CN106890341B - Phototherapy nano preparation based on chemical crosslinking and preparation method and application thereof - Google Patents
Phototherapy nano preparation based on chemical crosslinking and preparation method and application thereof Download PDFInfo
- Publication number
- CN106890341B CN106890341B CN201710142123.2A CN201710142123A CN106890341B CN 106890341 B CN106890341 B CN 106890341B CN 201710142123 A CN201710142123 A CN 201710142123A CN 106890341 B CN106890341 B CN 106890341B
- Authority
- CN
- China
- Prior art keywords
- photosensitizer
- phototherapy
- preparation
- nano
- solution
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
- A61K41/0057—Photodynamic therapy with a photosensitizer, i.e. agent able to produce reactive oxygen species upon exposure to light or radiation, e.g. UV or visible light; photocleavage of nucleic acids with an agent
- A61K41/0071—PDT with porphyrins having exactly 20 ring atoms, i.e. based on the non-expanded tetrapyrrolic ring system, e.g. bacteriochlorin, chlorin-e6, or phthalocyanines
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/001—Preparation for luminescence or biological staining
- A61K49/0013—Luminescence
- A61K49/0017—Fluorescence in vivo
- A61K49/005—Fluorescence in vivo characterised by the carrier molecule carrying the fluorescent agent
- A61K49/0054—Macromolecular compounds, i.e. oligomers, polymers, dendrimers
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/001—Preparation for luminescence or biological staining
- A61K49/0063—Preparation for luminescence or biological staining characterised by a special physical or galenical form, e.g. emulsions, microspheres
- A61K49/0069—Preparation for luminescence or biological staining characterised by a special physical or galenical form, e.g. emulsions, microspheres the agent being in a particular physical galenical form
- A61K49/0089—Particulate, powder, adsorbate, bead, sphere
- A61K49/0091—Microparticle, microcapsule, microbubble, microsphere, microbead, i.e. having a size or diameter higher or equal to 1 micrometer
- A61K49/0093—Nanoparticle, nanocapsule, nanobubble, nanosphere, nanobead, i.e. having a size or diameter smaller than 1 micrometer, e.g. polymeric nanoparticle
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0012—Galenical forms characterised by the site of application
- A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
Abstract
The invention discloses a preparation method and application of a novel phototherapy nano preparation based on polyhedral oligomeric silsesquioxane (POSS). The phototherapy nanometer preparation is prepared by carrying out chemical crosslinking reaction on amino of POSS and carboxyl of photosensitizer to form nanometer particles and then carrying out surface pegylation. The nanometer preparation uses a photosensitizer as a part by using a chemical crosslinking method, realizes ultrahigh drug loading, avoids the early leakage of the drug and inhibits the self-quenching effect of the photosensitizer. Meanwhile, the medicine has good water dispersibility, stability and biological safety. In addition, the phototherapeutic formulation is capable of being largely endocytosed by cancer cells and efficiently killing cancer cells under light conditions. In-vivo animal experiments prove that the phototherapy nano preparation can be enriched to a tumor area through passive targeting, and the imaging and photodynamic therapy of tumors are realized.
Description
Technical Field
The invention discloses a preparation method of a novel phototherapy nano preparation and application of the novel phototherapy nano preparation in tumor imaging and photodynamic therapy, and belongs to the technical field of nano materials and biology.
Background
Photodynamic therapy is an emerging cancer treatment means, avoids the defects of strong invasiveness, large systemic toxicity, incomplete treatment and the like of the traditional treatment modes (such as chemotherapy, radiotherapy and surgical excision), and has the advantages of strong specificity, small damage to normal tissues, no drug toxicity accumulation and the like. Photodynamic therapy mainly utilizes reactive oxygen species generated by photosensitizer molecules under light conditions to kill cancer cells. However, most photosensitizer molecules are poorly water soluble and tend to form aggregates in aqueous solutions, which greatly inhibit the singlet oxygen yield and therapeutic efficacy of the photosensitizer.
A currently common strategy is to load photosensitizers into nanoparticles, which mainly include liposomes, micelles, quantum dots, metal nanoparticles, polymer nanoparticles, graphene oxide, and the like, to improve the photodynamic therapy effect. Overall, existing drug delivery strategies can be largely divided into two categories, physical encapsulation and covalent grafting: the physical packaging method has the problems of low drug loading rate, early drug release and the like although the preparation is simple; the nanoparticles formed by the covalent grafting strategy have stable structures, and avoid the premature release of drugs, however, the excessive loading of the photosensitizer on the surfaces of the nanoparticles can affect the water solubility of the nanoparticles and cause the self-quenching of photosensitizer molecules. Therefore, there is a need to develop a photodynamic therapy drug with high drug loading, safety and efficacy.
In addition, the killing effect of photodynamic therapy on cancer cells is mainly due to the damage effect of singlet oxygen on organelles. The singlet oxygen has short service life and limited action distance, so the anticancer effect of the photosensitizer depends on the endocytosis efficiency and subcellular localization condition of the photosensitizer to a great extent. Generally speaking, photodynamic therapy localized to organelles such as mitochondria and endoplasmic reticulum is more effective, phototherapy applied to cell membranes can accelerate necrosis of cells, and phototherapy localized to organelles such as lysosomes is relatively less effective. Considering the factors, the improvement of the photodynamic therapy effect can be realized through two aspects, namely, the endocytosis efficiency and the endocytosis quantity of the photosensitizer are increased, and the subcellular localization condition of the photosensitizer is changed, so that the photosensitizer is mainly distributed in mitochondria, endoplasmic reticulum and other organelles which are more sensitive to the action of singlet oxygen.
In conclusion, it is necessary to develop a safe and effective phototherapy nano-preparation with high drug loading, stable structure, high endocytosis efficiency of cells and specific organelle targeting.
Disclosure of Invention
The purpose of the invention is as follows: in view of the above, in order to solve the above problems in the prior art, the applicant expects to provide a novel phototherapy nano-preparation with high drug loading rate, stable structure, large endocytosis amount of cells, and capable of being positioned in mitochondria and endoplasmic reticulum. It is a further object to provide a method of preparing the above-described phototherapy nano-formulation.
The technical scheme is as follows: the invention provides a novel phototherapy nano preparation based on polyhedral oligomeric silsesquioxane (POSS), which mainly comprises photosensitizer molecules with more than two (containing two) carboxyl groups, POSS and polyethylene glycol (PEG) molecules. Photosensitive molecules and POSS are crosslinked to form nanoparticles, and PEG molecules are connected to the nanoparticles to form a whole. The PEG is modified PEG, NHS-PEG-OMe. The preparation comprises the following components in proportion: 100 molar parts POSS, 30 to 500 molar parts photosensitizer, 100 to 400 molar parts NHS-PEG-OMe.
Wherein the molecular general formula of POSS is (RSiO)1.5)nAnd n is generally 6, 8, 10, 12, etc. Where R can be hydrogen, alkyl, epoxy, alkylene, phenyl, aryl, arylene, and the like, most typically with n =6 and 8. The POSS has a random structure, a trapezoidal structure, a cage structure, a bridge structure and the like.
Preferably, the photosensitizer molecule is protoporphyrin, chlorin e6 (Ce 6) or 5,10,15, 20-tetrakis (4-carboxyphenyl) porphyrin.
Preferably, the POSS comprises octaaminopropyl hexahedral silsesquioxane and octaaminoethyl hexahedral silsesquioxane, and further preferably, the POSS is octaaminopropyl hexahedral silsesquioxane.
Preferably, the PEG has a molecular weight of 500-10000. Further preferably, the PEG has a molecular weight of 2000.
Further, the invention also provides a preparation method of the novel phototherapy nano preparation based on POSS, which comprises the following steps:
(1) carboxyl group of activating photosensitizer: weighing photosensitizer, N-hydroxysuccinimide (NHS) and 1-ethyl- (3-dimethylaminopropyl) carbodiimide (EDC), and respectively dissolving in dimethyl sulfoxide (DMSO); sequentially adding the EDC solution and the NHS solution into the photosensitizer solution, and reacting at room temperature for 2-4 hours to obtain an activated photosensitizer solution;
(2) POSS is dissolved in DMSO, triethylamine is added, and the mixture is treated overnight at room temperature;
(3) mixing the activated photosensitizer with POSS treated by triethylamine, and reacting for 4-12 hours at room temperature;
(4) adding NHS-PEG-OMe into the solution, and reacting for 6-9 hours at room temperature; after the reaction, the sample was obtained by dialysis purification in DMSO, further dialysis purification with ultrapure water, and then freeze-drying.
Preferably, in the step 1, the molar ratio of the photosensitizer molecule, EDC and NHS is 1: (4-40): (6-60).
Further preferably, in the step 1, the molar ratio of the photosensitizer molecule, EDC and NHS is 1 (8-12) to (10-18). More preferably, the molar ratio of photosensitizer molecule, EDC and NHS is 1:10: 15.
Preferably, in the step 2, the molar ratio of POSS to triethylamine is 1 (8-96).
Further preferably, the molar ratio of the POSS to the triethylamine is 1 (18-30). More preferably, the molar ratio of POSS to triethylamine is 1: 24.
Preferably, in the step 3, the molar ratio of POSS to photosensitizer molecules is 1 (0.5-7).
Further preferably, in the step 3, the molar ratio of POSS to photosensitizer molecules is 1 (1-4). More preferably, the POSS is present in a 1:3 ratio with the photosensitizer molecule.
Preferably, in the step 4, the molar ratio of POSS to NHS-PEG-OMe molecules is 1 (2-6).
Further preferably, in the step 4, the molar ratio of POSS to NHS-PEG-OMe molecules is 1 (4-6). More preferably, the molar ratio of POSS to NHS-PEG-OMe molecules is 1: 5.
Furthermore, the invention also provides application of the phototherapy nano preparation in serving as or preparing a tumor imaging or photodynamic therapy medicine. Provides an application of a phototherapy nano preparation in preparing or serving as a tumor treatment medicine. Further provides an application of the phototherapy nano preparation in tumor imaging and photodynamic therapy. The phototherapy nanometer preparation comprises photosensitizer molecules, wherein the photosensitizer molecules and POSS can form nanoparticles through nonspecific covalent crosslinking, and then surface PEG modification is carried out to obtain the phototherapy nanometer preparation. The preparation method of the phototherapy nano-preparation is as described above and will not be described again here.
Preferably, the photosensitizer is protoporphyrin, Ce6 or 5,10,15, 20-tetrakis (4-carboxyphenyl) porphyrin.
Further preferably, the photosensitizer is Ce6, and the molecular weight of the PEG is 1000-10000.
Further preferably, the PEG has a molecular weight of 2000. Preferably, the molar ratio of POSS to photosensitizer molecule is 1 (0.5-7).
It is further preferred that the molar ratio of POSS to photosensitizer molecules is 1: 3.
Preferably, the molar ratio of POSS to NHS-PEG-OMe molecules is 1 (2-6).
Further preferably, the molar ratio of POSS to NHS-PEG-OMe molecules is 1: 5.
The novel phototherapy nano preparation provided by the invention is prepared by chemically crosslinking a plurality of carboxyl groups on the photosensitizer and a plurality of amino groups on the POSS, and has the advantages of mild reaction conditions, extremely high drug-loading rate, easiness in preparation and purification, and good water dispersibility and stability. The construction mode of covalent crosslinking can effectively avoid the problem of early release of the photosensitizer in vivo circulation. In addition, the unique rigid structure of the cage hexahedron of POSS effectively inhibits the self-quenching effect of the photosensitizer, thereby ensuring the photodynamic therapy effect thereof.
The technical effects are as follows: compared with the prior art, the invention has the following advantages:
(1) the synthesis is simple and convenient: the nano-particles are reacted at room temperature in the whole process, the reaction condition is mild, the preparation method is simple, and the feasibility is high.
(2) Good structural stability: the photosensitizer is loaded in a covalent crosslinking mode, so that the premature release of the drug is avoided.
(3) The drug loading is high: the photosensitizer is used as a part of the construction of the nano particles, so that the loading efficiency of the photosensitizer is greatly improved, reaches 19.8 wt%, and is higher than most of the current drug-carrying systems.
(4) Effectively inhibiting the self-quenching effect of the photosensitizer: the three-dimensional cage structure of POSS effectively inhibits the self-quenching effect of the photosensitizer, so that the fluorescence yield and the singlet oxygen yield of the photosensitizer are not affected, and the imaging and phototherapy effects of the photosensitizer are ensured.
(5) Large endocytosis capacity of cells, targeting multiple organelles: the phototherapeutic nano preparation has large uptake of cancer cells, and the photosensitizer is mainly positioned in mitochondria and endoplasmic reticulum, thereby effectively improving the photodynamic treatment effect.
(6) Diagnosis and treatment integration: can be injected into a mouse body through tail vein and is enriched to a tumor area through passive targeting, thereby realizing good tumor imaging and excellent photodynamic therapy effect.
(7) Good safety: under the condition of no light, the phototherapeutic nano preparation has very low cytotoxicity and almost no toxic or side effect on normal tissues.
Description of the drawings:
FIG. 1 is a schematic synthesis of the present invention.
FIGS. 2a to 2h are graphs showing the imaging effect of polyhedral oligomeric silsesquioxane-chlorin e 6-polyethylene glycol and chlorin e6 on human cervical cancer cells at different treatment times, respectively.
FIGS. 3a to 3o are diagrams of the localization of organelles of the present invention in human cervical cancer cells, endoplasmic reticulum, lysosome, and mitochondria, respectively.
FIG. 4 is a graph showing the effect of photodynamic therapy on human cervical cancer cells in accordance with the present invention.
FIG. 5 is the result of evaluating the dark toxicity of the present invention to human cervical cancer cells.
FIGS. 6a to 6j are graphs showing the in vivo imaging effect of chlorin e6 and polyhedral oligomeric silsesquioxane-chlorin e 6-polyethylene glycol on tumor-bearing mice at different treatment times, respectively.
FIG. 7 is a graph showing the photodynamic anti-cancer effect of the present invention on tumor-bearing mice.
Detailed Description
The present invention is further illustrated by the following figures and specific examples, which are to be construed as merely illustrative and not limitative of the remainder of the disclosure, and all changes and modifications that would be obvious to those skilled in the art are intended to be included within the scope of the present invention and the appended claims are intended to be embraced therein.
Example 1
The synthesis of POSS-Ce6-PEG is as follows (schematic shown in FIG. 1):
(1) firstly, Ce6, EDC and NHS are used as raw materials to activate carboxyl of Ce6 molecules. 10 mg Ce6, 26.0 mg EDC and 28.9 mg NHS were weighed out and dissolved in 1 mL DMSO respectively, Ce6 was mixed with EDC and NHS was added to react at room temperature (20-25 ℃ C. in this case) for 4 hours to complete the carboxyl activation.
(2) 6.55 mg of octaaminopropylhexahedral silsesquioxane (POSS) was weighed in a molar ratio of 1:3 of octaaminopropylhexahedral silsesquioxane to Ce6, 18.57. mu.L of triethylamine was co-dissolved in DMSO in a molar ratio of 1:24 of POSS to triethylamine, and mixed with the above reaction system to react overnight at room temperature.
(3) 58.6 mg of NHS-PEG-OMe was weighed out in DMSO in a molar ratio of POSS to NHS-PEG-OMe (wherein the molecular weight of PEG is 2000) of 1:5, mixed with the above reaction system, reacted at room temperature for 8 hours, dialyzed in DMSO for 2 days using a dialysis bag having a molecular weight cut-off of 10k, and then dialyzed in ultrapure water for 1 day for purification. And finally, freeze-drying in a freeze dryer to obtain POSS-Ce6-PEG dry powder, and freezing and storing at-20 ℃.
Examples 2 to 4
Example 2 the procedure for the preparation of phototherapy nanopreparations was the same as in example 1 except that the molar ratio of POSS to Ce6 in step (2) was 1: 0.5.
Example 3 the procedure for the preparation of phototherapy nanopreparations was the same as in example 1 except that the molar ratio of POSS to Ce6 in step (2) was 1: 4.
Example 4 phototherapeutic nanoformulations were prepared by the same procedure as in example 1, except that the molar ratio of POSS to Ce6 in step (2) was 1: 7.
Examples 5 to 7
Example 5 the procedure for the preparation of phototherapeutic nanoformulations was the same as in example 1, except that the reaction time of POSS with Ce6 in step (2) was 4 hours.
Example 6 the procedure for the preparation of phototherapy nanopreparations was the same as in example 1 except that the reaction time of POSS with Ce6 in step (2) was 8 hours.
Example 7 the procedure for the preparation of phototherapeutic nanoformulations was the same as in example 1, except that the reaction time of POSS with Ce6 in step (2) was 12 hours.
Examples 8 to 10
Example 8 the procedure for the preparation of phototherapy nanopreparation was the same as in example 1 except that in step (3) the molecular weight of PEG (herein referred to as viscosity average molecular weight) in the NHS-PEG-OMe molecule was 500.
Example 9 preparation of phototherapy nanopreparation the procedure was the same as in example 1 except that in step (3) the molecular weight of PEG in NHS-PEG-OMe molecule was 5000.
Example 10 preparation of phototherapy nanopreparation procedure was the same as in example 1 except that molecular weight of PEG in NHS-PEG-OMe molecule was 10000 in step (3).
Examples 11 to 13
Example 11 the procedure for the preparation of phototherapy nanopreparations was the same as in example 1, except that the molar ratio of POSS to NHS-PEG-OMe in step (3) was 1: 2.
Example 12 preparation of phototherapy nanopreparations the procedure was the same as in example 1 except that the molar ratio of POSS to NHS-PEG-OMe in step (3) was 1: 4.
Example 13 preparation of phototherapy nanopreparations the procedure was the same as in example 1 except that the molar ratio of POSS to NHS-PEG-OMe in step (3) was 1: 6.
Example 14
Phototherapeutic nanoformulations were prepared by the same procedure as in example 1, except that POSS was an octaaminoethyl hexahedral oligosilsesquioxane as in steps (2) and (3).
Example 15
The endocytosis efficiency of the phototherapy nano-preparation prepared in the example 1 by the human cervical cancer cells is evaluated by the following method:
(1) cell culture and plating: reviving human cervical cancer cells in DMEM complete medium at 37oC、5% CO2Culturing in the environment when the cell density is highWhen the growth reaches about 80%, cells are digested with pancreatin and pressed at 5 × 104Plating at cell density of one/mL, continuing at 37oC、5% CO2Culturing for 24 hours in the environment;
(2) drug interaction with cells: culturing human cervical cancer cells in 8-well plate, after 24 hr, dissolving phototherapy nanometer preparation and free Ce6 in DMEM complete culture medium, adding into 8-well plate (200 μ L/well), continuing at 37%oC、5% CO2Co-incubation in the environment;
(3) and (3) confocal fluorescence microscope imaging observation: at different time points of incubation of the drug and the cells, the phototherapy nano preparation emits red fluorescence by using laser with the wavelength of 552nm as exciting light, and observation is carried out.
The fluorescence imaging results are shown in fig. 2a to 2 h. As can be seen, the fluorescence intensity of POSS-Ce6-PEG in cells is much higher than that of free Ce6, and the fluorescence gradually increases with time. The phototherapy nano preparation is proved to be capable of being largely endocytosed by cancer cells.
Example 16
The situation of organelle localization of POSS-Ce6-PEG prepared in example 1 in human cervical carcinoma cells was observed as follows:
culturing human cervical cancer cells in 8-well plate, dispersing phototherapy nanometer preparation in DMEM complete culture medium according to Ce6 concentration of 3 μ g/mL after 24 hours, adding 8-well plate (200 μ L/well), adding into 37%oC、5% CO2After 2 hours of incubation in the environment, washing twice with PBS, adding organelle (endoplasmic reticulum, mitochondria and lysosome) staining reagent, incubating for half an hour, washing twice with PBS, adding cell nucleus staining reagent, washing twice with PBS after 10 minutes, and adding DMEM complete culture medium.
And (3) confocal fluorescence microscope imaging observation: at different time points of drug incubation with cells, observations were made with lasers of wavelengths 405, 488, and 552nm as excitation light. The cell nucleus staining reagent emits blue fluorescence under 405 nm exciting light, the organelle (endoplasmic reticulum, mitochondria and lysosome) staining reagent emits green fluorescence under 488 nm exciting light, and the phototherapy nanometer preparation emits red fluorescence under 552nm exciting light. The results are shown in FIGS. 3a to 3 o. As can be seen, phototherapeutic nanoformulations are mainly localized to the mitochondria and endoplasmic reticulum after being endocytosed by the cells. Considering the higher sensitivity of both mitochondria and endoplasmic reticulum to singlet oxygen, the phototherapeutic nanoformulation will have very good photodynamic therapy effect (see example 17 for specific results).
Example 17
The phototherapy effect of POSS-Ce6-PEG prepared in example 1 and free Ce6 on human cervical cancer cells was evaluated as follows:
(1) cell phototherapy: free Ce6 solution and POSS-Ce6-PEG solution were prepared in DMEM complete medium so that both finally contained Ce6 at a concentration of 2. mu.g/mL. Add 100. mu.L of each of the above solutions to the corresponding wells at 37oC、5% CO2The culture is carried out for 5 h in the environment. Subsequently, a 670 nm laser was used at 8 mW/cm2Respectively irradiating for 0-10 min at power density of (1), and transferring to (37)oC、5% CO2Culturing for 4 hours in the environment;
(2) and (3) cell survival rate detection: MTT solution was prepared at a concentration of 5 mg/mL, and 10. mu.L of thiazole blue (MTT) solution was added to each well of the cells at 37oC、5% CO2The culture was carried out for 4 h in the environment. The culture medium in each well was decanted, 150. mu.L of MSO was added, and the absorbance at 492 nm was measured using a microplate reader. The results are shown in FIG. 4.
As shown in the figure, the phototherapy nano-formulation prepared in example 1 has better photodynamic anticancer effect than free Ce 6.
Example 18
The phototherapeutic nanoformulations prepared in example 1 were evaluated for dark cytotoxicity by treating human cervical cancer cells as 5 × 104Inoculating the strain/mL of the strain in a 96-well plate, culturing for 24 hours, then incubating with phototherapy nano preparations with different concentrations for 24 hours in a dark place, and finally evaluating the toxicity of the phototherapy nano preparations on human cervical cancer cells by using an MTT detection method, wherein the result is shown in figure 5. The experimental result shows that the phototherapy nano preparation has almost no cytotoxicity under the condition of no illumination.
Example 19
The phototherapy nano-formulation prepared in example 1 was tested for its imaging effect in mice, and the procedure was as follows: several healthy 4-week-old BALA/c athymic female nude mice were collected and injected subcutaneously with murine cervical cancer cells (U14). When the tumor volume reaches 100 mm3On the other hand, Ce6 and the phototherapy nano-formulation obtained in example 1 were injected into nude mice by tail vein injection. And then placing the nude mouse after anesthesia by isoflurane gas into an imaging dark box platform, exciting by 600 nm exciting light and receiving 670 nm wave band emitting light in a living body imaging system of the small animal, wherein the exposure time is 750 ms. Imaging data were acquired at different time points (0, 2, 6, 12 and 24 hours) respectively and data analysis was performed. The experimental results are shown in FIGS. 6a to 6 j.
As can be seen, almost no fluorescence was observed in Ce6 after injection into nude mice, while the phototherapy nano-formulation obtained by the present invention was able to enrich in the tumor area, exhibited very significant in vivo fluorescence, and could be retained in the tumor area for at least 10 hours.
Example 20
The photodynamic anticancer effect of the phototherapy nano-formulation prepared in example 1 in mice was evaluated, and the procedure was as follows: when the tumor volume reaches 50 mm3At the left and right, Ce6 and the phototherapy nano preparation obtained in example 1 are injected into a nude mouse body in a tail vein injection mode, after injection is completed for 6 hours, a 670 nm red laser is used for irradiating a tumor area, and the laser power is 20 mW/cm2Tumor volume and body weight of nude mice were measured every other day, tumor volume 0.5 × tumor length × tumor width2The results are shown in FIG. 7. As can be seen from the figure, compared with physiological saline and Ce6, the phototherapy nano preparation obtained by the invention can obviously inhibit the tumor growth in a mouse body and has good tumor photodynamic curative effect.
Although the preferred embodiments of the present invention have been described in detail, the present invention is not limited to the details of the embodiments, and various equivalent modifications can be made within the technical spirit of the present invention, and the scope of the present invention is also within the scope of the present invention. It should be noted that the various features described in the above embodiments may be combined in any suitable manner without departing from the scope of the invention. The invention is not described in detail in order to avoid unnecessary repetition.
Claims (6)
1. A phototherapy nanometer preparation based on chemical crosslinking is characterized by comprising a plurality of nanoparticles formed by crosslinking a plurality of polyhedral oligomeric silsesquioxane molecules with a plurality of photosensitizers with more than two carboxyl groups and a plurality of polyethylene glycol molecules modified on the surfaces of the nanoparticles, wherein the photosensitizers are protoporphyrin, chlorin e6 or 5,10,15, 20-tetra (4-carboxyphenyl) porphyrin, the polyhedral oligomeric silsesquioxane is octaaminopropyl hexahedral silsesquioxane or octaaminoethyl hexahedral silsesquioxane, and the preparation method comprises the following steps:
(1) carboxyl group of activating photosensitizer: weighing photosensitizer, N-hydroxysuccinimide and 1-ethyl- (3-dimethylaminopropyl) carbodiimide, and respectively dissolving the photosensitizer, the N-hydroxysuccinimide and the 1-ethyl- (3-dimethylaminopropyl) carbodiimide in dimethyl sulfoxide to obtain corresponding solutions; sequentially adding a 1-ethyl- (3-dimethylaminopropyl) carbodiimide solution and an N-hydroxysuccinimide solution into a photosensitizer solution, and reacting at room temperature for 2-4 hours to obtain an activated photosensitizer solution;
(2) dissolving polyhedral oligomeric silsesquioxane in dimethyl sulfoxide, adding triethylamine in an amount which is 24 times that of the polyhedral oligomeric silsesquioxane, and treating at room temperature overnight;
(3) mixing the activated photosensitizer with polyhedral oligomeric silsesquioxane treated by triethylamine, and reacting for 4-12 hours at room temperature;
(4) adding N-hydroxysuccinimide ester-polyethylene glycol-methoxyl into the solution, and reacting for 6-9 hours at room temperature;
after the reaction is finished, dialyzing and purifying by dimethyl sulfoxide and ultrapure water respectively, and then freeze-drying to obtain the sample.
2. The phototherapy nano-formulation of claim 1, wherein the polyethylene glycol has a viscosity average molecular weight of 500-10000.
3. A method of preparing the phototherapy nano-formulation of claim 1, comprising the steps of:
(1) carboxyl group of activating photosensitizer: weighing photosensitizer, N-hydroxysuccinimide and 1-ethyl- (3-dimethylaminopropyl) carbodiimide, and respectively dissolving the photosensitizer, the N-hydroxysuccinimide and the 1-ethyl- (3-dimethylaminopropyl) carbodiimide in dimethyl sulfoxide to obtain corresponding solutions; sequentially adding a 1-ethyl- (3-dimethylaminopropyl) carbodiimide solution and an N-hydroxysuccinimide solution into a photosensitizer solution, and reacting at room temperature for 2-4 hours to obtain an activated photosensitizer solution;
(2) dissolving polyhedral oligomeric silsesquioxane in dimethyl sulfoxide, adding triethylamine in an amount which is 24 times that of the polyhedral oligomeric silsesquioxane, and treating at room temperature overnight;
(3) mixing the activated photosensitizer with polyhedral oligomeric silsesquioxane treated by triethylamine, and reacting for 4-12 hours at room temperature;
(4) adding N-hydroxysuccinimide ester-polyethylene glycol-methoxyl into the solution, and reacting for 6-9 hours at room temperature;
after the reaction is finished, dialyzing and purifying by dimethyl sulfoxide and ultrapure water respectively, and then freeze-drying to obtain the sample.
4. The method for preparing phototherapy nano-formulations as described in claim 3, wherein in the step (1), the molar ratio of the photosensitizer, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide and N-hydroxysuccinimide is 1 (4-40) to (6-60).
5. The method of preparing phototherapy nano-formulations as claimed in claim 3, wherein the molar ratio of polyhedral oligomeric silsesquioxane to photosensitizer is 1 (0.5-7).
6. The method of preparing phototherapy nano-formulations as claimed in claim 3, wherein the molar ratio of polyhedral oligomeric silsesquioxane to N-hydroxysuccinimide ester-polyethylene glycol-methoxy group is 1 (2-6).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710142123.2A CN106890341B (en) | 2017-03-10 | 2017-03-10 | Phototherapy nano preparation based on chemical crosslinking and preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710142123.2A CN106890341B (en) | 2017-03-10 | 2017-03-10 | Phototherapy nano preparation based on chemical crosslinking and preparation method and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106890341A CN106890341A (en) | 2017-06-27 |
| CN106890341B true CN106890341B (en) | 2020-09-11 |
Family
ID=59192756
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710142123.2A Active CN106890341B (en) | 2017-03-10 | 2017-03-10 | Phototherapy nano preparation based on chemical crosslinking and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106890341B (en) |
Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108295254B (en) * | 2018-01-19 | 2019-11-12 | 东南大学 | A kind of nanometer reagent and its preparation method and application for regulating cell nuclear membrane permeability |
| JP2022500439A (en) * | 2018-09-13 | 2022-01-04 | ザ リージェンツ オブ ザ ユニバーシティー オブ ミシガン | Small ultra-uniform nanopharmaceutical compositions for therapeutic, imaging, and seranostic applications |
| CN111569068B (en) * | 2020-05-14 | 2022-08-30 | 南京邮电大学 | Organic-inorganic hybrid photosensitizer and preparation method of hybrid nano diagnosis and treatment reagent |
| CN114573321B (en) * | 2022-03-21 | 2023-05-05 | 安徽紫朔环境工程技术有限公司 | Preparation method of catalytic ceramic filter tube based on 3D printing technology |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007147742A1 (en) * | 2006-06-19 | 2007-12-27 | Ciba Holding Inc. | Coloured silsesquioxanes |
| CN102212195A (en) * | 2010-04-07 | 2011-10-12 | 韩国科学技术研究院 | Photoactive group-bonded polysilsesquioxane having a ladder structure and a method for preparing the same |
| CN104888219A (en) * | 2015-06-10 | 2015-09-09 | 东南大学 | Tumor phototherapy reagent based on cell membrane coating, as well as preparation method and application of tumor phototherapy reagent |
| CN105969334A (en) * | 2016-05-06 | 2016-09-28 | 东南大学 | Fluorescent dyeing reagent for detecting bacterium and fungus survival state and its preparation method and use |
-
2017
- 2017-03-10 CN CN201710142123.2A patent/CN106890341B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007147742A1 (en) * | 2006-06-19 | 2007-12-27 | Ciba Holding Inc. | Coloured silsesquioxanes |
| CN102212195A (en) * | 2010-04-07 | 2011-10-12 | 韩国科学技术研究院 | Photoactive group-bonded polysilsesquioxane having a ladder structure and a method for preparing the same |
| CN104888219A (en) * | 2015-06-10 | 2015-09-09 | 东南大学 | Tumor phototherapy reagent based on cell membrane coating, as well as preparation method and application of tumor phototherapy reagent |
| CN105969334A (en) * | 2016-05-06 | 2016-09-28 | 东南大学 | Fluorescent dyeing reagent for detecting bacterium and fungus survival state and its preparation method and use |
Non-Patent Citations (2)
| Title |
|---|
| Redox-Responsive Porphyrin-Based Polysilsesquioxane Nanoparticles for Photodynamic Therapy of Cancer Cells;Daniel L. Vega 等;《International Journal of Molecular Sciences》;20161231;第17卷(第56期);第1-16页 * |
| 多面体低聚倍半硅氧烷（POSS）的合成与应用研究进展;马祥梅 等;《安徽化工》;20090228;第35卷(第1期);第14-16页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN106890341A (en) | 2017-06-27 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Liang et al. | Core-shell structured upconversion nanocrystal-dendrimer composite as a carrier for mitochondria targeting and catalase enhanced anti-cancer photodynamic therapy | |
| Sun et al. | One-pot fabrication of hollow porphyrinic MOF nanoparticles with ultrahigh drug loading toward controlled delivery and synergistic cancer therapy | |
| Yu et al. | Oxygen self-sufficient NIR-activatable liposomes for tumor hypoxia regulation and photodynamic therapy | |
| Taratula et al. | Dendrimer-encapsulated naphthalocyanine as a single agent-based theranostic nanoplatform for near-infrared fluorescence imaging and combinatorial anticancer phototherapy | |
| Gulzar et al. | Nano-graphene oxide-UCNP-Ce6 covalently constructed nanocomposites for NIR-mediated bioimaging and PTT/PDT combinatorial therapy | |
| Yang et al. | Biomineralization-inspired crystallization of manganese oxide on silk fibroin nanoparticles for in vivo MR/fluorescence imaging-assisted tri-modal therapy of cancer | |
| Zhao et al. | Insight into the efficiency of oxygen introduced photodynamic therapy (PDT) and deep PDT against cancers with various assembled nanocarriers | |
| Cui et al. | In vivo targeted deep-tissue photodynamic therapy based on near-infrared light triggered upconversion nanoconstruct | |
| CN106890341B (en) | Phototherapy nano preparation based on chemical crosslinking and preparation method and application thereof | |
| Fan et al. | Intranuclear biophotonics by smart design of nuclear-targeting photo-/radio-sensitizers co-loaded upconversion nanoparticles | |
| Huang et al. | Fluorescence resonance energy transfer-based drug delivery systems for enhanced photodynamic therapy | |
| Zhao et al. | An energy‐storing DNA‐based nanocomplex for laser‐free photodynamic therapy | |
| Ji et al. | Tandem activated photodynamic and chemotherapy: Using pH-Sensitive nanosystems to realize different tumour distributions of photosensitizer/prodrug for amplified combination therapy | |
| Cui et al. | Theranostic gold cluster nanoassembly for simultaneous enhanced cancer imaging and photodynamic therapy | |
| Sun et al. | Degradable hybrid CuS nanoparticles for imaging-guided synergistic cancer therapy via low-power NIR-II light excitation | |
| Zhao et al. | DUCNP@ Mn–MOF/FOE as a highly selective and bioavailable drug delivery system for synergistic combination cancer therapy | |
| Ai et al. | An upconversion nanoplatform with extracellular pH-driven tumor-targeting ability for improved photodynamic therapy | |
| Yao et al. | Tumor-targeted upconverting nanoplatform constructed by host-guest interaction for near-infrared-light-actuated synergistic photodynamic-/chemotherapy | |
| Hu et al. | Post-synthesis strategy to integrate porphyrinic metal–organic frameworks with CuS NPs for synergistic enhanced photo-therapy | |
| Sun et al. | Ce6-C6-TPZ co-loaded albumin nanoparticles for synergistic combined PDT-chemotherapy of cancer | |
| Chen et al. | Gadolinium–porphyrin based polymer nanotheranostics for fluorescence/magnetic resonance imaging guided photodynamic therapy | |
| Zheng et al. | Photoactivatable nanogenerators of reactive species for cancer therapy | |
| Ruan et al. | Redox-responsive prodrug-like PEGylated macrophotosensitizer nanoparticles for enhanced near-infrared imaging-guided photodynamic therapy | |
| Zheng et al. | Red blood cell membrane nanoparticles for tumor phototherapy | |
| Wang et al. | One stone, many birds: Recent advances in functional nanogels for cancer nanotheranostics |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |