CN106727284B - Magnetic reduction enhanced drug sensitive release nanogel and preparation and storage methods thereof - Google Patents
Magnetic reduction enhanced drug sensitive release nanogel and preparation and storage methods thereof Download PDFInfo
- Publication number
- CN106727284B CN106727284B CN201710012359.4A CN201710012359A CN106727284B CN 106727284 B CN106727284 B CN 106727284B CN 201710012359 A CN201710012359 A CN 201710012359A CN 106727284 B CN106727284 B CN 106727284B
- Authority
- CN
- China
- Prior art keywords
- nanogel
- solution
- drug
- sensitive release
- selenium
- 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.)
- Expired - Fee Related
Links
- 239000003814 drug Substances 0.000 title claims abstract description 45
- 229940079593 drug Drugs 0.000 title claims abstract description 43
- 230000009467 reduction Effects 0.000 title claims abstract description 31
- 238000000034 method Methods 0.000 title claims abstract description 22
- 238000002360 preparation method Methods 0.000 title claims abstract description 16
- 238000003860 storage Methods 0.000 title abstract description 6
- 239000002246 antineoplastic agent Substances 0.000 claims abstract description 10
- SZVJSHCCFOBDDC-UHFFFAOYSA-N ferrosoferric oxide Chemical compound O=[Fe]O[Fe]O[Fe]=O SZVJSHCCFOBDDC-UHFFFAOYSA-N 0.000 claims abstract description 10
- 239000002105 nanoparticle Substances 0.000 claims abstract description 10
- 229940041181 antineoplastic drug Drugs 0.000 claims abstract description 9
- 230000008569 process Effects 0.000 claims abstract description 4
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 24
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 20
- 229910021642 ultra pure water Inorganic materials 0.000 claims description 19
- 239000012498 ultrapure water Substances 0.000 claims description 19
- 238000006243 chemical reaction Methods 0.000 claims description 17
- IXPNQXFRVYWDDI-UHFFFAOYSA-N 1-methyl-2,4-dioxo-1,3-diazinane-5-carboximidamide Chemical compound CN1CC(C(N)=N)C(=O)NC1=O IXPNQXFRVYWDDI-UHFFFAOYSA-N 0.000 claims description 16
- 235000010413 sodium alginate Nutrition 0.000 claims description 16
- 229940005550 sodium alginate Drugs 0.000 claims description 16
- 239000000661 sodium alginate Substances 0.000 claims description 16
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 15
- LMDZBCPBFSXMTL-UHFFFAOYSA-N 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide Chemical compound CCN=C=NCCCN(C)C LMDZBCPBFSXMTL-UHFFFAOYSA-N 0.000 claims description 12
- 229920001661 Chitosan Polymers 0.000 claims description 12
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 12
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 claims description 10
- 229910052757 nitrogen Inorganic materials 0.000 claims description 10
- 238000003756 stirring Methods 0.000 claims description 9
- 238000000502 dialysis Methods 0.000 claims description 8
- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 claims description 7
- 229920002521 macromolecule Polymers 0.000 claims description 7
- QIJRTFXNRTXDIP-UHFFFAOYSA-N (1-carboxy-2-sulfanylethyl)azanium;chloride;hydrate Chemical compound O.Cl.SCC(N)C(O)=O QIJRTFXNRTXDIP-UHFFFAOYSA-N 0.000 claims description 6
- 229960001305 cysteine hydrochloride Drugs 0.000 claims description 6
- AXJLEYVRLPNPGF-UHFFFAOYSA-N 3-(2-carboxyethyldiselanyl)propanoic acid Chemical compound OC(=O)CC[Se][Se]CCC(O)=O AXJLEYVRLPNPGF-UHFFFAOYSA-N 0.000 claims description 5
- WORSJCPMKUOXAD-UHFFFAOYSA-N 4-(3-carboxypropyldiselanyl)butanoic acid Chemical compound OC(=O)CCC[Se][Se]CCCC(O)=O WORSJCPMKUOXAD-UHFFFAOYSA-N 0.000 claims description 5
- 239000002994 raw material Substances 0.000 claims description 5
- 230000002829 reductive effect Effects 0.000 claims description 5
- 229920001503 Glucan Polymers 0.000 claims description 4
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 claims description 4
- 150000001875 compounds Chemical class 0.000 claims description 4
- 238000001035 drying Methods 0.000 claims description 4
- 238000001914 filtration Methods 0.000 claims description 4
- 239000005457 ice water Substances 0.000 claims description 4
- 229920002307 Dextran Polymers 0.000 claims description 3
- NQTADLQHYWFPDB-UHFFFAOYSA-N N-Hydroxysuccinimide Chemical compound ON1C(=O)CCC1=O NQTADLQHYWFPDB-UHFFFAOYSA-N 0.000 claims description 3
- 125000003277 amino group Chemical group 0.000 claims description 3
- 125000000524 functional group Chemical group 0.000 claims description 3
- 229910052711 selenium Inorganic materials 0.000 claims description 3
- 239000011669 selenium Substances 0.000 claims description 3
- 108010006654 Bleomycin Proteins 0.000 claims description 2
- MWWSFMDVAYGXBV-RUELKSSGSA-N Doxorubicin hydrochloride Chemical compound Cl.O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(=O)CO)[C@H]1C[C@H](N)[C@H](O)[C@H](C)O1 MWWSFMDVAYGXBV-RUELKSSGSA-N 0.000 claims description 2
- FBOZXECLQNJBKD-ZDUSSCGKSA-N L-methotrexate Chemical compound C=1N=C2N=C(N)N=C(N)C2=NC=1CN(C)C1=CC=C(C(=O)N[C@@H](CCC(O)=O)C(O)=O)C=C1 FBOZXECLQNJBKD-ZDUSSCGKSA-N 0.000 claims description 2
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 2
- 229930192392 Mitomycin Natural products 0.000 claims description 2
- NWIBSHFKIJFRCO-WUDYKRTCSA-N Mytomycin Chemical compound C1N2C(C(C(C)=C(N)C3=O)=O)=C3[C@@H](COC(N)=O)[C@@]2(OC)[C@@H]2[C@H]1N2 NWIBSHFKIJFRCO-WUDYKRTCSA-N 0.000 claims description 2
- 230000003213 activating effect Effects 0.000 claims description 2
- 229960001561 bleomycin Drugs 0.000 claims description 2
- OYVAGSVQBOHSSS-UAPAGMARSA-O bleomycin A2 Chemical compound N([C@H](C(=O)N[C@H](C)[C@@H](O)[C@H](C)C(=O)N[C@@H]([C@H](O)C)C(=O)NCCC=1SC=C(N=1)C=1SC=C(N=1)C(=O)NCCC[S+](C)C)[C@@H](O[C@H]1[C@H]([C@@H](O)[C@H](O)[C@H](CO)O1)O[C@@H]1[C@H]([C@@H](OC(N)=O)[C@H](O)[C@@H](CO)O1)O)C=1N=CNC=1)C(=O)C1=NC([C@H](CC(N)=O)NC[C@H](N)C(N)=O)=NC(N)=C1C OYVAGSVQBOHSSS-UAPAGMARSA-O 0.000 claims description 2
- 229960002918 doxorubicin hydrochloride Drugs 0.000 claims description 2
- 238000007710 freezing Methods 0.000 claims description 2
- 230000008014 freezing Effects 0.000 claims description 2
- 229960000485 methotrexate Drugs 0.000 claims description 2
- 229960004857 mitomycin Drugs 0.000 claims description 2
- 238000002156 mixing Methods 0.000 claims description 2
- 239000000843 powder Substances 0.000 claims description 2
- 239000012429 reaction media Substances 0.000 claims description 2
- 238000002390 rotary evaporation Methods 0.000 claims description 2
- 229910000033 sodium borohydride Inorganic materials 0.000 claims description 2
- 239000012279 sodium borohydride Substances 0.000 claims description 2
- 229910000029 sodium carbonate Inorganic materials 0.000 claims description 2
- 239000007787 solid Substances 0.000 claims description 2
- VSNHCAURESNICA-NJFSPNSNSA-N 1-oxidanylurea Chemical compound N[14C](=O)NO VSNHCAURESNICA-NJFSPNSNSA-N 0.000 claims 1
- 238000010438 heat treatment Methods 0.000 claims 1
- 150000003573 thiols Chemical group 0.000 claims 1
- 231100000331 toxic Toxicity 0.000 abstract description 5
- 230000002588 toxic effect Effects 0.000 abstract description 5
- 239000000126 substance Substances 0.000 abstract description 3
- 230000008685 targeting Effects 0.000 abstract description 3
- 239000012467 final product Substances 0.000 abstract 1
- 230000002194 synthesizing effect Effects 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 28
- AOJJSUZBOXZQNB-TZSSRYMLSA-N Doxorubicin Chemical compound O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(=O)CO)[C@H]1C[C@H](N)[C@H](O)[C@H](C)O1 AOJJSUZBOXZQNB-TZSSRYMLSA-N 0.000 description 19
- 206010028980 Neoplasm Diseases 0.000 description 17
- 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 12
- 230000000694 effects Effects 0.000 description 11
- 241000699670 Mus sp. Species 0.000 description 10
- 239000003937 drug carrier Substances 0.000 description 7
- 229960003180 glutathione Drugs 0.000 description 6
- 229960004679 doxorubicin Drugs 0.000 description 5
- 239000002245 particle Substances 0.000 description 5
- 229940009456 adriamycin Drugs 0.000 description 4
- 201000011510 cancer Diseases 0.000 description 4
- 210000004027 cell Anatomy 0.000 description 4
- 230000005764 inhibitory process Effects 0.000 description 4
- 230000035945 sensitivity Effects 0.000 description 4
- 230000005540 biological transmission Effects 0.000 description 3
- 238000013270 controlled release Methods 0.000 description 3
- 230000004043 responsiveness Effects 0.000 description 3
- 241000699666 Mus <mouse, genus> Species 0.000 description 2
- 238000005576 amination reaction Methods 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
- 238000012925 biological evaluation Methods 0.000 description 2
- 239000012620 biological material Substances 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 230000001186 cumulative effect Effects 0.000 description 2
- 238000009826 distribution Methods 0.000 description 2
- 238000005538 encapsulation Methods 0.000 description 2
- 238000002347 injection Methods 0.000 description 2
- 239000007924 injection Substances 0.000 description 2
- 231100001231 less toxic Toxicity 0.000 description 2
- 238000011068 loading method Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 230000005012 migration Effects 0.000 description 2
- 238000013508 migration Methods 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 230000000638 stimulation Effects 0.000 description 2
- 125000003396 thiol group Chemical group [H]S* 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 208000031648 Body Weight Changes Diseases 0.000 description 1
- 108010024636 Glutathione Proteins 0.000 description 1
- VSNHCAURESNICA-UHFFFAOYSA-N Hydroxyurea Chemical compound NC(=O)NO VSNHCAURESNICA-UHFFFAOYSA-N 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 230000004579 body weight change Effects 0.000 description 1
- 229940044683 chemotherapy drug Drugs 0.000 description 1
- 238000004132 cross linking Methods 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
- 230000009977 dual effect Effects 0.000 description 1
- 230000009881 electrostatic interaction Effects 0.000 description 1
- 210000001163 endosome Anatomy 0.000 description 1
- 229960001330 hydroxycarbamide Drugs 0.000 description 1
- 239000003446 ligand Substances 0.000 description 1
- 210000003712 lysosome Anatomy 0.000 description 1
- 230000001868 lysosomic effect Effects 0.000 description 1
- 238000002595 magnetic resonance imaging Methods 0.000 description 1
- 230000005389 magnetism Effects 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 239000002086 nanomaterial Substances 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 235000018102 proteins Nutrition 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000007430 reference method Methods 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 239000012265 solid product Substances 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 229910052717 sulfur Chemical group 0.000 description 1
- 239000011593 sulfur Chemical group 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 229940126585 therapeutic drug Drugs 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/06—Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
- A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
- A61K31/704—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin attached to a condensed carbocyclic ring system, e.g. sennosides, thiocolchicosides, escin, daunorubicin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
-
- 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
- A61K47/02—Inorganic compounds
-
- 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
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0002—Galenical forms characterised by the drug release technique; Application systems commanded by energy
- A61K9/0009—Galenical forms characterised by the drug release technique; Application systems commanded by energy involving or responsive to electricity, magnetism or acoustic waves; Galenical aspects of sonophoresis, iontophoresis, electroporation or electroosmosis
Abstract
The invention relates to a magnetic reduction enhanced drug sensitive release nanogel and a preparation and storage method thereof. The magnetic reduction enhanced drug sensitive release nanogel is prepared from superparamagnetic ferroferric oxide nanoparticles with aminated surfaces, a high molecular derivative with a side chain containing a selenium-sulfur bond and an anticancer drug according to the mass ratio of (1-2): 1: (0.2-0.5). The preparation method specifically comprises the following steps: 1) synthesizing a macromolecular derivative with a side chain containing a selenium-sulfur bond; 2) adding SPION-NH2The anticancer drug and the macromolecular derivative with the side chain containing the selenium-sulfur bond obtained in the step 1) are self-assembled to obtain the final product. The method for storing the nanogel is to store the nanogel in an environment at 2-4 ℃. The preparation method has mild preparation conditions, no toxic and harmful substances are introduced, the process is simple, the repeatability is good, the prepared nanogel has strong reduction sensitive release, the burst release and the drug leakage in the normal physiological environment are effectively avoided, the active magnetic targeting property is realized, and the application prospect in the field of targeted antitumor drugs is wide.
Description
Technical Field
The invention relates to a magnetic reduction enhanced drug sensitive release nanogel and a preparation and storage method thereof, belonging to the field of nanomaterials and the technical field of drug controlled release.
Background
Nanogel refers to a three-dimensional network structure with nanometer (10-1000nm) size and formed by chemically or physically crosslinking macromolecules. The nanogel is used as a drug carrier, therapeutic drugs are loaded in an adsorption or embedding mode, and the drug carrier with active or passive targeting performance is designed through the combination between a targeting ligand and a receptor or the difference between normal tissues and the difference between the inside and the outside of cells and tumors. The EPR effect of the tumor part is combined, the larger cell gap and the looser structure of the tumor part are utilized to promote the nanometer carrier to pass through, so that the tumor part can be better enriched at the focus part, the bioavailability of the medicine and the conveying and releasing performance of the medicine in the body can be obviously improved, and the system is a safe system with potential biomedical application value.
Recently, multi-stimulus responsive drug delivery systems have been rapidly developed in the study of cancer diagnosis and treatment. It is known that the concentration of Glutathione (GSH) in the tumor extracellular environment is 2-10. mu.M, while the GSH values of endosomes and lysosomes are 2-10 mM, which differ by a factor of hundreds. GSH has reducibility, so chemical bonds which are reduced and broken and contain disulfide bonds, diselenide bonds and the like are designed in the nanogel structure, the reduction stimulation responsiveness of the nanogel can be realized, and the treatment effect of cancer is effectively improved.
The superparamagnetic ferroferric oxide nano particles can effectively enrich the carrier at a target site in an exogenous magnetic field, and the superparamagnetic ferroferric oxide nano particles have the properties of magnetocaloric curative effect, magnetic resonance imaging and the like, so that the superparamagnetic ferroferric oxide nano particles can be widely applied to the drug carrier.
Therefore, the preparation of the nano-drug carrier with good repeatability and good reduction sensitivity stimulation release under mild conditions is one of the problems which need to be solved urgently in the application of the current chemotherapeutic drugs.
Disclosure of Invention
Aiming at the problems in the prior art, the invention aims to provide a magnetic reduction enhanced drug sensitive release nanogel and a preparation and storage method thereof. The magnetic reduction enhanced drug sensitive release nanogel has higher reduction responsiveness sensitivity, can reduce the release amount of a carrier in normal tissues, thereby reducing toxic and side effects and improving the treatment efficiency, and the preparation method is mild and does not introduce toxic and harmful substances.
In order to achieve the purpose, the technical scheme adopted by the invention is as follows:
a magnetic reduction enhanced drug sensitive release nanogel is characterized by comprising the following raw materials in percentage by mass (1-2): 1: (0.2-0.5) preparation to obtain:
superparamagnetic ferroferric oxide nano particles with aminated surfaces, macromolecular derivatives with side chains containing selenium-sulfur bonds and anticancer drugs.
According to the scheme, preferably, the macromolecular derivative with the side chain containing the selenium-sulfur bond is prepared by the following steps:
under the protection of nitrogen at room temperature, taking a PBS solution with the concentration of 0.01mol/L as a reaction medium, and mixing the components in a mass ratio of 1-1.5: 1, taking the macromolecule with a sulfydryl functional group on the side chain and a compound with a double selenium bond as reaction raw materials, adjusting the pH value of a reaction system to be 6.5-7.4, stirring and reacting for 5-8 hours at 37 ℃, and dialyzing to obtain the macromolecule derivative with the selenium-sulfur bond on the side chain. Preferably, the medium of dialysis is ultrapure water.
According to the scheme, preferably, the macromolecule with the side chain containing the sulfydryl functional group is one of sulfhydrylated sodium alginate, sulfhydrylated dextran and sulfhydrylated chitosan.
The preparation method of the thiolated sodium alginate, the thiolated glucan and the thiolated chitosan comprises the following steps:
1) dissolving a proper amount of sodium alginate or aminated glucan or chitosan in a certain amount of ultrapure water; 2) dissolving an appropriate amount of 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) in an appropriate amount of ultrapure water; 3) under the protection of nitrogen, dropwise adding the EDC/NHS solution prepared in the step 2) into the sodium alginate or aminated glucan or chitosan solution prepared in the step 1), and activating for 30 min; 4) dissolving a proper amount of cysteine hydrochloride in a proper amount of ultrapure water, adjusting the pH to about 6-7 by using a 6mol/L sodium hydroxide solution, then dropwise adding the solution into the solution obtained in the step 3), continuously stirring for 12 hours, and carrying out light-shielding treatment in the whole process; after the reaction, the reaction solution was put into a dialysis bag and dialyzed for 2 days, followed by freezing and drying to obtain a solid powder. According to the scheme, preferably, the molar ratio of the carboxyl of the sodium alginate to the amino of the cysteine hydrochloride is 1: (1-1.2). According to the above scheme, preferably, the molar ratio of the amino group of the aminated dextran or chitosan to the carboxyl group of cysteine hydrochloride is 1: (1-1.2).
According to the scheme, preferably, the compound containing the diselenide bond is 3,3 '-diselenodipropionic acid or 4, 4' -diselenodibutyric acid. Among them, 3,3 '-diselenodipropionic acid can be prepared by reference to the literature (Langmuir,2006,22(13):5552-5565), (Int J Nanomed,2012,7(17):3991-4006), and 4, 4' -diselenodibutyric acid can be prepared by the following method:
respectively dissolving 3.95g of selenium powder and 2.00g of sodium hydroxide in 25mL of ice water; then 0.25g of sodium borohydride and 0.20g of sodium hydroxide are dissolved in 5mL of ice water and injected into the solution containing the selenium powder, and the solution is reacted at 0 ℃ under the protection of nitrogen in the whole process until the selenium powder is completely dissolved, and the solution is colorless. The solution was then allowed to warm to 90 ℃ and the reaction continued for 30min until the solution became reddish brown in color. 5.42g of 4-chloropropionic acid were dissolved in 20mL of ultrapure water, and after adjusting the pH to 8.0 with sodium carbonate, the solution was added to the above reddish brown solution and stirred overnight under nitrogen. After the reaction is finished, filtering the reaction solution to obtain a yellow solution, adjusting the pH value of the yellow solution to 3-4 by using 1.0mol/L hydrochloric acid, extracting twice by using ethyl acetate, drying by using anhydrous magnesium sulfate, filtering, performing rotary evaporation, and finally recrystallizing by using ethyl acetate to obtain a yellow solid product.
According to the scheme, preferably, the anti-cancer drug is one or more of doxorubicin hydrochloride, methotrexate, mitomycin, hydroxyurea and bleomycin.
The invention also provides a preparation method of the magnetic reduction enhanced type drug sensitive release nanogel, which is characterized by comprising the following steps of:
1) adding a macromolecular derivative with a side chain containing a selenium-sulfur bond into ultrapure water, and adjusting the pH value of the solution to 6-7.4;
2) dissolving superparamagnetic ferroferric oxide nano particles with aminated surfaces and an anticancer drug in hydrochloric acid, and controlling the pH value of the solution to be 4.5-5.5;
3) and (2) under the condition of keeping out of the sun, dropwise adding the solution obtained in the step 2) into the solution obtained in the step 1), adjusting the pH value of the solution to 7.2-7.4, stirring for 5-7h, and dialyzing to obtain the magnetic reduction enhanced type drug sensitive release nanogel.
According to the scheme, preferably, the dialyzed medium in the step 3) is ultrapure water, and the molecular weight cut-off of the dialysis bag is 10 KDa.
The invention also provides a storage method of the magnetic reduction enhanced type drug sensitive release nanogel, which is characterized by being stored in an environment at the temperature of 2-4 ℃.
Compared with the prior art, the invention has the beneficial effects that:
the invention has mild reaction conditions and low toxic and side effects of raw materials, realizes entrapment of the drug by means of electrostatic interaction, ensures strong reduction sensitivity of drug release by using the selenium-sulfur bond, effectively avoids adverse effects such as burst release and drug leakage in normal physiological environment, realizes rapid release of a large amount of drugs in reductive environment in cancer cells, and has low toxic and side effects and high treatment effect.
Drawings
Fig. 1 is a transmission electron microscope (a), a particle size distribution (b), a magnetic attraction figure (c) and a magnetic hysteresis loop (d) of the sodium alginate-based magnetic reduction-responsive nano-drug carrier prepared in example 1 of the present invention.
Fig. 2 is a drug release curve diagram of the sodium alginate-based magnetic reduction-responsive nano-drug carrier prepared in example 1 of the present invention in different environments.
Fig. 3 is a biological evaluation of mice with sodium alginate-based magnetic reduction-responsive nano-drug carriers prepared in example 1, a tumor tap (a), a tumor volume change graph (b), a tumor inhibition rate (c) and a mouse body weight change graph (d).
Detailed Description
The invention will be further illustrated with reference to the following specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention.
Example 1
1. Synthesis of proteins having surface amination by the reference method (Biomaterials,2009,30:4716-4722)The superparamagnetic ferroferric oxide nano particle (SPION-NH)2)。
2. The preparation method of the sodium alginate high molecular derivative (SA-SSe-COOH) with the side chain containing selenium and sulfur bonds comprises the following specific steps:
1) the side chain of sodium alginate is introduced into a thiol group by a method such as that described in references (J controlled release,2009,136:38-44), (Materials science and Engineering C,2015,46:41-51) to give a thiolated sodium alginate (SA-SH).
2) 3,3' -diselenodipropionic acid (DSeDPA) was synthesized by methods such as reference (Langmuir,2006,22(13):5552-5565), (Int J Nanomed,2012,7(17):3991-4006), and the like.
3) 0.1g of DSeDPA was weighed into a three-necked flask containing 30mL of PBS (0.01mol/L, pH6.56) and dissolved by stirring at room temperature under nitrogen. 0.125g of SA-SH was dissolved in 30mL of PBS (0.01mol of L-1, pH6.56) and injected into the DSeDPA solution under reaction conditions of 37 ℃ and stirred for 7 hours. The resulting solution was dialyzed against ultrapure water for 3 days to obtain 1.81mg/mL of SA-SSe-COOH.
3. 2.76mL of 1.81mg/mL SA-SSe-COOH was transferred into a three-necked flask containing 2.24mL of ultrapure water, and the pH thereof was adjusted to 6.5 with HCl; 0.908mL of 5.508mg/mL SPION-NH was taken2And 0.625mL of 4mg/mL doxorubicin was dissolved in 3.07mL of hydrochloric acid having pH 5.0, and added dropwise to the three-necked flask, and finally, the pH was adjusted to 7.4 with 0.1mol/L sodium hydroxide, and the mixture was stirred at room temperature for 5 hours, with light shielding over the whole course. Transferring the reaction solution into a dialysis bag, dialyzing with ultrapure water for 2 days, and storing the obtained nano-gel aqueous solution at 4 deg.C, wherein the cut-off molecular weight of the dialysis bag is 10 KDa. The drug loading rate and the encapsulation rate of the obtained magnetic nanogel are respectively 18.2 percent and 95.6 percent.
Measurement of properties of the magnetic nanogel obtained in this example:
the nanogel prepared in this example had a particle size of 110. + -.15 nm as measured by a laser particle sizer (DLS), and the results are shown in FIG. 1-a. The measured particle size is consistent with the result of the transmission electron microscope. The morphology and size distribution of the phases were observed using a Transmission Electron Microscope (TEM) and the results are shown in FIG. 1-b. The prepared particles have uniform and stable appearance.
The migration behavior, hysteresis loop and saturation magnetic strength of the prepared magnetic drug-loaded nanogel with time were analyzed by using a permanent magnet migration and an electromagnet Vibration Sample Magnetometer (VSM), respectively, and the results are shown in FIGS. 1-c and 1-d. The magnetic reduction enhancement type nanogel prepared by the method has good magnetism and can be rapidly enriched near a magnet. The carrier has no obvious magnetic hysteresis, has superparamagnetism, and has the saturation magnetic strength of 53.4emu/g Fe.
The drug-loaded nanogel prepared by the embodiment has a remarkable reduction response release characteristic, as shown in fig. 2. The release reached 97.5% in the pH 5.0PBS and 10mM GSH environments, with the cumulative release being the highest in all environments. And the cumulative release amount was 54.2% under the condition of PBS of pH 5.0, indicating that the prepared vector had significant reduction sensitivity. In addition, the carrier also exhibits significant pH responsiveness. Under the dual actions of pH and GSH, the release amount of the magnetic drug-loaded nanogel is remarkably improved.
The nanogel prepared in this example was subjected to biological evaluation using an experimental mouse as a model, as shown in fig. 3. Firstly, the cancer cells are implanted into the mice subcutaneously until the tumor grows to 100mm3On the left and right, mice were divided into a blank control group (PBS control group), an doxorubicin control group, and a magnetic drug-loaded nanogel group. The injection amount of adriamycin in the adriamycin control group and the magnetic drug-loaded nanogel group is 3.3mg/kg/week through tail vein injection, and is compared with that in the PBS control group without the drug. The body weight of mice treated with magnetic drug-loaded nanogels and PBS did not change significantly after 21 days of observation, while doxorubicin-treated mice lost significantly the next day (figure 3-a). In subsequent observation, compared with the PBS control group, the mice in the doxorubicin group grew more slowly, while the mice in the magnetic drug-loaded nanogel group grew less greatly than the mice in the PBS control group, and the magnetic drug-loaded nanogel group had less toxic and side effects. On the other hand, the tumor volume of the mice of the magnetic drug-loaded nanogel group is obviously increased at a lower speed than that of the mice of the adriamycin group (figure 3-b), the tumor inhibition rate of the adriamycin is 52.6 percent, and the tumor inhibition rate of the magnetic drug-loaded nanogel is 76.6 percent (figure3-c and d). The result shows that the prepared magnetic drug-loaded nano gel has better tumor inhibition effect than free adriamycin and generates less toxic and side effects.
Example 2
1. Superparamagnetic ferroferric oxide nanoparticles (SPION-NH) with surface amination were synthesized by reference (Biomaterials,2009,30:4716-4722) method2)。
2. The CS-SSe-COOH is prepared by the following specific steps:
1) the side chain of the aminated chitosan is introduced into a thiol group by a method such as that described in references (J controlled release,2009,136:38-44), (Materials science and engineering C,2015,46:41-51) to give a thiolated chitosan (CS-SH).
2) 3,3' -diselenodipropionic acid (DSeDPA) was synthesized by methods such as reference (Langmuir,2006,22(13):5552-5565), (Int J Nanomed,2012,7(17):3991-4006), and the like.
3) 0.1g of DSeDPA was weighed into a three-necked flask containing 30mL of PBS (0.01mol/L, pH6.56) and dissolved by stirring at room temperature under nitrogen. 0.215g of CS-SH was dissolved in 30mL of PBS (0.01mol of L-1, pH6.56) and injected into the DSeDPA solution under reaction conditions of 37 ℃ and stirred for 7 hours. The solution obtained by the reaction was dialyzed against ultrapure water for 3 days to obtain 2.10mg/mL of CS-SSe-COOH.
3. 2.50mL of 2.10mg/mL CS-SSe-COOH was transferred into a three-necked flask containing 2.50mL of ultrapure water, and its pH was adjusted to 5.0 with 1.0mol/L hydrochloric acid; 0.454mL of 5.508mg/mL SPION-NH was taken2And 0.625mL of 4mg/mL doxorubicin was dissolved in 3.524mL of hydrochloric acid having a pH of 5.0, and added dropwise to the three-necked flask, and finally the pH was adjusted to 7.4 with 0.1mol/L sodium hydroxide, and the mixture was stirred at room temperature for 5 hours, with light shielding over the whole period. Transferring the reaction solution into a dialysis bag, dialyzing with ultrapure water for 2 days, and storing the obtained nano-gel aqueous solution at 4 deg.C, wherein the cut-off molecular weight of the dialysis bag is 10 KDa. The drug loading rate and the encapsulation rate of the obtained magnetic nanogel are respectively 10.8 percent and 86.2 percent.
Claims (11)
1. A magnetic reduction enhanced drug sensitive release nanogel is characterized by comprising the following raw materials in percentage by mass (1-2): 1: (0.2-0.5) preparation to obtain:
the surface-aminated superparamagnetic ferroferric oxide nanoparticle, a macromolecular derivative with a side chain containing a selenium-sulfur bond and an anticancer drug, wherein the macromolecular derivative with the side chain containing the selenium-sulfur bond is prepared by the following steps:
under the protection of nitrogen at room temperature, taking a PBS solution with the concentration of 0.01mol/L as a reaction medium, and mixing the components in a mass ratio of 1-1.5: 1, taking the macromolecule with a sulfydryl functional group on the side chain and a compound with a double selenium bond as reaction raw materials, adjusting the pH value of a reaction system to be 6.5-7.4, stirring and reacting for 5-8 hours at 37 ℃, and dialyzing to obtain the macromolecule derivative with the selenium-sulfur bond on the side chain.
2. The magnetic reduction-enhanced drug-sensitive release nanogel according to claim 1, wherein the macromolecule having a thiol functional group on a side chain is one of thiolated sodium alginate, thiolated dextran, and thiolated chitosan.
3. The magnetic reduction-enhanced drug-sensitive release nanogel according to claim 2, wherein the preparation method of the thiolated sodium alginate, the thiolated dextran and the thiolated chitosan comprises the following steps:
1) dissolving sodium alginate or aminated dextran or chitosan in ultrapure water;
2) dissolving 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide in ultrapure water to obtain an EDC/NHS solution;
3) under the protection of nitrogen, dropwise adding the EDC/NHS solution prepared in the step 2) into the sodium alginate or aminated glucan or chitosan solution prepared in the step 1), and activating for 30 min;
4) dissolving cysteine hydrochloride in ultrapure water, adjusting the pH value to 6-7 by using 6mol/L sodium hydroxide solution, then dropwise adding the solution obtained in the step 3), stirring for 12 hours, dialyzing, freezing and drying to obtain solid powder.
4. The magnetically-reduced enhanced drug-sensitive release nanogel of claim 3, wherein the molar ratio of the carboxyl group of sodium alginate to the amino group of cysteine hydrochloride is 1: (1-1.2).
5. The magnetically-reduced-enhanced drug-sensitive release nanogel of claim 3, wherein the molar ratio of the amino group of the aminated dextran or chitosan to the carboxyl group of cysteine hydrochloride is 1: (1-1.2).
6. The magnetic reduction-enhanced drug-sensitive release nanogel according to claim 1, wherein the compound containing a diselenide bond is 3,3 '-diselenodipropionic acid or 4, 4' -diselenodibutyric acid.
7. The magnetic reduction-enhanced drug-sensitive release nanogel according to claim 6, wherein the 4, 4' -diselenodibutyric acid can be prepared by the following method:
1) dissolving 3.95g of selenium powder and 2.00g of sodium hydroxide in 25mL of ice water, dissolving 0.25g of sodium borohydride and 0.20g of sodium hydroxide in 5mL of ice water, injecting the solution into the selenium powder-containing solution, reacting at 0 ℃ under the protection of nitrogen in the whole process until the selenium powder is completely dissolved, heating to 90 ℃, and continuing to react for 30min until the color of the solution becomes reddish brown;
2) dissolving 4-chloropropionic acid 5.42g in ultrapure water 20mL, adjusting the pH value to 8.0 by using sodium carbonate, adding the solution into the reddish brown solution prepared in the step 1), stirring overnight under the condition of nitrogen, filtering the reaction solution after the reaction is finished, adjusting the pH value to 3-4 by using hydrochloric acid 1.0mol/L, extracting twice by using ethyl acetate, drying by using anhydrous magnesium sulfate, filtering, performing rotary evaporation, and finally recrystallizing by using ethyl acetate to obtain the 4, 4' -diseleno-dibutyrate.
8. The magnetic reduction-enhanced drug susceptible release nanogel according to claim 1, wherein the anticancer drug is one or more of doxorubicin hydrochloride, methotrexate, mitomycin, hydroxyurea, bleomycin.
9. The method for preparing a magnetically reduced enhanced drug-sensitive release nanogel according to any one of claims 1 to 8, comprising the steps of:
1) adding a macromolecular derivative with a side chain containing a selenium-sulfur bond into ultrapure water, and adjusting the pH value of the solution to 6-7.4;
2) dissolving superparamagnetic ferroferric oxide nano particles with aminated surfaces and an anticancer drug in hydrochloric acid, and controlling the pH value of the solution to be 4.5-5.5;
3) and (2) under the condition of keeping out of the sun, dropwise adding the solution obtained in the step 2) into the solution obtained in the step 1), adjusting the pH value of the solution to 7.2-7.4, stirring for 5-7h, and dialyzing to obtain the magnetic reduction enhanced type drug sensitive release nanogel.
10. The method for preparing magnetic reduction-enhanced drug-susceptible release nanogel according to claim 9, wherein the dialyzed medium in step 3) is ultrapure water, and the cut-off molecular weight of the dialysis bag is 10 KDa.
11. The method for storing the magnetic reduction-enhanced drug sensitive release nanogel according to any one of claims 1 to 8, wherein the nanogel is stored in an environment at 2 to 4 ℃.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710012359.4A CN106727284B (en) | 2017-01-09 | 2017-01-09 | Magnetic reduction enhanced drug sensitive release nanogel and preparation and storage methods thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710012359.4A CN106727284B (en) | 2017-01-09 | 2017-01-09 | Magnetic reduction enhanced drug sensitive release nanogel and preparation and storage methods thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106727284A CN106727284A (en) | 2017-05-31 |
| CN106727284B true CN106727284B (en) | 2020-06-02 |
Family
ID=58950278
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710012359.4A Expired - Fee Related CN106727284B (en) | 2017-01-09 | 2017-01-09 | Magnetic reduction enhanced drug sensitive release nanogel and preparation and storage methods thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106727284B (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112641759B (en) * | 2020-12-31 | 2022-06-10 | 中国农业科学院油料作物研究所 | Redox-enhanced drug sensitive release mesoporous silica nanoparticle and preparation method thereof |
| CN116023681A (en) * | 2023-01-31 | 2023-04-28 | 福州大学 | Preparation method and application of acoustic-thermal double-response hydrogel |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102250365A (en) * | 2011-05-09 | 2011-11-23 | 武汉理工大学 | PH-sensitive reduction responsive nanogel and preparation method thereof |
| CN102935236A (en) * | 2012-11-21 | 2013-02-20 | 武汉平华生物医药科技有限公司 | Antineoplastic prodrug with P-glycoprotein inhibition function |
| CN103505746A (en) * | 2013-06-05 | 2014-01-15 | 华中科技大学 | Contrast medium used for glioma targeted magnetic resonance and fluorescence dual modality imaging, and preparation method thereof |
| CN104606687A (en) * | 2015-01-16 | 2015-05-13 | 东华大学 | Preparation method of ferric oxide nanoparticle supported sodium alginate nanogel |
| CN104940951A (en) * | 2015-07-20 | 2015-09-30 | 武汉工程大学 | Preparation method of anti-tumor drug carrier with magnetic reduction responsiveness |
| CN105012959A (en) * | 2015-07-20 | 2015-11-04 | 武汉工程大学 | PH-responsive sodium alginate nano gel and production method thereof |
| CN105327356A (en) * | 2014-08-11 | 2016-02-17 | 南京理工大学 | Magnetic polysaccharide nanometer gel material preparation method |
-
2017
- 2017-01-09 CN CN201710012359.4A patent/CN106727284B/en not_active Expired - Fee Related
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102250365A (en) * | 2011-05-09 | 2011-11-23 | 武汉理工大学 | PH-sensitive reduction responsive nanogel and preparation method thereof |
| CN102935236A (en) * | 2012-11-21 | 2013-02-20 | 武汉平华生物医药科技有限公司 | Antineoplastic prodrug with P-glycoprotein inhibition function |
| CN103505746A (en) * | 2013-06-05 | 2014-01-15 | 华中科技大学 | Contrast medium used for glioma targeted magnetic resonance and fluorescence dual modality imaging, and preparation method thereof |
| CN105327356A (en) * | 2014-08-11 | 2016-02-17 | 南京理工大学 | Magnetic polysaccharide nanometer gel material preparation method |
| CN104606687A (en) * | 2015-01-16 | 2015-05-13 | 东华大学 | Preparation method of ferric oxide nanoparticle supported sodium alginate nanogel |
| CN104940951A (en) * | 2015-07-20 | 2015-09-30 | 武汉工程大学 | Preparation method of anti-tumor drug carrier with magnetic reduction responsiveness |
| CN105012959A (en) * | 2015-07-20 | 2015-11-04 | 武汉工程大学 | PH-responsive sodium alginate nano gel and production method thereof |
Non-Patent Citations (1)
| Title |
|---|
| 《Efficient reduction and pH co-triggered DOX-loaded magnetic nanogel carrier using disulfide crosslinking》;Juan Huang等;《Materials Science and Engineering C》;20141005;第46卷;摘要，43页2.3-2.7 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN106727284A (en) | 2017-05-31 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Wang et al. | Synthesis and modification of ZIF-8 and its application in drug delivery and tumor therapy | |
| Manatunga et al. | pH responsive controlled release of anti-cancer hydrophobic drugs from sodium alginate and hydroxyapatite bi-coated iron oxide nanoparticles | |
| Nosrati et al. | Preparation of magnetic albumin nanoparticles via a simple and one-pot desolvation and co-precipitation method for medical and pharmaceutical applications | |
| Rahimi et al. | Biocompatible magnetic tris (2-aminoethyl) amine functionalized nanocrystalline cellulose as a novel nanocarrier for anticancer drug delivery of methotrexate | |
| Ghadi et al. | Synthesis and optimization of chitosan nanoparticles: Potential applications in nanomedicine and biomedical engineering | |
| Wu et al. | Synthesis and characterization of mesoporous magnetic nanocomposites wrapped with chitosan gatekeepers for pH-sensitive controlled release of doxorubicin | |
| Javanbakht et al. | Preparation of Fe3O4@ SiO2@ Tannic acid double core-shell magnetic nanoparticles via the Ugi multicomponent reaction strategy as a pH-responsive co-delivery of doxorubicin and methotrexate | |
| Ding et al. | Chitosan-based magnetic/fluorescent nanocomposites for cell labelling and controlled drug release | |
| Chee et al. | Superparamagnetic iron oxide nanoparticles for drug delivery | |
| Sivakumar et al. | Bacterial exopolysaccharide based magnetic nanoparticles: a versatile nanotool for cancer cell imaging, targeted drug delivery and synergistic effect of drug and hyperthermia mediated cancer therapy | |
| WO2013075394A1 (en) | Drug carrier with chelating composite micell and use thereof | |
| Mukwaya et al. | Saccharide‐based nanocarriers for targeted therapeutic and diagnostic applications | |
| Wu et al. | Superparamagnetic chitosan nanocomplexes for colorectal tumor-targeted delivery of irinotecan | |
| CN106727284B (en) | Magnetic reduction enhanced drug sensitive release nanogel and preparation and storage methods thereof | |
| Thakur et al. | Nanoparticles as an emerging tool to alter the gene expression: Preparation and conjugation methods | |
| Qasim et al. | Investigation of novel superparamagnetic Ni0. 5Zn0. 5Fe2O4@ albumen nanoparticles for controlled delivery of anticancer drug | |
| ES2505890T3 (en) | Conjugates comprising nanoparticles coated with platinum-containing compounds | |
| Khoee et al. | Preparation of dual-targeted redox-responsive nanogels based on pegylated sorbitan for targeted and antitumor drug delivery | |
| Christodoulou et al. | Metal-organic frameworks for drug delivery: Degradation mechanism and in vivo fate | |
| TWI791640B (en) | Nanovectors, methods for their preparation, uses thereof, and injectable pharmaceutical solution comprising the same | |
| Cornell et al. | Green-light-responsive metal–organic frameworks for colorectal cancer treatment | |
| US20120121717A1 (en) | DRUG-LOADED POLYSACCHARIDE-COATED GOLDMAG PARTICLES (DPGPs) AND ITS SYNTHESIS METHOD | |
| CN104940951A (en) | Preparation method of anti-tumor drug carrier with magnetic reduction responsiveness | |
| Varalakshmi et al. | Nickel sulfide and dysprosium-doped nickel sulfide nanoparticles: Dysprosium-induced variation in properties, in vitro chemo-photothermal behavior, and antibacterial activity | |
| Ruman et al. | Synthesis and Characterization of Chitosan-Based Nanodelivery Systems to Enhance the Anticancer Effect of Sorafenib Drug in Hepatocellular Carcinoma and Colorectal Adenocarcinoma Cells. Nanomaterials 2021, 11, 497 |
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 | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20200602 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |