CN114904011B - Non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material, and preparation method and application thereof - Google Patents

Non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material, and preparation method and application thereof Download PDF

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CN114904011B
CN114904011B CN202110763296.2A CN202110763296A CN114904011B CN 114904011 B CN114904011 B CN 114904011B CN 202110763296 A CN202110763296 A CN 202110763296A CN 114904011 B CN114904011 B CN 114904011B
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林翰
吴陈瑶
施剑林
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Shanghai Institute of Ceramics of CAS
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Abstract

The invention discloses a non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material, and a preparation method and application thereof. The non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material comprises cobalt molybdate-phosphomolybdic acid composite nano-sheets and biocompatible polymers with surfaces modified with the composite nano-sheets; the mass ratio of cobalt molybdate to phosphomolybdic acid is 1:2-1:4, a step of; the mass ratio of the cobalt molybdate-phosphomolybdic acid composite nano-sheet to the biocompatible polymer is 1:5-1:15. the non-iron-based glutathione consumption synergistic active oxygen species enhanced composite material can react with hydrogen peroxide to generate same active OH, and meanwhile GSH antioxidants in tumor environments can be reduced to achieve effective lipid peroxide accumulation.

Description

Non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material, and preparation method and application thereof
Technical Field
The invention belongs to the technical field of biological nano materials, and particularly relates to a non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material, and a preparation method and application thereof.
Background
The modes of cell death mainly include apoptosis and non-apoptosis. The anti-apoptotic capacity of the tumor itself and the tumor growth require more iron accumulation and lipid metabolism, so iron death as a non-apoptotic means of cell death is of more interest. Oncogenic signals stimulate oxidative activity-related enzymes such that Reactive Oxygen Species (ROS) are increased, often accompanied by the metabolism of Glutathione (GSH) with antioxidant capacity to eliminate highly expressed reactive oxygen species. Utilizing hydrogen peroxide rich in tumor microenvironment, fe 2+ The directed Fenton (Fenton) reaction produces highly toxic hydroxyl radicals (. OH), which promote the production of lipid peroxides and thus lead to iron death. However, the presence of GSH with reactive oxygen scavenging capability prevents the use of treatments such as ROS-induced chemical kinetics, acoustic kinetics, and photodynamic, and iron-based materials are prone to cause normal tissue iron allergy. And the presence of GSH maintains the activity of glutathione peroxidase 4 (GPX 4) to reduce toxic lipid peroxides to low toxicity fatty alcohols, resulting in reduced iron death effects. Therefore, the development of other metal compound alternatives to enable their use in iron death therapy is a technical problem that is currently in urgent need to be addressed.
Disclosure of Invention
In view of the above problems, the invention provides a non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material, a preparation method and application thereof, wherein the non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material can react with hydrogen peroxide to generate same-activity OH, and meanwhile GSH antioxidants in tumor environments can be reduced to achieve effective accumulation of lipid peroxides.
In a first aspect, the present invention provides a non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite. The non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material comprises cobalt molybdate-phosphomolybdic acid composite nano-sheets and biocompatible polymers with surfaces modified with the composite nano-sheets; the mass ratio of cobalt molybdate to phosphomolybdic acid is 1:2-1:4, a step of; the mass ratio of the cobalt molybdate-phosphomolybdic acid composite nano-sheet to the biocompatible polymer is 1:5-1:15. by controlling the mass ratio within this range, the biocompatibility of the material and the performance of the glutathione consumption synergistic active oxygen species can be ensured.
Preferably, the biocompatible polymer comprises at least one of polyether F127, soybean phospholipid, distearyl phosphatidylethanolamine-polyethylene glycol, preferably polyether F127.
Preferably, the cobalt molybdate-phosphomolybdic acid composite nano-sheet is a composite uniformly dispersed in the nano-sheet and interwoven with each other.
Preferably, the non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material has both glutathione consumption capacity and active oxygen production capacity; preferably, the non-iron-based glutathione depletion synergizes with the redox reaction of the reactive oxygen species-enhanced composite material with the glutathione overexpressed in the tumor to eliminate the glutathione and increase the production of reactive oxygen species during its redox reaction with the glutathione.
Preferably, the product of the non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material and glutathione subjected to oxidation-reduction reaction is further reacted with hydrogen peroxide to generate singlet oxygen through a roxithromycin mechanism 1 O 2 Reactive oxygen species.
In a second aspect, the present invention provides a method for preparing a non-iron-based glutathione consumption synergistic active oxygen species enhanced composite material as described in any one of the preceding claims. The preparation method comprises the following steps: mixing a molybdenum source, a cobalt source and an organic solvent to obtain a mixed solution; transferring the mixed solution into an autoclave for reaction, centrifugally washing after the reaction is finished, and collecting a reaction product; and (3) dispersing and mixing the reaction product and a biocompatible polymer to obtain the non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material.
Preferably, the molybdenum source is polyoxometalate containing molybdenum, and the concentration of the molybdenum source is 0.01-0.02mM; the cobalt source is cobalt salt, and the concentration of the cobalt source is 0.06-0.07mM; the organic solvent is a mixture of acetone, ethanolamine and oleylamine, preferably, the volume ratio of the acetone, the ethanolamine and the oleylamine is (1-4): (1-4): (1-4).
Preferably, the reaction temperature is 160-200 ℃ and the reaction time is 3-12h.
Preferably, the reaction product is mixed with the biocompatible polymer in an organic solvent and the organic solvent is subsequently removed; the organic solvent comprises at least one of dichloromethane, ethanol, chloroform and cyclohexane.
In a third aspect, the present invention provides the biological use of a non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite of any of the above as an iron death inducer.
Drawings
FIG. 1 is a transmission electron micrograph of cobalt molybdate-phosphomolybdic acid composite nanoplatelets (CPMNSs);
FIG. 2 is a scanning electron micrograph of a cobalt molybdate-phosphomolybdic acid composite nanoplatelet;
FIG. 3 is a Mo, P, co, O spectrum elemental analysis spectrum of a cobalt molybdate-phosphomolybdic acid composite nano-sheet; the length of the scale in each figure is consistent with the scale unit, and the scale unit is 2.5 mu m;
FIG. 4 is an X-ray diffraction (XRD) spectrum of a cobalt molybdate-phosphomolybdic acid composite nano-sheet;
FIG. 5 (a) is a schematic diagram of electron paramagnetic resonance spectrum detection of hydroxyl radical of cobalt molybdate-phosphomolybdic acid composite nano-sheet, (b) is CPMS detected by 5,5' -dithiobis (2-nitrobenzoic acid (DTNB) indicator and GSH content in GSH reaction system, wherein the content is characterized by absorbance, (c) is ultraviolet absorption spectrum of Methylene Blue (MB) degradation caused by active oxygen, each curve represents only MB, 2mM GSH, 0mM GSH, 0.5mM GSH and 1mM GSH from top to bottom in sequence, (d) is cumulative release amount of cobalt ions at different time points under GSH condition and under GSH-free condition;
fig. 6 (a) is an X-ray photoelectron (XPS) spectrum of Mo valence state change after GSH reduction, and (b) is a schematic diagram of ESR spectrum detection of generation of singlet oxygen by reaction of reduced CPMNSs (ReCPMNSs) with hydrogen peroxide;
FIG. 7 (a) shows the effect of iron death inhibitor Ferrostatin-1 (Fer-1) on the cell viability of CPMNSs-induced mouse breast cancer (4T 1), and (b) shows the effect of iron death inhibitor Liproxstatin-1 (Lip-1) on the cell viability of CPMNSs-induced mouse breast cancer (4T 1); (c) The intracellular GSH content measurement after the CPMS is cultivated for different concentrations, and the intracellular GSH content measurement after the CPMS is cultivated for different times;
FIG. 8 (a) shows intracellular GPX4 activity measurement after CPMS incubation at different concentrations, (b) shows intracellular GPX4 activity measurement after CPMS incubation at different times, (c) shows intracellular Malondialdehyde (MDA) content measurement after CPMS incubation at different concentrations, and (d) shows intracellular Malondialdehyde (MDA) content measurement after CPMS incubation at different times;
fig. 9 is a plot of tumor volume versus time for four treatment groups Control, CPMNSs i.v., cpms i.t., and cpms i.t. + DFOM; the Control group does not perform any processing; CPMNSs i.v. group 10mg/kg CPMNSs was injected at day 0 and day 7 tail vein (i.v.); CPMNSs i.t. group 10mg/kg CPMNSs were injected in tail tumors (i.t.) on days 0 and 7; CPMNSs i.t. + DFOM group 20mg/kg iron death inhibitor deferoxamine mesylate (DFOM) was intraperitoneally injected on days 0, 4, 8, 12 based on CPMNSs i.t. group;
FIG. 10 is intratumoral Reactive Oxygen Species (ROS) and lipid peroxidation Level (LPO) assays for four treatment groups Control, CPMNSs i.v., CPNSs i.t., and CPNSs i.t. + DFOM; the length of the scale in each figure is consistent with the unit of the scale, and the unit of the scale is 200 mu m;
FIG. 11 is an intratumoral GPX4 immunohistochemical section of four treatment groups Control, CPMNSs i.v., CPNSs i.t., and CPNSs i.t. + DFOM; the scale units of the figures are kept consistent and are 100 μm;
FIG. 12 is a graph of tumor hematoxylin-eosin staining (H & E), deoxyribonucleotide terminal transferase (TdT) mediated notch end labeling (TUNEL) and Ki-67 sections of four treatment groups Control, CPMNSs i.v., CPMNs i.t., and CPMNs i.t. + DFOM after treatment is completed; the scale units of the figures are kept consistent and are 100 μm;
fig. 13 is a record of experimental mouse growth curves, i.e., growth rate versus time, for four treatment groups Control, CPMNSs i.v., cpms i.t., and cpms i.t. + DFOM.
Detailed Description
The invention is further illustrated by the following embodiments, which are to be understood as merely illustrative of the invention and not limiting thereof. Unless otherwise specified, each percentage refers to a mass percent.
The present disclosure provides a non-iron-based glutathione consuming synergistic reactive oxygen species reinforced composite. Glutathione depletion and active oxygen species enhanced cobalt molybdate (CoMoO), which may also be referred to as tumor microenvironment response 4 ) Phosphomolybdic acid (H) 3 PMo 12 O 40 ) The composite material, or the degradable glutathione consumes synergistic active oxygen species to strengthen the composite material. The non-iron-based glutathione consumption synergistic active oxygen species enhanced composite includes cobalt molybdate (CoMoO) with glutathione consumption and active oxygen species enhancement 4 ) Phosphomolybdic acid (H) 3 PMo 12 O 40 ) Composite nanoplatelets and biocompatible polymers that modify the composite nanoplatelets. The surface-modified biocompatible polymer is critical to impart biocompatibility and colloidal stability to the composite nanoplatelets.
Cobalt molybdate (CoMoO) 4 ) Phosphomolybdic acid (H) 3 PMo 12 O 40 ) The composite nano-sheet is structurally characterized by uniformly dispersed and mutually interwoven two-dimensional nano-sheets. The cobalt provided by the composite nano-sheet can generate high-toxicity hydroxyl free radical (OH) with hydrogen peroxide which is overexpressed in the tumor microenvironment, so as to promote the generation of cell membrane lipid peroxide; the molybdenum provided can perform oxidation-reduction reaction with the GSH which is over-expressed in the tumor to eliminate the GSH, inhibit GPX4 enzyme activity and prevent lipid peroxide from being eliminated. The efficient accumulation of lipid peroxides promotes the execution of iron death.
In the experimental process, the change of the valence state of the molybdenum element in the reduction process of molybdenum and glutathione leads to the degradation of the nano-sheet caused by the change of the chemical structure of the material, promotes the release of cobalt source and increases the generation of active oxygen. In particular, hexavalent molybdenum (in the polymetallic phosphomolybdate structure) is reduced to pentavalent, thereby being capable of further reacting with hydrogen peroxide to generate another singlet oxygen through a roxen mechanism 1 O 2 ) Active oxygen species, increases the source of active oxygen.
It is also described herein that the dual GSH consumption and active oxygen production of cobalt molybdate and phosphomolybdic acid exacerbates the lipid peroxidation consequences, allowing iron death to occur more effectively than with a single component.
The following also illustrates the preparation method of the non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material.
Mixing a molybdenum source, a cobalt source and an organic solvent. The molybdenum source is a polyoxometalate containing molybdenum at a concentration of 0.01-0.02mM, preferably 0.016mM, of phosphomolybdic acid. The cobalt source is cobalt salt at a concentration of 0.6-0.7mM, preferably 0.68mM cobalt acetate. The specific chemical composition of cobalt acetate and phosphomolybdic acid is advantageous for the formation of cobalt molybdate (CoMoO 4 ) Phosphomolybdic acid (H) 3 PMo 12 O 40 ) Two-dimensional nanoplatelet structures of composite nanoplatelets. Wherein, partial phosphomolybdic acid reacts with cobalt acetate to form cobalt molybdate, and other phosphomolybdic acids participate in the construction of the two-dimensional nano-sheet structure. The organic solvent is a mixture of acetone, ethanolamine and oleylamine. The organic solvent of the composition has the function of maintaining the directional growth of the two-dimensional nano-sheets. Preferably, the volume ratio of acetone, ethanolamine and oleylamine is 1:1:1.
transferring the mixed solution into a high-pressure reaction kettle for sealing and reacting. The reaction temperature is 160-200 ℃; the reaction time is 3 to 12 hours, preferably 6 to 12 hours. After the reaction, the reaction mixture is centrifugally washed to clean the organic solvent remained on the surface of the product. The product was washed with ethanol and cyclohexane, respectively. The number of washes may be 3-5 each.
The resulting product is mixed with a biocompatible polymer in an organic solvent. The effect is that the surface of the product is effectively wrapped by the biocompatible polymer, thus ensuring good biocompatibility and stability of the system. The organic solvent comprises at least one of dichloromethane, ethanol, chloroform and cyclohexane. For example, dichloromethane. The biocompatible polymer includes, but is not limited to, any of polyether F127, soybean phospholipid, distearoyl phosphatidylethanolamine-polyethylene glycol (DSPE-PEG), preferably polyether F127. The organic solvent was then removed by rotary evaporation. The rotary evaporation temperature can be 40-60 ℃, and the evaporation time can be 1-2h.
In some embodiments, the product after spin-steaming is dispersed in a physiological buffer solution. For example, the product is dispersed in a physiological buffer solution of sodium dihydrogen phosphate/disodium hydrogen phosphate at a pH of 7.4.
The non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material solves the technical problem of insufficient accumulation of lipid peroxide in tumor iron death treatment, and provides cobalt molybdate (CoMoO) with GSH consumption and active oxygen production capacity 4 ) Phosphomolybdic acid (H) 3 PMo 12 O 40 ) The non-iron-based composite material of the composite nano-sheet is used as an iron death inducer for relevant application of tumor treatment.
The present invention will be further illustrated by the following examples. It is also to be understood that the following examples are given solely for the purpose of illustration and are not to be construed as limitations upon the scope of the invention, since numerous insubstantial modifications and variations will now occur to those skilled in the art in light of the foregoing disclosure. The specific process parameters and the like described below are also merely examples of suitable ranges, i.e., one skilled in the art can make a suitable selection from the description herein and are not intended to be limited to the specific values described below.
Example 1
CoMoO 4 -H 3 PMo 12 O 40 Preparation of composite nanosheets: 0.016mM phosphomolybdic acid, 0.68mM cobalt acetate, 3mL acetone, 3mL oleylamine, and 3mL ethanolamine were mixed and stirred for 10 minutes. The mixed solution was transferred to a 25mL stainless steel reaction vessel lined with para-polyphenyl, and reacted at 180℃for 6 hours in a sealed manner. After the reaction was completed, the reaction product was centrifugally washed with ethanol and cyclohexane at 10000r/min for 3min. Collecting the product after centrifugal separation to obtain CoMoO 4 -H 3 PMo 12 O 40 Composite nanoplatelets (CPMNSs).
CPMS was dispersed in ethanol and dropped on a copper mesh to observe morphology by TEM, and it can be seen from FIGS. 1 and 2 that CPMS presents an irregular nanoplatelet morphology. The thickness of the composite nano sheet is 3-4nm.
Dispersing CPMS in ethanol, and dripping on a silicon wafer for element energy spectrum analysis. From fig. 3 it can be seen that Mo, P, co and O are uniformly distributed in cpms nanoplatelets.
CPMS was freeze-dried for XRD measurement. XRD analysis of FIG. 4 shows CoMoO 4 -H 3 PMo 12 O 40 Composite nano sheet with CoMoO 4 And H 3 PMo 12 O 40 The complex exists in the form of a solid.
Modification of CoMoO 4 -H 3 PMo 12 O 40 Composite nanosheets: 5mg of CoMoO 4 -H 3 PMo 12 O 40 Composite nanoplatelets) were mixed well with 50mg of polyether F127 and added to 10mL of dichloromethane and rotary evaporated under vacuum at 45 ℃ for 2h. After the rotary evaporation was completed, the resulting material was dispersed in a phosphate buffer solution (ph=7.4).
Characterization of cpms. Modification of biocompatible polymers (e.g., polyether F127) does not affect performance evaluation of cpms. Without specific explanation, performance measurements were performed in phosphate buffered saline at pH 6.5 (simulating tumor microenvironment).
CPMS (100. Mu.g/mL) was taken and reacted with 4mM hydrogen peroxide and 10mM sodium bicarbonate for 10 minutes, and the resulting OH was captured with 5, 5-dimethyl-1-pyrroline-N-oxide (DMPO) as a capture reagent and detected by ESR spectroscopy. A typical 1 can be observed in fig. 5 (a): 2:2: the presence of the signal peak of 1 indicates that OH is generated. And HCO with rich physiological condition is added 3 - After this, the radical signal is enhanced.
After 3h reaction of 1mM GSH with CPMS (50. Mu.g/mL), GSH content was measured with DTNB (5, 5' -dithiobis (2-nitrobenzoic acid) (0.5 mM). As can be seen in FIG. 5 (b), after reaction with CPMS, the UV characteristic absorbance of DTNB at 412nm decreased, indicating GSH depletion.
Reduced ReCPMNSs and H after 30min reaction with different concentrations of GSH 2 O 2 (10 mM) and HCO 3 - (25 mM) for 10min, and the presence of active oxygen was detected by the degradation of Methylene Blue (MB) (10. Mu.g/mL). As can be seen from FIG. 5 (c), when GSH concentration is 0.5mM or 1mM, the methylene blue degradation capacity of ReCPMS is stronger than that of CPMS, indicating that the addition of GSH accelerates the release of cobalt ions, leading to cobalt ions and H 2 O 2 Class of (2)The Fenton reaction is enhanced. Whereas when the GSH concentration is 2mM, the methylene blue degradation ability is reduced because the excessive unreacted GSH having the antioxidant ability eliminates the active oxygen generated.
After a set time point (10 min, 0.5h, 1h, 2h, 4h, 6h, 12h, 24h, 36h, 48h, 72 h) was reached by reacting 5mg CPMNSs with 15mL PBS (pH=6.5, 10 mM) and GSH (10 mM), 1mL of the reaction solution was taken. The resulting solution was digested with aqua regia and the released content of Co ions was detected by Inductively Coupled Plasma (ICP). As can be seen from FIG. 5 (d), the addition of GSH accelerates the release of Co ions to make them more effective with H 2 O 2 The reaction produces active oxygen.
CPMS (100. Mu.g/mL) was freeze-dried and XPS analyzed after redox reaction with GSH (1 mM). As can be seen from fig. 6 (a), after GSH reduction, a low valence Mo peak, represented by a low binding energy, appears, indicating that hexavalent molybdenum is reduced to pentavalent molybdenum.
The reduced ReCPMNSs (100. Mu.g/mL) were reacted with H 2 O 2 (10 mM) reaction 10min,2, 6-Tetramethylpiperidine (TEMP) as a capture reagent, and detection of singlet oxygen production using ESR spectroscopy. From fig. 6 (b), it can be seen that the reaction of recmnss with hydrogen peroxide produces a distinct 1 representing singlet oxygen: 1: triplet of 1. This is because phosphomolybdic acid and hydrogen peroxide are further converted to singlet oxygen by the formation of tetraoxide through the roxen mechanism.
Intracellular iron death assessment
4T1 cells were seeded in 96-well plates containing 5% CO by volume 2 Incubate in a 37℃incubator for 24h. Adherent cells were washed with PBS and then incubated with CPMS (150. Mu.g/mL) and iron death inhibitor-containing medium for 24h, and cell viability was measured using CCK-8 kit.
It can be seen from FIGS. 7 (a) and (b) that CPMS induced cell death was significantly inhibited after co-incubation with either Fer-1 or Lip-1 iron death inhibitors.
4T1 cells were seeded in 6-well plates for 24 hours and then incubated with CPMNSs (0, 50. Mu.g/mL, 100. Mu.g/mL, 150. Mu.g/mL) at different concentrations for 12 hours or CPMNSs (150. Mu.g/mL) at the same concentration for different times (0, 3 hours, 6 hours, 9 hours) after which 4T1 cells were lysed with lysate and then assayed for GSH content and GPX4 activity with a total glutathione assay kit (Shanghai Biyun Biotechnology Co., ltd.).
From FIGS. 7 (c) and (d), GSH content decreases with increasing concentration or with increasing incubation time. As can be seen from FIGS. 8 (a) - (b), GPX4 activity decreased with increasing concentration or with increasing incubation time. The content of Malondialdehyde (MDA) which is a lipid peroxidation product under the same cell processing conditions was detected by a lipid oxidation (MDA) detection kit (shanghai bi yunshan biotechnology limited). From (c) and (d) in 8, it can be seen that the cellular MDA content increases with increasing CPMNSs concentration and prolonged incubation time, indicating an increase in lipid peroxidation.
Evaluation of therapeutic efficacy of CPMNSs in iron death in vivo
4T1 cells were plated subcutaneously in Balb/c nude mice when tumor volume reached about 100mm 3 After that, the mice were randomly divided into four groups. The first group does not perform any processing (Control); the second group was injected with 10mg/kg CPMNSs (CPMNSs i.v.) at day 0 and day 7 tail vein (i.v.); the third group injected 10mg/kg cpms (cpms i.t.) in tail tumors on days 0 and 7 (i.t.); the fourth group was intraperitoneally injected with 20mg/kg iron death inhibitor deferoxamine mesylate (DFOM) (CPMNSs i.t. +dfom) on days 0, 4, 8, 12 based on the third group. Mice were fed and analyzed for 16 days.
As can be seen from fig. 9, cpms treated mice inhibited tumor to a different extent than control group, and the inhibition effect of cpms on tumor growth was significantly reduced after the iron death inhibitor DFOM was introduced.
To observe the in vivo peroxidation of active oxygen and lipid by CPMS, mice were injected intratumorally or by tail vein injection of CPMS (10 mg/kg) for 12h, intratumorally with 100. Mu.L DCFH-DA dye (active oxygen indicator reagent, 10. Mu.M) or 100. Mu. L C11-BODIPY 581/591 The dye (lipid peroxidation indicator reagent, 10. Mu.M) was incubated for 30min, and then the tumors were harvested for sectioning and fluorescence observation using a confocal microscope. The activity can be seen in FIG. 10Oxygen and lipid peroxidation fluorescence increases with the addition of cpms, indicating that cpms can cause the production of active oxygen within tumors and lipid peroxidation.
At the end of treatment, tumors were harvested for GPX4 immunohistochemistry, hematoxylin-eosin staining (H & E), transferase mediated deoxyguanosine triphosphate-biotin nick end labeling (TUNEL) and Ki-67 section analysis. GPX4 expression was significantly reduced in CPMNSs treated groups compared to control groups (FIG. 11) and tumor tissue was damaged, apoptosis occurred, and cell proliferation capacity was reduced (FIG. 12).
To further evaluate the effect of CPMS tumor treatment, mice survival curves were recorded, mice tumor volumes were recorded every two days, when tumor volumes reached 1500mm 3 When it is regarded as dead. As can be seen in fig. 13, cpms showed greater survival in mice treated than untreated controls and decreased survival in mice after iron death inhibition with DFOM, demonstrating that cpms can act as an effective iron death inducer.

Claims (12)

1. The non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material is characterized by comprising cobalt molybdate-phosphomolybdic acid composite nano-sheets and biocompatible polymers with surfaces modified by the composite nano-sheets; the biocompatible polymer comprises at least one of polyether F127, soybean phospholipid and distearoyl phosphatidylethanolamine-polyethylene glycol; the mass ratio of cobalt molybdate to phosphomolybdic acid is 1:2-1:4, a step of; the mass ratio of the cobalt molybdate-phosphomolybdic acid composite nano-sheet to the biocompatible polymer is 1:5-1:15.
2. the non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite of claim 1, wherein the biocompatible polymer is polyether F127.
3. The non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite of claim 1, wherein the cobalt molybdate-phosphomolybdic acid composite nanoplatelets are uniformly dispersed and interwoven composites in nanoplatelets.
4. The non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite of claim 1, wherein the non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite has both glutathione consuming and reactive oxygen species generating capabilities.
5. The non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite of claim 4, wherein the non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite undergoes a redox reaction with the glutathione overexpressed in the tumor to eliminate glutathione and increase reactive oxygen species production during its redox reaction with glutathione.
6. The non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite material of claim 4, wherein the product of the redox reaction of the non-iron-based glutathione consuming synergistic reactive oxygen species enhanced composite material with glutathione is further reacted with hydrogen peroxide to produce singlet oxygen via the ro mechanism 1 O 2 Reactive oxygen species.
7. The method for producing a non-iron-based glutathione consumption synergistic reactive oxygen species enhanced composite according to any one of claims 1 to 6, characterized in that the method for producing comprises: mixing a molybdenum source, a cobalt source and an organic solvent to obtain a mixed solution; transferring the mixed solution into an autoclave for reaction, centrifugally washing after the reaction is finished, and collecting a reaction product; and (3) dispersing and mixing the reaction product and a biocompatible polymer to obtain the non-iron-based glutathione consumption synergistic active oxygen species reinforced composite material.
8. The method of claim 7, wherein the molybdenum source is a polyoxometalate comprising molybdenum, and the concentration of the molybdenum source is 0.01-0.02mM; the cobalt source is cobalt salt, and the concentration of the cobalt source is 0.06-0.07mM; the organic solvent is a mixture of acetone, ethanolamine and oleylamine.
9. The method according to claim 8, wherein the volume ratio of acetone, ethanolamine and oleylamine is (1-4): (1-4): (1-4).
10. The process according to claim 7, wherein the reaction temperature is 160 to 200 o C, the reaction time is 3-12h.
11. The method of claim 7, wherein the reaction product is dispersed and mixed with the biocompatible polymer in an organic solvent and then the organic solvent is removed; the organic solvent comprises at least one of dichloromethane, ethanol, chloroform and cyclohexane.
12. The biological use of the non-iron-based glutathione consuming synergistic active oxygen species enhanced composite of any of claims 1 to 6 in the manufacture of a medicament for the treatment of cancer.
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Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104511290A (en) * 2014-12-31 2015-04-15 中国地质大学（武汉） Preparation method of visible-light-driven photocatalyst nano spherical MoSe2 material
CN108133829A (en) * 2017-12-21 2018-06-08 哈尔滨师范大学 Co(OH)2@CoMoO4The preparation method of composite nano plate
WO2019079841A1 (en) * 2017-10-26 2019-05-02 University Of South Australia Nanoparticle cancer therapy
CN110585238A (en) * 2018-06-11 2019-12-20 沈阳药科大学 Antitumor drug composition with synergistic effect and application thereof
CN111017996A (en) * 2019-09-25 2020-04-17 青岛大学 Synthesis of MoO with double simulated enzyme activity3-XMethod for producing antimicrobial material

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2017100162A1 (en) * 2015-12-07 2017-06-15 General Oncology, Inc. Combination for the effective treatment of metastatic cancer in patients

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104511290A (en) * 2014-12-31 2015-04-15 中国地质大学（武汉） Preparation method of visible-light-driven photocatalyst nano spherical MoSe2 material
WO2019079841A1 (en) * 2017-10-26 2019-05-02 University Of South Australia Nanoparticle cancer therapy
CN108133829A (en) * 2017-12-21 2018-06-08 哈尔滨师范大学 Co(OH)2@CoMoO4The preparation method of composite nano plate
CN110585238A (en) * 2018-06-11 2019-12-20 沈阳药科大学 Antitumor drug composition with synergistic effect and application thereof
CN111017996A (en) * 2019-09-25 2020-04-17 青岛大学 Synthesis of MoO with double simulated enzyme activity3-XMethod for producing antimicrobial material

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
Maqsood A. Siddiqui et al.Molybdenum nanoparticles-induced cytotoxicity, oxidative stress,G2/Marrest, and DNA damage in mouse skin fibroblast cells (L929).《Colloids and Surfaces B: Biointerfaces》.2014,第125卷(第2015期),第73-81页. *
Recent advances in MoS 2 -based photothermal therapy for cancer and infectious disease treatment;Jinping Shi et al;《Journal of Materials Chemistry B》;第8卷;第5793--5807页 *
氯化钴通过活性氧诱导PC12细胞凋亡并伴有AP-1的活化;邹卫国等;《中国生物化学与分子生物学会第八届会员代表大会暨全国学术会议论文摘要集》;第421-422页 *

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