CN109125742A - A kind of preparation method of novel oxidized Fe nanometer particles and its application in cancer target diagnosis and treatment - Google Patents

A kind of preparation method of novel oxidized Fe nanometer particles and its application in cancer target diagnosis and treatment Download PDF

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CN109125742A
CN109125742A CN201811059714.4A CN201811059714A CN109125742A CN 109125742 A CN109125742 A CN 109125742A CN 201811059714 A CN201811059714 A CN 201811059714A CN 109125742 A CN109125742 A CN 109125742A
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nanometer particles
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CN109125742B (en
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熊非
单秀红
顾宁
钱炜
柳迎昭
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Zhenjiang First Peoples Hospital
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Abstract

The present invention provides a kind of preparation methods of novel oxidized Fe nanometer particles: polylactic acid-polyglycol and dimethylformamide are placed in water；1-3- dimethylamino-propyl -3- ethyl-carbodiimide hydrochloride and n-hydroxysuccinimide are separately added into above-mentioned reaction system；Aminoglucose hydrochloride is added into it again；Then dialysis purification filtering is carried out；After oleic acid is conjugated taxol, with PLA-PEG-CO-NHC6H11O5With equimolar than mixing；Take Fe3O4Ethyl alcohol is added in@OA NPs, carries out adsorbing separation to the system with magnet, then tetrahydrofuran is added into it and carries out sonic oscillation；OA-PTX-OA-PLA-PEG-CO-NHC is added into it again6H11O5Carry out sonic oscillation；Sonic oscillation is carried out after obtained reaction system is added drop-wise in water again, and dialysis purification filters again.The invention also includes the targeting diagnosis and treatment that nanoparticle made from any of the above-described kind of preparation method is applied to tumour.Novel oxidized Fe nanometer particles provided by the invention have both oncotherapy and tracer dual function simultaneously；High sensitivity and high specificity, penetrability are strong.

Description

The preparation method of a kind of novel oxidized Fe nanometer particles and its in cancer target diagnosis and treatment Application
Technical field
The invention belongs to medical field of antineoplastic medicaments, in particular to a kind of preparation method of novel oxidized Fe nanometer particles And its application in cancer target diagnosis and treatment.
Background technique
In in the past few decades, various anti-tumor drugs are widely used in the treatment of tumour.But anti-tumor drug cannot Tumour cell in accurate target tumor pathogenic site, to reduce its anticancer effect and lead to serious toxicity and secondary work Generation.By anticancer drug be covalently attached to can the ligand of selectively targeted combination tumour cell will will increase controlling for tumour Therapeutic effect.The real-time of oncotherapy effect all has weight for prognosis of adjustment therapeutic scheme and tumour etc. in body evaluation simultaneously The meaning wanted.Prior art such as fluorescence imaging etc. evaluates chemotherapeutics, but the safety of its fluorescent reagent itself is deposited In problem, certain toxic side effect may be generated to body.And technology needs to connect chemotherapeutics and preparation mostly It connects, and directly develops the diagnosis and treatment integration probe that can have both treatment and imaging function, the technology without extra drug load is also It is rarely reported.
Glucose is for maintaining human normal function to play an important role, as a kind of essential energy source, The glycolysis speed of malignant tumour and the demand to glucose are significantly higher than normal tissue.Glucose transporter-4 (GLUT1) is Adjust the main carriers of glucose uptake.Therefore, the overexpression that GLUT1 is caused to the increase in demand of glucose, can be used as The targeting ligand of drug.Pass through radiolabeled tracer such as [18F] fluoro- 2- deoxidation-D glucose (18F-FDG), [14C] It is living that 2-deoxy-D-glucose and [14C] or [3H] 3-O- methyl-D-glucose can monitor the GLUT1 in mammalian cell Property.18F-FDG is the increased radioactive tracer of positron e mission computed tomography (PET) the most common glucose metabolism Agent, for detecting activity and the position of tumour.Although this method has the characteristics that sensitive and quantitative, 18F- may be implemented Half-life period FDG shorter because of its isotope is not suitable for clinical high-throughput testing requirement.In addition, radioactive tracer is to body Often there is radiotoxicity in organ.
Summary of the invention
Technical problem: in order to solve the defects of prior art, the present invention provides a kind of novel oxidized Fe nanometer particles Preparation method and its application in cancer target diagnosis and treatment.
A kind of technical solution: preparation method of novel oxidized Fe nanometer particles provided by the invention, comprising the following steps:
Step 1: taking 2.53-253mg polylactic acid-polyglycol (PLA-PEG), place it in the dimethyl methyl of 0.2-20mL In amide (DMF), then said mixture is placed in 0.6-60mL water and stands 3h；
Step 2: by the 1-3- dimethylamino-propyl -3- ethyl-carbodiimide hydrochloride (EDCHCl) of 55-550mg and After the n-hydroxysuccinimide (NHS) of 37-370mg is added separately to the reaction system of above-mentioned steps 1,20min is stirred；
Step 3: after the aminoglucose hydrochloride (DGH) of 35-350mg is added to the reaction system of step 2, stirring 12h；
Step 4: reaction system obtained in step 3 is subjected to dialysis purification and crosses 0.22 μm of filter membrane progress bacteriological filtration afterwards for 24 hours, Obtain PLA-PEG-CO-NHC6H11O5；
Step 5: after oleic acid (OA) is conjugated taxol (PTX), with PLA-PEG-CO-NHC6H11O5With equimolar than mixing Obtain OA-PTX-OA-PLA-PEG-CO-NHC6H11O5；
Step 6: taking 2-300mg Fe3O40.2-30mL ethyl alcohol is added in@OA NPs, carries out absorption point to the system with magnet From rear, 0.4-40mL tetrahydrofuran (THF) is added into it and supersonic oscillations are carried out to reaction system；
Step 7: continuing that 2-200mg OA-PTX-OA-PLA-PEG-CO- is added into the reaction system of step 6 NHC6H11O5, gained mixture is then subjected to supersonic oscillations 5min；
Step 8: after carrying out supersonic oscillations 3min after gained mixture is added drop-wise in water dialysis purification for 24 hours, last mistake 0.22 μm of filter membrane bacteriological filtration is to obtain final products: novel oxidized Fe nanometer particles.
As a kind of prioritization scheme: in step 8 power of supersonic oscillations be 750W, pulse open and close the time be 2 seconds.
As advanced optimizing scheme: the environment temperature of all reaction process is between 20-30 DEG C.
Technical solution of the present invention further includes the target that nanoparticle made from any of the above-described kind of preparation method is applied to tumour To diagnosis and treatment.
The utility model has the advantages that novel oxidized Fe nanometer particles provided by the invention have both oncotherapy and the dual function of tracer simultaneously Energy；The oleic acid layer of particle surface can be conjugated fat-soluble anti-tumor drug, further increase its antitumous effect；Used magnetic Resonance image-forming tracer technique, which has, exempts ionising radiation, improves the advantage of sensitivity and specificity, while having multiple plane imaging The features such as ability, penetration into tissue is strong, and structure imaging may be implemented；Prepared ferric oxide nano particles may be implemented swollen in vivo The efficient treatment of tumor and synchronous imaging.
Detailed description of the invention
Fig. 1 is that the preparation method of the novel oxidized Fe nanometer particles of the present invention chemically reacts schematic diagram；
Fig. 2 is tumour cell after tumour cell experimental comparison group prussian blue staining of the present invention detects incubated cell 30 minutes Absorb the schematic diagram of nanoparticle；
Fig. 3 is tumour cell after tumour cell experiments experiment group prussian blue staining of the present invention detects incubated cell 30 minutes Absorb the schematic diagram of nanoparticle；
Fig. 4 is that tumor-bearing mice experimental tumor histotomy prussian blue staining of the present invention shows that the absorption of control group nanoparticle is shown It is intended to；
Fig. 5 is that tumor-bearing mice experimental tumor histotomy prussian blue staining of the present invention shows that the absorption of experimental group nanoparticle is shown It is intended to；
Fig. 6 be the present invention targeting mouse knurl magnetic resonance test experience in magnetic resonance magnetic susceptibility weighting sequence mIP picture and SWI as shown in experimental group intravenous injection before, 6h and for 24 hours after magnetic resonance imaging (1.5T MR) nanoparticle aggregation schematic diagram；
Fig. 7 be the present invention targeting mouse knurl magnetic resonance test experience in magnetic resonance magnetic susceptibility weighting sequence mIP picture and SWI as shown in control group intravenous injection before, 6h and for 24 hours after magnetic resonance imaging (1.5T MR) nanoparticle aggregation schematic diagram；
Fig. 8 is that targeted therapy curative effect magnetic resonance of the present invention monitoring experiment injection in the 1st, 3,6 day compares agent knurl before the injection Volume schematic diagram；
Fig. 9 is that targeted therapy curative effect magnetic resonance of the present invention monitoring experiment injection in the 1st, 3,6 day compares agent knurl after injection 15d volume schematic diagram；
Figure 10 is that targeted therapy curative effect magnetic resonance of the present invention monitoring experiment injection in the 1st, 3,6 day tests agent knurl before the injection Volume schematic diagram；
Figure 11 is that targeted therapy curative effect magnetic resonance of the present invention monitoring experiment injection in the 1st, 3,6 day tests agent knurl after injection 15d volume schematic diagram.
Specific embodiment
In the following with reference to the drawings and specific embodiments, the present invention is furture elucidated, it should be understood that these embodiments are merely to illustrate It the present invention rather than limits the scope of the invention, after the present invention has been read, those skilled in the art are to of the invention each The modification of kind equivalent form falls within the application range as defined in the appended claims.
Embodiment
A kind of preparation method of novel oxidized Fe nanometer particles provided by the invention, follows the steps below specific reality It tests:
Step 1: taking 25.3mg polylactic acid-polyglycol (PLA-PEG), place it in the dimethylformamide (DMF) of 2mL In, then said mixture is placed in 6mL water and stands 3h；
Step 2: by the 1-3- dimethylamino-propyl -3- ethyl-carbodiimide hydrochloride (EDCHCl) and 370mg of 550mg N-hydroxysuccinimide (NHS) be added separately to the reaction system of above-mentioned steps 1 after, stir 20min；
Step 3: after the aminoglucose hydrochloride (DGH) of 350mg is added to the reaction system of step 2, stirring 12h；
Step 4: reaction system obtained in step 3 is subjected to dialysis purification and crosses 0.22 μm of filter membrane progress bacteriological filtration afterwards for 24 hours, Obtain PLA-PEG-CO-NHC6H11O5；
Step 5: after oleic acid (OA) is conjugated taxol (PTX), with PLA-PEG-CO-NHC6H11O5With equimolar than mixing Obtain OA-PTX-OA-PLA-PEG-CO-NHC6H11O5；
Step 6: taking 20mg Fe3O42mL ethyl alcohol, after carrying out adsorbing separation to the system with magnet, Xiang Qi is added in@OA NPs Interior addition 4mL tetrahydrofuran (THF) simultaneously carries out supersonic oscillations to reaction system；
Step 7: continuing that 20mg OA-PTX-OA-PLA-PEG-CO-NHC is added into reaction system6H11O5, then by institute It obtains mixture and carries out supersonic oscillations 5min；
Step 8: by gained mixture be added drop-wise in water after carry out supersonic oscillations 3min after dialysis purification for 24 hours, ultrasonic wave The power of oscillation is 750W, and the pulse opening and closing time is 2 seconds, and it is final to obtain finally to cross 0.22 μm of filter membrane bacteriological filtration Product: novel oxidized Fe nanometer particles.
The environment temperature of all of above reaction process is room temperature, between 20-30 DEG C.
The preparation involved in the present invention for obtaining a kind of novel oxidized Fe nanometer particles, during the preparation process, by small molecule amino Glucose modified is directly used in oncotherapy on ferric oxide nano particles surface, using the magnetic resonance imaging of noninvasive on-radiation Method tracking evaluation therapeutic effect.The ferric oxide nano particles of connection Glucosamine in surface prepared by the present invention have both swollen simultaneously The dual function of tumor treatment and tracer, the present invention are conjugated fat-soluble chemotherapy in the rouge layer of iron oxide nano-granule surfacing simultaneously Agent makes it further increase antitumor action, and used magnetic resonance imaging tracer technique has a Non-ionizing radiation, more planes at The features such as picture ability, higher sensitivity and specificity, and penetration into tissue are strong and structure imaging may be implemented.Based on this, institute Efficient treatment and the synchronous imaging of tumour may be implemented in the ferric oxide nano particles of preparation.
Polylactic acid (PLA) is connect with polyethylene glycol (PEG) as shown in Figure 1:, and the side PEG and aminoguanidine hydrochloride deoxyglucose contract It closes, polylactic acid side connects the oleic acid for being conjugated taxol (PTX), finally marks molecule OA-PTX-OA-PLA-PEG-CO-DG To oleic acid oxidation iron (Fe3O4@OA) up to Fe3O4@OA-PTX-PLA-PEG-DG cancer target diagnosis and treatment nanoparticle.Wherein aoxidize Iron (Fe3O4) it is magnetic resonance imaging contrast agent, magnetic resonance can in real time show the site specific of vivo oxidation iron aggregation；DG is More the cancer target ligand of wide spectrum, the glucose transporter of targets neoplastic cells film, while DG enters energy after tumour cell Inhibit cell metabolism, and there is antitumor action；The PTX that nanoparticle enters after cell the release that is decomposed simultaneously further enhances Antitumor action.
For the actual efficacy for verifying drug, medicament is subjected to mouse experiment, targets diagnosis and treatment agent Fe3O4@OA-PTX-PLA- PEG-DG is in application, medicament is sealed after being sterilized in the form of freeze-dried powder；Before being injected in Mice Body in 30 minutes, use Sterile water pharmaceutical dissolution simultaneously water bath sonicator 3-20 seconds, medicine of the concentration of ordinary dissolution between 0.1-1.0mgFe/ml was configured as needed Object, mouse administration route can be intravenous injection or intraperitoneal injection.The injection of experimental group is label targeting ligand DG (Fe3O4@OA-PTX-PLA-PEG-DG)；Control group is set simultaneously, and the injection of control group is not mark targeting ligand DG (Fe3O4@OA-PTX-PLA-PEG)。
Experiment 1: tumour cell experiment: tumour cell absorbs nanoparticle after prussian blue staining detects incubated cell 30 minutes Difference, blue particle is iron-containing nanoparticle: as shown in Fig. 2, control group has no that obvious nanoparticle absorbs, as shown in figure 3, real The visible obvious nanoparticle of group is tested to absorb.
Experiment 2: tumor-bearing mice experiment: tumor tissue section's prussian blue staining shows that experimental group and control group nanoparticle are inhaled Astigmat is different.As shown in figure 4, nanoparticle is had no in control group tumor tissues, as shown in figure 5, tumour is thin in experimental group tumor tissues Then nanoparticle absorbs obviously born of the same parents.
Experiment 3: magnetic resonance detection (targeting diagnosis) experiment of targeting mouse knurl: magnetic resonance magnetic susceptibility weighting sequence mIP Aggregation as showing this nanoparticle is more sensitive, and the left side is mIP picture, and the right is SWI picture；Experimental group: before intravenous injection, after injection 6h and for 24 hours rear magnetic resonance imaging (1.5T MR) picture are believed for 24 hours after injection as shown in fig. 6, signal decline in 6 hours after injection Number further decline；Control group: before intravenous injection, injection after 6h and (1.5T MR) picture of magnetic resonance imaging for 24 hours as shown in fig. 7, 6 hours signals increase after injection, and 24 hours signals still increase after injection.
Experiment 4: agent and experiment agent targeted therapy curative effect magnetic resonance monitoring experiment: are compareed respectively at injection in the 1st, 3,6 day；It is right According to group: as shown in Figure 8 and Figure 9, knurl is gradually increased, 15d volume balanced growth 60.5%；Experimental group: such as Figure 10 and Figure 11 institute Show, hereafter 1-3 days volumes of knurl are gradually reduced without significant change, volume averagely reduces about 36.2% after injecting 15d.
The manufacturer and model of drug used in the embodiment of the present invention and reagent are listed in the table below respectively:

Claims (4)

1. a kind of preparation method of novel oxidized Fe nanometer particles, it is characterised in that: the following steps are included:
Step 1: taking 2.53-253mg polylactic acid-polyglycol, place it in the dimethylformamide of 0.2-20mL, then will Said mixture is placed in 0.6-60mL water and stands 3h；
Step 2: by the 1-3- dimethylamino-propyl -3- ethyl-carbodiimide hydrochloride of 55-550mg and the N- hydroxyl of 37-370mg After succinimide is added separately to the reaction system of above-mentioned steps 1,20min is stirred；
Step 3: after the aminoglucose hydrochloride of 35-350mg is added to the reaction system of step 2, stirring 12h；
Step 4: reaction system obtained in step 3 being subjected to dialysis purification and crosses 0.22 μm of filter membrane progress bacteriological filtration afterwards for 24 hours, is obtained PLA-PEG-CO-NHC6H11O5；
Step 5: after oleic acid is conjugated taxol, with PLA-PEG-CO-NHC6H11O5OA-PTX-OA- is mixed to obtain with equimolar ratio PLA-PEG-CO-NHC6H11O5；
Step 6: taking 2-300mg Fe3O40.2-30mL ethyl alcohol is added in@OA NPs, after carrying out adsorbing separation to the system with magnet, 0.4-40mL tetrahydrofuran is added into it and supersonic oscillations are carried out to reaction system；
Step 7: continuing that 2-200mg OA-PTX-OA-PLA-PEG-CO-NHC is added into the reaction system of step 66H11O5, so Gained mixture is subjected to supersonic oscillations 5min afterwards；
Step 8: dialysis purification for 24 hours, finally crosses 0.22 μ after carrying out supersonic oscillations 3min after gained mixture is added drop-wise in water The filter membrane bacteriological filtration of m is to obtain final products: novel oxidized Fe nanometer particles.
2. a kind of preparation method of novel oxidized Fe nanometer particles according to claim 1, it is characterised in that: the step The power of supersonic oscillations is 750W in 8, and the pulse opening and closing time is 2 seconds.
3. a kind of preparation method of novel oxidized Fe nanometer particles according to claim 1, it is characterised in that: all reactions The environment temperature of process is between 20-30 DEG C.
4. nanoparticle made from method described in -3 any claims according to claim 1, it is characterised in that: applied In the targeting diagnosis and treatment of tumour.
CN201811059714.4A 2018-09-12 2018-09-12 Preparation method of iron oxide nanoparticles and application of iron oxide nanoparticles in tumor targeted diagnosis and treatment Expired - Fee Related CN109125742B (en)

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114099719A (en) * 2021-11-30 2022-03-01 东华大学 Dendritic macromolecule coated nano gold particle composite material and preparation and application thereof

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Title
FEI XIONG等: ""A Functional Iron Oxide Nanoparticles Modified with PLA-PEG-DG as Tumor-Targeted MRI Contrast Agent"", 《PHARM RES》 *
JILAI TIAN等: ""Paclitaxel loaded magnetic nanoparticles: Synthesis, characterization and application in targeting"", 《JOURNAL OF PHARMACEUTICAL SCIENCES》 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114099719A (en) * 2021-11-30 2022-03-01 东华大学 Dendritic macromolecule coated nano gold particle composite material and preparation and application thereof

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