CN113209046B - CoSe @ BSA nanoparticle pharmaceutical composition and preparation method and application thereof - Google Patents
CoSe @ BSA nanoparticle pharmaceutical composition and preparation method and application thereof Download PDFInfo
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- CN113209046B CN113209046B CN202110501733.3A CN202110501733A CN113209046B CN 113209046 B CN113209046 B CN 113209046B CN 202110501733 A CN202110501733 A CN 202110501733A CN 113209046 B CN113209046 B CN 113209046B
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
- A61K9/51—Nanocapsules; Nanoparticles
- A61K9/5107—Excipients; Inactive ingredients
- A61K9/513—Organic macromolecular compounds; Dendrimers
- A61K9/5169—Proteins, e.g. albumin, gelatin
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K33/00—Medicinal preparations containing inorganic active ingredients
- A61K33/04—Sulfur, selenium or tellurium; Compounds thereof
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P39/00—General protective or antinoxious agents
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P39/00—General protective or antinoxious agents
- A61P39/06—Free radical scavengers or antioxidants
Abstract
The invention belongs to the research field of anti-radiation medicines, and particularly relates to a CoSe @ BSA nanoparticle medicine composition and a preparation method and application thereof. The CoSe @ BSA nanoparticle medicine prepared by the invention overcomes the defects of instability, easy enzymatic degradation, short half-life period in vivo, single performance, difficult metabolism in vivo, strong side effect and the like of traditional anti-radiation medicines such as chemical micromolecules, proteins and the like, can eliminate excessive ROS in vivo, promotes bone marrow to restore hematopoiesis and the like, realizes anti-radiation, and plays the anti-radiation effect to the maximum extent; the BSA-mediated active bone marrow targeting and the particle size-mediated nanoparticle drug passive bone marrow targeting are realized, so that the active targeting and the passive targeting of the bone marrow are combined; and can be degraded in vivo, and the degraded product can be metabolized and discharged out of the body, so that the biological safety is better.
Description
Technical Field
The invention belongs to the research field of anti-radiation medicines, and particularly relates to a CoSe @ BSA nanoparticle medicine composition and a preparation method and application thereof.
Background
The radiation damage caused by nuclear radiation to human body mainly comes from direct action and indirect action, wherein the radiation damage caused by indirect action accounts for about 80%. Indirect action means that nuclear radiation directly acts on water to ionize water molecules through Compton effect, Auger effect and the like to generate high Reactive Oxygen Species (ROS), such as O 2- 、H 2 O 2 、HO 2· And OH, etc. The human body contains about 80 percent of water, a large amount of ROS can be generated in the human body after the human body is subjected to nuclear radiation, and the nuclear radiation can inhibit the activities of antioxidants such as SOD, GSH-PX, CAT, MDA and the like in the human body, so that excessive ROS can be accumulated in the body. If the excessive ROS can not be timely, effectively and clearly seen, the excessive ROS can attack biological macromolecules with functions, such as nucleic acid, protein, saccharides and the like, so that cells generate oxidative stress reaction, the body is oxidized and damaged, and finally the body is subjected to radioactive damage. Thus, the elimination of excess ROS in the body is a critical step in the treatment of radiation injury. In recent years, some compounds are proved to be capable of eliminating ROS and have radiation-resistant effect, such as sulfur-containing compounds, estrogens, amine compounds, sulfydryl-containing compounds and the like, but most of the radiation-resistant compounds have the defects of large side effect, short half-life in vivo, unobvious effect and the like. Therefore, the development of low-toxicity and effective ROS-scavenging drugs is a key problem to be solved urgently for exploring radiopharmaceuticals. In addition, bone marrow is sensitive to nuclear radiation and is subject to systemic or local dosesAfter nuclear radiation irradiation, hematopoietic damage of bone marrow is caused, which is manifested by the rapid decrease of whole blood cells such as white blood cells, red blood cells, platelets and the like, and human bleeding and infection are caused, thereby influencing the survival and recovery of organisms. Thus, promotion of restoration of bone marrow hematopoiesis is another key step in the treatment of radiation injury. At present, few medicines for clinically promoting the recovery of bone marrow hematopoiesis exist, and most medicines also have the defects of weak bone marrow targeting, large dosage, large side effect, slow effect taking and the like. Therefore, the development of radiopharmaceuticals that promote the restoration of bone marrow hematopoiesis is becoming an increasingly interesting research direction. The above studies show that two key links of radiation resistance are: (1) effectively eliminating excessive ROS in the body, and (2) promoting the recovery of bone marrow hematopoiesis.
Selenium (Se) is one of trace elements necessary for maintaining normal biological metabolism of a human body and cannot be synthesized in the human body. Selenium has the function of improving the immune function of human body, so that the specific immunity and the nonspecific immunity, the humoral immunity and the cellular immunity of the human body are in a relative balance state; selenium can effectively relieve the toxic and side effects of cancer radiotherapy and chemotherapy, increase the dosage of anticancer drugs and be beneficial to the treatment of cancer; selenium is also an essential component constituting antioxidant substances such as glutathione peroxidase, superoxide dismutase and catalase. After a human body is exposed to nuclear radiation, the activity of antioxidant substances in the human body is inhibited, and the activity of the antioxidant substances in the human body can be improved by supplementing a proper amount of selenium, so that peroxidation is inhibited, peroxides are decomposed, excessive ROS in the body is eliminated, and the balance of ROS in the body after nuclear radiation is realized, thereby protecting the human body from being damaged by the excessive ROS. In addition, nuclear radiation in the human body causes increased osteoblast apoptosis and overactivity of osteoclast function, resulting in bone damage. Supplementing proper amount of cobalt ion (divalent cobalt ion, without radioactivity), can promote the expression of osteogenic factor (TNF alpha, OPG, TRAF6) and angiogenesis promoting factor (VEGFA, PDGFa, alpha-SMA), reduce the expression of IFN gamma and RANKL, reestablish dynamic balance between bone absorption and new bone formation, and realize bone reconstruction after nuclear radiation. Meanwhile, the cobalt ions can stimulate the hematopoietic system of human bone marrow, promote the synthesis of hemoglobin and increase the number of red blood cells; cobalt ions can promote high expression of HIF-1 alpha of cells, inhibit respiratory enzymes in the cells and ensure that the histiocytes are anoxic, thereby ensuring that the histiocytes are radiation-resistant; the cobalt ion can promote spleen to release red blood cells, thereby promoting hematopoiesis. And the cobalt ion is an auxiliary factor of 14 enzymes such as DNA polymerase, RNA polymerase, reverse transcriptase and the like, can maintain the normal metabolism of nucleic acid, and can interfere the replication of DNA when lacking, so that errors are generated, and certain functions of cells are degraded, the metabolism is abnormal and even the cells die. The research proves that a proper amount of selenium can effectively eliminate excessive ROS in the body, and a proper amount of cobalt ions can promote the recovery of bone marrow hematopoiesis. The excellent properties of selenium and cobalt ions are just two key links for treating radioactive injury. However, most studies tend to focus on only one of the key links. If the two key links are combined into a whole, new eosin is provided for exploring novel anti-radiation medicines. How to combine the two key links into a whole, namely integrating the excellent performances of selenium and cobalt ions into a whole is a difficult point for preparing multifunctional anti-radiation medicines. In recent years, with the rapid development of nanoscience, the above difficulties can be solved well. The research reports on the aspect of eliminating ROS of the nano material are also provided, but the reports on the application of the nano material in promoting the recovery of the hematopoietic function of the bone marrow are few, and the research reports on the research of the nano material into the anti-radiation medicine which can effectively eliminate excessive ROS and promote the recovery of the hematopoietic function of the bone marrow are less.
Disclosure of Invention
In view of the above, one of the objectives of the present invention is to provide a method for preparing a CoSe @ BSA nanoparticle drug; the second purpose of the invention is to provide a CoSe @ BSA nano-particle drug; the invention also aims to provide the application of the CoSe @ BSA nano-particle drug in radiation resistance; the third purpose of the invention is to provide a pharmaceutical preparation; the fourth purpose of the invention is to provide the application of the pharmaceutical preparation in the preparation of the anti-radiation medicine.
In order to achieve the purpose, the invention provides the following technical scheme:
1. a process for preparing CoSe @ BSA nanoparticle medicine includes such steps as dissolving BSA in water, adding ascorbic acid, reaction for 10-30min, adding Na 2 SeO 3 The reaction is carried out for 1 to 3 hours,obtaining a reaction solution I, adding ascorbic acid and CoCl into the reaction solution I 2 Reacting for 3-5h, adjusting pH to 8-11, reacting for 10-14h, dialyzing, and drying.
As one of the preferable technical schemes, BSA, ascorbic acid and Na are contained in the reaction solution I 2 SeO 3 In a mass molar ratio of 40: 5X 10 -4 ：2×10 -4 ，mg：mol：mol。
According to one preferable technical scheme, the volume ratio of the reaction liquid I to a mixed liquid is 1-3: 1-2, and the mixed liquid contains ascorbic acid and CoCl 2 In a molar ratio of 1: 1.
as one of preferable embodiments, the drying is freeze drying.
2. The CoSe @ BSA nanoparticle drug prepared by the preparation method.
3. The CoSe @ BSA nanoparticle drug is applied to radiation resistance.
4. A pharmaceutical formulation comprising a CoSe @ BSA nanoparticle drug and one or more pharmaceutically acceptable carriers or excipients.
5. The application of the medicinal preparation in preparing the anti-radiation medicament.
The invention has the beneficial effects that:
the CoSe @ BSA nanoparticle medicine prepared by the invention overcomes the defects that traditional anti-radiation medicines such as chemical micromolecules, proteins and the like are unstable, are easy to be degraded by enzyme, have short half-life in vivo, have single performance, are difficult to metabolize in vivo, have strong side effects and the like. Has multiple functions: firstly, the nano particle medicine can realize radiation resistance through various ways such as eliminating excessive ROS in a body and promoting bone marrow to restore hematopoiesis, and the like, and play the radiation resistance role to the maximum extent; secondly, the nano particle drug has BSA mediated active bone marrow targeting and is suitable for particle size mediated nano particle drug passive bone marrow targeting, so that the active targeting and the passive targeting of the bone marrow are combined; finally, the nano particle medicine can be degraded in vivo, and the degraded product can be metabolized and discharged out of the body, so that the nano particle medicine has good biological safety.
Drawings
FIG. 1 is a Transmission Electron Micrograph (TEM) of CoSe @ BSA, wherein A is a bright field image, B, C is an elemental analysis of CoSe @ BSA, and D is a superimposed image of B, C.
FIG. 2 shows that 100ul of 2mg/kg CoSe @ BSA nanoparticle drug or physiological saline with the same volume is injected into the tail vein of a male C57BL/6 mouse, the drug is continuously administered for 4 times every 7 days, and after one month, the mouse organ is taken for HE staining.
FIG. 3 is a graph of the results of intracellular excess ROS performance test after CoSe @ BSA nanoparticle drug scavenging irradiation, A, B, C, D is a peak graph of flow test of ROS content in each group of cells, and E is a quantitative graph of ROS content in each group of cells.
FIG. 4 is a graph showing the result of the test of the radiation resistance of CoSe @ BSA nanoparticle drug, A, B, C, D is a distribution graph of apoptosis of each group of flow test, and E is a quantitative graph of apoptosis of each group of cells.
Detailed Description
The following embodiments of the present invention are provided by way of specific examples, and other advantages and effects of the present invention will be readily apparent to those skilled in the art from the disclosure herein. The invention is capable of other and different embodiments and of being practiced or of being carried out in various ways, and its several details are capable of modification in various respects, all without departing from the spirit and scope of the present invention.
Example 1
Preparation and physical and chemical property analysis of CoSe @ BSA nanoparticle medicine
40mg BSA was dissolved in 10ml water and stirred for 1h, 10ml 0.05M ascorbic acid solution was added and reacted for 0.5h, 4ml 0.05M Na was added 2 SeO 3 The solution was reacted for 1h, 10ml of 0.05M ascorbic acid solution and 10ml of 0.05M CoCl were added 2 Reacting the mixed solution of the solution for 3h, adjusting the pH value to 8, reacting for 10h, dialyzing in a dialysis bag of 14000 for 2 days, and freeze-drying to obtain the CoSe @ BSA nano-particle drug. The prepared CoSe @ BSA nano-particle drug is detected by an electron microscope, and the result is shown as A-D in figure 1, the nano-particle drug is spherical, has good dispersibility in water, and has a hydrated particle size of about 80 nm.
Example 2
Evaluation of radiation resistance of CoSe @ BSA nanoparticle drug
1) Drug safety evaluation
The male C57BL/6 mouse tail vein is injected with 100ul 2mg/kg CoSe @ BSA nanoparticle drug or physiological saline with the same volume, the drug is continuously administered for 4 times every 7 days, after one month, the survival rate and the weight change of the mouse are observed, the visceral organs and peripheral blood of the mouse are respectively taken for HE staining and blood routine analysis, the result is shown in figure 2, the CoSe @ BSA nanoparticle drug has no damage to the main visceral organs of the mouse, and has good biological safety in vivo, and a good foundation is laid for the in vivo anti-radiation application of the CoSe @ BSA nanoparticle drug.
2) Ability of CoSe @ BSA nanoparticle drug to scavenge ROS in vitro
IEC-6 cells were seeded into 96-well plates in log phase, 7000 cells per well were seeded overnight. Cells were irradiated (Rad for short) and the experiments were grouped into Control, CoSe @ BSA, Rad, Rad + CoSe @ BSA groups. 50ug/ml of nanoparticle drug is given to the CoSe @ BSA group, 10Gy irradiation is given to the Rad group, 50ug/ml of nanoparticle drug is given to the Rad + CoSe @ BSA group after 10Gy irradiation, and the ROS content of each group is detected by a flow cytometer after 24 h. The results are shown in FIG. 3A-E: the ROS content of the cells of the Rad group can be increased after 24 hours of irradiation treatment, and the ROS content of the cells of the Rad + CoSe @ BSA group can be reduced to a normal value after 50ug/ml of CoSe @ BSA nano-particle medicine is given for 24 hours after irradiation treatment. The appropriate amount of CoSe @ BSA nanoparticle drug has excellent performance of eliminating excessive ROS in cells after irradiation.
3) Radiation resistance of CoSe @ BSA nanoparticle drug
IEC-6 cells were seeded into 96-well plates in log phase, 7000 cells per well were seeded overnight. Cells were irradiated (Rad for short) and the experiments were grouped into Control, CoSe @ BSA, Rad, Rad + CoSe @ BSA groups. 50ug/ml of nanoparticle drug is given to the CoSe @ BSA group, 10Gy irradiation is given to the Rad group, 50ug/ml of nanoparticle drug is given to the Rad + CoSe @ BSA group after 10Gy irradiation, and the cell survival rate of each group is tested by a flow cytometer after 24 h. The results are shown in FIG. 4A-E: the survival rate of IEC-6 cells of the Rad group after 24 hours of irradiation treatment is 76.7%, and the survival rate of IEC-6 cells of the Rad + CoSe @ BSA group after 24 hours of irradiation treatment with 50ug/ml CoSe @ BSA nanoparticle drug is 92.2%. The CoSe @ BSA nanoparticle medicament with a proper amount has good radiation resistance.
Finally, the above embodiments are only intended to illustrate the technical solutions of the present invention and not to limit the present invention, and although the present invention has been described in detail with reference to the preferred embodiments, it will be understood by those skilled in the art that modifications or equivalent substitutions may be made on the technical solutions of the present invention without departing from the spirit and scope of the technical solutions, and all of them should be covered by the claims of the present invention.
Claims (8)
1. A preparation method of CoSe @ BSA nanoparticle medicine is characterized in that the preparation method comprises the steps of dissolving BSA in water, adding ascorbic acid, reacting for 10-30min, and then adding Na 2 SeO 3 Reacting for 1-3h to obtain reaction liquid I, and adding ascorbic acid and CoCl into the reaction liquid I 2 Reacting for 3-5h, adjusting pH to 8-11, reacting for 10-14h, dialyzing, and drying.
2. The method according to claim 1, wherein BSA, ascorbic acid and Na are contained in the reaction solution I 2 SeO 3 The mass molar ratio of (a) is 40: 5X 10 -4 ：2×10 -4 ，mg：mol：mol。
3. The method according to claim 1, wherein the volume ratio of the reaction solution I to the mixed solution is 1 to 3:1 to 2, and the mixed solution contains ascorbic acid and CoCl 2 In a molar ratio of 1: 1.
4. the method of claim 1, wherein the drying is freeze-drying.
5. A CoSe @ BSA nanoparticle drug prepared by the method of any one of claims 1-4.
6. The use of a CoSe @ BSA nanoparticle drug of claim 5 in the preparation of an anti-radiation drug.
7. A pharmaceutical formulation comprising a nanoparticle medicament of CoSe @ BSA as claimed in claim 5 and one or more pharmaceutically acceptable carriers or adjuvants.
8. Use of a pharmaceutical formulation according to claim 7 for the preparation of a radiation resistant medicament.
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