CN115054613A - Multifunctional nano catalyst and preparation method thereof - Google Patents

Multifunctional nano catalyst and preparation method thereof Download PDF

Info

Publication number
CN115054613A
CN115054613A CN202210682380.6A CN202210682380A CN115054613A CN 115054613 A CN115054613 A CN 115054613A CN 202210682380 A CN202210682380 A CN 202210682380A CN 115054613 A CN115054613 A CN 115054613A
Authority
CN
China
Prior art keywords
msn
dox
cuo
catalyst
nano
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.)
Granted
Application number
CN202210682380.6A
Other languages
Chinese (zh)
Other versions
CN115054613B (en
Inventor
唐昭敏
金秋野
江舒婷
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Southwest Petroleum University
Original Assignee
Southwest Petroleum University
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Southwest Petroleum University filed Critical Southwest Petroleum University
Priority to CN202210682380.6A priority Critical patent/CN115054613B/en
Publication of CN115054613A publication Critical patent/CN115054613A/en
Application granted granted Critical
Publication of CN115054613B publication Critical patent/CN115054613B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A61K33/24—Heavy metals; Compounds thereof
    • A61K33/34—Copper; Compounds thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
    • A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
    • A61K31/704—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin attached to a condensed carbocyclic ring system, e.g. sennosides, thiocolchicosides, escin, daunorubicin
    • 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
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5107—Excipients; Inactive ingredients
    • A61K9/5115—Inorganic compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y40/00—Manufacture or treatment of nanostructures
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y5/00—Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Nanotechnology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • General Health & Medical Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Epidemiology (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Inorganic Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Molecular Biology (AREA)
  • Optics & Photonics (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Medical Informatics (AREA)
  • Biophysics (AREA)
  • Biomedical Technology (AREA)
  • Condensed Matter Physics & Semiconductors (AREA)
  • General Physics & Mathematics (AREA)
  • Manufacturing & Machinery (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

The invention discloses a multifunctional nano catalyst and a preparation method thereof, which mainly solve the problem of H in the existing cancer cells in the prior art 2 O 2 Too low a content of OH can not produce enough OH to kill cancer cells, resulting in poor effect of chemo-kinetic therapy and limitation of monotherapy. The multifunctional nano catalyst is a loadCopper peroxide nanodot modified mesoporous silica nanocatalyst DOX @ MSN @ CuO of DOX 2 . Through the scheme, the invention produces H by itself 2 O 2 ，Cu 2+ GSH is consumed, a large amount of OH and loaded chemotherapeutic drug DOX are generated, so that the nano-catalyst can effectively inhibit tumor growth, and CDT and chemotherapy are used for treating cancers in a synergistic manner.

Description

Multifunctional nano catalyst and preparation method thereof
Technical Field
The invention relates to the technical field of nano-catalysts, in particular to a multifunctional nano-catalyst and a preparation method thereof.
Background
Cancer is the second leading cause of death worldwide, and with the development of advanced nanotechnology, the chemokinetic treatment method CDT becomes an emerging catalytic anticancer therapy, which utilizes transition metal ions to catalyze endogenous hydrogen peroxide H 2 O 2 The generation of a fenton or fenton-like reaction that generates a cytotoxic hydroxyl radical OH to kill cancer cells is a promising cancer treatment strategy.
Reactive oxygen ROS from oxygen O 2 Is required for the normal metabolism of all aerobic organisms; cancer cells produce more reactive oxygen species than normal cells, and low concentrations of reactive oxygen species in normal cells play an important role in maintaining the cell life cycle, but high concentrations of reactive oxygen species in cancer cells can effectively destroy cancer cell components. Among all the active oxygen, H 2 O 2 Is the most stable and abundant non-free radical active oxygen in cancer cells, can easily diffuse to a biological membrane, and is converted into OH through Fenton-like reaction to kill the cancer cells; however, H in cancer cells 2 O 2 Too low a content of OH can not produce enough OH to kill cancer cells, resulting in undesirable effects of chemokinetic therapy and limitations of monotherapy. Thus, increase of H in cancer cells 2 O 2 The content of (A) is not very slow.
Disclosure of Invention
The invention aims to provide a multifunctional nano catalyst and a preparation method thereof, aiming at solving the problem of H in the existing cancer cells 2 O 2 Too low a content of OH does not produce enough OH to kill cancer cells, resulting in poor effect of chemo-kinetic therapy and limitations of monotherapy.
In order to solve the above problems, the present invention provides the following technical solutions:
multifunctional nano catalyst is DOX-loaded copper peroxide nano dot modified mesoporous silica nano catalyst DOX @ MSN @ CuO 2 ；DOX@MSN@CuO 2 After being endocytosed into tumor cells, the tumor cells are firstly decomposed into Cu in a weakly acidic specific tumor microenvironment TME 2+ And exogenous H 2 O 2 Simultaneously releasing DOX; subsequent Cu 2+ Reacting with local glutathione to deplete glutathione and deplete Cu 2+ Reduction to Cu + (ii) a Final Cu + With exogenous H 2 O 2 The Fenton-like reaction is carried out to generate toxic OH combined with DOX to cause the apoptosis of tumor cells, and the reaction rate of the process is fast.
The preparation method of the multifunctional nano catalyst comprises the following steps:
s1, synthesizing mesoporous silica nano-particles MSN;
s2, loading chemotherapeutic drug DOX on the MSN in the step S1 to obtain DOX @ MSN;
s3, DOX @ MSN and CuNO as copper nitrate trihydrate through step S2 3 ·3H 2 Preparation of DOX @ MSN @ CuO from O and NaOH 2 。
Further, the specific process of step S1 is: reacting NH 3 ·H 2 Dissolving O in deionized water to form a solution, and continuously stirring at 80 ℃; then, adding an ethanol solution containing tetraethoxysilane, Cetyl Trimethyl Ammonium Bromide (CTAB) and 3-Aminopropyltriethoxysilane (APTES) into the solution, and reacting for 10 hours; HCl was added to remove the template and the product MSN was collected by high speed centrifugation.
Further, the specific process of step S2 is: dispersing MSN and DOX & HCI in dimethyl sulfoxide DMSO according to a mass ratio of 4:1, adding triethylamine TEA, and continuously stirring for 24 hours at room temperature in the dark; free DOX was removed by repeated centrifugation and DOX @ MSN was collected by freeze drying.
Further, the specific process of step S3 is: adding DOX @ MSN and CuNO 3 ·3H 2 O and NaOH are dispersed in deionized water in sequence for continuous reactionAdding H 2 O 2 Then stirring for 1 hour at normal temperature, and finally collecting DOX @ MSN @ CuO by freeze drying 2 。
An application of multifunctional nano catalyst in preparing the medicines for treating cancer is disclosed.
Compared with the prior art, the invention has the following beneficial effects:
(1) DOX @ MSN @ CuO in the invention 2 Capable of self-producing exogenous H 2 O 2 And reacting in a specific tumor microenvironment TME; DOX @ MSN @ CuO 2 After being endocytosed into tumor cells, the tumor cells are firstly decomposed into Cu in a weakly acidic specific tumor microenvironment TME 2+ And exogenous H 2 O 2 (ii) a Subsequent Cu 2+ Reacting with local glutathione to deplete glutathione and deplete Cu 2+ Reduction to Cu + (ii) a Final Cu + With exogenous H 2 O 2 The tumor cell apoptosis is caused by generating toxic OH through Fenton-like reaction, and the toxic OH is combined with DOX to further accelerate the tumor cell apoptosis.
(2) The DOX-loaded copper peroxide nanodot modified mesoporous silica nanocatalyst DOX @ MSN @ CuO synthesized by the invention 2 Realizes the integration of chemo-kinetic treatment and chemotherapy and can solve the problem of H 2 O 2 Limited and insufficient efficacy of chemokinetic therapy CDT due to overexpression of glutathione GSH; in which H is self-supplied 2 O 2 And Cu 2+ GSH depletion combined with CDT enhancement and combined with DOX-induced chemotherapy results in DOX @ MSN @ CuO 2 Can effectively inhibit the growth of tumors in vivo and has small side effect.
(3) DOX @ MSN @ CuO in the invention 2 Can decompose and generate exogenous H in specific tumor microenvironment TME 2 O 2 While using Cu 2+ Effectively catalyze Fenton reaction, and solve the problems of low reaction efficiency and H in chemokinetic therapy 2 O 2 The problem of insufficient concentration; meanwhile, the successful release of the adriamycin DOX realizes the synergistic effect of the chemodynamic therapy CDT and chemotherapy, and improves the treatment effect of tumors.
(4) DOX @ MSN @ CuO in the invention 2 Has the advantages ofGood biocompatibility and in vivo stability, and can position the target tumor region by enhancing permeability and retention effect; depletion of glutathione GSH to produce Cu upon entry into cancer cells + Catalyzing exogenous H 2 O 2 Generating virulent OH; the release of DOX, depletion of glutathione GSH and production of OH together amplify the cascade of anti-tumor effects.
Drawings
FIG. 1 shows DOX @ MSN @ CuO 2 And a schematic diagram of the process of synthesis and killing of cancer cells.
FIG. 2 shows DOX @ MSN @ CuO 2 Schematic representation of the synergistic chemo-kinetic therapy and chemotherapy in cancer cells in combination to kill cancer cells.
Fig. 3 is a schematic diagram of 4T1 tumor xenograft establishment.
Fig. 4 is a photograph of tumors in nine groups after treatment.
FIG. 5 is MSN @ CuO 2 An ESR pattern of OH was generated at different times.
FIG. 6 is MSN @ CuO 2 Generated at different times 1 O 2 ESR graph of (d).
FIG. 7 is CuO 2 ESR patterns of OH were generated at different times.
FIG. 8 is a graph of the concentration of GSH measured by MSN @ CuO using DTNB 2 Consumption situation diagram.
FIG. 9 is DOX @ MSN and DOX @ MSN @ CuO 2 Cumulative release profile of drug at different pH values.
Fig. 10 is a graph showing the release of copper ions at different pH values of the solutions (pH 7.4 and 5.0).
FIG. 11 shows 4T1 cells at different concentrations of MSN and MSN @ CuO 2 Cytotoxicity profile after incubation.
Detailed Description
The present invention is further illustrated by the following figures and examples, which include, but are not limited to, the following examples.
Example 1
As shown in figure 1 and figure 2, the multifunctional nano-catalyst is DOX-supported copper peroxide nano-dot modified mesoporous silica nano-catalyst DOX @ MSN @ CuO 2 Can be fromProducing exogenous H 2 O 2 And reacts in the specific tumor microenvironment TME.
DOX@MSN@CuO 2 After endocytosis into tumor cells, first, Cu is decomposed in weakly acidic TME 2+ And exogenous H 2 O 2 (ii) a Subsequent Cu 2+ Reacting with local glutathione to deplete glutathione and deplete Cu 2+ Reduction to Cu + (ii) a Final Cu + With exogenous H 2 O 2 The Fenton-like reaction is carried out to generate toxic OH, so that the tumor cell apoptosis is caused, and the reaction rate of the process is fast. Meanwhile, the combination of chemokinetic treatment and chemotherapy is realized with the loaded adriamycin DOX; can solve the problem of H 2 O 2 Limited and glutathione GSH overexpression causes insufficient efficacy of chemokinetic therapy CDT and limitations of monotherapy methods. Self-supplying H 2 O 2 (ii) potentiation of chemokinetic therapy CDT in combination with GSH depletion and combination with DOX-induced chemotherapy to produce DOX @ MSN @ CuO 2 Has the characteristics of effective inhibition of tumor growth and small side effect in vivo.
Example 2
As shown in fig. 1 and 2, a preparation method of a multifunctional nano catalyst is to synthesize mesoporous silica nanoparticles MSN first; then loading the chemotherapy drug DOX on the MSN to obtain DOX @ MSN; finally, the mixture is processed by DOX @ MSN and copper nitrate trihydrate CuNO 3 ·3H 2 Preparation of DOX @ MSN @ CuO from O and NaOH 2 。
Material
Ammonium hydroxide (NH) 3 ·H 2 O), absolute ethyl alcohol (EtOH), Cetyl Trimethyl Ammonium Bromide (CTAB), Tetraethoxysilane (TEOS), 3-Aminopropyltriethoxysilane (APTES), hydrochloric acid (HCl), Triethylamine (TEA), copper nitrate trihydrate (CuNO) 3 ·3H 2 O), sodium hydroxide (NaOH), hydrogen peroxide (H) 2 O 2 ) Hydrochloric acid (HCl), dimethyl sulfoxide (DMSO), potassium permanganate (KMnO) 4 ) 3,3,5, 5-Tetramethylbenzidine (TMB) was purchased from Doxolone chemical Co., Ltd. 5,5' -Dithiobenzimide (2-nitrobenzoic acid) (DTNB), Glutathione (GSH), Doxorubicin hydrochloride (HCl-DOX) and 5, 5-dimethyl-1-pyrroline oxide (DMPO) were purchased fromFrom alatin (shanghai, china). 3- (4, 5-dimethylthiazol-2-yl) -2, 5-diphenyltetrazolium ammonium bromide (MTT), Calcein acetoxymethyl ester (Calcein-AM), Propidium Iodide (PI), 4 ', 6-diamino-2-phenylindole Dihydrochloride (DAPI), 2 ', 7 ' -dichlorofluorescein diacetate (DCFH-DA), an apoptosis kit with annexin V-FITC and PI, and a glutathione/oxidized glutathione (GSH/GSSG) assay kit were purchased from Bizungtian (Shanghai, China).
Example 3
The MSN synthesis process comprises the following steps: first, 2g, 0.5mmol of NH 3 ·H 2 Dissolving O in deionized water, and continuously stirring at 80 ℃; then, to the above solution was added an ethanol solution containing 3g, 0.58mmol of ethyl orthosilicate, 1g, 0.1mmol of CTAB and 500. mu.L of APTES; after 10 hours of reaction, HCl was added to ethanol to remove the template and the product was collected by high speed centrifugation.
Example 4
The synthesis process of DOX @ MSN is as follows: dispersing MSN and DOX & HCI in DMSO according to a mass ratio of 4:1, adding triethylamine TEA, and continuously stirring for 24 hours at room temperature in the dark; free DOX was removed by repeated centrifugation and DOX @ MSN was collected by freeze drying.
Example 5
DOX@MSN@CuO 2 The synthesis process comprises the following steps: 500mg DOX @ MSN, 50mg CuNO 3 ·3H 2 O and 30mg NaOH were dispersed in 40mL of deionized water in this order to continue the reaction, and 2.5mL of 3% H was added 2 O 2 Stirring for 1 hour at normal temperature after reaction, and finally collecting DOX @ MSN @ CuO by freeze drying 2 。
Characterization of
DOX@MSN@CuO 2 As shown in fig. 1, the MSN nanoparticles having a particle size of about 80nm and CuO were shown to have a particle size of about 80nm by Scanning Electron Microscope (SEM) and Transmission Electron Microscope (TEM) images 2 The particle size of the coated MSN was about 100nm and X-ray Dispersion Spectroscopy (EDS) images showed MSN @ CuO 2 Consisting of Cu, O, Si, etc., indicating CuO 2 Successfully loaded to the MSN.
Detection of oxidizability
MSN @ CuO by X-ray photoelectron spectroscopy (XPS) 2 The appearance of distinct O1s, Cu 2p and N1s peaks in the sample was observed; MSN @ CuO 2 The peaks of O1s near 531.5eV and 533.2eV are attributed to the Cu-O bond and the O-O bond, demonstrating the presence of peroxy groups. The XPS spectrum of Cu 2p has two main peaks at 952.4eV and 933.4eV and two satellite peaks at 962.5eV and 941.2eV, indicating that copper is in a divalent oxidation state (Cu) 2+ ) (ii) a The unique N1s peak was attributed to the amino groups on the MSN surface.
To determine the ability to generate active oxygen, CuO was examined by Electron Spin Resonance (ESR) 2 And MSN @ CuO 2 The ability to catalyze the production of OH, as shown in FIG. 5, a unique quartet (1:2:2:1) can be seen to indicate that OH is produced, and the intensity of the peak becomes more pronounced with the increase of the culture time; the unique three peaks in FIG. 6 confirm singlet oxygen: ( 1 O 2 ) Forming; FIG. 7 shows CuO 2 Has the capability of generating OH, and CuO is increased by the existence of MSN 2 The surface active site of (1) improves the catalytic activity, so that the MSN @ CuO 2 A large amount of OH can be generated within 5 minutes.
In addition, acidic potassium permanganate (KMnO) 4 ) Can be used to demonstrate H 2 O 2 Forming; MSN @ CuO is added 2 After that, acidic KMnO 4 The purple color of (1) disappears, its effect is equal to H 2 O 2 The same; interpreted as MSN @ CuO 2 In acidic solution there is H production 2 O 2 In contrast, MSN nanoparticles do not cause a color change.
MSN @ CuO was measured by 3,3,5, 5-Tetramethylbenzylamine (TMB) color development 2 Generating ROS that oxidize a colorless TMB substrate to blue ox-TMB with a maximum absorbance occurring at 650 nm; the results show MSN @ CuO 2 +GSH+H 2 O 2 And MSN @ CuO 2 + GSH enables the conversion of colorless TMB to blue ox-TMB, with Cu added separately 2+ Or H 2 O 2 No color change occurs; indicating MSN @ CuO 2 Has excellent catalytic activity in an acidic environment and can be decomposed into Cu 2+ And H 2 O 2 (ii) a Subsequently, Cu 2+ Reaction with GSH to Cu + Formed of Cu + And H 2 O 2 A Fenton-like reaction was performed to generate OH, which induces a rapid oxidation of TMB, showing a strong absorbance at 650 nm.
Glutathione (GSH) is an important intracellular antioxidant (0.1-10mM), widely exists in organisms, and can protect cells from high reactivity OH damage. The presence of GSH in tumor cells impairs the effect of reactive oxygen species, and therefore DTNB was used to detect MSN @ CuO 2 The depletion effect on GSH is very essential. Coupling GSH with DTNB to produce diphenyl 3-thio-6-nitrobenzoic acid (TNB) 2- ) Has obvious absorption peak at 412nm, and gradually becomes MSN @ CuO along with GSH 2 Consumption, TNB 2- Is reconverted to DTNB. As shown in FIG. 8, MSN @ CuO 2 GSH was consumed even at a concentration of 10mM, and its excellent oxidation properties were fully exhibited.
In vitro acid-induced DOX and copper ion release
The drug Loading Capacity (LC) and drug Loading Efficiency (LE) of MSN were calculated to be 15% and 68%, respectively, according to the following equations.
Figure BDA0003696771680000081
Figure BDA0003696771680000082
MSN has no absorption peak in the ultraviolet spectrum, and DOX @ MSN @ CuO 2 Shows obvious absorbance at 480nm, and indicates that DOX is successfully encapsulated into DOX @ MSN and DOX @ MSN @ CuO 2 In (1).
Adding MSN @ CuO to dialysis bag (MWCO ═ 1000) 2 (2mL, 1mg/mL) were immersed in buffers at different pH values (7.4 or 5.0). Then, the shaker was set to 37 ℃ and the rotation speed was set to 100 rpm. Collecting the buffer at a predetermined time (30 minutes, 1 hour, 2 hours, 4 hours, 8 hours, 12 hours, 24 hours, 36 hours, 48 hours) to detect the released copper ions by inductively coupled plasma emission spectroscopy (ICP-OES); as shown in the figureAs shown in fig. 9, approximately 50% of DOX was released in acidic buffer (pH 5.0) compared to less than 10% in neutral buffer (pH 7.4), indicating that DOX release is pH dependent, which contributes to DOX @ MSN @ CuO 2 Maintaining structural integrity in the blood circulation.
Measurement of MSN @ CuO by ICP-OES 2 Cu in (1) 2+ Release behavior, as shown in FIG. 10, MSN @ CuO 2 Nearly 90% of Cu was released in ABS buffer (pH 5.0) 2+ While the copper ion content in PBS buffer (pH 7.4) was negligible, further confirming that the acidic environment initiated CuO 2 Thereby liberating Cu 2+ 。
In vitro cytotoxicity
Vascular Endothelial Cells (EC), mouse breast cancer cells (4T1), and mouse melanoma cells (B16) were obtained from southwestern university of transportation. Mouse colon cancer cells (CT26) were obtained from the institute of adult medical science. 4T1 cells and CT26 cells were cultured in RPMI 1640 supplemented with 10% Fetal Bovine Serum (FBS), and B16 cells were cultured in DEME medium supplemented with 10% FBS at 37 deg.C under CO 2 The concentration was 5% and the humidity was 100%.
The MTT method detects the biocompatibility of the nanoparticles to tumor cells. EC. 4T1, B16 and CT26 cells at 5X 10 cells/well, respectively 3 Individual cells were seeded at density in 96-well plates. Cu in MSN @ CuO 2 The content in (B) was determined to be 3% by ICP-OES. After overnight incubation, MSN and MSN @ CuO were used separately 2 Cells were co-cultured for 24 hours at various concentrations (material: 0, 50, 100, 200, 400, 600, 800 mg/mL. Cu: 0, 1.5, 3, 6, 12, 18, 24 mg/mL). Then, the cells were incubated with MTT for 4 hours. After removal of the medium and addition of DMSO, OD values were obtained using a microplate reader.
The formation amount of MTT crystal is in direct proportion to the number of living cells in a certain range; as shown in FIG. 11, MSN @ CuO 2 The toxicity to 4T1 cells is strongest, and when the concentration is increased to 800mg/mL, the cell survival rate is reduced to below 20 percent; however, the survival rate of EC after 24 hours of culture was maintained above 60% even at a high concentration of 800 mg/mL. This is due to the presence of normal cellsNeutral pH value of MSN @ CuO 2 Decomposing to make MSN @ CuO 2 Has good cell compatibility with normal cells; the two groups of tumor cells used as controls, free DOX and DOX @ MSN, have reduced cell death, which indicates that CDT has superiority compared with chemotherapy; indicating DOX @ MSN @ CuO 2 The compound has good cytotoxicity on tumor cells even under low concentration, which shows that the CDT combined chemotherapy has good treatment effect in tumor treatment and wide prospect; the results show MSN @ CuO 2 Has general toxicity and can kill other cancer cells, such as B16 and CT26 cells.
The AM/PI cell double staining method is used for observing the survival condition of the cells under the visualization condition. It is clear that DOX @ MSN @ CuO 2 Induced tumor cell apoptosis was greater than control DOX @ MSN and MSN @ CuO 2 This is due to the synergistic effect of CDT in combination with chemotherapy being better than CDT alone or chemotherapy alone, and the results are equally applicable to B16 cells and CT26 cells.
Flow cytometry results for 4T1 cells showed MSN @ CuO after various time treatments (1 hour, 3 hours, 6 hours) 2 The most apoptosis was induced within 6 hours, indicating a good treatment effect for CDT.
GSH consumption and reactive oxygen species generation
To verify whether CDT drugs can reduce GSH, MSN @ CuO was studied at the cellular level 2 Has GSH scavenging effect. At different concentrations of MSN @ CuO 2 After (0, 50, 100, 200, 400, 800mg/mL) co-culture, the intracellular GSH content in 4T1 cells decreased significantly as expected, further confirming that MSN @ CuO was present prior to ROS production 2 GSH can be consumed.
DCFH-DA is a cell membrane permeability indicator of reactive oxygen species; when it crosses the cell membrane, it is converted to non-fluorescent DCFH, which, after oxidation by OH, produces the strongly fluorescent product 2 ', 7' -Dichlorofluorescein (DCF), which can be detected by fluorescence microscopy. The results show that 4T1 cells and DOX @ MSN @ CuO 2 Shows the strongest green fluorescence, indicating DOX @ MSN @ CuO 2 A large amount of. OH can be produced by Fenton reaction.
Intracellular uptake studies
Electronic display by biological transmissionDirect microscopic (Bio-TEM) observation of 4T1 cells at different time intervals (1 hour, 3 hours, and 6 hours) versus MSN @ CuO 2 The phagocytic process of (1). MSN @ CuO was observed after 1 hour of co-incubation 2 Present near the microvilli of tumor cells; MSN @ CuO was observed over 3 hours 2 Gradually endocytosed by the tumor cells; a large amount of apoptosis was observed at 6 hours, with complete destruction of the structure. These results indicate MSN @ CuO 2 Has high cytotoxicity to tumor cells, and is expected to play a role in subsequent treatment.
To assess nanoparticle uptake by cells, 4T1 cells were incubated with free DOX and FITC @ DOX @ MSN @ CuO 2 After co-incubation for 1 hour, 3 hours and 6 hours, observation was carried out by laser Confocal (CLSM). Among them, DOX fluoresces spontaneously in red, and MSN is labeled as green fluorescence by FITC. At 1 hour, the fluorescence intensity of the cells after the culture of free DOX is obviously higher than FITC @ DOX @ MSN @ CuO due to the fact that DOX enters the cells through diffusion 2 . The time increased to 6 hours, FITC @ DOX @ MSN @ CuO was observed 2 The red fluorescence intensity of (A) is higher than that of free DOX, because FITC @ DOX @ MSN @ CuO 2 Can be effectively absorbed by tumor cells, and the pH responsive nano catalyst promotes the release of DOX.
In vivo antitumor effect
45 tumor-bearing female BALB/c mice were randomly divided into 9 groups (n-5). Tumor growth to 50mm 3 Thereafter, the tail vein and intratumoral injection of PBS, free DOX, DOX @ MSN, MSN @ CuO, was administered to the mice 2 ,DOX@MSN@CuO 2 (shown in FIG. 3). The body weight and the tumor volume are measured every two days, and after treatment for 15 days by intravenous injection and intratumoral injection, the body weight average of each group of mice is not changed obviously, but the change of the tumor volume is different obviously, and the tumor volume of the PBS group and the DOX group is increased obviously. At the end of the experiment, tumor tissue was removed from the mice and weighed. As shown in fig. 4, intratumoral injection showed better antitumor effect than intravenous injection. The difference between the PBS group and the free DOX group is not significant, which indicates that the treatment of the free DOX has no obvious anti-tumor effect, and the MSN @ CuO 2 CDT with moderate antitumor potency DOX @ MSN @ CuO 2 The best antitumor efficacy was shown in all groups, indicating an enhanced efficacy of the combination chemotherapy and CDT treatment。
In vivo biosafety study
The blood biochemical analysis shows that tail vein injection and intratumoral injection DOX @ MSN @ CuO 2 The contents of CK, CK-MB, LDH, ALP, AST, UREA and CREA are not influenced, and the material has the minimum systemic toxicity and obvious anti-tumor effect.
H of copper peroxide coated mesoporous silica for cooperating CDT and chemotherapy 2 O 2 The self-supply and GSH consumption nano catalyst has the characteristics of high drug loading, strong stability, excellent anti-tumor capability and the like; DOX @ MSN @ CuO 2 After entering tumor cells, CuO is reacted under the action of acidic TME 2 Degradation to Cu 2+ And oxidation-reduction reaction is carried out between the cancer cell and GSH, so that the oxidation resistance of the tumor is effectively relieved, the cancer cell is more easily damaged by OH, and the curative effect of CDT is further improved. Meanwhile, the chemotherapeutic drug DOX inhibits the growth of tumor cells through DNA damage and other ways. Self-produced H 2 O 2 ，Cu 2+ GSH is consumed, a large amount of OH and loaded chemotherapeutic drug DOX are generated, so that the nano-catalyst can effectively inhibit tumor growth, and CDT and chemotherapy are used for treating cancers in a synergistic manner.
The invention is well implemented in accordance with the above-described embodiments. It should be noted that, based on the above structural design, in order to solve the same technical problems, even if some insubstantial modifications or colorings are made on the present invention, the adopted technical solution is still the same as the present invention, and therefore, the technical solution should be within the protection scope of the present invention.

Claims (6)

1. The multifunctional nano catalyst is characterized in that the catalyst is DOX-loaded copper peroxide nano dot modified mesoporous silica nano catalyst DOX @ MSN @ CuO 2 。
2. The method for preparing the multifunctional nano catalyst as recited in claim 1, comprising the steps of:
s1, synthesizing mesoporous silica nano-particles MSN;
s2, loading chemotherapeutic drug DOX on the MSN in the step S1 to obtain DOX @ MSN;
s3, DOX @ MSN and CuNO as copper nitrate trihydrate through step S2 3 ·3H 2 Preparation of DOX @ MSN @ CuO from O and sodium hydroxide NaOH 2 。
3. The method for preparing the multifunctional nano-catalyst according to claim 2, wherein the specific process of the step S1 is as follows: reacting NH 3 ·H 2 Dissolving O in deionized water to form a solution, and continuously stirring at 80 ℃; then, adding an ethanol solution containing tetraethoxysilane, Cetyl Trimethyl Ammonium Bromide (CTAB) and 3-Aminopropyltriethoxysilane (APTES) into the solution, and reacting for 10 hours; HCl was added thereto to remove the template, and the product MSN was collected by high speed centrifugation.
4. The method for preparing the multifunctional nano-catalyst according to claim 3, wherein the specific process of the step S2 is as follows: dispersing MSN and DOX & HCI in dimethyl sulfoxide DMSO according to a mass ratio of 4:1, adding triethylamine TEA, and continuously stirring for 24 hours at room temperature in the dark; free DOX was removed by repeated centrifugation and DOX @ MSN was collected by freeze drying.
5. The method for preparing the multifunctional nano-catalyst according to claim 4, wherein the specific process of the step S3 is as follows: adding DOX @ MSN and CuNO 3 ·3H 2 Dispersing O and NaOH in deionized water in sequence for continuous reaction, and adding H 2 O 2 Then stirring for 1 hour at normal temperature, and finally collecting DOX @ MSN @ CuO by freeze drying 2 。
6. Use of a multifunctional nanocatalyst according to any one of claims 1 to 5 in a medicament for the treatment of cancer.
CN202210682380.6A 2022-06-15 2022-06-15 Multifunctional nano catalyst and preparation method thereof Active CN115054613B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN202210682380.6A CN115054613B (en) 2022-06-15 2022-06-15 Multifunctional nano catalyst and preparation method thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202210682380.6A CN115054613B (en) 2022-06-15 2022-06-15 Multifunctional nano catalyst and preparation method thereof

Publications (2)

Publication Number Publication Date
CN115054613A true CN115054613A (en) 2022-09-16
CN115054613B CN115054613B (en) 2023-05-09

Family

ID=83200320

Family Applications (1)

Application Number Title Priority Date Filing Date
CN202210682380.6A Active CN115054613B (en) 2022-06-15 2022-06-15 Multifunctional nano catalyst and preparation method thereof

Country Status (1)

Country Link
CN (1) CN115054613B (en)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116327979A (en) * 2023-05-25 2023-06-27 西南石油大学 Transition metal-based mesoporous nano catalytic medicine, preparation method and application
CN117160455A (en) * 2023-06-21 2023-12-05 中国科学院上海硅酸盐研究所 P-d orbit hybridization enhanced Fenton-like inorganic nano material and preparation method and application thereof

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
LI-SEN LIN等: "Synthesis of Copper Peroxide Nanodots for H2O2 Self-Supplying Chemodynamic Therapy" *
楼佳栋等: "Ｃｕ２＋ 螯合封孔的介孔硅载药纳米颗粒的制备及应用" *

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116327979A (en) * 2023-05-25 2023-06-27 西南石油大学 Transition metal-based mesoporous nano catalytic medicine, preparation method and application
CN117160455A (en) * 2023-06-21 2023-12-05 中国科学院上海硅酸盐研究所 P-d orbit hybridization enhanced Fenton-like inorganic nano material and preparation method and application thereof
CN117160455B (en) * 2023-06-21 2024-05-10 中国科学院上海硅酸盐研究所 P-d orbit hybridization enhanced Fenton-like inorganic nano material and preparation method and application thereof

Also Published As

Publication number Publication date
CN115054613B (en) 2023-05-09

Similar Documents

Publication Publication Date Title
He et al. A solid lipid coated calcium peroxide nanocarrier enables combined cancer chemo/chemodynamic therapy with O2/H2O2 self-sufficiency
Barahuie et al. Graphene oxide as a nanocarrier for controlled release and targeted delivery of an anticancer active agent, chlorogenic acid
CN115054613B (en) Multifunctional nano catalyst and preparation method thereof
Ko et al. Graphene quantum dot-based theranostic agents for active targeting of breast cancer
Dumur et al. Controlled spontaneous generation of gold nanoparticles assisted by dual reducing and capping agents
Xiao et al. Core–shell structured 5-FU@ ZIF-90@ ZnO as a biodegradable nanoplatform for synergistic cancer therapy
CN111253581A (en) Metal organic framework material for enhancing combination of chemical power therapy and hunger therapy, preparation method and application
Gao et al. Tumor-targeted biocatalyst with self-accelerated cascade reactions for enhanced synergistic starvation and photodynamic therapy
US11638700B2 (en) Iron/shikonin nano-composite and use thereof and method for preparing the same by supermolecular self-assembly
Gu et al. Dual catalytic cascaded nanoplatform for photo/chemodynamic/starvation synergistic therapy
Song et al. Fabrication of the biomimetic DOX/Au@ Pt nanoparticles hybrid nanostructures for the combinational chemo/photothermal cancer therapy
Cui et al. Metal–organic framework-encapsulated nanoparticles for synergetic chemo/chemodynamic therapy with targeted H 2 O 2 self-supply
Cheng et al. Multimodal therapies: glucose oxidase-triggered tumor starvation-induced synergism with enhanced chemodynamic therapy and chemotherapy
Zhang et al. Catalyzing Generation and Stabilization of Oxygen Vacancies on CeO2− x Nanorods by Pt Nanoclusters as Nanozymes for Catalytic Therapy
Shaik et al. Optical emissions of chitosan modified LaAlO3: Bi3+, Tb3+ nanoparticles for bio labeling and drug delivery to breast cancer cells
CN114732795B (en) Preparation method of long-circulation multifunctional metal organic framework nano preparation
CN116327979B (en) Transition metal-based mesoporous nano catalytic medicine, preparation method and application
CN117045823B (en) Mesoporous hollow copper single-atom carbon sphere, preparation method and application
Sheykhisarem et al. In vitro biocompatibility evaluations of pH-sensitive Bi2MoO6/NH2-GO conjugated polyethylene glycol for release of daunorubicin in cancer therapy
Alamer et al. Synergizing effects of chemodynamic therapy and chemotherapy against breast cancer by oxaliplatin-loaded polydopamine/BSA@ copper ferrite
Shao et al. Efficient free radical generation against cancer cells by low-dose X-ray irradiation with a functional SPC delivery nanosystem
Liang et al. H2O2/O2 self-supply and Ca2+ overloading MOF-based nanoplatform for cascade-amplified chemodynamic and photodynamic therapy
Zhao et al. Cu3-xP nanocrystals filled halloysite nanotubes for chemodynamic therapy of breast cancer
CN113952470B (en) Nanoparticle of dual active oxygen generator and preparation method and application thereof
CN111450081B (en) Protein nanoparticle for enhanced dynamic therapy, preparation method and application thereof

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant