i.e. nanocomposites with X-ray mediated photodynamic therapeutic effect:
to further enhance the nano-meterBiocompatibility of the material, and the LAMCCS nano composite material is obtained by a method of hydrolyzing tetraethyl silicate (TEOS). LAMCC (100 mg) was dispersed in 20mL of ethanol, and Cetyltrimethylammonium (CTAB) was dissolved in warm water (10 mg/mL concentration) at 40 ℃ and added dropwise to the LAMCC ethanol dispersion solution at a rate of 5mL/min. The solution was heated to 80 ℃ to evaporate excess ethanol and centrifuged to give a reaction mass. Deionized water (40 mL), ethanol (6 mL), naOH solution (300. Mu.L, 2M) were mixed and placed in a three-neck flask, heated to 80 ℃ and the LAMCC reactant and TEOS (300. Mu.L) were added dropwise to the flask and reacted for 10 minutes. After cooling, centrifugation and repeated washing with ethanol gave lamcscs. Finally, NH is added 4 NO 3 (0.8 g) was dispersed in 50mL of ethanol with LAMCCS and reacted at 50 ℃ for 2 hours to remove unreacted CTAB. After the reaction was complete, the LAMCCS was washed 3 times with water and lyophilized for subsequent use.
FIG. 1 is a representation of the Ce6 starting material and the intermediate and end products LAMCCS of example 1; wherein a is an ultraviolet-visible light absorption spectrum of the monomer LAMCC and Ce6, b is a Fourier transform infrared spectrum of the monomer LAMC and the monomer LAMCC, c is an X-ray diffraction spectrum of the monomer LAMC and the monomer LAMCC, d is a particle size and Zeta potential detection diagram of the monomer LAMC, the monomer LAMCC and the monomer LAMCCS, and e is an X-photoluminescence spectrum of the monomer LAMC, the monomer LAMCCS and an ultraviolet-visible light absorption spectrum of Ce 6.
In FIG. 1, a shows UV-VIS absorption spectra of LAMCC and Ce6 dispersed in ethanol and measured by U-4100 spectrophotometer (HITACHI, japan); the results showed that the absorption peak of Ce6 was around 670nm and that of LAMCC around 670nm, demonstrating that Ce6 is loaded on LAMCC.
From FIG. 1 b is a record of 4000-500 cm by a Nicolet 670 FTIR spectrometer (Thomas Nicolet, USA) -1 Fourier transform infrared spectroscopy of the spectral range; as can be seen, 3488 cm -1 、2358 cm -1 And 1641 cm -1 The absorption peak is generated by the symmetric stretching vibration and bending vibration of water molecules of the combined water, and the absorbed water is mutually combined through H-O, so that the inertia moment and the molecular cluster are relatively large. Meanwhile, the positions of the absorption peaks of N-H and O-H are asymmetrically superposed. 2358 cm of -1 、1394 cm -1 And 1096 cm -1 The absorption peak is due to stretching vibration and bending vibration of the carbonyl group of Ce 6. These demonstrate that Ce6 is loaded onto LAMC.
In FIG. 1, c represents the crystal structure of the material studied by X-ray diffractometry using Cu- -K.alpha.as the target, and the 2 theta angle measured was in the range of 10 to 80 degrees, showing the crystallinity of LAMC and LAMCCS and Lu 3 Al 5 O 12 In line, the invention successfully synthesizes the LAMCCS composite nano material through electrostatic and chemical adsorption.
In FIG. 1 d shows that the LAMCCS has a particle size of about 125 nm; due to Mn 4+ /Ca 2+ For Lu 3 Al 5 O 12 Lu in matrix 3+ As an inequivalent alternative, LAMC showed a potential of +36.8 mV, which made Ce6 (-21.2 mV) more loaded onto LAMC, coating SiO 2 The potential of LAMCCS was-18.6 mV.
FIG. 1, e, is an X-ray photoluminescence spectrum for detecting LAMC, LAMCS at room temperature using a fluorescence spectrometer (FP-6500, JASCO Co., japan) equipped with a phi 60-mm integrating sphere (ISF-513, JASCO, tokyo, japan) and a 150-W xenon lamp as excitation light sources, and an ultraviolet-visible absorption spectrum for detecting Ce6 using a U-4100 spectrophotometer (HITACHI, japan). Under the X-ray excitation of 6Gy, the LAMC has emission peaks in 670, 686 and 705 nanometer wave bands. Generation of emission peak at 686 nm based on Mn 4+ ( 2 E- 4 A2 Red radiation transition, generation of emission peaks at 670, 705 based on Mn 4+ Interaction of phonon and electron transitions. Ce6 has an absorption peak at 660 nm, and the absorption spectrum of Ce6 and the light-induced spectrum of LAMC have large-area coincidence at a red light position, so that the rationality of exciting Ce6 by using LAMC is proved.
FIG. 2 is a representation of the end product LAMCCS of example 1; wherein a is a transmission electron microscope and high-resolution transmission electron microscope microscopic image of the LAMCCS, b is an energy dispersion spectrum of the LAMCCS, and c is an element positioning image of the LAMCCS.
As can be seen from a in FIG. 2, transmission electron microscopy shows the form of the soft agglomerates (soft agglomerates) of LAMCC, and high resolution transmission electron microscopy shows the interplanar spacing of the materialInterplanar spacing of 0.484 nm, lu 3 Al 5 O 12 The lattice planes (lattice planes) of (1) are uniform.
By observing the energy dispersion spectrum and the element distribution of b and c in FIG. 2, the successful doping of Mn/Ca into Lu is shown 3 Al 5 O 12 And the surface of the LAMCC is coated with SiO successfully 2 A housing.
FIG. 3 is a graph of the X-ray mediated photodynamic effects of LAMC, LAMCCS in example 1; wherein a is the signal intensity of detection indexes excited by singlet oxygen generated by LAMC and LAMCCS under different conditions, and b is the active oxygen level generated by HeLa cells under different stimulation conditions.
In FIG. 3, a is a diagram illustrating the amount of Singlet Oxygen generated when the reaction system receives 10Gy X-ray (X-ray) through a specific indicator Singlet Oxygen fluorescent probe (SOSG). The concentrations of nanoparticles and SOSG in the reaction system were 100. Mu.g/mL and 100. Mu.M, respectively. 6 groups (Control, LAMC, LAMCCS, X-ray, LAMC + X-ray, LAMCCS + X-ray) were set. After irradiation, placing the reaction system on a low-speed shaker for incubation for 12 hours in a dark place at 30 ℃, then centrifuging for 2 minutes at 12000 r, and detecting the fluorescence intensity of the solution when the excitation wavelength is 488 nm by a fluorescence spectrophotometer; the results show that LAMCCS has higher fluorescence intensity and higher singlet oxygen conversion efficiency than the LAMC group.
In FIG. 3, b is HeLa cells seeded in 24-well plates (8X 10) 4 One/well), 6 groups were set: (1) a control group, (2) LAMC, (3) LAMCCS, (4) X-ray, (5) LAMC + X-ray, (6) LAMCCS + X-ray; after 24 hours of cell inoculation, adding 500 mu L of culture medium into each hole, wherein the concentration of the nanoparticles is 100 mu g/mL; the irradiation dose of the X-ray irradiation group was 5Gy. After 2 hours of irradiation, serum-free medium containing 10 μ M DCFH-DA probe was added to each well, incubation was performed at 37 ℃ for 2 hours, PBS was washed 3 times, and fluorescence images were taken by a fluorescence microscope under 488 nanometer excitation light. The results show that lamcscs can significantly increase the reactive oxygen species produced by cells under X-ray irradiation.
FIG. 4 shows the in vitro tumor killing effect of LAMC, LAMCCS in example 1; wherein a is a cell plate cloning experiment of the HeLa cells under different condition stimulations, b is a DNA damage level (blue is cell nucleus and green is DNA double-chain damage) of the HeLa cells under different condition stimulations, and c is an apoptosis level of the HeLa cells under different condition stimulations.
In fig. 4, a is a 6-well plate (1000/well) in which HeLa cells were inoculated for 24 hours, and 6 groups were set: and (1) a control group, (2) LAMC, (3) LAMCCS, (4) X-ray, (5) LAMC + X-ray and (6) LAMCCS + X-ray, wherein 1mL of culture medium is added into each hole, the concentration of the nanoparticles is 100 microgram/mL, the incubation is carried out for 12 hours, and the irradiation dose of the group receiving X-ray irradiation is 2Gy. After the culture medium is replaced and incubated for 7 days, 4% paraformaldehyde is fixed and stained with crystal violet, and the cell population number is counted; the results show that LAMCCS can significantly reduce the proliferative capacity of cells after X-ray irradiation.
In FIG. 4, b is HeLa cells seeded in 24-well plates (5X 10) 4 One/well) were incubated for 24 hours, 6 groups were set: and (1) a control group, (2) LAMC, (3) LAMCCS, (4) X-ray, (5) LAMC + X-ray and (6) LAMCCS + X-ray, wherein 500 mu L of culture medium is added into each hole, the concentration of the nanoparticles is 100 mu g/mL, the incubation is carried out for 12 hours, and the irradiation dose of the group receiving X-ray irradiation is 5Gy. After the culture medium is replaced and incubated for 12 hours, fixing by 4% paraformaldehyde, penetrating a membrane by 0.2% Triton-X100, blocking by BSA solution, incubating an anti-gamma-H2 AX antibody for 1 hour at room temperature, incubating a fluorescent secondary antibody for 1 hour after PBS (phosphate buffered saline) is washed, staining nuclei by DAPI (Dapi) after the PBS is washed, and observing by a fluorescent microscope; the results show that lamcscs significantly increase the level of cellular DNA double strand damage after X-ray irradiation.
In FIG. 4, c is the inoculation of HeLa cells into 6-well plates (1X 10) 5 One/well) were incubated for 24 hours, and 6 groups were set: and (1) a control group, (2) LAMC, (3) LAMCCS, (4) X-ray, (5) LAMC + X-ray and (6) LAMCCS + X-ray, wherein 1mL of culture medium is added into each hole, the concentration of the nanoparticles is 100 microgram/mL, the incubation is carried out for 12 hours, and the irradiation dose of the group receiving X-ray irradiation is 5Gy. After 48 hours incubation with fresh medium, cells were collected and stained with annexin V-FITC/PI apoptosis detection kit and detected by flow cytometry. The results show that LAMCCS can significantly increase apoptosis after X-ray irradiation。
FIG. 5 shows the in vivo tumor killing effect of LAMC, LAMCCS in example 1; wherein a is the tumor image of the mice of different treatment groups, b is the tumor volume growth curve of the mice of different treatment groups, and c is the weight change curve of the mice of different treatment groups.
Constructing a nude mouse model of cervical cancer, and dividing 1 × 10 7 The HaLa cells are inoculated to the right groin of a BALB/C female nude mouse at the age of 6 weeks, and the nude mice are randomly divided into 6 groups after 1 week: (1) a control group, (2) LAMC, (3) LAMCCS, (4) X-ray, (5) LAMC + X-ray, (6) LAMCCS + X-ray, 25 mu L PBS or nanoparticle solution (nanoparticle concentration is 2 mg/mL) is injected into tumor, the group receiving X-ray irradiation is respectively given tumor local X-ray irradiation Gy 5 on days 1, 4 and 7 after injection, the mouse is killed on day 15, and tumor tissues are taken; tumor images of mice in different treatment groups are shown in fig. 5 a; the tumor size was measured every 3 days after the mice were grouped and injected with the drug, and the tumor volume was calculated (calculation formula: volume =1/2 × length × width) 2 ) A tumor volume growth curve is drawn, shown in fig. 5 b, and the result shows that the LAMCCS can obviously inhibit the tumor growth after X-ray radiotherapy; the body weight of the mice is measured once every 3 days after the mice are grouped and injected with the medicament, a body weight change curve of the mice is drawn, and the result is shown in c in figure 5, which shows that the body weight of the mice does not obviously change and has no obvious difference among different treatment groups, and the LAMCCS prepared by the invention has good biocompatibility.
The above-described embodiments are merely illustrative of the preferred embodiments of the present invention, and do not limit the scope of the present invention, and various modifications and improvements of the technical solutions of the present invention can be made by those skilled in the art without departing from the spirit of the present invention, and the technical solutions of the present invention are within the scope of the present invention defined by the claims.

Claims (10)

1. A method for preparing a nanocomposite material having an X-ray mediated photodynamic therapy effect, comprising the steps of:
（1）Lu 3 Al 5 O 12 the synthesis of Mn/Ca nano scintillators:
will contain Lu 3+ 、Al 3+ 、Mn 2+ And Ca 2+ Is added dropwise to the NH 4 HCO 3 Reacting in the solution, drying and calcining the precipitate obtained after the reaction is finished, and adding the calcined product and polyethylene glycol into water for reaction to obtain Lu 3 Al 5 O 12 Mn/Ca nano-scintillators;
（2）Lu 3 Al 5 O 12 synthesizing a Mn/Ca-Ce6 nano composite material:
mixing the Lu 3 Al 5 O 12 Mixing Mn/Ca nano scintillator with chlorin e6, N-hydroxysuccinimide, N- (3-dimethylaminopropyl) -N' -ethylcarbodiimide hydrochloride in solvent, reacting in dark, centrifuging, precipitating and drying to obtain Lu 3 Al 5 O 12 Mn/Ca-Ce6 nanocomposites;
（3）Lu 3 Al 5 O 12 :Mn/Ca-Ce6@SiO 2 synthesis of the nano composite material:
the Lu is mixed 3 Al 5 O 12 Dispersing a Mn/Ca-Ce6 nano composite material in a solvent, dropwise adding a hexadecyl trimethyl ammonium aqueous solution for reaction, and removing the solvent after the reaction is finished; adding the precipitate and tetraethyl silicate into an ethanol water solution of sodium hydroxide, heating for reaction, centrifuging and washing the precipitate to obtain Lu 3 Al 5 O 12 :Mn/Ca-Ce6@SiO 2 A nanocomposite material.
2. The method according to claim 1, wherein the Lu in step (1) 3+ 、Al 3+ 、Mn 2+ And Ca 2+ 3; the NH 4 HCO 3 The concentration of the solution was 2M; the drying temperature is 80 ℃; the calcining temperature is 1000 ℃, and the time is 4h; the polyethylene glycol is PEG600, and the mass-to-volume ratio of the calcined product to the PEG600 is 10mg.
3. The method according to claim 1, wherein the Lu-containing solution in step (1) 3+ 、Al 3+ 、Mn 2+ And Ca 2+ With said NH and a precursor salt solution of 4 HCO 3 The reaction time of the solution is 30min; the reaction time of the calcined product and the polyethylene glycol is 12h.
4. The method according to claim 1, wherein the Lu in step (2) 3 Al 5 O 12 The mass ratio of Mn/Ca nano-scintillator, chlorin e6, N-hydroxysuccinimide and N- (3-dimethylaminopropyl) -N' -ethylcarbodiimide hydrochloride is (100); the reaction time is 24h.
5. The method according to claim 1, wherein the Lu in step (3) 3 Al 5 O 12 The mass to volume ratio of the Mn/Ca-Ce6 nanocomposite to the tetraethyl silicate is 100mg.
6. The preparation method according to claim 1, wherein the aqueous ethanol solution of sodium hydroxide in the step (3) is prepared by mixing water, ethanol and 2M sodium hydroxide solution in a volume ratio of 40.
7. The method of claim 1, wherein step (3) further comprises subjecting the Lu to 3 Al 5 O 12 :Mn/Ca-Ce6@SiO 2 Removing impurities from the nano composite material, specifically, removing impurities from the Lu 3 Al 5 O 12 :Mn/Ca-Ce6@SiO 2 Nanocomposite and NH 4 NO 3 Dispersing in ethanol, reacting at 50 deg.C for 2 hr, washing with water, and drying to remove impurities.
8. A nanocomposite material having an X-ray-mediated photodynamic therapy effect, which is produced by the production method for a nanocomposite material having an X-ray-mediated photodynamic therapy effect according to any one of claims 1 to 7.
9. Use of the nanocomposite material with X-ray mediated photodynamic therapeutic effect according to claim 8 for the preparation of a medicament for radiotherapy.
10. The use of claim 9, wherein the radiotherapeutic agent is an X-ray mediated adjunctive therapy agent.
CN202211263799.4A 2022-10-17 2022-10-17 Nanocomposite with X-ray mediated photodynamic therapy effect and preparation method and application thereof Pending CN115317608A (en)

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Citations (2)

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Publication number Priority date Publication date Assignee Title
US20060086311A1 (en) * 2003-11-04 2006-04-27 Zagumennyi Alexander I Scintillation substances (variants)
CN112057617A (en) * 2020-09-14 2020-12-11 中国人民解放军陆军军医大学第一附属医院 Preparation method of core-shell structured porphyrin MOFs/scintillator composite nano material, product and application thereof

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20060086311A1 (en) * 2003-11-04 2006-04-27 Zagumennyi Alexander I Scintillation substances (variants)
CN112057617A (en) * 2020-09-14 2020-12-11 中国人民解放军陆军军医大学第一附属医院 Preparation method of core-shell structured porphyrin MOFs/scintillator composite nano material, product and application thereof

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Application publication date: 20221111