CN113082226B - Preparation method of drug-loaded nanocage and application of drug-loaded nanocage in targeting circulating tumor cells to release triptolide - Google Patents
Preparation method of drug-loaded nanocage and application of drug-loaded nanocage in targeting circulating tumor cells to release triptolide Download PDFInfo
- Publication number
- CN113082226B CN113082226B CN202110459985.4A CN202110459985A CN113082226B CN 113082226 B CN113082226 B CN 113082226B CN 202110459985 A CN202110459985 A CN 202110459985A CN 113082226 B CN113082226 B CN 113082226B
- Authority
- CN
- China
- Prior art keywords
- nanocage
- drug
- loaded
- sequence
- triptolide
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 239000003814 drug Substances 0.000 title claims abstract description 62
- 239000002091 nanocage Substances 0.000 title claims abstract description 62
- 229940079593 drug Drugs 0.000 title claims abstract description 31
- DFBIRQPKNDILPW-CIVMWXNOSA-N Triptolide Chemical compound O=C1OCC([C@@H]2C3)=C1CC[C@]2(C)[C@]12O[C@H]1[C@@H]1O[C@]1(C(C)C)[C@@H](O)[C@]21[C@H]3O1 DFBIRQPKNDILPW-CIVMWXNOSA-N 0.000 title claims abstract description 20
- YKUJZZHGTWVWHA-UHFFFAOYSA-N triptolide Natural products COC12CC3OC3(C(C)C)C(O)C14OC4CC5C6=C(CCC25C)C(=O)OC6 YKUJZZHGTWVWHA-UHFFFAOYSA-N 0.000 title claims abstract description 20
- 238000002360 preparation method Methods 0.000 title claims abstract description 12
- 208000005443 Circulating Neoplastic Cells Diseases 0.000 title abstract description 38
- 230000008685 targeting Effects 0.000 title abstract description 9
- XUIMIQQOPSSXEZ-UHFFFAOYSA-N Silicon Chemical compound [Si] XUIMIQQOPSSXEZ-UHFFFAOYSA-N 0.000 claims abstract description 15
- 229910052710 silicon Inorganic materials 0.000 claims abstract description 15
- 239000010703 silicon Substances 0.000 claims abstract description 15
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 claims abstract description 11
- 108091028043 Nucleic acid sequence Proteins 0.000 claims abstract description 10
- 230000001093 anti-cancer Effects 0.000 claims abstract description 7
- 239000007788 liquid Substances 0.000 claims abstract description 5
- 239000006185 dispersion Substances 0.000 claims abstract description 4
- 238000003756 stirring Methods 0.000 claims abstract description 4
- 238000004140 cleaning Methods 0.000 claims abstract description 3
- 238000005538 encapsulation Methods 0.000 claims abstract description 3
- 238000006243 chemical reaction Methods 0.000 claims description 8
- 238000000034 method Methods 0.000 claims description 8
- 238000009396 hybridization Methods 0.000 claims description 4
- 238000001556 precipitation Methods 0.000 claims description 3
- 230000001376 precipitating effect Effects 0.000 abstract 1
- RWSXRVCMGQZWBV-WDSKDSINSA-N glutathione Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@@H](CS)C(=O)NCC(O)=O RWSXRVCMGQZWBV-WDSKDSINSA-N 0.000 description 23
- 238000003380 quartz crystal microbalance Methods 0.000 description 23
- 210000004027 cell Anatomy 0.000 description 19
- 108010024636 Glutathione Proteins 0.000 description 11
- 206010028980 Neoplasm Diseases 0.000 description 8
- 238000001514 detection method Methods 0.000 description 8
- 108020004414 DNA Proteins 0.000 description 7
- 229960003180 glutathione Drugs 0.000 description 7
- 230000035945 sensitivity Effects 0.000 description 6
- 201000010099 disease Diseases 0.000 description 4
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 4
- 108091023037 Aptamer Proteins 0.000 description 3
- 108010053070 Glutathione Disulfide Proteins 0.000 description 3
- 230000009471 action Effects 0.000 description 3
- 239000002246 antineoplastic agent Substances 0.000 description 3
- 229940041181 antineoplastic drug Drugs 0.000 description 3
- YPZRWBKMTBYPTK-BJDJZHNGSA-N glutathione disulfide Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@H](C(=O)NCC(O)=O)CSSC[C@@H](C(=O)NCC(O)=O)NC(=O)CC[C@H](N)C(O)=O YPZRWBKMTBYPTK-BJDJZHNGSA-N 0.000 description 3
- 238000003384 imaging method Methods 0.000 description 3
- 230000003834 intracellular effect Effects 0.000 description 3
- 230000036542 oxidative stress Effects 0.000 description 3
- 239000000523 sample Substances 0.000 description 3
- -1 sulfhydryl compound Chemical class 0.000 description 3
- 150000003573 thiols Chemical group 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 238000001727 in vivo Methods 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 125000003396 thiol group Chemical group [H]S* 0.000 description 2
- JEOQACOXAOEPLX-WCCKRBBISA-N (2s)-2-amino-5-(diaminomethylideneamino)pentanoic acid;1,3-thiazolidine-4-carboxylic acid Chemical compound OC(=O)C1CSCN1.OC(=O)[C@@H](N)CCCN=C(N)N JEOQACOXAOEPLX-WCCKRBBISA-N 0.000 description 1
- WYTZZXDRDKSJID-UHFFFAOYSA-N (3-aminopropyl)triethoxysilane Chemical compound CCO[Si](OCC)(OCC)CCCN WYTZZXDRDKSJID-UHFFFAOYSA-N 0.000 description 1
- BWGNESOTFCXPMA-UHFFFAOYSA-N Dihydrogen disulfide Chemical compound SS BWGNESOTFCXPMA-UHFFFAOYSA-N 0.000 description 1
- 208000035240 Disease Resistance Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- WHUUTDBJXJRKMK-UHFFFAOYSA-N Glutamic acid Natural products OC(=O)C(N)CCC(O)=O WHUUTDBJXJRKMK-UHFFFAOYSA-N 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- XUJNEKJLAYXESH-REOHCLBHSA-N L-Cysteine Chemical compound SC[C@H](N)C(O)=O XUJNEKJLAYXESH-REOHCLBHSA-N 0.000 description 1
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 1
- 208000012902 Nervous system disease Diseases 0.000 description 1
- 208000025966 Neurological disease Diseases 0.000 description 1
- 101710147059 Nicking endonuclease Proteins 0.000 description 1
- 208000006011 Stroke Diseases 0.000 description 1
- BOTDANWDWHJENH-UHFFFAOYSA-N Tetraethyl orthosilicate Chemical compound CCO[Si](OCC)(OCC)OCC BOTDANWDWHJENH-UHFFFAOYSA-N 0.000 description 1
- GSEJCLTVZPLZKY-UHFFFAOYSA-N Triethanolamine Chemical compound OCCN(CCO)CCO GSEJCLTVZPLZKY-UHFFFAOYSA-N 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000003963 antioxidant agent Substances 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 230000019522 cellular metabolic process Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 1
- 235000018417 cysteine Nutrition 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000001212 derivatisation Methods 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 230000001627 detrimental effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000012377 drug delivery Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000005518 electrochemistry Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000001917 fluorescence detection Methods 0.000 description 1
- 238000000799 fluorescence microscopy Methods 0.000 description 1
- 235000013922 glutamic acid Nutrition 0.000 description 1
- 239000004220 glutamic acid Substances 0.000 description 1
- 208000019622 heart disease Diseases 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 238000012417 linear regression Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000001000 micrograph Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000002539 nanocarrier Substances 0.000 description 1
- 239000002086 nanomaterial Substances 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 235000018102 proteins Nutrition 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 239000013074 reference sample Substances 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000002798 spectrophotometry method Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 229910021642 ultra pure water Inorganic materials 0.000 description 1
- 239000012498 ultrapure water Substances 0.000 description 1
- 230000000007 visual effect Effects 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
- A61K47/6949—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit inclusion complexes, e.g. clathrates, cavitates or fullerenes
-
- 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
- A61K31/56—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
- A61K31/58—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids containing heterocyclic rings, e.g. danazol, stanozolol, pancuronium or digitogenin
- A61K31/585—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids containing heterocyclic rings, e.g. danazol, stanozolol, pancuronium or digitogenin containing lactone rings, e.g. oxandrolone, bufalin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
- A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
- G01N33/5008—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
- G01N33/5011—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics for testing antineoplastic activity
Abstract
The invention discloses a preparation method of a medicine carrying nano cage with a mesoporous structure, which comprises the following steps: the preparation method comprises the steps of preparing an amino mesoporous silicon nanocage, continuously stirring a functionalized DNA sequence, the amino mesoporous silicon nanocage and the triptolide in a dispersion liquid, precipitating, centrifugally cleaning for two times, and realizing the embedding of the anti-cancer triptolide in the mesoporous encapsulation of the amino mesoporous silicon nanocage, so that the drug-loaded nanocage can be obtained, and the application of the drug-loaded nanocage for targeting circulating tumor cells to release the triptolide is provided. The invention belongs to the technical field of medicine, and particularly relates to a preparation method of a medicine-carrying nanocage and application of the medicine-carrying nanocage in targeting circulating tumor cells to release triptolide.
Description
Technical Field
The invention belongs to the technical field of medicine, and particularly relates to a preparation method of a medicine-carrying nanocage and application of the medicine-carrying nanocage in targeting circulating tumor cells to release triptolide.
Background
Reduced Glutathione (GSH) is a tripeptide containing gamma-amide bonds and sulfhydryl groups, which is formed by condensing glutamic acid, cysteine and glycine, and is also a main non-protein sulfhydryl compound found in cells, and is the most abundant antioxidant factor in cells. Reduced Glutathione (GSH) plays an important role in cellular metabolism, cellular proliferation, intracellular environmental stability, and disease resistance. Glutathione is present in the redox balance prior to disulfide (GSSG oxidized) and sulfhydryl (GSH reduced) compounds, and the levels of reduced glutathione within living cells significantly alter the corresponding oxidative stress that has been implicated in many diseases and accelerated aging processes. Upon oxidative stress, reduced glutathione may be converted to oxidized form (GSSG) to protect cells from oxidative stress and to assist in capturing that radical that damages RNA or DNA. As a sensitive indicator, any change in the optimal ratio of intracellular GSH and GSSG can lead to diseases in humans such as: cancer, heart disease, stroke, and many neurological disorders. In particular, in some cancer cells, the concentration of intracellular reduced glutathione is generally higher than in normal cells. Therefore, detection of glutathione in vivo plays a very important role in disease diagnosis.
Currently, fluorescence detection, electrochemistry, high performance liquid chromatography, chemiluminescence and spectrophotometry are widely used to study GSH. However, these methods have derivatization, complex experimental procedures, time consuming, limited in vivo applications, which are detrimental to the development of practical procedures, and difficult to achieve for effective targeted drug delivery. Quartz Crystal Microbalance (QCM) has been receiving more and more attention from the scientific community due to its high sensitivity to mass changes, and the sensor has advantages of good specificity, high sensitivity, low cost and simple operation, and has a very good application potential in the field of disease analysis and detection. In addition, in research in recent years, mesoporous silicon nanocages as nanocarriers are attracting attention, and the surfaces of the nanocages can be chemically modified, have a high specific surface area, a characteristic nanostructure, biocompatibility and a high-load function. As a nano-scale molecule, DNA is active in nano science, bioscience, and material science with various advantages of its own programmability, structural diversity, and controllability of variation.
Therefore, it is necessary to design a DNA functionalized nanocage by utilizing the precise pairing of bases and the designable sequence characteristics, so as to realize the detection of high specificity and high sensitivity of the circulating tumor cells and the release of anticancer drugs into the circulating tumor cells by targeting reduced glutathione.
Disclosure of Invention
Aiming at the situation, in order to make up the existing defects, the invention prepares a medicine carrying nano-cage with a mesoporous structure based on a DNA functionalized nano-cage and taking the nano-cage as an anticancer medicine carrier; the invention provides a mesoporous structure drug-loaded nano cage which is simple in preparation method, high in sensitivity and high in drug-targeted performance for detecting the circulating tumor cells, and is suitable for rapid detection and targeted drug release of the circulating tumor cells in various human bodies.
The invention provides the following technical scheme:
the preparation method of the drug-loaded nano cage specifically comprises the following steps: preparing an amino mesoporous silicon nanocage, continuously stirring the functionalized DNA sequence, the amino mesoporous silicon nanocage and the medicine triptolide in a dispersion liquid, and performing precipitation and centrifugal cleaning twice to realize mesoporous encapsulation of the amino mesoporous silicon nanocage and embedding of the anticancer medicine triptolide, thus obtaining the medicine carrying nanocage.
Further, the functionalized DNA sequence is a disulfide bond-containing chain hybridization product of three DNA sequences subjected to chain hybridization reaction.
Further, the three DNA sequences are sequence one, sequence two and sequence three, respectively, the sequence one is: 5'-ATC AGA CTG ATG TTG A CAA AGT-/HS-SH/-T CAA CAT CAG TCT- (BHQ) -GAT AAG CTA-3'; the second sequence is: 5'-ACT TTG TCA ACA TCA GTC TGA T TAG CT-/HS-SH/-T ATCA GAC TGA TGT TGA-3', the sequence three is: 5'-COOH-TTT TTT TTT TTT TAG CTT ATC AGA CTG ATG TTG A-3'.
Preferably, the drug-carrying nanocages are mesoporous structure drug-carrying nanocages.
The application of the drug-loaded nanocage to release triptolide from circulating tumor cells specifically comprises the following steps:
(1) The medicine carrying nano-cage is used as a functional probe, the medicine carrying nano-cage with a mesoporous structure is used for modifying functional nucleic acid on the quartz crystal microbalance chip, incubating the medicine carrying nano-cage with a mesoporous structure into circulating tumor cells, capturing the cells based on an aptamer recognition principle, capturing the circulating tumor cells by the quartz crystal microbalance chip, improving the sensitivity of detecting the circulating tumor cells by the quartz crystal microbalance, and realizing high-sensitivity detection of the circulating tumor cells;
(2) And adding a quartz crystal microbalance chip combined with the circulating tumor cells into the incision enzyme, and carrying out reaction between the medicine carrying nano cage with a mesoporous structure and reduced glutathione in the circulating tumor cells of the tested object based on thiol exchange reaction, so as to release the anticancer medicine triptolide in the circulating tumor cells in a targeted manner.
The beneficial effects obtained by the invention by adopting the structure are as follows: compared with the prior art, the invention has the main innovation and superiority as follows:
1. the invention adopts a two-step method to prepare the medicine carrying nano cage with the mesoporous structure, and the preparation method is simple to operate;
2. the invention takes the medicine carrying nano cage with a mesoporous structure as a probe, improves the sensitivity of Quartz Crystal Microbalance (QCM) for detecting the circulating tumor cells, and realizes the high-sensitivity detection of the circulating tumor cells;
3. the invention takes the medicine carrying nano-cage with mesoporous structure as the medicine carrying probe, realizes the release of the anticancer medicine in the circulating tumor cells with high specific selectivity, and reduces the unnecessary damage to the normal cells.
Drawings
FIG. 1 is a scanning electron microscope image of a drug-loaded nano cage with a mesoporous structure;
FIG. 2 is a graph showing the response of the number of circulating tumor cells to a Quartz Crystal Microbalance (QCM) signal in accordance with the present invention;
FIG. 3 is a graph showing the linear relationship between the number of circulating tumor cells and the frequency intensity of a Quartz Crystal Microbalance (QCM) according to the present invention;
FIG. 4 is a graph showing the comparison of fluorescence intensity of drug release amounts of drug-loaded nanocages (a), normal cells (b) and circulating tumor cells (c) with mesoporous structures according to the present invention;
FIG. 5 is an imaging view of the mesoporous structured drug-loaded nanocages of the present invention for targeted delivery of drugs to circulating tumor cells.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and fully with reference to the accompanying drawings, in which it is evident that the embodiments described are only some, but not all embodiments of the invention; all other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
1. Preparation of a medicine carrying nano cage with a mesoporous structure:
(1) 70 g of triethanolamine, 390 g of hexadecyl trimethyl benzene sulfonic acid, 200mL of ultrapure water, organic amine small molecule solution and 2200g of tetraethoxysilane are mixed and stirred for 3 hours at 80 ℃;
(2) Filtering and washing the mixture obtained in the step (1), and baking the mixture in an oven at 95 ℃ overnight for 24 hours to synthesize mesoporous silicon;
(3) Weighing 10 g of synthesized mesoporous silicon, dissolving the mesoporous silicon in 1000 ml of absolute ethyl alcohol, adding 3-aminopropyl triethoxysilane, and stirring for 6 hours at 37 ℃;
(4) Filtering the product prepared in the step (3), washing with absolute ethyl alcohol, and drying at 60 ℃ to obtain an amino mesoporous silicon nano cage;
(5) The DNA sequence is hybridized in chain, the hybridized product is continuously stirred with amino mesoporous silicon nano cage (30 mg/ml) and medicine triptolide at 37 ℃ for 3 hours in dispersion liquid, and the suspension of medicine carrying nano cage with mesoporous structure is obtained after precipitation and centrifugal washing twice.
2. Quartz Crystal Microbalance (QCM) detection of circulating tumor cells:
modifying an aptamer on a QCM chip, incubating a suspension (20 mg/ml) of a medicine carrying nano cage with a mesoporous structure into a circulating tumor cell MDA-MB-231, capturing the circulating tumor cell MDA-MB-231 by the QCM chip based on an aptamer recognition principle in a QCM reaction tank at 37 ℃, and carrying out QCM test, wherein the experimental result is shown in figures 2 and 3.
As shown in FIG. 2, the QCM frequency intensity increased with increasing cell number of the circulating tumor MDA-MB-231.
As shown in fig. 3, when the number of the MDA-MB-231 cells of the circulating tumor is in the range of 10 to 9000, the QCM intensity response is in a linear relationship with the number of the MDA-MB-231 cells of the circulating tumor, the linear regression equation is Δf= 137.18lgN-122.19, where N is the number of the MDA-MB-231 cells of the circulating tumor, Δf=f-f0, F is the QCM frequency intensity after the MDA-MB-231 cells of the circulating tumor act, F0 is the frequency intensity before the MDA-MB-231 cells of the circulating tumor act, and r=0.992.
3. Fluorescence intensity contrast of released drug:
the fluorescence intensity of triptolide released by the medicine carrying nano-cage with a mesoporous structure (20 mg/ml) is used as a reference sample by a fluorescence spectrophotometer detection technology, the fluorescence intensity of triptolide released by the medicine carrying nano-cage with a mesoporous structure in normal cells and circulating tumor cells MDA-MB-231 is matched and compared, the medicine carrying nano-cage with the mesoporous structure can target reduced glutathione in the circulating tumor cells, a thiol exchange reaction is initiated, the release of the anti-cancer medicine triptolide in the circulating tumor cells is realized, and the experimental result is shown in figure 4.
The data in fig. 4 shows fluorescence intensity contrast of three drug release amounts of mesoporous structure drug-loaded nanocages (a), mesoporous structure drug-loaded nanocages in normal cells (b) and mesoporous structure drug-loaded nanocages in circulating tumor cells MDA-MB-231 (c), and the contrast data shows that: the mesoporous structure drug-loaded nano cage can target reduced glutathione in circulating tumor cells to trigger thiol exchange reaction, so that the anti-cancer drug triptolide in the circulating tumor cells can be effectively released, the drug release amount to normal cells is small, and the damage to the normal cells is reduced.
4. Targeting circulating tumor cells releases drugs:
and 3 mu L of nicking endonuclease is added into a QCM chip reaction tank combined with the circulating tumor cells MDA-MB-231, the QCM chip is separated from the liquid to obtain a product, and a laser confocal microscope imaging test is carried out, wherein the experimental result is shown in figure 5.
Fig. 5 shows a fluorescence imaging diagram of drug-loaded nanocage targeting circulating tumor cells MDA-MB-231 with mesoporous structure to release drugs, and the visual imaging technology shows that a large amount of anticancer drugs triptolide are released in the circulating tumor cells.
It is noted that relational terms such as first and second, and the like are used solely to distinguish one entity or action from another entity or action without necessarily requiring or implying any actual such relationship or order between such entities or actions. Moreover, the terms "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or apparatus.
Although embodiments of the present invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made therein without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
SEQUENCE LISTING
<110> university of Shandong traditional Chinese medicine
<120> preparation method of drug-loaded nanocage and application of drug-loaded nanocage in targeting circulating tumor cells to release triptolide
<130> university of Shandong traditional Chinese medicine
<160> 3
<170> PatentIn version 3.5
<210> 1
<211> 44
<212> DNA
<213> sequence 1
<400> 1
atcagactga tgttgacaaa gttcaacatc agtctgataa gcta 44
<210> 2
<211> 44
<212> DNA
<213> sequence 2
<400> 2
actttgtcaa catcagtctg attagcttat cagactgatg ttga 44
<210> 3
<211> 34
<212> DNA
<213> sequence 3
<400> 3
tttttttttt tttagcttat cagactgatg ttga 34
Claims (2)
1. The preparation method of the drug-loaded nano cage is characterized by comprising the following steps of: preparing an amino mesoporous silicon nanocage, continuously stirring the functionalized DNA sequence, the amino mesoporous silicon nanocage and the medicine triptolide in a dispersion liquid, and performing precipitation and centrifugal cleaning twice to realize mesoporous encapsulation of the amino mesoporous silicon nanocage and embedding of the anticancer medicine triptolide, thus obtaining the medicine carrying nanocage;
the functionalized DNA sequence is a chain hybridization product containing disulfide bonds, wherein the chain hybridization reaction of the three DNA sequences is carried out;
the three DNA sequences are respectively a sequence I, a sequence II and a sequence III, wherein the sequence I is as follows: 5'-ATC AGA CTG ATG TTG A CAA AGT-/HS-SH/-T CAA CAT CAG TCT- (BHQ) -GAT AAG CTA-3'; the second sequence is: 5'-ACT TTG TCA ACA TCA GTC TGA T TAG CT-/HS-SH/-T A TCA GAC TGA TGT TGA-3', the sequence three is: 5'-COOH-TTT TTT TTT TTT TAG CTT ATC AGA CTG ATG TTG A-3'.
2. The method for preparing the drug-loaded nanocages according to claim 1, wherein the drug-loaded nanocages are mesoporous drug-loaded nanocages.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110459985.4A CN113082226B (en) | 2021-04-27 | 2021-04-27 | Preparation method of drug-loaded nanocage and application of drug-loaded nanocage in targeting circulating tumor cells to release triptolide |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110459985.4A CN113082226B (en) | 2021-04-27 | 2021-04-27 | Preparation method of drug-loaded nanocage and application of drug-loaded nanocage in targeting circulating tumor cells to release triptolide |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN113082226A CN113082226A (en) | 2021-07-09 |
| CN113082226B true CN113082226B (en) | 2023-12-15 |
Family
ID=76680329
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110459985.4A Active CN113082226B (en) | 2021-04-27 | 2021-04-27 | Preparation method of drug-loaded nanocage and application of drug-loaded nanocage in targeting circulating tumor cells to release triptolide |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113082226B (en) |
Citations (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102657598A (en) * | 2012-05-09 | 2012-09-12 | 上海交通大学 | Porous inorganic material based oral preparation of secondary-dispersion insoluble drug and preparation method thereof |
| CN104013965A (en) * | 2014-05-06 | 2014-09-03 | 重庆大学 | Method for preparing meso-porous silicon nano medicine carrier with cell specificity target, reduction responsiveness and triple anticancer treatment effects |
| CN105125510A (en) * | 2015-09-30 | 2015-12-09 | 福州大学 | Antibody coupled mesoporous silica/mifepristone nanometer preparation |
| CN106890340A (en) * | 2017-02-28 | 2017-06-27 | 福州大学 | A kind of pair of mesoporous silicon oxide targeted medicament carrying nano grain of aptamers modification |
| CN109212211A (en) * | 2017-07-07 | 2019-01-15 | 叶健 | It is a kind of for detecting the chip of circulating tumor cell |
| CN109715802A (en) * | 2016-03-18 | 2019-05-03 | 卡里斯科学公司 | Oligonucleotide probe and application thereof |
| CN110257520A (en) * | 2019-07-04 | 2019-09-20 | 福州大学 | A technique for utilizing miRNA in short capillary high speed electrophoresis detection lung carcinoma cell |
| CN110478497A (en) * | 2019-09-03 | 2019-11-22 | 青岛大学 | A kind of dual drug-loading system and preparation method and application based on targeting paranuclein |
| CN113189070A (en) * | 2021-04-28 | 2021-07-30 | 香港理工大学 | Nano probe, preparation method and application thereof, and detection system for miRNAs of circulating tumor cells |
| CN114438215A (en) * | 2022-04-02 | 2022-05-06 | 山东中医药大学 | SERS (surface enhanced Raman Scattering) probe and kit for detecting lung cancer marker based on DNA (deoxyribonucleic acid) hyperbranched self-assembly |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CA2852064A1 (en) * | 2011-10-14 | 2013-04-18 | Stc.Unm | Porous nanoparticle-supported lipid bilayers (protocells) for targeted delivery including transdermal delivery of cargo and methods thereof |
-
2021
- 2021-04-27 CN CN202110459985.4A patent/CN113082226B/en active Active
Patent Citations (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102657598A (en) * | 2012-05-09 | 2012-09-12 | 上海交通大学 | Porous inorganic material based oral preparation of secondary-dispersion insoluble drug and preparation method thereof |
| CN104013965A (en) * | 2014-05-06 | 2014-09-03 | 重庆大学 | Method for preparing meso-porous silicon nano medicine carrier with cell specificity target, reduction responsiveness and triple anticancer treatment effects |
| CN105125510A (en) * | 2015-09-30 | 2015-12-09 | 福州大学 | Antibody coupled mesoporous silica/mifepristone nanometer preparation |
| CN109715802A (en) * | 2016-03-18 | 2019-05-03 | 卡里斯科学公司 | Oligonucleotide probe and application thereof |
| CN106890340A (en) * | 2017-02-28 | 2017-06-27 | 福州大学 | A kind of pair of mesoporous silicon oxide targeted medicament carrying nano grain of aptamers modification |
| CN109212211A (en) * | 2017-07-07 | 2019-01-15 | 叶健 | It is a kind of for detecting the chip of circulating tumor cell |
| CN110257520A (en) * | 2019-07-04 | 2019-09-20 | 福州大学 | A technique for utilizing miRNA in short capillary high speed electrophoresis detection lung carcinoma cell |
| CN110478497A (en) * | 2019-09-03 | 2019-11-22 | 青岛大学 | A kind of dual drug-loading system and preparation method and application based on targeting paranuclein |
| CN113189070A (en) * | 2021-04-28 | 2021-07-30 | 香港理工大学 | Nano probe, preparation method and application thereof, and detection system for miRNAs of circulating tumor cells |
| CN114438215A (en) * | 2022-04-02 | 2022-05-06 | 山东中医药大学 | SERS (surface enhanced Raman Scattering) probe and kit for detecting lung cancer marker based on DNA (deoxyribonucleic acid) hyperbranched self-assembly |
Non-Patent Citations (4)
| Title |
|---|
| Combination of microfluidic chips and biosensing for the enrichment of circulating tumor cells;Jiaju Shi，等;Biosensors and Bioelectronics;第202卷;1-9 * |
| Mesoporous silica nanoparticles with surface transformation ability for prostate cancer treatment;Chang-Ming Liu，等;Colloids and Surfaces A: Physicochemical and Engineering Aspects;第621卷;1-10 * |
| Nano Meets Micro-Translational Nanotechnology in Medicine: Nano-Based Applications for Early Tumor Detection and Therapy;Svenja Siemer，等;Nanomaterials;第10卷(第383期);1-28 * |
| 氨基表面修饰有序介孔硅对槲皮素的载药性能及药物释放影响研究;胡坪，等;中国药学杂志;第48卷(第11期);894-898 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN113082226A (en) | 2021-07-09 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Copur et al. | Nanopaper-based photoluminescent enantioselective sensing of L-Lysine by L-Cysteine modified carbon quantum dots | |
| KR101066159B1 (en) | Method of purifying/concentrating sugar chain with sugar chain-trapping molecule and method of analyzing sugar chain structure | |
| Fu et al. | Melamine-induced decomposition and anti-FRET effect from a self-assembled complex of rhodamine 6G and DNA-stabilized silver nanoclusters used for dual-emitting ratiometric and naked-eye-visible fluorescence detection | |
| Sun et al. | Size-dependent property and cell labeling of semiconducting polymer dots | |
| Li et al. | A hydrogel microsphere-based sensor for dual and highly selective detection of Al3+ and Hg2+ | |
| Sangubotla et al. | Fiber-optic biosensor based on the laccase immobilization on silica-functionalized fluorescent carbon dots for the detection of dopamine and multi-color imaging applications in neuroblastoma cells | |
| CN106947811B (en) | Method, probe set and kit for detecting miRNAs-21 | |
| CN106908429B (en) | Method for detecting glutathione | |
| Qin et al. | Aggregation-induced emission-active fluorescent polymer: multi-targeted sensor and ROS scavenger | |
| CN106892947B (en) | One kind containing the complex of iridium and its preparation method and application of (Hydrazinocarbonyl) ferrocene ligands | |
| US20210269421A1 (en) | Water-soluble fluorescent probe and nanoparticals with aggregation-induced emission effect for ovarian cancer and preparation method and use thereof | |
| CN110684523A (en) | Near-infrared fluorescent molecular probe for detecting hydrogen sulfide and preparation method and application thereof | |
| CN106932371B (en) | Fluorescence imaging method of intracellular glutathione | |
| Jia et al. | Target-triggered and controlled release plasmon-enhanced fluorescent AIE probe for conformational monitoring of insulin fibrillation | |
| Ji et al. | DNAzyme-functionalized porous carbon nanospheres serve as a fluorescent nanoprobe for imaging detection of microRNA-21 and zinc ion in living cells | |
| CN113082226B (en) | Preparation method of drug-loaded nanocage and application of drug-loaded nanocage in targeting circulating tumor cells to release triptolide | |
| CN106243244B (en) | A kind of carboxyl chitosan namo fluorescence probe and preparation method thereof with aggregation-induced emission characteristic | |
| CN109705129B (en) | Gold nano-cell fluorescent probe capable of improving cell activity and preparation method and application thereof | |
| CN115572335B (en) | Chitosan-based fluorescent probe for formaldehyde monitoring and preparation method and application thereof | |
| CN106674315A (en) | Pentaerythritol-based Janus lipid and intermediate thereof, preparation method and application | |
| CN114853921A (en) | Preparation method of modified chitosan and application of modified chitosan in detection of chromium ion pollutants | |
| CN109444103B (en) | Preparation method of PEI functionalized green fluorescent carbon dots and thrombin detection method based on carbon dots | |
| CN106802294B (en) | Glutathione fluorescence sensor based on controlled release technology and preparation method thereof | |
| CN110554012A (en) | carbon dot fluorescent probe for glutathione detection and preparation method thereof | |
| US10711094B2 (en) | Conjugated polymer nanoparticles, methods of using, and methods of making |
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 |